A history of significant steroid discoveries and developments originating at the Schering Corporation (USA) since 1948. A firsthand historical account of some of the significant contributions of the steroid research group at the Schering Corporation (Bloomfield, NJ, USA) to the discovery and/or development of important therapeutic agents is presented. These include the discovery of the antiinflammatory corticosteroid drugs prednisone, prednisolone, and betamethasone, all of which, more than 30 years after their introduction, continue to enjoy wide use in human and animal medical practice throughout the world.


[Diagnosis of the obstructed pulmonary abscesses]. Only spot bronchography, and if its results are negative, transthoracal puncture permit a correct diagnosis of blocked or partially blocked pulmonary abscess in 2/3 of cases. This result is of importance for the choice of the treatment strategy in such patients. Use of thin or super-thin needles for puncture helped to do without serious complications and obtain reliable diagnostic information in 90% of cases.


Recovery of mononuclear cell subsets after bone marrow transplantation: overabundance of CD4+CD8+ dual-positive T cells reminiscent of ontogeny. Patients after bone marrow transplantation are immunodeficient for months to years. To understand better the pathogenesis of this immunodeficiency, we studied quantitative reconstitution of blood monocytes, natural killer (NK) cells, T cells, and B cells at 2-22 months post-transplant. The results indicate monocyte and NK counts generally recover within 2 months, followed by CD4-CD8+ T cell, B cell, and finally (after > 1 year) CD4+CD8- T cell numbers. Dual-positive CD4+CD8+ T cells (which were barely detectable in normal adults), CD4-CD8+ T cells and B cells transiently reached supranormal levels during recovery. Both CD4+CD8- and CD4-CD8+ T cells were larger than controls throughout the 2-year follow up. Comparison with neonatal and infant mononuclear cell subsets suggested the reconstitution of CD4+CD8+ T cells and B cells is similar to ontogeny. In contrast, the reconstitution of CD4+CD8- T cells did not resemble ontogeny.


Purging with 4-hydroperoxycyclophosphamide. Over the past 10 years, very-high-dose cytotoxic therapy with rescue of marrow function by autologous bone marrow transplantation (BMT) has been shown to be effective treatment for patients with lymphomas and acute leukemias. However, a major concern has been that infusion of occult tumor cells with the autologous marrow will contribute to relapse. Accordingly, multiple techniques for treating autologous marrow grafts in vitro, to remove or "purge" occult tumor, have been developed. Despite controversy regarding the necessity and efficacy of purging, most autologous transplants for acute leukemia and many for lymphoma are currently purged. Most studies have used immunologic or cytotoxic agents for purging. The most widely used cytotoxic purging agents have been the cyclophosphamide congeners, 4-hydroperoxycyclophosphamide (4-HC) and mafosfamide. Since the first 4-HC-purged autologous marrow transplant was performed in 1980, over 700 patients have received 4-HC-purged autologous bone marrow transplants at Johns Hopkins alone. This review will chronicle the development of 4-HC as a purging agent.


[Effect of A, E, C, P vitamin complex on erythrocyte hemocoagulation properties and platelet aggregation in thrombinemia]. The protective effect of A, E, C, P vitamin complex has been experimentally proven to display in a decrease of animal death rate and in limiting the intensity of hemocoagulation shifts in thrombinemia. This effect is stipulated by a decrease in thrombocyte aggregation and in coagulation activity and by the increase of erythrocyte deformation properties. These changes seem to be caused by the effect of vitamins on the phospholipid spectrum, electrolyte transport of ATPases and erythrocyte membrane stability.


Use of the Terumo SteriCell for the processing of bone marrow and peripheral blood stem cells. The SteriCell cell processing instrument is a good choice for a stem cell processing laboratory that is of sufficient size that they cannot share an apheresis machine with the blood bank. It is a laboratory instrument, with no facility for patient connection. Because of its minimal size and weight, it is easily stored in a cramped laboratory. Its automated programs are appropriate for processing of bone marrow and peripheral blood stem cells, and it is quite easy to learn how to use (in our laboratory, most individuals have been completely facile with the SteriCell after fewer than six processings). Based on reported results from other instruments, the SteriCell provides cell yields that are comparable to competing instruments. Service (provided by Haemonetics) has been satisfactory, and support from Terumo has been excellent. We can recommend this instrument to any other laboratory.


Use of the polymerase chain reaction for the detection of tumor cell involvement of bone marrow and peripheral blood: implications for purging. Bone marrow purging is being performed increasingly in an effort to deplete residual tumor cells from the graft prior to reinfusion. Several studies have suggested that the removal of tumor cells is an important clinical goal. In this review the utility of the polymerase chain reaction (PCR) for the detection of small numbers of tumor cells in bone marrow and peripheral blood is discussed. Using sensitive assays such as PCR, it is expected that the efficacy of bone marrow purging strategies will be improved and this will hopefully result in decreased relapse rates following autologous bone marrow transplantation.


Oncogenes, molecular medicine, and bone marrow transplantation. Retroviruses are known to carry specific genes that are likely to be responsible for induction of the malignant phenotype in the cells they infect. These genes, termed viral oncogenes (v-onc), have subsequently been shown to be derived from highly conserved, normal cellular genes commonly referred to as proto-oncogenes (c-onc). Proto-oncogenes are thought to be intimately involved in the processes of cell proliferation and differentiation. Therefore, any c-onc amplification, mutation, structural alteration, or change in transcriptional regulation might lead to, or be associated with, induction of a malignant phenotype. Targeted disruption of these genes may therefore be of therapeutic value. We discuss the role of antisense DNA in carrying out such therapy.


Design of large-scale separation systems for positive and negative immunomagnetic selection of cells using superparamagnetic microspheres. The ex vivo selective separation of cells from bone marrow and peripheral blood stem cell preparations is increasingly used as an adjunct to hematopoietic rescue following high-dose therapy for refractory cancer. Immunomagnetic separation, in which the target cells are identified using monoclonal antibodies and separated by attachment to paramagnetic particles and passage through a magnetic field, is widely used for both negative and positive cell selection. In this paper, we discuss the factors that should be considered when developing a magnetic separation device for purging tumor cells and selecting stem cells from bone marrow using superparamagnetic microspheres.


Multiple elements may be used for regulation of the GAP-43 gene in different cell-types. Recent evidence suggests that GAP-43 expression is not restricted to the nervous system, but may also occur outside the neural cell lineage. Two distinct patterns of GAP-43 regulation can therefore be distinguished. The first is the regulation of GAP-43 expression in multiple cell-types, and the second is the gene's temporal modulation within one specific cell-type. The latter type is well documented for neurons, where GAP-43 regulation is regulated in a fashion that is dependent on axon integrity. Results from partial analysis of the GAP-43 promoter/enhancer region indicate that at least some of these aspects of GAP-43 gene regulation may be accounted for by distinct cis-acting elements. For example, the expression of a rat GAP-43 promoter fusion gene in epidermal cells of transgenic zebrafish is dependent on an enhancer element, that is clearly distinct from the minimal neural-specific promoter. Characterization of specific GAP-43 regulatory elements responsible for particular aspects of its regulation may provide insight to signal pathways also utilized by other genes during development. Ultimately, a better understanding of the molecular events during development could help to define more precisely the complex sequences necessary for the establishment of an intact organism.


Functional domains of neuromodulin (GAP-43). Although neuromodulin (GAP-43, B50, F1, pp46, protein 4) was first identified over a decade ago, the physiological function(s) of the protein and the molecular mechanism(s) for its biological activities are still an area of active investigation. Neuromodulin has been implicated in several biological processes in neurons, including growth and regeneration, synaptic plasticity and neurotransmitter release. The molecular mechanisms underlying these implied physiological roles have not been elucidated, but there are several molecular properties of neuromodulin that may be important for its function in neurons. In this review, we will discuss research which has defined several of the functional domains of neuromodulin, including its phosphorylation sites, calmodulin binding domain, membrane binding domain and growth cone targeting domain. We will also suggest possible molecular functions of neuromodulin based on its biochemical properties.


GAP-43 expression in macroglial cells: potential functional significance. Although growth-associated protein-43 (GAP-43) was initially considered to be neuron-specific, it has more recently been found in astrocytes, oligodendrocytes, Schwann cells, glial cell equivalents in embryonic Drosophila and other non-neuronal cells. Here I summarize evidence for the presence of GAP-43 in macroglial cells (i.e., astrocytes and oligodendrocytes) cultured from neonatal rat cortex and describe its developmental expression in a lineage-specific and cell-type-specific manner. These and other data suggest that GAP-43 is a multifunctional protein involved in the synthesis of membranes associated with the growth of various types of cellular processes, including those of macroglial cells.


GAP-43 in non-neuronal cells of the embryonic chick limb: clues to function. The expression of GAP-43 in developing and regenerating neurons has been well characterized, but the function of this membrane-bound phosphoprotein is still unclear. Although GAP-43 is considered to be neuron-specific, it is also expressed in various glial cells of the peripheral and central nervous systems and in at least two populations of mesenchymal cells in the developing chick limb. GAP-43 mRNA is expressed transiently in developing limbs, which contain axons of spinal cord and dorsal root ganglion neurons, but do not contain neuronal cell bodies. This expression is correlated temporally with the in-growth of neurites and axons to the limbs, but appears to be independent of nerves. In some regions of the limb, GAP-43 immunoreactivity co-localizes in cells that are also immunoreactive for meromyosin, a muscle-specific marker. In addition, GAP-43 mRNA and protein are particularly abundant in the interdigital mesenchyme that undergoes apoptosis, or programmed cell death. GAP-43 has been postulated to mediate rapid changes in cell shape and the extension of processes in neuronal growth cones and elongating axons. We suggest here that GAP-43 may serve a similar function in glial cells, in myoblasts fusing to form myotubes, and in apoptotic and phagocytic cells of the interdigital mesenchyme.


[Laparoscope and hernia surgery].

[Laparoscope and hernia surgery]. Attempts are now progressing to improve laparoscopic hernia surgery, which need our interest and reflexion. Widely informed on these new techniques, though not directly dealing with them, the author proposes, here, his own remarks.


[Distal arterial bypass surgery for lower limb salvage]. From 1988 to 1990, 34 operations for distal arterial revascularization aimed at lower limb salvage were carried out in 29 patients with arteritis lesions at stage IV with distal necrosis (52%), severe stage III arteritis (10%), severe acute or subacute ischemia (38%). The indications and therapeutic modalities are described and discussed. The results are compared with the data found in the literature. This is an availability and emergency surgery, the last means to avoid life-saving amputation. The rate of patent revascularization after one year can be as high as 85 to 90%. Almost 9 extremities out of 10 can be saved.


