Hematopoietic target cells of anemogenic subgroup C versus nonanemogenic subgroup A feline leukemia virus. Feline leukemia viruses (FeLVs) belonging to interference subgroup C induce fatal anemia resembling human pure red cell aplasia (PRCA). Subgroup A FeLVs, although closely related genetically to FeLVs of subgroup C, do not induce PRCA. The determinants for PRCA induction by a molecularly cloned prototype subgroup C virus (FeLV-Sarma-C [FSC]) have been localized to the N-terminal 241 amino acids of the surface glycoprotein (SU) gp70. To investigate whether the anemogenic activity of FSC reflects a unique capacity to infect erythroid progenitor cells, we used correlative immunogold, immunofluorescence, and cytological staining to study prospectively the hemopoietic cell populations infected by either FSC or FeLV-FAIDS-61E-A (F6A), a prototype of subgroup A virus. The results demonstrated that although only FSC-infected animals developed erythrocyte aplasia, the env SU and the major core protein (p27) were expressed in a surprisingly large fraction of the lymphoid, erythroid, and myeloid lineage marrow cells in both FSC- and F6A-infected cats. Between days 8 and 17 postinoculation, gp70 and p27 were detected in 43 to 73% of erythroid, 25 to 75% of lymphoid, and 35 to 50% of myeloid lineage cells, regardless of whether the cats were infected with FSC or F6A. Thus, anemogenic subgroup C and nonanemogenic subgroup A FeLVs have similar hemopoietic cell tropism and infection kinetics, despite their divergent effects on erythroid progenitor cell function. Acute anemia induction by subgroup C FeLV, therefore, does not reflect a unique tropism for marrow erythroid cells but rather indicates a unique cytopathic effect of the SU on erythroid progenitor cells.


Identification of an African swine fever virus gene with similarity to a myeloid differentiation primary response gene and a neurovirulence-associated gene of herpes simplex virus. Here we describe an open reading frame (LMW23-NL) in the African swine fever virus genome that possesses striking similarity to a murine myeloid differentiation primary response gene (MyD116) and the neurovirulence-associated gene (ICP34.5) of herpes simplex virus. In all three proteins, a centrally located acidic region precedes a highly conserved, hydrophilic 56-amino-acid domain located at the carboxy terminus. LMW23-NL predicts a highly basic protein of 184 amino acids with an estimated molecular mass of 21.3 kDa. The similarity of LMW23-NL to genes involved in myeloid cell differentiation and viral host range suggests a role for it in African swine fever virus host range.


Partial elimination of Epstein-Barr virus plasmids from Burkitt's lymphoma cells by transfecting the BZLF1 gene. Epstein-Barr virus (EBV) nonproducer Raji cells stably maintain approximately 45 copies of the EBV genome per cell, depending on the presence of the EBV-determined nuclear antigen 1 (EBNA-1) protein. We found that transfection of the EBV BZLF1 gene causes the disappearance of EBNA proteins on Western blots (immunoblots). On the basis of these results, we attempted to eliminate EBV plasmids in Raji cells by transfecting a BZLF1 plasmid. Among 33 clones that were cotransfected with a BZLF1 plasmid and a hygromycin B resistance plasmid and selected resistant for hygromycin B, 24 clones had decreased numbers of EBV plasmids, as revealed by the decrease in the intensity of the EBV band on Southern blots compared with that of nontransfected Raji cells.


Multimerization of ICP0, a herpes simplex virus immediate-early protein. ICP0, a herpes simplex virus immediate-early gene product, is a highly phosphorylated nuclear protein that is a potent activator of virus and host genes. Using biochemical and genetic assays employing plasmids encoding mutant forms of ICP0 and a recombinant adenovirus that expresses ICP0, we mutant forms of ICP0 and a recombinant adenovirus that expresses ICP0, we provide evidence that the protein multimerizes. Some mutant forms of ICP0 were transdominant and interfered with activation of a target reporter gene or with complementation of an ICP0-minus virus.


Identification and characterization of a novel herpes simplex virus glycoprotein, gK, involved in cell fusion. Antipeptide sera were used to identify a novel glycoprotein encoded by the UL53 gene of herpes simplex virus type 1 (HSV-1). The UL53 gene product is thought to play a central role in regulating membrane fusion because mutations giving rise to the syncytial phenotype, wherein cells are extensively fused, frequently map to this gene. A single 40-kDa protein, designated gK (the ninth HSV-1 glycoprotein to be described), was detected with antipeptide sera in cells infected with both wild-type and syncytial strains of HSV-1 which were labelled with [35S]methionine and [35S]cysteine or with [3H]glucosamine, and this protein was sensitive to treatment of cells with tunicamycin. With all other HSV glycoproteins studied to date, at least two glycosylated species, often differing substantially in electrophoretic mobility, have been observed in infected cells; thus, gK is unusual in this respect. The 40-kDa gK protein was also immunoprecipitated from cells infected with a recombinant adenovirus vector carrying the UL53 gene. Two glycosylated species of 39 and 41 kDa were produced when UL53 mRNA was translated in vitro in the presence of microsomes, and these proteins differed from gK produced in infected cells not only because they possessed different electrophoretic mobilities but also because they were unable to enter gels after being heated. In addition, a 36-kDa protein was detected in extracts from cells infected with HSV-2 with use of these sera.


The cDNA of UL15, a highly conserved herpes simplex virus 1 gene, effectively replaces the two exons of the wild-type virus. The UL15 gene of herpes simplex virus 1 (HSV-1) is encoded by two or more exons in all herpesvirus genomes sequenced to date. The UL15 coding region is highly conserved, and the intron invariably encodes other genes transcribed antisense to the UL15 coding region. Previously we reported that we deleted the intron domain encoding UL16 but were unable to delete UL15 (J. D. Baines and B. Roizman, J. Virol. 65:938-944, 1991). Here we report that we replaced exon I of UL15 with an unspliced cDNA copy of UL15 in HSV-1 DNA and deleted 58% of the carboxyl-terminal sequences of the natural copy of exon II, including the polyadenylation signal. The yields of infectious virus obtained upon infection with viruses containing the cDNA copy of UL15 were similar to those of an isogenic virus with a wild-type UL15 gene. We therefore conclude that the separation of the two exons of UL15 by an intron encoding two genes is not essential for the replication of HSV, at least in cell culture.


Characterization of the deletion and rearrangement in the BamHI C region of the X50-7 Epstein-Barr virus genome, a mutant viral strain which exhibits constitutive BamHI W promoter activity. Epstein-Barr virus infection of peripheral B lymphocytes predominantly results in a latent infection, with a concomitant growth transformation of the infected cells. These cells express six nuclear antigens (EBNAs) and three membrane antigens. Transcription of all the EBNA genes is driven by one of two promoters, Cp or Wp, located near the left end of the viral genome, and the activities of these promoters are mutually exclusive. We have previously shown that Wp is exclusively used during the initial stages of B-cell immortalization, followed by a switch to Cp usage. However, several cell lines which appear to have failed to switch from Wp to Cp usage and which exhibit constitutive Wp activity have been identified. In two cases, we have shown that this failure to switch is the result of a deletion of approximately 3.5 kb, spanning Cp. In this paper, we characterize the deletion in one of these cell lines, X50-7, and demonstrate not only that the viral genome in this cell line has sustained a deletion in the region of Cp, but also that there has been a rearrangement into the BamHI C region of viral sequences from the BamHI W and Y fragments.


Adeno-associated virus sensitizes HeLa cell tumors to gamma rays. Infection with the helper virus-dependent human parvovirus adeno-associated virus (AAV) is known to interfere with cellular transformation in vitro and oncogenesis in vivo. Here we report on sensitization to gamma irradiation by AAV infection of cells in culture and of tumors established from HeLa cells grafted into immunodeficient (nude) mice: infection of HeLa cells with AAV type 2 enhanced cell killing and reduced plating efficiency after irradiation compared with uninfected cells. Similarly, HeLa cell tumors in nude mice displayed a reduced growth rate and were more sensitive to gamma irradiation when the animals were infected with AAV type 2 prior to or after tumor cell inoculation. Since no pathogenicity is known for AAV, the ability of this virus to render radiotherapy of human tumor cells more efficient may up open novel approaches in cancer treatment.


Packaging cells for avian leukosis virus-based vectors with various host ranges. Using our previously described Haydée semipackaging cell line (F. L. Cosset, C. Legras, Y. Chebloune, P. Savatier, P. Thoraval, J. L. Thomas, J. Samarut, V. M. Nigon, and G. Verdier, J. Virol. 64:1070-1078, 1990) which produces avian leukosis virus gag and pol proteins, we have constructed packaging cells with subgroups B, C, and E envelope specificities. This allows us to produce helper-free avian leukosis virus particles carrying the lacZ reporter gene and the A, B, C, or E subgroup specificities. Titers of the recombinant lacZ virus are shown to be dependent upon the type of the env subgroup and the target avian cell.


Epstein-Barr virus DNA recombines via latent origin of replication with the human genome in the lymphoblastoid cell line RGN1. We show here that in a lymphoblastoid cell line Epstein-Barr virus DNA recombines with the human genome. The genetic exchange involves the oriP region of the virus. A junction between viral and human DNA from this line has been cloned and sequenced. The results indicate that the integration of Epstein-Barr virus DNA involves a region of the human genome which contains internal short repetition. An 800-bp probe has been isolated from the human part of the junction. This probe has been used to show that the human region exists as a duplication in normal cells.