[Osteosynthesis using resorbable plates in maxillofacial surgery: hopes and disappointments]. The use of absorbable osteosynthesis material has two advantages in maxillofacial surgery: suppressing the possible biological effects of implanted metallic foreign bodies, making second surgery for material removal useless. Although the biotolerance of currently used materials is well known, the authors intended to test their use in the specific conditions of maxillofacial surgery, in which simply extrapolating the data collected in general surgery leads to erroneous conclusion. The authors report the results of a preliminary clinical study on 21 cases of osteosynthesis of the upper third of the face using absorbable plates (18 fractures of the zygomatic bone, 3 maxillary osteotomies). The biotolerance of the material is very good, after more than 3 years for the oldest cases. The mechanical stability of the assembly is sufficient for the selected indications. As regards disappointments, these include the thickness of the plates, the difficulty to adapt them to bony surfaces, the fragility of the screws and the slow resorption of the material.


[Immunotherapy of severe infectious states]. Although symptomatic treatments have been optimized, mortality in severe infectious conditions still exceeds 50% in many instances. Excessive activation of inflammation mediators, the endogenous relay of the initial stimulus, may account for the secondary occurrence of polyvisceral failure. This "internal" conception of severe infectious conditions opens new therapeutic possibilities including the modulation of the host's response. However, this new treatment of severe septic conditions, which complements antibiotic therapy and symptomatic measures, encounters several difficulties: a) proinflammatory mediators also have beneficial effects; b) as the mediator cascade is activated within a few hours, ordering immunity-modulating treatments early appears to be a logical step; c) monoclonal antibodies and recombining molecules are very expensive. Costs may be further increased by the probable need to use "cocktails" of antiinflammatory molecules. All these reservations show how technically blameless future clinical trials have to be. Cost efficiency and cost/benefit studies are also required. We may wonder whether society will make the financial choice to use these new treatments.


[Gangrene of the perineum. Plea for a standardized therapeutic management apropos of 50 cases]. From 1988 to 1992, 50 cases of perineal gangrene were treated with a therapeutic protocol combining: a) repeated extensive excisions, b) hyperbaric oxygen therapy, even before surgery if this was possible, and c) intensive care. The mortality rate was 24% (12/50). It was even higher in patients admitted more than 6 hours after diagnosis. The average stay in hospital was of 20 (+/- 2) days. Four patients presented with residual signs. Twenty-eight (56%) had had colostomy for lesions originating in the rectum or threatening the anal margin; 9 of these patients died, while gastrointestinal continuity was restored in another 17 cases. There were three predictive factors of survival in this series: a) early diagnosis and treatment, b) severity index on admission, c) some associations of bacteria.


[Role of hyperbaric oxygenation in the treatment of acute anaerobic infections]. Hyperbaric oxygenation (HBO), as an adjuvant treatment in necrotizing soft tissues infections and in gas gangrene is reviewed, including its specific effects, experimental studies and clinical reports. With a global therapeutic management, considered as an emergency state, including surgery, antibiotic regimens and HBO, we can obtained 50% mortality and morbidity decrease with a more conservative surgery. Finally, favorable outcome is associated with early therapeutics and localization of the initial necrotizing infection.


[Relation between oxygen delivery and consumption during septic states. Value of an early dobutamine test]. From previous studies it has been hypothesized that multiple organ failure and high level of mortality, seen in critically ill septic patients, may be due to defective oxygen extraction and tissue hypoxia occurring early in the course of sepsis. Oxygen flux test has been proposed as a method of revealing an occult oxygen debt. We used a one hour dobutamine infusion test, in septic patients, without increase in blood lactate. Fifty patients with sepsis syndrome entered a multicentric prospective study. After fluid loading to increase pulmonary artery occlusion pressure (Paop) to a minimum value of 10 mmHg, all the patients were given 10 mcg/kg.min of dobutamine for one hour. Hemodynamic and metabolic variables were recorded before, HO, and after the test, H1 (cardiac index, Paop, oxygen deliver, DO2, and consumption, VO2, oxygen extraction ratio, (OER), blood lactate). The dobutamine test allowed to identify responders (R) who increased VO2 by more than 15% and non-responders. R and NR differed significantly in mortality (8.5% vs 44.4%). The test has a good predictive value for surviving. Without respect of the result of the test, the patients were randomized in two groups. The group D+ was given conventional therapy and dobutamine at the same rate of infusion for 9 consecutive days and the D- group received only conventional therapy. The RD+ patients improved more rapidly when compared with RD-, NRD+, NRD-. We concluded that a one hour dobutamine test is able to identify R and NR critically ill septic patients. The response is associated with significant difference in outcome.(ABSTRACT TRUNCATED AT 250 WORDS)


[Perioperative parenteral nutrition. For whom? How?]. Preoperative artificial feeding (for 7 to 15 days) through an enteral or parenteral route is justified only in a subgroup of patients suffering form severe denutrition, who are scheduled for surgery with high morbidity or mortality. Postoperatively, the field of indication is broader, and it includes any patients who cannot soon feed themselves enough to catch their resting energy expenditure plus 20%. The administration route should use the technique with the lowest morbidity (enteral feeding) and the lowest costs. The role of new substrates (structured lipids, glutamine, trace elements) on immunity should be specified.


[Post-stress acute hemorrhagic ulcers of the cecum. Apropos of 3 cases]. Three cases of acute ulcers of the cecum causing massive bleeding with hemodynamic impact are reported. These three cases involve young subjects (28, 31 and 51 years old). Bleeding occurs after an aggression (post-traumatic context in 2 cases, postoperative in 1 case). For these three patients, the angiographic location of arterial bleeding allows elective colic exeresis without rebleeding. From a pathogenetic point of view, the most likely hypothesis seems to be that of ischemic lesions following an aggression.


[Free myocutaneous flap of the latissimus dorsi in cervicofacial surgery]. Free myocutaneous latissimus dorsi transplants are exceptionally used in cervicofacial surgery. The authors have performed an anatomical study of 23 non-embalmed subjects, using injections of neoprene latex and barium sulfate into the axillary artery and dye injections. Dissections, arteriographs and corrosion show that the lower scapular pedicle is constant, with an average length of 9.5 cm and a caliber that is sufficient for vasuclar microsurgery. Eight patients with large T4 cervicocephalic neoplasms were operated, including two with tongue tumors, one with a tumor of the oropharynx, two oromandibular lesions, two lesions of the maxillary sinus and a neuroblastoma involving the middle level of the facial structures. A free myocutaneous latissimus dorsi transplant was used. The transplant was revascularised by neck vessels using microsurgical techniques. Complete success was obtained in all eight patients. Good functional, cosmetic and morphological results were obtained as a rule. In cervicofacial surgery, the authors have thus chosen and used a free myocutaneous latissimus dorsi transplant in three topographic indications: for the oropharynx, to fill large cavities, especially the maxillary sinus, and to fill the middle level of the facial structures and of the base of the skull.


[Treatment of spontaneous pneumothorax under videosurgery, 32 cases (with videofilm presentation)]. From September, 1991, to June, 1992, 32 cases of pneumothorax were operated with thoracoscopy (video surgery). The indication was established for second recurrence in 6 cases, first recurrence in 14 cases, a persistent bulla or a lung failing to return to the wall after a first pneumothorax in 5 cases, and in the presence of a large pulmonary bulla on radiographs or CT scans during an initial episode in the last 7 cases. Thoracic CT was performed in 18 cases and demonstrated a system of bullae in 14 (13 in the apical segment and 1 in the segmentum apicale). The procedure included exeresis of the bullae on endo-GIA with apical and posterolateral parietal pleurectomy. In two cases, conversion into axillary thoracotomy was required because of extensive pleural adhesion in one case and of a technical problem in the other. The average duration of surgery was 72 mn. The thoracic drains were removed on the 2nd and 3rd postoperative days. Partial pleural detachment occurred in two cases, one on the 4th day and the other on the 5th day after surgery, with spontaneous return to the wall on the 8th day in both cases. The average stay in hospital was of 6 days. All patients were examined 15 days after discharge with a control radiograph, which was normal in all cases. No patient complained of parietal pain when no conversion into thoracotomy was made.


[Role of herpes virus simplex and cytomegalovirus as cofactors of papillomavirus in dysplastic and cancerous lesions of the uterine cervix]. Human papillomaviruses (HPV) play an important role in cervical carcinogenesis but are probably not the only factors. To assess the role of HPV 6 and 11, of herpes simplex virus (HSV) and of cytomegalovirus (CMV) as cofactors for HPV 16 and HPV 18, we used polymerase chain reaction (PCR). The genomes of these six viruses was searched for in cervical biopsies, inbred in paraffin, from 22 normal cervices, 21 low grade cervical intraepithelial neoplasia (CIN), 21 high grade CIN and 20 squamous cancers. HPV 16 DNA had the highest prevalence in cervical lesions. Its frequency reached 24% in low grade CIN, 57% in high grade CIN, 50% in cancers and 9% in normal cervices. The simultaneous presence of HPV 16 and/or HPV 18 DNA and CMV and/or HSV DNA was significantly more frequent in high grade CIN and cancers than in normal cervices. On the other hand the presence of HPV 16 and/or HPV 18 DNA in samples lacking the other viral genomes did not significantly differ between the groups. Consequently the disease-viral infection relation was not apparent when these two "potentially oncogenic" HPV are isolated without coinfection by HSV or CMV. Our results are in agreement with the hypothesis of the role of herpesviruses as cofactors in cervical carcinogenesis. These viruses may disturb the control mechanisms of cellular growth and differentiation in HPV infected cells.


[Reduction of atherogenesis in an arterialized venous graft by using an external sleeve]. The parietal thickening of a vein under hemodynamical conditions in the arterial system can be reduced when supporting the vein with a rigid external sleeve. To assess the role of thickness reduction in atherogenesis of an arterialized vein graft, a 4 mm thick external sleeve was implemented around the proximal half of a carotid-carotid vein graft in 16 New Zealand rabbits. Eleven rabbits were submitted to a hypercholesterolemic diet (HC group), while five others were submitted to a normal cholesterol diet (NC group). After eight weeks, a statistically significant difference in thickness was observed between free and sleeved segments, in the NC group with 105 +/- 9 microns versus 65 +/- 6 microns (p < 10.0001) respectively as well as in the HC group with 180 +/- 37 microns versus 99 +/- 35 microns (p < 10.0001) respectively. Studying the extent of soudanophilic lesions showed a statistically significant difference according to the use or not of an external sleeve where average extent is 68 +/- 17% in free segments versus 31 +/- 26% (p < 10.0001) is sleeved segments. The reduction in vein overfullness using an external constrictive sleeve prevents structural parietal changes in the vein and allows reducing atherogenesis of the arterialized vein graft.