Presence of transplantable T-lymphoid cells in C57BL/6 mice infected with murine AIDS virus. The Duplan strain of murine leukemia virus induces murine AIDS in C57BL/6 mice. When spleen cells from C57BL/6 mice infected with the virus were transplanted into nude mice, subcutaneous solid tumors at the transplanted sites were formed and splenomegaly and lymphadenopathy were induced. These transplantable cells were Thy-1- CD4+ alpha-beta T-cell receptor-positive T cells and integrated with the pathogenic defective viral genome. These results indicate that neoplastic cells of T-cell lineage were induced by infecting C57BL/6 mice with murine AIDS virus.


[Study of sonographic findings of small hepatocellular carcinoma based on its pathologic findings]. Sonograms of 282 cases of hepatocellular carcinoma (HCC) less than 5 cm in size were examined. Among these, 73 cases of resected or biopsied HCCs were compared in terms of their pathologic findings and sonograms. Low echoic pattern was the more common among smaller HCCs, and low echoic periphery pattern tended to prevail with increasing size. The pathologic factors of fatty change and clear cell formation are responsible for elevating the echo level. Among HCCs less than 2 cm, the low echoic group is more differentiated than the iso-echoic group by Edmondson's classification. "Lateral acoustic shadow", "nodule in nodule", and "septum" are characteristic findings in HCC by sonography, and they correspond to the pathologic findings. However, "posterior echo enhancement" was not seen to be specific for HCC, as it was also observed with similarly frequency in hemangiomas.


Synthesis and metabolism of cysteinyl leukotrienes by the isolated pig kidney. The metabolism and synthesis of cysteinyl leukotrienes by the isolated perfused pig kidney has been investigated. Kidneys were maintained for up to six hours in a recirculating perfusion system by using an oxygenated Krebs-Henseleit buffer containing albumin and the perfluorinated oxygen carrier, FC-43. Perfusion pressure was maintained at 12-13.5 kPa, with perfusion flow rates of 150-250 ml/min resulting in a urine output of between 20-180 ml/hr. Infusion of 3H-LTC4 into the renal artery resulted in rapid and complete metabolism, with the major urinary metabolites comprising LTE4, omega-hydroxy-LTE4, omega-carboxy-LTE4 and N-acetyl-omega-hydroxy-LTE4. The capacity of the isolated kidney to synthesize cysteinyl leukotrienes was monitored by measuring urinary LTE4 excretion; there was a basal urinary excretion of LTE4 (median 43 pg/min, range 8-470 pg/min). Neither lipopolysaccharide or human recombinant tumor necrosis factor alpha had any effect on basal excretion. Treatment with the calcium ionophore A23187, however, resulted in a 38.1 +/- 9.6-fold increase in urinary LTE4 excretion. We conclude that the isolated pig kidney, in the absence of circulating cells, can synthesize cysteinyl leukotrienes in the absence of circulating cells, which can then undergo extensive oxidative metabolism.


Rapid oxygen-dependent changes in erythropoietin mRNA in perfused rat kidneys: evidence against mediation by cAMP. Erythropoietin (EPO) is mainly produced in the kidneys and is regulated by blood oxygen availability. Studies with isolated perfused kidneys have established that an oxygen-sensing system exists intrarenally but the mechanisms involved are poorly understood. Using a quantitative RNase protection assay, we have demonstrated oxygen-dependent EPO mRNA production in isolated perfused rat kidneys, with EPO mRNA levels increasing 30-fold when perfusate pO2 was reduced from 474 to 25 mm Hg. To determine if the high amplitude changes in EPO mRNA levels in response to hypoxia are mediated by cyclic AMP, four agents, which activate the cyclic AMP system in different ways, were administered to isolated kidneys perfused over a range of perfusate pO2. Salbutamol and N6-ethyl carboxamidoadenosine, which activate adenylate cyclase, dibutyryl cyclic AMP (a cyclic AMP analogue) and forskolin did not augment EPO mRNA production, and no significant differences in the regression of log (EPO mRNA) on perfusate pO2, were found between experimental groups exposed to each of these compounds and controls. We conclude that the rapid increase in EPO mRNA levels in response to hypoxia is not mediated or substantially modulated by a cyclic AMP-dependent mechanism.


Endothelin-1 receptor antagonist: effects on endothelin- and cyclosporine-treated mesangial cells. Endothelin-1 (Et) has profound effects on glomerular microcirculation and mesangial cell contraction. A parameter of mesangial cell contraction was examined by measuring myosin light chain phosphorylation (MLCP) in glomerular mesangial cells in the presence and absence of a newly developed endothelin-1 receptor antagonist (EtA). Addition of Et alone (10 nM) caused a marked increase in MLCP, which, on average, rose by 53 +/- 6% above the level in cells exposed to vehicle (P less than 0.0005). This effect was shown to continue for at least one hour; MLCP at 60 minutes was 64 +/- 12% higher than controls, (P less than 0.025), constituting a unique observation of an in vitro parameter which parallels the characteristic in vivo effect of Et. Treatment of cells with EtA virtually abolished this Et-induced increase in MLCP, which rose by only 2 +/- 3% and -1 +/- 4% for doses of EtA of 44 nM and 66 nM, respectively. Examination of the intracellular calcium concentration, [Ca2+]i, revealed that EtA almost completely abolished the transient increase in [Ca2+]i evoked by Et and also suppressed the early portions of the sustained increase in [Ca2+]i. EtA was ineffective in abolishing [Ca2+]i increase in response to arginine vasopressin. Finally, to evaluate EtA's efficacy in a pathophysiologic setting, we also studied mesangial cells exposed to cyclosporine (Cs). Exposure of mesangial cells to Cs (10(-5) M) for 60 minutes caused a significant increase in MLCP, on average, by 38 +/- 6% above control (P less than 0.0005), while cells exposed to Cs in the presence of EtA increased MLCP significantly less, by only 15 +/- 9%. These data provide further evidence for Et's long-lasting cellular actions, and demonstrate inhibitory effects of an Et receptor antagonist after direct cellular exposure to Et and also after Cs exposure, a pathophysiologic setting which likely involves Et.