[Cyclosporin A metabolism and induction of cytochrome P-450 in orthoptic hepatic transplantation in rats]. The aim of our study were 1) to establish that cyclosporin (CsA) metabolism was correlated with the rate of cytochrome P4503A (cyt.) in Wistar rats induced with dexamethasone (Dex.), 2) to demonstrate that the induction of cyt. with Dex. in liver "rat donor" was transmissible to "recipient rat" after liver transplantation. Sixty rats were divided in 5 groups. In group T, a single dose of CsA (10 mg/kg) was administered intravenously in 10 rats; in group D, 10 rats were treated with Dex (300 mg/kg daily for 4 days) and then received CsA as above; in group BN 5 rats were treated with beta-naphthoflavone. Thirty five rats underwent a liver transplantation either from "non induced donors" (group G, n = 11) or from "induced donors with Dex." (group GD, n = 24) followed by CsA injection the next day. For each rat, CsA plasma levels were determined by radioimmunoassay in 6 samples. Liver microsomes cyt. from samples of the liver of donor rats (group G and GD) or after sacrifice (group T, D, BN) were quantitated by immunoblot analysis and estimated from densitometric analysis of the blot. Mean maximal plasma concentration (Cmax) were 2,822 +/- 997 ng/ml in group T, 1,447 +/- -458 ng/ml in group D, 2,685 +/- 1,383 ng/ml in group G, 1,337 +/- 713 ng/ml in group GD and 3,094 +/- 685 ng/ml in group BN. Considering the Cmax and the ASC (area under curve), there was a significant difference between all groups and separately between groups T and D, G and GD.(ABSTRACT TRUNCATED AT 250 WORDS)


[Extracorporeal bio-artificial liver using isolated hepatocytes. An experimental study in the rat]. A new type of bioartificial liver using isolated hepatocytes immobilized in alginate beads was developed and its capacity of correcting the metabolic deficiency of bilirubin conjugation in Gunn rats was assessed. Hepatocytes were isolated from Sprague-Dawley rats using the in situ collagenase perfusion technique, and they were then immobilized in Calcium alginate beads. The capacity of these immobilized hepatocytes to conjugate in vitro bilirubin was checked as compared to monolayer hepatocyte culture. The bioartificial liver consisted in a cylindrical bioreactor containing either alginate beads and hepatocytes (test group), or only alginate beads (control group). Gunn rats were connected to this bioreactor through and extracorporeal circulation system, and "bile samples were collected" every hour. Bilirubin mono and biconjugates were dosed in the bile using the high-performance liquid chromatography technique. The viability of alginate immobilized hepatocytes, determined before and after each experiment, was stable at 75%. In the test group, the total conjugate concentration rapidly increased to reach a maximum value of 204 +/- 16 microns after 3 hours, while in the control group, there were only conjugate traces (1 micron). These results show that the bioartificial liver is an efficient means to temporarily correct genetic deficiency in Gunn rats. Such a system could be of therapeutic interest in case of acute hepatic insufficiency.


[Beneficial effects of perfusion of atrial natriuretic factor in acute post-ischemic renal insufficiency in the rat]. Owing to its capacity of increasing glomerular filtration, potential beneficial effects of atrial natriuretic factor (ANF) were assessed during a post ischemic acute renal insufficiency in rats. Renal insufficiency was obtained by clamping the renal artery during 30 min., and by performing a reperfusion during 2 hours in uninephrectomized rats. Three groups were defined: a control group where animals were submitted to an operation procedure without renal artery clamping, a control group were animals received a physiological serum perfusion (1.5 ml/h) during the renal reperfusion time and an experimental group where animals were administrated a rat 1-28 ANF perfusion (5 microns/ml in NaCl 0.9%, 1.5 ml/hour) during the reperfusion time. Insulin clearance (1.0 + 0.05 ml/h vs 0.7 + 0.05 ml/h, p < 0.01), and diuresis (32.9 +/- 3.6 microliters/min. vs 7.5 +/- 0.23 microliters/min., p < 0.01) were significantly higher in rats which were administrated a NaCl 0.9% perfusion. Histologically, a significant decrease in kidney weight and in he percentage of diseases nephrons was observed after reperfusion in ANF treated rats. The results obtained demonstrate that ANF perfusion in case of post ischemic acute renal insufficiency in rats improves the recovery of renal function and reduces the renal histologic lesions.


[Experimental sclerosing cholangitis following intrabiliary injection of formol in the rat]. Our study was aimed at assessing the effect on the biliary epithelium of formol and hypertonic salt serum solutions used as parasiticide for hydatid cysts of the liver in rats. One hundred and forty six rats were administrated an intrabiliary injection of isotonic salt serum (control group), of 20% hypertonic salt serum and of 0.5% and 2% formol solutions. A histological study performed 3 months later did not show any abnormality in the rat control group (n = 11). In the hypertonic salt serum group (n = 14) and 0.5% formol group (n = 12), moderate lesions of biliary epithelium were observed. After a 2% formol injection, a periductal sclerosis was observed in 11 cases out of 16, associated with pseudocirrhosis lesions in 4 cases. A sequential histo-immunochemical study did not show any histological abnormalities in the rat control group (n = 24). In rats which were administrated a 2% formol solution (n = 36), first abnormalities were observed as soon as the 7th day with a IA antigen expression at the biliary epithelium level. Fibrosis lesions were observed after 2 months. After 9 months, the infiltration included only T lymphocytes in direct contact with biliary tracts. A cholangiographic study showed a normal aspect in the rat control group (n = 11) or in rats having received hypertonic salt serum (n = 11), and stenoses of biliary tracts, mainly intrahepatic, in 10 rats out of the 11 of the 2% formol group.(ABSTRACT TRUNCATED AT 250 WORDS)


[Rupture of the female urethra in fractures of the pelvic bones. Apropos of 2 new cases]. Female urethra ruptures associated with pelvic ring fractures are exceptionally observed. The authors compared these 2 cases (Besançon and Toulon) to already published cases. This diagnosis must be contemplated and established by the surgeon or in his/her presence using urethral catheterization. The surgical approach (most of time abdominoperineal) allows an accurate injury repair and prevents from fearsome complications due to late diagnosis and treatment. An active attitude following this urological act is highly required, in case of osteoarticular injuries, whose instability and rules are currently known.


[Surgery for lumbar disk hernia after chemonucleolysis. Results and analysis. Apropos of 100 cases]. We studied the clinical results of 100 patients operated after failure of chemonucleolytic treatment with chemopapain for lumbosciatica caused by disk herniation. Secondary surgical procedures were performed because of persistence of sciatic pain and disk herniation visible on CT scan. Post-operative follow-up was done at a minimum of one year. Clinical results were analyzed according to the criteria of Mac-Naff. Out of 100 patients who were re-operated, there were: 14% excellent results, 38% good results, 26 mediocre results, and lastly 22% failures or worsening of the condition. Thus 52% satisfactory compared to 75-95% when surgery is done immediately; 48% of the results were unsatisfactory or worse than before. We analyzed the mechanism of action of papain and suggested a pathophysiological explanation for these results. We concluded that chemonucleolysis with chemopapain cannot be an intermediary step between systematic medical treatment and surgical treatment.


[Prosthesis used in the surgery of the abdominal wall. Focus on a comparative experimental protocol: difficulties and limits]. During many years the use of prosthesis for abdominal wall repair was not accepted by all surgeons. Now the laparoscopic repair has brought some surgeons to do it nearly systematically and industry proposes new materials. To allow comparing new materials the authors have developed an experimental proceeding of which they analyse technical difficulties and limits.


[Colonic perforation during colonoscopy. 100 cases]. The analysis of 100 cases of colon perforation during colposcopic examinations highly demonstrates such a statement. The perforation risk during colposcopies is generally of the order of 0.2% for a diagnosis coloscopy. According to the statistic data used, it can reach 0.5 to 3% in therapy coloscopy. This is a risk inherent to the technique used. It is thus required to analyse the causes and take the appropriate measures to reduce it to a minimum. Mortality due to such a complication remains high (14%), i.e about 0.015 to 0.1% (#2/10000) of all colposcopies. In 11% of the patients, serious sequelae are to be observed. This demonstrates the significance of the medico-legal problem set by these perforations during colposcopies. The whole personnel responsibility can be involved: colposcopist, surgeon, anesthetist and hospital unit.


[Surgical treatment of ulcerative-necrotizing enterocolitis in premature infants. Indications and results; apropos of 50 cases]. From 1984 to 1991, 50 premature infants needed surgery for necrotizing enterocolitis. In 36 cases, surgery was necessary in emergency (weight 700 to 3,000 g, mean term: 29 weeks). Surgical treatment consisted either in intestinal resection associated with enterostomy (n = 26, or enterostomy alone (n = 10). The results were as follows: 5 early deaths, 4 late deaths (3 due to extra-digestive causes), 22 good results with nutritional recovery after closure of enterostomy. At the present time, 5 children need parenteral nutrition or are waiting for closure of enterostomy. 14 infants needed a surgical treatment for late intestinal stricture (3 to 6 weeks) by resection with enterostomy (n = 7) or with immediate anastomosis (n = 7). 13 are alive without digestive sequelae and 1 died of neurological disease. According to our experience, early surgical treatment consists in enterostomy associated whenever possible with resection of necrotic intestine. Late strictures are at best managed by resection with immediate anastomosis.


[Distal splenorenal shunt without deconnection in the prevention of recurrent digestive hemorrhage in the cirrhotic patient]. The emergent treatment of gastrointestinal hemorrhage caused by the rupture of esophageal varices in cirrhotic patients is based on sclerotherapy. The prevention of frequent recurrence may be an indication of portocaval shunting. Over an 8-year period, 72 patients were operated with a distal splenorenal shunt without deconnection aimed at preventing gastrointestinal rebleeding. This was non-emergent surgery. The Child-Pugh grade was 41 A and 31 B. All patients had an angiography, which demonstrated the lack of arterioportal reflux. Operative mortality was 2.7%. Actuarial survival at 5 years was of 67%, respectively 71% for grade A and 60% for grade B. Patency of the shunt was estimated to be 90%. Persistence of hepatopetal flow on control arteriography has been established in 65% of cases. Rebleeding was observed in 10% of cases, and episodes of encephalopathy in 10% as well out of 60 studied cases, 29 presented with a chronic increase in ammoniemia (48%). Two risk factors of mortality have been demonstrated: age higher than 60 years, and relapse of ethylic intoxication, which has been observed in 40% of cases. Later hepatic transplantation has been performed in one case, without success. These results are similar to those obtained with Warren's procedure. Further development of hepatic transplantation may limit its indications.