Simultaneous expression of 70 kilodalton type IV collagenase and type IV collagen alpha 1 (IV) chain genes by cells of early human placenta and gestational endometrium. In this study we used in situ hybridization to investigate the expression of the genes 70 kilodalton (kd) collagenase and the alpha 1(IV) collagen chain of type IV collagen in cells of early human placenta and gestational endometrium. The aim was to study the spatial distribution of these gene expressions within a developing tissue which possesses physiologic invasive potential. The results obtained for the 70 kd type IV collagenase mRNA expression were also compared with the immunohistochemical distribution of the corresponding antigen. Expression of mRNAs for these proteins was found in cells of trophoblastic columns, stromal cells of villi and in cells of decidua and endometrial stroma. The only differences between the expressions was the lower level of signals for 70 kd type IV collagenase in fibroblastic stromal cells and endothelial cells of villi and in the pericytic cells of spiral arteries. Otherwise the results for both types of mRNA were comparable. We also studied the immunohistochemical distribution of the 70 kd type IV collagenase using specific monoclonal antibodies against the enzyme. Immunohistochemistry supported well the findings obtained by in situ hybridization. The results indicate that the genes for the 70 kd type IV collagenase and for the alpha 1(IV) collagen chain are simultaneously active in cells of placenta and gestational endometrium and the same cells which produce type IV collagen also can produce the cleaving enzyme, the 70 kd type IV collagenase. The results also show that the cytotrophoblastic cells, which during early pregnancy invade the extracellular matrix and spiral arteries of uterine wall contain significant amount of mRNA for the 70 kd type IV collagenase. This finding supports the concept that the 70 kd type IV collagenase would be important for invasion, and in the case of this study, also for the physiologic invasion of placental cytotrophoblasts.


Heat-shock protein induction in adriamycin and picornavirus-infected cardiocytes. Both chemicals, such as the chemotherapy agent adriamycin, and viruses, such as the picornaviruses coxsackievirus B3 and encephalomyocarditis virus, cause metabolic injury in myocardial cells. This injury includes depression of cellular RNA and protein synthesis and production of oxygen free radicals which are known to induce increased expression of "heat-shock" or stress proteins (hsp). These hsp can stimulate potent T lymphocyte responses that may then contribute to cardiac damage associated with adriamycin therapy and picornavirus infections. To determine whether adriamycin and the picornaviruses stimulate hsp expression, cultured neonatal myocardial cells from BALB/c Cum mice were treated with heat-shock, adriamycin and either infectious or ultraviolet irradiated (noninfectious) CVB3. The treated myocardial cell homogenates were subjected to polyacrylamide gel electrophoresis and Western blot analysis for 70 kilodalton hsp. To determine whether these treatments stimulated T lymphocyte responses (presumably to hsp), BALB/c Cum mice were injected with 0.1 ml complete Freund's adjuvant containing 0.1 mg of heat-killed mycobacterium tuberculosis, 10 mg/kg adriamycin or 5 x 10(4) plaque-forming units CVB3 intraperitoneally. Splenic lymphocytes obtained 7 days later from these animals were evaluated in a 51Cr release cytotoxicity assay to cultured myocytes treated with heat-shock, adriamycin or virus. Cytolytic T lymphocytes (CTL) were characterized as to T cell subset and T cell receptor (TcR) utilization treating CTL populations with anti-CD4 or anti-CD8 antibodies or with anti-alpha/beta TcR or anti-gamma/delta TcR antibodies. Both adriamycin and infectious virus treatment of myocardial cells stimulated increased hsp expression. Ultraviolet irradiation of the virus prevents virus replication and failed to elicit hsp production in heart cells. Two types of CTL were detected. Animals injected with complete Freund's adjuvant virus, and adriamycin produced CD8+, gamma/delta TcR+ CTL that were not major histocompatibility complex antigen restricted since both CBA (H-2k) and BALB/c (H-2d) myocardial cells were lysed. Virus-specific CTL, belonging to the CD4+, alpha/beta TcR+ population, were detected in CVB3-infected mice. Different agents that metabolically injure myocardial cells can induce increased expression of one or more hsp. CTL presumably directed to these hsp can cross-reactively lyse targets treated with any of the hsp inducing agents. This observation raises the question whether multiple exposures of individuals to dramatically different hsp-inducing agents might result in increasingly damaging immune responses.