[Indications and role of surgery in painful dysfunction syndromes of the masticatory apparatus]. Among the numerous causes of facial neuralgias, the painful dysfunction syndrome of the manducatory apparatus corresponds to a frequent etiology whose thorough study carried out by a multidisciplinary team over about 500 cases allowed their splitting into several evolutive clinical forms. At present, a number of these forms which are particularly painful, require a disk surgery (dislocation reduction, perforation suture, etc.) and if necessary an articular facet surgery (modelling condylectomy, surfacing, etc.). This study specifies the indications and prognosis for these surgical operations.


[Proposal of environmental and biological monitoring and health surveillance of the exposed to inhalation anesthetics. Consensus. A Study Group on Occupational Exposure to Inhalation Anesthetics]. The Study Group on Occupational Exposure to Inhalation Anaesthetics of the Lombardy Association of Occupational Health and Industrial Hygiene prepared a document that was discussed during the Congress "Occupational Risks due to Inhalation Anaesthetics", held in Brescia, Italy, on May 12, 1992. The same document was then approved by the Directory Council of the Lombardy Association of Occupational Health and Industrial Hygiene. Data on environmental concentrations of Nitrous Oxide collected from 1989 to 1991 in 269 operating rooms of 47 hospitals in Lombardy are reported. The measured levels are considerably lower than those collected from the same Study Group from 1985 to 1987 in 111 operating rooms. The methodologies for exposure control are discussed, regarding both environmental and biological monitoring. These two techniques are complementary and can be used with standardized methods. The review of the literature on the early effects showed, even with some uncertainty at the current exposure levels, effects on liver and Central Nervous System. Still controversial are the data regarding the reproductive toxicity. Health surveillance programs have been organized in the last 5 years in 18 Lombardy hospitals and they indicate no cases of pathologies due to inhalation anesthetics on 1498 subjects. Operative proposals are suggested on the methodology and the frequency of environmental/biological monitoring and health surveillance. The "technical limit values" reported in the document from the Italian Ministry of Health are also discussed. Finally, research topics are suggested in order to assess the early effects of exposure.


[Measurements and environmental and biological monitoring of occupational exposure to inhalation anesthetics]. This paper reports the data of nitrous oxide (N2O) environmental pollution in 269 operating rooms of 47 hospitals in Italy in 1989-91. In 40% of the operating rooms the N2O concentrations are lower than 50 ppm, limit value proposed by Health Council for new operating rooms. In 65.4% of the operating room studied, N2O mean environmental concentrations are lower than 100 ppm, value proposed by the above-mentioned Health Council as limit value for the already existing operating rooms. Concerning the biological monitoring, the authors report several N2O data in urine (2193), whose levels confirm the data obtained with environmental monitoring. The authors believe that they presently have reliable methods to perform biological and environmental monitoring: the two techniques are complementary in the assessment of the exposure. The method of measuring N2O concentrations as exposure index, both for the environmental and biological monitoring, is considered very useful to simplify the performance of the analyses. In order to assess exposure more precisely, it is however necessary also to determine the environmental and/or biological measure of the other different anaesthetics used.


[Early effects of exposure to anesthetic gases and vapors. A critical review on the suitability of several indicators of the effect]. Apart from a risk excess of liver disease among operating theatre personnel and of spontaneous abortion in women exposed during pregnancy, no definitive conclusion can be drawn regarding health impairment among anaesthesiology staff. However, many studies pointed out that several adverse effects occur as a consequence of experimental, therapeutic and occupational exposure to inhalation anaesthetics. This paper reviews the early changes of the organ systems (hepatobiliary, renal, cardiac, hematopoietic, reproductive systems; immunologic functions; cytogenetic effects), which are considered to follow anaesthetics exposure, to evaluate their possible use as biological indices of effect. The liver microsomal enzyme system has received particular attention with the aim of clarifying the mechanisms involved in anaesthetics hepatotoxicity. An increased microsomal enzyme activity was observed in experimental conditions and in humans. This inductive effect, which reflects metabolic changes affecting liver function, is commonly considered the earliest sign caused by exposure to several chemicals (other than anaesthetics) and may be evaluated by means of biomarkers, among which the measurement of urinary D-glucaric acid excretion is a well established non-invasive tool. Urinary D-glucaric acid excretion represents the most promising early metabolic effect of the exposure to anaesthetics. However, its measurement is not yet suitable as an index of effect for use in biomonitoring practice. The main aspects to be studied in the future are the following: (i) the evaluation of urinary D-glucaric acid excretion in the acute and chronic exposure to low-dose anaesthetics to check the existence of a dose-response relationship; (ii) the study of other parameters (urinary enzymes, immunological profile, chromosome aberrations) in selected groups of exposed and control subjects, in which both exposure and confounding factors (age, gender, life style, former diseases) as well as concomitant occupational exposures should be carefully taken into account.


[Occupational exposure to volatile anesthetics and reproductive effects: a bibliographic review of epidemiological studies]. Up to the early eighties, a variety of epidemiological studies suggest that chronic exposure to low doses of anesthetic gases, as occurs in operating rooms, is an occupational risk factor for spontaneous abortion and congenital defects. Numerous and more recent epidemiologic studies are reviewed; currently it is suggested that there is inadequate evidence to conclude that occupational exposure to anesthetic gases causes increased rates of spontaneous abortion or congenital anomalies. The improvement of environmental conditions, that has reduced airborne levels of anesthetic gases in operating rooms, has been critical in reducing the risk of abortion and congenital defects. In agreement with recent epidemiological reports, we believe that the health surveillance of exposed workers must include the study of pregnancy outcome.


[Health surveillance of workers].

[Health surveillance of workers]. The main purposes of the health surveillance of workers exposed to anaesthetic gases are: a) the finding of disorders either reliable to the exposure or incompatible with the activity in the operating theatre; b) the epidemiological evaluation of the late effects. Simple and rigorous methods to estimate the previous and the current risk are suggested. The main data to be collected during the medical examination are indicated. The registration of the data of the reproductive function by a protocol of recording is proposed. For the health surveillance of workers in Health Care Institution it's advisable to establish an interdisciplinary group, referring to a specialist on occupational medicine.


[Neurobehavioral effects of inhalation anesthetics used occupationally: a critical analysis]. Unlike other apparatuses, the potential effects on the nervous system of occupational exposure to anaesthetic gases have not been exhaustively reviewed. Because of the relevance of these effects, their significance of the quality of life of the exposed subjects and of the increased risk inherent in their delicate work tasks, these themes deserve the greatest attention. This paper briefly examines the data from the international literature as well as the neurobehavioral methods employed, underlines the existing gaps of knowledge and eventually proposes strategies aimed at filling these gaps of knowledge.


[Evaluation of the risk of anesthetic gases to exposed subjects with different tasks in operating rooms]. A research was carried out on risk evaluation of workers with different tasks in operating theatre. The airborne concentrations were determined by an IR analyzer with two distinct sampling lines: the first was placed in surgical zone and the second one in anesthesiological zone. The anesthetists resulted more at risk than surgical team. Nevertheless when the evaluation was based on N2O urinary concentration and data were stratified according to task a prevalent inhalation absorption resulted for non medical staff (instrumentist and professional nurse). Our data concerning N2O ranged between 95-764 ppm indoor and between 3-92 mcg/l in the urines of exposed subjects.


[Pollution by nitrous dioxide during diagnostic laparoscopy interventions]. Here we have outlined the data relative to an environmental survey carried out in a laparoscopy clinic at the hospital of Padua, to determine the environmental concentration of nitrous dioxide (N2O) used for intra-abdominal inflation during diagnostic laparoscopy interventions. The data obtained have revealed cases of considerable N2O environmental pollution. This above all, due to spontaneous loss of gas during the intervention and to the lack of adequate ventilation systems.


[Cooperation between the occupational health department and the technical assistance office to confront the problem of pollution by anesthetics]. In the past years the problem related to occupational exposure to anaesthetic gases in operating rooms has been dealt with in different ways by each hospital administration, and frequently with little cooperation between the health departments involved. At the Spedali Civili of Brescia in the five year period, 1987-1991, a monitoring activity of nitrous oxide (N2O) and halogenated anaesthetics was performed in 30 operating rooms. This lead to a noticeable reduction of environmental concentrations of these substances. Favourable results have been achieved principally by maintenance and/or substitution of anesthesiological machine parts. In order to organize and facilitate this program a close collaboration was accomplished between the Medical Direction of the Hospital and the Occupational Health Department, with the support of the Technical Assistance Department.


[Neurobehavioral effects of exposure to anesthetic gases]. Neurobehavioral studies on operating room personnel exposed to anaesthetic gases in hospitals of the Region Lombardia have confirmed findings already reported in the literature. There was high subjective symptomatology and reduced efficiency at psychologic testing among exposed people with protracted exposure. In this group, reaction time was worse at the end of the workshift with respect to the beginning. These changes were reversible after removal from exposure. It was impossible to establish a clear dose/response relationship due to the difficulty to control an important confounding factor, such as stress, in field studies.


[Anesthetic gases and neuropsychological tests]. The Authors observed, in a group of 15 subjects working in operating theatres and occupationally exposed to anesthetic gases, an increase of simple reaction average times and an increase of the latency of the wave V of the brainstem auditory evoked potentials. The small number of subjects doesn't allow to draw conclusions. Nevertheless the findings agree with first analogous studies. The Authors, therefore, think opportune to continue the research to acquire further evidence and to ascertain the validity of the current TLV.


[Role of inhalation anesthetics in current anesthesiological practice]. Inhalational anesthetic agents represented for long time the sole or mainly anesthetic technique in general anesthesia. In the last twenty years, however, their use has been also related to toxicity among operating room personnel, chronically exposed to the volatile agents. This toxicity could induce the anesthesiologist to give up the volatile anesthetic agents in favour of a total intravenous anesthetic technique. Nevertheless well constructed prospective studies have clearly demonstrated that there is no significant correlation between morbidity index and chronic exposure to inhalational anesthetics. Furthermore a valid air exchange in the operating room, as well as more appropriate anesthesiologic procedures, have significantly reduced the concentration of volatile anesthetic agents in the operating areas. Nowadays inhalational anesthetic agents represent one of the different choices to perform general anesthesia. The anesthesiologist, however, according to patient's physical status and surgical procedure, must choose the anesthetic technique compatible with minimal risk both for patient and operating room personnel.