Sino-bronchial reflex, asthma and sinusitis. Sinusitis should be suspected in cases of chronic, difficult to control asthma or other pulmonary diseases. Appropriate measures to diagnose and treat sinus disease are listed, and an upright Waters roentgenogram may be all that is required for diagnosis. A true sino-bronchial reflex is proposed in these patients. Nasal receptors and reflexes are effective in the physiology of the nose, and in many cases, the diagnosis and treatment of rhinitis and sinusitis results in the improvement of various chronic pulmonary conditions.


The use of intracellular dialysis to study signal transduction coupling in the squid giant axon. The squid giant axon has proved a useful model in the study of ionic channel gating, intracellular homeostasis and receptor-mediated signal transduction leading to generation of intracellular second messengers. In the latter category, previous studies on activation of adenylate or guanylate cyclase have used intact and intracellularly perfused axons to investigate the effects of extra- and intracellular agents on the transduction processes. However, the perfusion of the axon interior washes out many factors which may be important in the processes under study. We introduce here the use of porous cellulose dialysis tubing as a means to circumvent these problems. We find that this dialysis technique is a simple procedure to set-up, and the serotonin/G-protein/adenylate cyclase system can readily be studied in the dialysed axon. This approach should allow investigation under conditions which retain asymmetric transmembrane conditions.


Bronchodilator action of an agonist for histamine H3-receptors in guinea pig perfused bronchioles and lung parenchymal strips. Isolated guinea pig perfused bronchioles and lung parenchymal strips were examined as an in vitro model for assessment of the direct effect of pharmacologic agents on the airway smooth muscle. The experiments were performed with a perfusion technique in bronchioles, the input pressure being measured as an index of the state of dilation, while changes in tension of the lung parenchymal strips were measured with an isometric force transducer. In both preparations, histamine and acetylcholine elicited dose-related contractile responses whereas fenoterol induced a concentration-dependent relaxation. After the 3 agonists' activities were compared in these 2 preparations, we tested the intrinsic effects of a specific H3 agonist, (R) alpha-methylhistamine ([R] alpha-MeHA). Statistical analysis was by Student's t test on the Emax (expressed as a percentage of 10(-4) M papaverine relaxation), EC50, and slopes of regression lines calculated from the concentration-response curves plotted for (R) alpha-MeHA alone or in presence of antagonists. Our results showed that (R) alpha-MeHA induced a concentration-dependent relaxation of perfused bronchioles and lung parenchymal strips competitively inhibited by 10(-7) M thioperamide (H3-antagonist), whereas 10(-5) M cimetidine (H2-antagonist) failed to prevent this effect. These results suggest the presence of H3-histaminergic dilatory receptors in the guinea pig airway.


2-(125I) iodomelatonin binding sites in rat adrenals: pharmacological characteristics and subcellular distribution. Specific binding sites for 2-[125I] iodomelatonin, a selective radiolabeled melatonin receptor ligand, were detected and characterized in rat adrenal membranes. Saturation studies demonstrated that 2-[125I]iodomelatonin binds to a single class of sites with an affinity constant (Kd) of 541 pM and a total binding capacity (Bmax) of 3.23 fmol/mg protein. Competition experiments revealed that the relative order of potency of compounds tested was as follows: 6-chloromelatonin greater than 2-iodomelatonin greater than melatonin greater than 5-methoxytryptamine greater than 5-methoxytryptophol. The highest density of binding sites was found in membranes from nuclear (0.76 fmol/mg protein) and mitochondrial (1.82 fmol/mg protein) subcellular fractions.


Solid-phase extraction and HPLC-UV confirmation of drugs of abuse in urine. A series of six liquid chromatographic methods were developed to confirm the presence of six classes of drugs of abuse in urine. The chromatographic separations were performed with a reversed-phase C8 column, except in the case of morphine, which was separated on a normal phase column. Isocratic and gradient elutions, ion pair technique, and UV detection were employed. Sample pretreatment involved the extensive application of solid-phase extractions and liquid-liquid extractions on solid supports. The specificity and sensitivity enabled the confirmation of morphine, benzoylecgonine, THC-COOH, amphetamine, and methamphetamine, six barbiturates, and nine benzodiazepines screened positive by EMIT in urine.


Nifedipine-resistant Ca(++)-induced contraction in tail artery of spontaneously hypertensive rats. Nifedipine-resistant Ca(++)-induced contractions (NR-Ca(++)-contraction) were compared in the tail arteries from SHRs and WKYs (5 and 13 week old). NR-Ca(++)-contraction of tail artery was defined as follows: Ca(++)-induced contraction in the presence of norepinephrine (NE) (10(-5) M) or 5-hydroxytryptamine (5-HT) (10(-5) M) in Ca(++)-free medium containing EGTA (0.1 mM) and nifedipine (10(-6) M). NR-Ca(++)-contractions in arteries from 5 week old SHRs and WKYs were not different. In contrast, NR-Ca(++)-contractions in arteries from 13 week old SHRs were about 2-fold greater than in arteries from 13 week old WKYs. In arteries from 13 week old WKYs and SHRs, nitroglycerin (10(-5) M) significantly reduced the NR-Ca(++)-contraction in the presence of 5-HT but not in the presence of NE. The reduction was inhibited by the presence of methylene blue (3 x 10(-6) M). 8-Bromo-cGMP (10(-4) M) reduced significantly the NR-Ca(++)-contraction in the presence of 5-HT in arteries from 13 week old SHRs and WKYs. The present experiments clearly demonstrated that the NR-Ca(++)-contractions (both in the presence of NE and 5-HT) in 13 week old SHRs were significantly greater than those in arteries from 13 week old WKYs. These results suggest that in addition to an increase in voltage-operated Ca++ mobilization reported by others, an increase in NR-Ca++ mobilization may contribute to the development of hypertension in SHR.


Metabolism of tetrahydrocannabinol in frequent and infrequent marijuana users. delta 9-tetrahydrocannabinol (THC), 11-nor-9-carboxy-delta 9- tetrahydrocannabinol (THC-COOH), and its O-ester glucuronide were measured in plasma by GC/MS and in urine by GC/MS and enzyme immunoassay after frequent and infrequent marijuana users were given 5 mg THC intravenously. Plasma THC concentrations were detectable 4-5 h after infusion using solid-phase columns for drug extraction and gas chromatography/mass spectrometry (GC/MS) for detection and quantification. THC-COOH and its O-ester glucuronide were analyzed for 12 days. Concentrations were higher in the plasma and urine of frequent marijuana users. The ratio of THC-COOH to its O-ester glucuronide in plasma was greater than 2 in both frequent and infrequent users 2 to 30 min postinfusion. Ratios for the subsequent 12 days were less than 2. A ratio of less than 1 for total THC-COOH/THC occurred only before 45 min for both frequent and infrequent users. Physiological, psychological, and pharmacokinetic data revealed few differences between frequent and infrequent marijuana users.


Argyrophilic nucleolar organizer regions in neoplastic and non-neoplastic hepatocytes bearing Mallory bodies. The proliferative activity of Mallory bodies (MB)-positive hepatocytes (neoplastic and non-neoplastic) was examined by counting the argyrophilic nucleolar organizer regions (AgNORs). Among 19 cases of hepatocellular carcinoma, the mean number of AgNORs was lower in the MB-positive carcinoma cells than in the negative ones in nine cases, higher in six, and there was no difference in four. In non-neoplastic cases (seven cases of advanced primary biliary cirrhosis and seven cases of alcoholic or nutritional liver injury), the mean number of AgNORs was lower in the MB-positive hepatocytes than that in the negative ones in eight cases, and approximately equal in number in six cases. These findings imply that MB formation does not directly represent the level of proliferative activity of hepatocytes, regardless of whether they are malignant or not.


Prospective controlled study of post-transfusion hepatitis after cardiac surgery in a large referral hospital in India. We studied the risk of post-transfusion hepatitis (PTH) in recipients of blood collected from voluntary donors screened for HBsAg. Two hundred and fifty patients without any previous history of liver disease or transfusion were followed up for 12 months subsequent to cardiac surgery. Thirty-five of them had closed-heart surgery without receiving transfusion and served as controls. The remaining 215 patients received single-point transfusions (mean 4 +/- 2.4 units). None of the controls and 15 (6.9%) blood recipients developed PTH. Three (20%) patients had hepatitis-B-virus-induced hepatitis while the remainder (80%) had non A, non B (NANB) hepatitis. The number of units of blood transfused and surrogate markers for development of PTH (donor alanine aminotransferase, anti-HBc and anti-HBs antibody) were not associated with the occurrence of PTH (p greater than 0.05). Nine (60%) of the 15 patients developing PTH were asymptomatic. All the patients recovered from the PTH, except one who died of fulminant hepatitis. At the end of 1 year of follow-up, none of the patients had evidence of chronic hepatitis. Only three (25%) of the patients with NANB-PTH developed anti-hepatitis C virus (HCV) antibody during the follow-up. We conclude that the incidence of PTH in India is similar to other parts of the world and NANB virus was the major cause of the PTH. The absence of chronicity and lack of seroconversion to anti-HCV antibody in the majority of the patients after 1 year of follow-up may suggest the possibility of a NANB virus other than HCV as the major cause of PTH in India.