[Evaluation of several neuropsychological parameters in subjects occupationally exposed to anesthetics]. 51 workers, occupationally exposed to anaesthetic gases and vapours (nitrous oxide, halothane, and isoflurane), were studied monitoring their environmental and biological exposure. Moreover, they were tested for visual reaction times and neurobehavioural batteries. There was no evidence of important neurotoxic effects nor of neurobehavioural problems with low concentrations of anaesthetics.


[Risk of infectious hepatitis and risk of anesthetic gases in operating rooms: considerations on 3 cases of chronic hepatic pathology]. Three subject with professional activity in operating theatre and with a diagnosis of chronic viral "C" hepatitis are considered. All cases (Two surgeons and an anesthetist) had been continuously exposed to low level of airborne anesthetic compounds (Nitrous oxide and halogenated compounds). According to possible, but not proved, synergic effects, two cases with a clinical picture of active chronic hepatitis were classified as unqualified to specific work. The third case, affected by a persistent chronic hepatitis, was admitted to operating activities only for four hours a week, i.e. only an operating session a week. Hepatitis B and C markers must be always monitored during the medical surveillance in exposed people of operating theatre.


Magnetic resonance imaging of degenerative diseases of the central nervous system. MRI enables a better assessment than CT of the bulk loss, i.e. atrophy, which is a characteristic feature of all the degenerative diseases of CNS, at least in their advanced phases. Moreover, in several of these disorders, proton density, balanced and T2 weighted MR images can show symmetric areas of abnormally low or high signal intensity in the deep gray nuclei or white matter. Since these signal abnormalities are not specific of degenerative diseases of the CNS, their shape and distribution have to match those of the histopathological changes characteristic of each disease, before they could represent useful ancillary signs. Combination of the above MRI findings with appropriate clinical and laboratory features will however be crucial to the diagnosis in any single case.


[Prevalence of changes in performance, subjective symptoms and occupational dermatitis before and after carrying out improvements in the surgical block of the Modena Polyclinic]. The Preventive Medicine Unit for Personnel in the USL N. 16 of Modena has conducted a study to evaluate the prevalence of subjective symptoms and professional dermatitis among nurses working in a general surgery operating theatre of the Modena "Policlinico" Hospital (36 and 41 employees were studied respectively in 1990 and 1991), before and after the installation of gas evacuators and a modification in the use of detergent and disinfectant substances. The study demonstrates that installation of gas evacuating systems, if not supported by an intervention of air-conditioning plant, does not sufficiently reduce alterations in subjective symptoms. On the other hand, the modifications in the use of detergent and disinfectants has demonstrated a favourable reduction in the prevalence of professional dermatitis (from 43% to 18%; chi2 = 4.35, p = 0.037).


[Urinary measurement of D-glucaric acid in operating room personnel in the Modena USL 16 hospitals]. The Authors describe the results of a research conducted in order to evaluate the amount of D-glucaric acid in the urine of 131 operating theatre workers exposed to anesthetic gases and in a control group of 25 non-exposed hospital personnel. The comparison of the results of the two groups points out a significant difference (in the exposed members 60.7 +/- 2.4 mumol/1 to 46.7 +/- 4.3 mumol/l [+/- s.e.] in the control group; t-test: p = 0.016). The research also considered the influence of certain variables such as age, sex, smoking, alcohol, drug use and liver disorders. None of these variables has proven to be of significant influence on the D-glucaric acid urinary concentration.


Magnetic resonance imaging in multiple sclerosis: an overview. In less than a decade, Magnetic Resonance Imaging (MRI) has become the examination of choice in patients with suspected multiple sclerosis (MS). It is the best paraclinical test in assessing dissemination in space of lesions. With serial MRI scans, even dissemination in time can be detected. Using serial Gd-DTPA-enhanced MRI scans, the evolution of lesion can be easily followed. MRI studies in MS patients have contributed to shape current ideas about the pathogenesis of the disease showing that focal breakdown of the blood-brain barrier (BBB) is an early event, if not the first, in the evolution of MS lesions. A number of asymptomatic lesions can be detected by MRI in MS patients, suggesting an ongoing disease activity independent of the clinical appearance. Thus, besides its diagnostic usefulness, MRI will represent the best tool to evaluate effectiveness of treatments in therapeutical trials in MS.


[Health surveillance of the exposed to anesthetics in the environment: a complete program of preventive medicine for health personnel]. Hospital workers often can be simultaneously exposed to several potentially hazardous (biological, chemical, and physical) agents. Consequently the environmental controls and the periodic health assessments must be overall and systematic, but not too frequent. The authors suggest the establishment of an interdisciplinary group, including: Hospital Management, Technical Staff, Occupational Physician, Occupational Health Nurses, Environmental Hygienists.


Magnetic resonance imaging in the diagnosis and follow-up of patients with multiple sclerosis. Magnetic resonance imaging (MRI) has had a major impact on the diagnosis of multiple sclerosis (MS). In addition serial MRI studies help to reveal a new aspect of measurable activity in MS. The degree of activity shown by serial MRI studies is considerably greater than the degree of activity indicated by history and physical examination. In addition, the extent of the MRI abnormalities can be measured by outlining the lesions and summing the areas of abnormality slice by slice. In the future careful, however, clinical follow-up studies must be done in order to identify the prognostic implications of these MRI data. At this time MRI evaluation techniques are considered a necessary adjunct method of assessment of disease activity for MS therapeutic trials. MRI methods are complementary to clinical methods and measure an index of extent of disease that is undetectable to clinical methods.


Cognitive and brain imaging measures of multiple sclerosis. In this review we will describe the cognitive deficiency in Multiple Sclerosis (MS) and analyze the relationship between the performance on neuropsychological tests and the anatomofunctional findings assessed by neuroimaging techniques. Memory, abstract reasoning, and visuospatial abilities impairments are correlated with lesion extension and with corpus callosum atrophy, quantified on MRI. On the other hand, in MS patients with cognitive disturbance, PET and SPET studies show metabolic alterations and perfusion deficits at the cortical level, particularly in the left hemisphere and in the frontal and temporal lobes.


Notes on the therapy of multiple sclerosis (MS). MRI, neuroimmunology and neurobiology have provided significant contributions to the rationale of the therapeutic approach in multiple sclerosis. It is clear that the ignorance of the etiologic factor has lead to a double treatment policy: the first is aimed to control the acute phase of the disease while the second strives to block the pathogenetic mechanisms underlying the disease itself and the relapses. The author reviews and discusses the latest issues on this topic based on his own clinical and laboratory experience.


[Proposal of a health education program]. Since 1987 at the Spedali Civili of Brescia the problem of anaesthetic gases pollution in operating surgery rooms was managed by means of periodical environmental monitoring and maintenance of the anaesthesiological equipments. The mean nitrous oxide concentrations were generally reduced to values below the limits indicated by the Italian Ministry of Health. On the contrary, a low participation of the operating room personnel was observed in the health surveillance program and this phenomenon is clearly evidenced by the progressive decrease of the number of medical examinations. In order to clarify the usefulness of health surveillance programs and to promote the adoption of "less pollutant" behaviour, a Health Education Program was planned by the Medical Administration, in collaboration with the Occupational Health Service and the Department of Anesthesiology and Intensive Care. This program will be performed at different times, including the administration of a questionnaire, the participation at information meetings, the distribution of brochures and the evaluation of environmental and biological exposure data before and after the education program.


The immunopathogenesis of a viral model of multiple sclerosis: Theiler's virus induced demyelination. Most studies suggest that the pathogenesis of Multiple Sclerosis (MS) is based on immune mechanisms of myelin injury. In addition, a viral infection, possibly established in the first few years of the patient's life, may be crucial in setting the stage for such immune mediated injury when acting on the appropriate genetic background. Of the several models of virus-induced demyelination described in the last two decades, Theiler's murine encephalomyelitis virus (TMEV) infection has emerged as one of the best models for MS.


White matter lesions in Creutzfeldt-Jakob disease. A short review. This short review takes in consideration the role played by the cerebral white matter in Creutzfeldt-Jakob disease (CJD) and analyzes three different hypotheses on the meaning of the involvement of the white matter only as secondary phenomenon or as primary neuropathological damage related to the causative agent(s) of the disease.


HTLV-I in neurological diseases.

HTLV-I in neurological diseases. The human lymphotropic retrovirus type I (HTLV-I) has been recently associated with neurological diseases. Antibodies against HTLV-I were found in the sera and in the CSF of patients affected by Tropic Spastic Paraparesis (TSP), diffused in tropical areas such as Caraibi, south America, Seychelles. A similar clinical pattern was found in Japan and was named Human myelopathy (HAM). The virus was isolated from mononuclear cells either of the peripheral blood, and of the CSF. Molecular studies have shown that the "neurotropic" HTLV-I is similar to that associated to T cell leukemia. In vitro studies have shown that tumoral and fetal astroglial cells are susceptible to HTLV-I entry. Actually after 7 days, cells exposed to HTLV-I showed the virus core protein p19 together with an high expression of class II antigens and a disorganization of the GFAP. Multiple sclerosis (MS) has also been associated with HTLV-I infection, on the basis of finding antibodies against HTLV-I in the sera and in the CSF of some patients. However the presence of HTLV-I genome detected by PCR analysis within mononuclear cells from peripheral blood lymphocytes of MS patients is still a controversial question. The aim of the present review is to critically analyze the role of a lymphotropic retrovirus in demyelinating diseases.


Issues in multiple sclerosis. A focused disease oriented research program. This report presents a brief overview of our own focused disease oriented research program, and rationale, to find the etiopathogenesis of multiple sclerosis (MS). Our hypothesis-driven research proposes that MS is caused by a persistent virus located at the edge of active plaques. Further, we propose that the immune system has eradicated the virus from inactive plaques or that the putative virus has become latent. In 1988 polymerase chain reaction (PCR) was reported and it has turned out to be the most sensitive and specific method to detect viral nucleic acid sequences in body fluids and tissues. This technique may be a breakthrough to test the MS viral hypothesis. Our search is further enhanced by the availability of a large collection (from MS Neurospecimen Bank) of cryopreserved MS brains and other neurological diseases suitable for PCR. An enigma is the specificity of the elevation of intrathecal IgG synthesis (rate and oligoclonal bands) that is found in over 99% of clinically definite MS cases. Our 2 dimensional electrophoresis of cerebrospinal fluid (CSF) IgG shows it to be temporally and clonally stable, evidence that intrathecal IgG synthesis is not non-specific. Intrathecal IgG synthesis is a marker of IgG synthesizing plasma cells in the central nervous system (CNS) and especially in active plaques. Another issue is the inclusion in all clinical trials of our objective quantitative examination of neurologic function (instrumented tests of functions which patients want improved) as well as CSF examination before and after the trial to determine the effect of the putative treatment on the polyphasic CNS inflammation.