Transcription of bovine leukemia virus in peripheral blood cells obtained during early infection in vivo. Bovine leukemia virus (BLV) is transcriptionally silent in most circulating peripheral blood mononuclear cells (PBMCs) of animals with well-established infections. Using PBMCs from a newly infected sheep, we asked whether viral transcription proceeded differently during the initial months of infection, when the prevalence of BLV-infected cells and the host's immunological response change markedly. Shortly after being injected with BLV, the animal displayed a characteristic, transient increase in PBMCs that transcribed BLV when cultured. Even when transcriptionally competent PBMCs were most prevalent (1.2%), only rare cells in the circulation (1 in 50,000) contained enough BLV transcripts to be identified readily by in situ hybridization. However, at one point several weeks later, some PBMCs appeared to contain small amounts of BLV RNA as soon as they had been purified from blood. Throughout this period, BLV-transcribing PBMCs greatly outnumbered virus-producing cells, which were counted using a new infectious centers assay. Its viscous medium reduced cell to cell contact among PBMCs, enabling increased detection of BLV-producing cells at a time when virus-specific killer cells might be active. Early infection was polyclonal, and most infected PBMCs transcribed BLV upon being cultured. By 2 months after infection, provirus-containing cells were as abundant as they had been earlier, but few cells transcribed BLV. These results suggest that BLV-infected cells are more easily stimulated to transcribe the provirus and produce infectious virus during the early months of a new infection.


Laboratory diagnosis of rotavirus infection in diarrhoeal patients by immunoenzymatic and latex-agglutination assays. One hundred stool samples from children with acute diarrhoea were examined by six commercial latex and immunoenzymatic assays for the diagnosis of rotavirus infection in four different laboratories. Samples were also analyzed by solid-phase immune electron microscopy using a rabbit anti-group A rotavirus antiserum. With electron microscopy as a basis for comparison, sensitivity and specificity for the latex and ELISA assays varied from 91.1 to 92.9% and from 94.2 to 99.4%, respectively. Statistically significant differences were revealed in the confirmation rate of electron microscopy-negative samples between different commercial assays. Significant variability was also found between results obtained by the laboratories taking part in the study.


Prevalence of genital papilloma virus infections in asymptomatic and symptomatic women, studied with a combined dot-blot and Southern blot procedure. A technique for the detection and typing of genital infections with human papilloma virus (HPV) is described. Following a non-invasive sampling and a simplified preparation procedure, the analysis was performed as a combined dot-blot and Southern blot analysis, where the former test was used to exclude cases without demonstrable content of HPV DNA. The subgenomic probes used in the Southern blot analyses have a higher specificity than can be achieved with genomic ones, and only one band is obtained for each of HPV types 6, 11, 16, 18, 31, 33 and 35. Extra bands indicating the presence of cross-reacting HPV of undetermined type occurred in only 0.7% of the 1,268 cases tested. HPV was demonstrated in 7.8% of the 612 health control samples and in 16% of 634 consecutive samples from patients with symptoms. All but type 18 occurred more commonly in symptomatic than in asymptomatic patients. Presence of HPV also correlated to cervical intraepithelial neoplasia (CIN) in simultaneously collected smears. The HPV was demonstrated in 13% of patients with normal cytology, in 45% with CIN and in 82% of patients with invasive carcinoma. All types of HPV occurred more commonly in CIN patients, while only types 16, 18, 31 and 33 were found in the patients with cancer.


Viral susceptibility of an established cell line of swine embryo kidney. The viral susceptibility of a cell line, named KSEK6, newly established from the kidney cortex of swine embryo was tested for the indication of CPE occurrences and also plaque formations. The multiplication of porcine adenoviruses was considerably high in the cells among the virus strains tested though plaques of these viruses were hardly visible under agar overlay medium. Two strains of swine enteroviruses, Aujeszky's disease virus and hemagglutinating encephalomyelitis virus also multiplied well in a similar order to those received in the other cells employed.


Studies into secretions of Tetrahymena: enzymes secreted into inorganic medium. The peptides secreted by Tetrahymena cells into inorganic medium were chromatographed. Six fractions showing a marked enzyme-like activity were examined for influence on certain physiological parameters of Tetrahymena. The enzymatically active fractions increased the phagocytic activity of Tetrahymena and decreased its binding capacity for lectins and hormone (insulin), but enhanced insulin imprinting at primary interaction. It remains to be clarified whether these effects were due to the enzymatic or other components of the fractions investigated, or to lack of the compensatory influence of the fractions not studied.


A unique type of GABA binding by Mycobacterium leprae. Neurotropism is one of the unusual properties of Mycobacterium leprae. The organism contains glutamic acid decarboxylase that generates gamma-amino-butyric acid (GABA) which is an inhibitory neurotransmitter. The binding of GABA by M. leprae in vitro was studied by using 3H-GABA as substrate. The bacteria had high-affinity binding sites for the amino acid. The uptake was a specific saturable process with a Km of 66.7 pM, pH optimum of 7.3 and a temperature optimum of 37 degrees C. The binding did not seem to be time-dependent, being complete in about 5 min. None of the known antagonists and agonists of GABA uptake by neurons, showed any significant effect on M. leprae; the receptors in the bacteria are apparently of a non-neuronal type, and different from those reported in spermatozoa and Pseudomonas.