Blood-brain barrier changes in multiple sclerosis. We review the salient published data suggesting blood-brain barrier (BBB) damage in multiple sclerosis (MS). We focus attention specifically on: 1) the cerebral microvessel involvement in demyelination plaques, 2) the presence of dynamic changes in BBB permeability depending on the phase of the disease course, 3) the primary role of cerebral endothelial cells in the development of the intracerebral immune response.


On the role of interleukin-2 (IL-2) in multiple sclerosis (MS). IL-2-mediated endothelial cell activation. Increased serum levels of IL-2 and sIL-2R can be demonstrated in patients with active MS, a finding that strongly supports the hypothesis of a systemic T cell activation in such patients. However, IL-2 may play a crucial role in MS immunopathology by activating endothelial cells, as suggested by the vascular leak syndrome complicating hrIL-2 therapy. Early BBB impairment and focal perivascular edema that characterize MS lesions may be the effect of an IL-2-induced cytokine cascade.


Intrathecal synthesis of proteins: from immunoglobulins to prealbumin. The assessment of proteins intrathecal synthesis (ITS) is an essential step in the CSF analysis. It can be established qualitatively by different ratios and quantitatively by empirical formulae. Schuller and Sagar's formula was proposed 10 years ago for the calculation of IgG ITS. From this calculation, the antibody specific activity of intrathecal immunoglobulins may be also evaluated. The same principle may be used for complement components and for different other CSF proteins. Two examples (concerning Fibronectin and prealbumin ITS) demonstrate the usefulness of this approach, which can be programmed by a computer.


Tumour necrosis factor-alpha synthesis by cerebrospinal-fluid-derived T cell clones from patients with multiple sclerosis. T-cell clones derived from the CSF (cerebrospinal fluid) of MS (multiple sclerosis) patients have been analysed for the production of interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2) and interleukin-4 (IL-4). Each CSF-T clone (both CD4+ and CD8+) produced substantial amounts of IFN-gamma and especially TNF-alpha compared with autologous peripheral T clones and liver-infiltrating T clones from patients with chronic active hepatitis. The large quantities of TNF produced by CSF-T cell clones suggest an important role for this cytokine in MS immunopathogenesis.


Neurological complications of Lyme borreliosis. Lyme disease, like syphilis, a spirochetal infection, can appear with exacerbations and remissions in different stages. The clinical picture is marked by dermatological, neurological, rheumatic and cardiological complications. PNS complications appear in the second and third stage. Tick bite meningoradiculoneuritis neuritis (Garin-Bujadoux-Bannwarth-Syndrome), characterized by painful asymmetrical sensory and motor dysfunctions and inflamed CSF, is a typical manifestation of the second stage. Mononeuritis multiplex appearing in conjunction with acrodermatitis chronica atrophicans is a typical PNS manifestation of the third stage. CNS involvement may also occur in early and late stages of Lyme-Borreliosis, presenting as myelitis or progressive encephalomyelitis. Lyme-Borreliosis is a treatable condition, which should not be missed in the differential diagnosis of PNS and CNS disorders.


Neurobiological aspects of glial cells: astrocyte subtypes and the 0-2A glial cell lineage. Cell culture studies have shown the existence, in the rat CNS, of two populations of astrocytes, generally named type-1 and type-2 astrocytes, differing in a number of morphological, antigenic and functional features, and belonging to two different cell lineages. Type-2 astrocytes seem to derive from a bipotential glial progenitor cell (0-2A progenitor) which gives rise also to oligodendrocytes. Several epigenetic factors influence the proliferation and the differentiation destiny of 0-2A progenitors. Among such factors, heterotypic and homotypic glial cell interactions appear to be particularly important. Interaction with factors present on the surface of type-1 astrocytes or in their extracellular matrix leads to an increased proliferation of 0-2A progenitors and facilitates their oligodendroglial differentiation, while homotypic interactions among the progenitors induce them to differentiate into oligodendrocytes through the secretion of autocrine non mitogenic factors.


Basics of the magnetic resonance phenomenon. Magnetic Resonance is increasing its importance as a diagnostic mean. For an effective and accurate MRI practice, new fundamental principles, which are at the basis of this phenomenon, must be acquired. In this chapter an extremely simplified overview of the physical basics of MRI is introduced, to help the readers understand the principles underlying this complex technique. Basic physics, T1 and T2 relaxation times, spin echo sequences, which can be acquired with the now available units, and tissue characteristics are illustrated. For those who are interested in a further and more detailed reading in this field, a bibliography is also quoted.


An in vivo and post mortem MRI study in multiple sclerosis with pathological correlation. A young woman affected by multiple sclerosis (MS) was examined by magnetic resonance (MRI) during a relapse. Three months later the patient died from acute pulmonary embolism. An imaging and quantitative MRI study was performed on the formalin-fixed brain. Finally, the left hemisphere was examined by light microscopy after histological and immunocytochemical staining. While fixation significantly reduced T1 and T2 relaxation times, MRI signal and image contrast of the fixed brain were satisfactory. Lesion distribution was very similar in corresponding MRI slices and histological sections. The post mortem MRI scan and pathological study detected several new lesions, as expected from the patient's clinical course. Thus, it was possible to evaluate the age of lesions by comparing the MRI scans. In this study, signal intensity of MS lesions varied according to their histological features, i.e. to their age.


Chronical administration of anti-DNA antibodies by subcutaneous injection of hybridoma clones in BALB/c and NZBxNZW/F1 mice. Experimental model of the pathogenic role of DNA in acute lupus disease. Several sets of data suggest that specific classes of anti-DNA antibodies could be implicated in the genesis of glomerular lesions in SLE. The goal of this work is to investigate if this pathogenic role could be related to the antibodies' genetic origin--from BALB/c or NZBxNZW/F1 mice--or to their physiological origin--induced either by DNA or by polyclonal B cell activation in normal mice. For this purpose, anti-DNA antibody hybridoma clones produced from different origins were subcutaneously injected in BALB/c or NZBxNZW/F1 female mice, followed by studies of immunological parameters and kidney lesions. Results concur that the induced anti-DNA antibodies can play a role in fatal disease development, related to clonal specificity but not to the way of stimulation which was either polyclonal B cell activation or DNA immunization. Also, they emphasize the possible very lethal role of serum circulating DNA.


Reciprocal changes in serum levels of immunoglobulins (IgG, IgA, IgM) and complements (C3, C4) in normal pregnancy and after delivery. Sequential changes in serum levels of IgG, IgA, and IgM, and C3 and C4 during and after pregnancy were studied in 8 healthy women. Serum IgG decreased gradually during pregnancy, but increased markedly during the six months following delivery. Serum IgA and IgM levels also showed patterns similar to IgG. In contrast, C3 and C4 levels increased significantly and reached maximum levels in the last trimester during pregnancy, but decreased gradually for six months after delivery. Reciprocal changes between immunoglobulins and complements were clarified for the first time, and were suggestive of a compensatory autoregulatory mechanism in the suppression of the humoral immune system during pregnancy.


Medical reports on persons claiming compensation for personal injury. An audit of one insurance company's files on all employer's liability and third party motor claims settled over two years for 5000 pounds or more presented an opportunity to review the medical reports on the patients involved. A stratified random sample of files on 203 patients contained 602 reports prepared by 400 consultants. Content analysis was undertaken to evaluate compliance with published guidance on reports prepared for medico-legal purposes and to ascertain how well reports met recipients' requirements. While clinical topics were well covered, generally to a high standard, other functional, psychosocial and occupational topics, reflecting the wider clinical and non-clinical frame of reference within which lawyers and insurers normally seek information and advice, were covered less frequently, extensively and comprehensively--leaving considerable scope to improve these aspects of assessment and reporting. Further review of this aspect of professional practice should include attention to the appropriateness of existing guidance, postgraduate training requirements and the involvement of other agencies or professions in some aspects of assessment for medico-legal purposes.


Natural history of insect sting allergy: relationship of severity of symptoms of initial sting anaphylaxis to re-sting reactions. To examine the postulate that the nature of the symptoms of initial insect sting anaphylaxis is related to the risk and severity of subsequent sting reactions, the results of field re-stings were analyzed in 220 patients who had had venom anaphylaxis and did not receive venom immunotherapy. The incidence of a reaction after the first re-sting was 56% in the total group, was more frequent in adults (74%) than in children (40%), and was unrelated to the time interval since the initial sting reaction. When re-sting reactions did occur, the nature of the symptoms was similar to the symptoms of the initial sting reaction. Reactions to repeated re-stings tended to be similar. Overall, more severe reactions to re-stings occurred eventually in 24 patients. These observations confirm the frequent self-limiting course of insect sting allergy, especially in children, and the repetitive nature of specific anaphylactic symptoms, and the observations thus suggest that patients with mild to moderate anaphylactic symptoms probably do not require venom immunotherapy.


A comparison of angioplasty with medical therapy in the treatment of single-vessel coronary artery disease. Veterans Affairs ACME Investigators. Despite the widespread use of percutaneous transluminal coronary angioplasty (PTCA), only a few prospective trials have assessed its efficacy. We compared the effects of PTCA with those of medical therapy on angina and exercise tolerance in patients with stable single-vessel coronary artery disease. Patients with 70 to 99 percent stenosis of one epicardial coronary artery and with exercise-induced myocardial ischemia were randomly assigned either to undergo PTCA or to receive medical therapy and were evaluated monthly. The patients assigned to PTCA were urged to have repeat angioplasty if their symptoms suggested restenosis. After six months, all the patients had repeat exercise testing and coronary angiography. A total of 107 patients were randomly assigned to medical therapy and 105 to PTCA. PTCA was clinically successful in 80 of the 100 patients who actually had the procedure, with an initial reduction in mean percent stenosis from 76 to 36 percent. Two patients in the PTCA group required emergency coronary-artery bypass surgery. By six months after the procedure, 16 patients had had repeat PTCA. Myocardial infarction occurred in five patients assigned to PTCA and in three patients assigned to medical therapy. At six months 64 percent of the patients in the PTCA group (61 of 96) were free of angina, as compared with 46 percent of the medically treated patients (47 of 102; P less than 0.01). The patients in the PTCA group were able to increase their total duration of exercise more than the medical patients (2.1 vs. 0.5 minutes, P less than 0.0001) and were able to exercise longer without angina on treadmill testing (P less than 0.01). For patients with single-vessel coronary artery disease, PTCA offers earlier and more complete relief of angina than medical therapy and is associated with better performance on the exercise test. However, PTCA initially costs more than medical treatment and is associated with a higher frequency of complications.


Combination therapy with zidovudine and dideoxycytidine in patients with advanced human immunodeficiency virus infection. A phase I/II study. To evaluate the safety and immunologic and antiviral effects of combination therapy with zidovudine and dideoxycytidine (ddC) in patients with advanced human immunodeficiency virus type 1 (HIV) infection. A phase I/II open-label, dose-ranging study. Two AIDS Clinical Trials Group units. Patients (56) with advanced HIV disease. Patients were randomly assigned to one of three paired regimens of zidovudine and ddC. We evaluated six dosing regimens, each involving oral administration of the study drugs at 8-hour intervals. Pharmacokinetics, toxicity, CD4 counts, p24 antigenemia and clinical end points. The median follow-up period was 40.6 weeks (range, 0.3 to 70 weeks). Neither drug affected the pharmacokinetic profile of the other. Episodes of serious hematologic toxicity were infrequent, occurring in only 17.9% of patients, and did not differ among the regimens (P = 0.15). Severe sensory peripheral neuropathy occurred in two patients (one patient each in regimens 1 and 4). One patient receiving regimen 4 died. The mean maximal increase in CD4 counts exceeded 109 cells/mm3, and 69% of patients receiving combinations containing 300 or 600 mg of zidovudine daily had an increase in CD4 counts of 50 cells/mm3 or greater. Regimens containing 600 mg of zidovudine daily (regimens 2 and 5) were also more likely to result in persistent increases in CD4 counts above pretreatment values than were the two lowest dose regimens (P = 0.003). The decline in CD4 counts was more rapid, and the suppression of the p24 antigenemia was less rapid and less sustained in patients receiving the lowest zidovudine dose alone (regimen 6). The addition of ddC to regimen 6 (regimen 3) resulted in a slower decline in the CD4 counts (P = 0.06). Combination therapy with zidovudine and ddC at the doses tested was well tolerated and did not result in toxicity. A daily oral dose of 150 mg of zidovudine appeared to produce a suboptimal effect on p24 antigenemia and CD4 counts. Combination therapy with ddC and higher doses of zidovudine produced greater and more persistent effects in patients with advanced HIV infection compared with other study regimens and with the results of previous trials of zidovudine monotherapy.


Administration of excitatory amino acid antagonists via microdialysis attenuates the increase in glucose utilization seen following concussive brain injury. Immediately following concussive brain injury, cells exhibit an increase of energy demand represented by the activation of glucose utilization. We have proposed that this trauma-induced hypermetabolism reflects the effort of cells to restore normal ionic balance disrupted by massive ionic fluxes through transmitter-gated ion channels. In the present study, changes in local CMRglc following fluid-percussion concussive injury were determined using [14C]2-deoxy-D-glucose autoradiography, and the effects of in situ administration (via microdialysis) of excitatory amino acid (EAA) antagonists [kynurenic acid (KYN), 2-amino-5-phosphonovaleric acid (APV; 100 microM, 1 mM, and 10 mM), and 6-cyano-7-nitroquinoxaline-2,3-dine (CNQX; 300 microM, 1 mM, and 10 mM] on glucose utilization were investigated. Animals that did not receive dialysis showed a remarkable increase (up to 181% of normal control) in cortical glucose utilization following injury. In contrast, this high demand for glucose was reduced in areas infiltrated with KYN, APV, and CNQX. These results indicate that EAA-activated ion channels are involved in the posttraumatic increase in glucose utilization, reflecting the energy demand of cells required to drive pumping mechanisms against an ionic perturbation seen immediately following the concussive injury. The effects of KYN, APV, and CNQX suggest that although all subtypes of the glutamate receptor appear to be involved in this phenomenon, N-methyl-D-aspartate-activated channels may play a major role.


Postischemic blockade of AMPA but not NMDA receptors mitigates neuronal damage in the rat brain following transient severe cerebral ischemia. Glutamatergic transmission is an important factor in the development of neuronal death following transient cerebral ischemia. In this investigation the effects of N-methyl-D-aspartate (NMDA) and non-NMDA receptor antagonists on neuronal damage were studied in rats exposed to 10 min of transient cerebral ischemia induced by bilateral common carotid occlusion combined with hypotension. The animals were treated with a blocker of the ionotropic quisqualate or alpha-amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) receptor, 2.3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)quinoxaline (NBQX), given postischemia as an intraperitoneal bolus dose of 30 mg kg-1 followed by an intravenous infusion of 75 micrograms min-1 for 6 h, or with the noncompetitive NMDA receptor blocker dizocilpine (MK-801) given 1 mg kg-1 i.p. at recirculation and 3 h postischemia, or with the competitive NMDA receptor antagonist DL-(E)-2-amino-4-methyl-5-phosphono-3-pentenoic acid (CGP 40116), 5 mg kg-1, given intraperitoneally at recirculation. Treatment with NBQX provided a significant reduction of neuronal damage in the hippocampal CA1 area by 44-69%, with the largest relative decrease in the temporal part of the hippocampus. In neocortex a significant decrease in the number of necrotic neurons was also noted. No protection could be seen following postischemic treatment with dizocilpine or CGP 40116. Our data demonstrate that AMPA but not NMDA receptor antagonists decrease neuronal damage following transient severe cerebral ischemia in the rat and that the protection by NBQX may be dependent on the severity of the ischemic insult. We propose that the AMPA receptor-mediated neurotoxicity could be due to ischemia-induced changes in the control mechanisms of AMPA receptor-coupled processes or to changes of AMPA receptor characteristics.


Reduction in focal cerebral ischemia by agents acting at imidazole receptors. Treatment with the alpha 2-adrenergic antagonist idazoxan (IDA) can provide protection from global cerebral ischemia. However, IDA also recognizes another class of receptors, termed imidazole (IM) receptors, which differ from alpha 2-adrenergic receptors and are responsible for the hypotensive actions of some centrally acting agents such as the oxazole rilmenidine (RIL). We therefore sought to determine whether RIL, an agent highly selective for IM receptors, offered protection from focal cerebral ischemia elicited in rat by ligation of the middle cerebral artery (MCA). We compared the effects of RIL with the effects of IDA and the selective non-IM alpha 2-antagonist SKF 86466 (SKF). In addition, we examined whether the neuroprotective effects of RIL and IDA could be attributed to changes in local CBF (LCBF). The MCA was occluded and animals either received immediate administration of drug while arterial pressure was maintained for 1 h or had local CBF increased to 200% of control for 1 h by hypercapnia or hypertension. RIL elicited a significant dose-dependent preservation of tissue to 33% of control at optimal dose (0.75 mg/kg). IDA (3 mg/kg) significantly reduced the size of ischemic infarction by 22%. In contrast, SKF (15 mg/kg) as well as doubling of LCBF did not preserve ischemic tissue. We conclude that both RIL and IDA can reduce focal ischemic infarction but that the mechanism does not appear secondary to antagonism of alpha 2-adrenergic receptors or elevation of LCBF. Occupation of IM receptors, either in the ischemic zone or at remote brain sites, may be responsible for neuroprotection of RIL and IDA.


Dihydrolipoate reduces neuronal injury after cerebral ischemia. It has been shown in vitro that dihydrolipoate (DL-6,8-dithioloctanoic acid) has antioxidant activity against microsomal lipid peroxidation. We tested dihydrolipoate for its neuroprotective activity using models of hypoxic and excitotoxic neuronal damage in vitro and rodent models of cerebral ischemia in vivo. In vitro, neuronal damage was induced in primary neuronal cultures derived form 7-day-old chick embryo telencephalon by adding either 1 mM cyanide or 1 mM glutamate to the cultures. Cyanide-exposed and dihydrolipoate-treated (10(-9)-10(-7) M) cultures showed an increased protein and ATP content compared with controls. The glutamate-exposed cultures treated with dihydrolipoate (10(-7)-10(-5) M) showed a decreased number of damaged neurons. In vivo, dihydrolipoate treatment (50 and 100 mg/kg) reduced brain infarction after permanent middle cerebral artery occlusion in mice and rats. Dihydrolipoate treatment (50 and 100 mg/kg) could not ameliorate neuronal damage in the rat hippocampus or cortex caused by 10 min of forebrain ischemia. A comparable neuroprotection was obtained by using dimethylthiourea, both in vitro (10(-7) and 10(-6) M) and at a dose of 750 mg/kg in the focal ischemia models. Lipoate, the oxidized form of dihydrolipoate, failed to reduce neuronal injury in any model tested. We conclude that dihydrolipoate, similarly to dimethylthiourea, is able to protect neurons against ischemic damage by diminishing the accumulation of reactive oxygen species within the cerebral tissue.


Metabolic disorders of the brain in chronic hepatic encephalopathy detected with H-1 MR spectroscopy. Proton magnetic resonance (MR) spectroscopy of the brain was performed in 11 patients with chronic hepatic encephalopathy (CHE), and the results were compared with those of patients with liver disease but without CHE; clinical control subjects with diabetes, uremia, or cortical atrophy; and healthy subjects. The technique of water-suppressed stimulated-echo hydrogen-1 MR spectroscopy for detection of cerebral glutamate, glutamine, glucose, N-acetylaspartate, choline metabolites, (phospho)creatine, and myo-inositol is described. Specific changes in the brain of CHE patients included the anticipated elevation in cerebral glutamine levels (P less than or equal to .0001), a 23% reduction in choline metabolite levels (P less than or equal to .0001), and a more than 50% reduction in cerebral myo-inositol levels (P less than or equal to .0001). In four of the 15 patients with liver disease but without clinical CHE, a significant reduction in the myo-inositol level was detected, and in two of these patients an elevation in the glutamine concentration was also observed. These findings indicate a role for image-guided H-1 MR spectroscopy in the diagnosis and monitoring of both overt and preclinical CHE.


Analysis of the PvuII restriction fragment-length polymorphism and exon structure of the estrogen receptor gene in breast cancer and peripheral blood. The presence of estrogen receptor (ER) is a well-known predictor of clinical outcome in human breast cancer. We examined the ER gene in 26 primary breast cancers (14 ER-positive, 12 ER-negative) to determine if alterations of the gene are associated with the ER-negative status. In tumor biopsies and peripheral blood DNA obtained from the same patients we analyzed the ER exon structure using polymerase chain reaction amplification, restriction endonuclease digestion, and agarose gel electrophoresis. All blood and tumor samples, regardless of ER status, showed a complete set of eight exons of normal sizes, ruling out deletions or rearrangements of the ER gene in excess of +/- 20 nucleotides. Previous reports indicate that the two-allele ER PvuII polymorphism could be associated with ER expression in breast cancer (Hill et al., Cancer Res., 49: 145-148, 1989) as well as with patient age at time of tumor diagnosis (Parl et al., Breast Cancer Res. Treat., 14: 57-64, 1989). We localized the PvuII polymorphism in intron 1, 0.4 kilobase upstream of exon 2. Sequence analysis showed the polymorphism to result from a point mutation (T----C) at the fifth position of the restriction site (CATCTG). We determined the PvuII restriction fragment-length polymorphism genotype in 257 primary breast cancers and 140 peripheral blood DNA samples obtained from women without breast cancer. The results indicate that the PvuII polymorphism is not associated with ER content or patient age at tumor diagnosis.


Physiological levels of ammonia regulate glutamine synthesis from extracellular glutamate in astrocyte cultures. The effect of ammonia on glutamate accumulation and metabolism was examined in astrocyte cultures prepared from neonatal rat cortices. Intact astrocytes were incubated with 70 microM L-[14C(U)]glutamate and varying amounts of ammonium chloride. The media and cells were analyzed separately by HPLC for amino acids and labelled metabolites. Extracellular glutamate was reduced to 8 microM by 60 min. Removal of glutamate from the extracellular space was not altered by addition of ammonia. The rate of glutamine synthesis was increased from 3.6 to 9.3 nmol/mg of protein/min by addition of 100 microM ammonia, and intracellular glutamate was reduced from 262 to 86 nmol/mg of protein after 30 min. The metabolism of accumulated glutamate was matched nearly perfectly by the synthesis of glutamine, and both processes were proportional to the amount of added ammonia. The transamination and deamination products of glutamate were minor metabolites that either decreased or remained unchanged with increasing ammonia. Thus, ammonia addition stimulates the conversion of glutamate to glutamine in intact astrocyte cultures. At physiological concentrations of ammonia, glutamine synthesis appears to be limited by the rate of glutamate accumulation and the activity of competing reactions and not by the activity of glutamine synthetase.


Presynaptic glutamate receptors regulate noradrenaline release from isolated nerve terminals. The wide-ranging neuronal actions of excitatory amino acids, such as glutamate, are thought to be mediated mainly by postsynaptic N-methyl-D-aspartate (NMDA) and non-NMDA receptors. We now report the existence of presynaptic glutamate receptors in isolated nerve terminals (synaptosomes) prepared from hippocampus, olfactory bulb, and cerebral cortex. Activation of these receptors by NMDA or non-NMDA agonists, in a concentration-dependent manner, resulted in Ca(2+)-dependent release of noradrenaline from vesicular transmitter stores. The NMDA-stimulated release was potentiated by glycine and was blocked by Mg2+ and selective NMDA antagonists. In contrast, release stimulated by selective non-NMDA agonists was blocked by 6-cyano-7-nitroquinoxaline-2,3- dione, but not by Mg2+ or NMDA antagonists. Our data suggest that the presynaptic glutamate receptors can be classified pharmacologically as both the NMDA and non-NMDA types. These receptors, localized on nerve terminals of the locus ceruleus noradrenergic neurons, may play an important role in interactions between noradrenaline and glutamate.


Transmitter-like basal and K(+)-evoked release of 3,4-dihydroxyphenylalanine from the striatum in conscious rats studied by microdialysis. Using microdialysis and HPLC, characteristics of the release of endogenous 3,4-dihydroxyphenylalanine (DOPA) from striatum in conscious rats were studied in comparison with those of 3,4-dihydroxyphenylethylamine (dopamine; DA). Purified L-aromatic amino acid decarboxylase (AADC) converted a putative peak of DOPA to DA. The retention time of DOPA differed from that of DA and major metabolites of DA and norepinephrine. The DOPA peak of dialysates comigrated with that of authentic DOPA when the pH of the HPLC buffer was modified. The ratio of the basal release of DOPA:DA was 1:2. 3-Hydroxybenzylhydrazine (NSD-1015; 100 mg/kg, i.p.), an AADC inhibitor, markedly increased the basal release of DOPA but produced no effect on DA. The basal release of DOPA was markedly decreased by alpha-methyl-p-tyrosine (200 mg/kg, i.p.), substantially tetrodotoxin (1 microM) sensitive, and Ca2+ (removal plus 12.5 mM Mg2+ addition) dependent. Fifty millimolar K+ released DOPA and this release was also Ca2+ dependent. These characteristics of the basal and evoked release of DOPA were similar to those of DA. The ratio of the evoked release of DOPA:DA was 1:3. These results indicate that DOPA is released under physiological conditions and by K(+)-induced depolarization in a manner similar to that for transmitter DA from striatum in freely moving rats.


Enhanced tyrosine hydroxylation in hippocampus of chronically stressed rats upon exposure to a novel stressor. We have used microdialysis to measure the in vivo level of tyrosine hydroxylation in hippocampus of the freely moving rat. An inhibitor of aromatic amino acid decarboxylase, NSD-1015, was administered through the dialysis probe and the resulting accumulation of 3,4-dihydroxyphenylalanine (DOPA) in extracellular fluid of hippocampus was quantified. Administration of the tyrosine hydroxylase inhibitor, alpha-methyl-p-tyrosine, decreased extracellular DOPA to undetectable level. In addition, both systemic and local application of clonidine, an alpha 2-adrenergic agonist, produced a decrease in extracellular DOPA. In response to acute tail shock, a significant increase in extracellular DOPA was observed. Thus, it appears that in vivo accumulation of DOPA after local administration of NSD-1015 provides a reliable index of hippocampal tyrosine hydroxylation. We have used this technique to investigate whether prior exposure to chronic stress alters the in vivo level of tyrosine hydroxylation in hippocampus under basal conditions as well as in response to a novel stressor. In rats previously exposed to chronic cold stress, the basal accumulation of extracellular DOPA did not differ from naive controls. Acute tail shock, however, produced a significantly greater and more prolonged elevation in extracellular DOPA of chronically stressed rats. These data suggest that enhanced biosynthetic capacity of noradrenergic terminals may be one mechanism underlying adaptation to chronic stress.


An activation of synaptosomal Na+,K(+)-ATPase by a novel dibenzoxazepine derivative (BY-1949) in the rat brain: its functional role in the neurotransmitter uptake systems. In search of factors mitigating the final outcome of ischemic and epileptic brain damage, we tested a novel dibenzoxazepine derivative (BY-1949), as the compound has been shown to be effective under these two conditions. First, using rat brain, we assessed whether or not BY-1949 affects the Na+,K(+)-ATPase activity. Although in vitro applications of either BY-1949 or its three major metabolites did not cause any apparent effects, both acute and chronic oral administrations of the compound (10 mg/kg) invariably increased the Na+,K(+)-ATPase activity in the synaptosomal plasma membranes by increasing Vmax values. Second, it was shown by this study that the drug treatment caused marked increases in the uptake of both glutamic acid and gamma-aminobutyric acid into the synaptosomes. These results suggest that the activity against ischemic/epileptic brain damage by BY-1949 is explicable, at least partly, in terms of improvement of ionic derangements across the neural membranes via Na+,K(+)-ATPase activation.


Alzheimer beta/A4-amyloid precursor protein: evidence for putative amyloidogenic fragment. Recombinant baculovirus was used to overexpress human Alzheimer beta/A4-amyloid precursor protein (APP) in Spodoptera frugiperda (Sf9) cells. Lysates of these cells were then analyzed for the presence of carboxyl-terminal fragments of APP by an immunoblotting assay using either an antibody against the APP cytoplasmic domain (rabbit anti-human 695APP645-694) or an antibody against the amino terminus of beta/A4-amyloid (rabbit anti-human 695APP586-606). Anti-human 695APP645-694 identified APP holoprotein, a 25-kDa species, and a prominent group of carboxyl-terminal fragments of 17, 16, and 14 kDa, whereas anti-human 695APP586-606 identified APP holoprotein and a single prominent low-molecular-mass protein species comigrating with the 17-kDa carboxyl-terminal fragment identified by anti-human 695APP645-694. No immunoreactive species was detected at these molecular mass positions when either antibody was used for analysis of lysates of either uninfected Sf9 cells or Sf9 cells infected with wild-type Autographa californica baculovirus. For each antibody, specific immunoreactivity was abolished by preabsorption with the corresponding peptide immunogen. The incorporation of a beta/A4-amyloid amino-terminal epitope into a 17-kDa fragment of APP suggests that, in the baculoviral overexpression system, the electrophoretic microheterogeneity of APP carboxyl-terminal fragments is due, at least in part, to alternative proteolysis of APP. If such carboxyl-terminal fragments of APP containing an intact beta/A4-amyloid domain are produced in human brain, then they may represent intermediates in the conversion of APP to deposited beta/A4-amyloid.(ABSTRACT TRUNCATED AT 250 WORDS)


Activation and multiple-site phosphorylation of tyrosine hydroxylase in perfused rat adrenal glands. Tryptic digestion of tyrosine hydroxylase (TH) isolated from rat adrenal glands labeled with 32Pi produced five phosphopeptides. Based on the correspondence of these phosphopeptides with those identified in TH from rat pheochromocytoma cells, four phosphorylation sites (Ser8, Ser19, Ser31, and Ser40) were inferred. Field stimulation of the splanchnic nerves at either 1 or 10 Hz (300 pulses) increased 32P incorporation into TH. At 10 Hz, the phosphorylation of Ser19 and Ser40 was increased, whereas at 1 Hz, Ser19, Ser31, and Ser40 phosphorylation was increased. Stimulation at either 1 or 10 Hz also increased the catalytic activity of TH, as measured in vitro (pH 7.2) at either 30 or 300 microM tetrahydrobiopterin. Nicotine (3 microM, 3 min) increased Ser19 phosphorylation, vasoactive intestinal polypeptide (10 microM, 3 min) increased Ser40 phosphorylation, and muscarine (100 microM, 3 min) increased TH phosphorylation primarily at Ser19 and Ser31. Vasoactive intestinal polypeptide, but not nicotine or muscarine, mimicked the effects of field stimulation on TH activity. Thus, the regulation of rat adrenal medullary TH phosphorylation by nerve impulses is mediated by multiple first and second messenger systems, as previously shown for catecholamine secretion. However, different sets of second messengers are involved in the two processes. The action of vasoactive intestinal polypeptide as a secretagogue involves the mobilization of intracellular calcium, whereas its effects on TH phosphorylation are mediated by cyclic AMP. This latter effect of vasoactive intestinal polypeptide and the consequent increase in Ser40 phosphorylation appear to be responsible for the rapid activation of TH by splanchnic nerve stimulation.