Wallerian degeneration in peripheral nerve disease. This article first reviews the different classes of nerve injury. The temporal evolution of electrophysiologic changes occurring during Wallerian degeneration in humans is then described, followed by a description of sequential structural changes and cellular responses that occur after nerve transection. The final section focuses on the basis for resurgent research interest in Wallerian degeneration and the different research approaches being taken to answer basic science and clinical questions.


Inflammatory-demyelinating polyneuropathies. Recent advances in the pathogenesis and management of Guillain-Barré syndrome (GBS) and chronic inflammatory demyelinating polyneuropathy (CIDP) are presented. These diseases are both diagnosable and treatable. The role of Campylobacter infection preceding GBS and the use of intravenous immunoglobulin to treat CIDP are discussed.


Peripheral neuropathies associated with human immunodeficiency virus infection. In the 1990s, HIV has replaced syphilis as the "great masquerader." Virtually every level of the neuraxis may be affected in a patient with HIV infection. The superimposition of multiple levels of neuropathology further complicate the bedside neurologic diagnosis of an AIDS patient. This article has reviewed the variety of forms of peripheral neuropathy that may be associated with HIV infection and its treatment. Distal symmetrical polyneuropathy may be produced in patients with HIV infection by neurotoxic drugs (e.g., vincristine, INH, ddC, or ddI) or by vitamin B12 deficiency or may develop in the later stages of HIV infection without identifiable cause. GBS and CIDP occur with increased frequency in early HIV infection owing to presumed autoimmunity, and these IDPs respond to plasmapheresis or prednisone, similar to HIV-seronegative patients. A limited distribution of mononeuropathy simplex or multiplex occurs in patients with CD4 counts greater than 200; the neuropathy will usually spontaneously improve in these patients. Widespread mononeuropathy multiplex may occur in patients with AIDS and CD4 counts less than 50 and is then usually caused by CMV infections; those neuropathies are usually progressive unless antiviral treatment is given. Progressive polyradiculopathy usually occurs in patients with AIDS and low CD4 counts. If the cerebrospinal fluid has a polymorphonuclear pleocytosis, CMV infection is almost always present, and progression is expected unless ganciclovir therapy is promptly started. Finally, mild autonomic neuropathy is commonly present in HIV-infected patients. Protocols for the evaluation and therapy of cranial and peripheral neuropathies are presented (Figs. 6 and 7). It is unfortunate but likely that increasing numbers of "neuro-AIDS" patients will be encountered, not only in urban medical centers but also in general community practice. The pace at which research in the field of HIV research has proceeded is unprecedented. It is, therefore, important that neurologists stay at the forefront of investigation and clinical care of these complex disorders.


Monoclonal proteins in neuropathy.

Monoclonal proteins in neuropathy. The presence of a monoclonal protein in the serum of a patient with peripheral neuropathy raises the suspicion of systemic amyloidosis, POEMS syndrome, macroglobulinemia, multiple myeloma, or lymphoma. If these conditions are excluded, the patient is classified as having a monoclonal gammopathy of undetermined significance (MGUS) with an associated neuropathy. Approximately one half of patients with peripheral neuropathy and IgM monoclonal gammopathy have IgM antibodies that bind to a myelin-associated glycoprotein (MAG). In addition to MAG, other antigens such as glycolipids or gangliosides represent other target antigens. Monoclonal gammopathies and motor neuron diseases have also been reported. Sensorimotor peripheral neuropathy occurs in about 15% of patients with primary amyloidosis but is uncommon in multiple myeloma.


Inflammatory sensory polyganglionopathies. The inflammatory sensory polyganglionopathies are a group of uncommon neurologic disorders primarily affecting dorsal root ganglion cells and their processes. Subdivided into malignant and nonmalignant inflammatory sensory polyganglionopathy, these conditions are important in the differential diagnosis of sensory neuropathy. The clinical significance of malignant inflammatory sensory polyganglionopathy, a paraneoplastic syndrome, rests in the discovery of subclinical cancer, which may produce antineuronal nuclear antibodies directed against the dorsal root ganglion cell and other neurons resulting in characteristic neurologic deficits. Prompt recognition of these distinctive signs and symptoms may result in early diagnosis and improved patient survival and lead to a better understanding of immune-mediated neurologic disease.


Vasculitic neuropathy.

Vasculitic neuropathy. Peripheral neuropathy is common in many vasculitic syndromes and may be the only manifestation of the underlying vasculitic disease. Although traditional teaching has been that a true multiple mononeuropathy is the classic clinical presentation of vasculitic neuropathy, an overlapping (or extensive) multiple mononeuropathy or a distal symmetric polyneuropathy is commonly encountered. Similarly, the leukocytoclastic reaction has traditionally been considered the primary mechanism of vessel injury in these diseases, although more recent evidence suggests that cellular-mediated mechanisms may be more important in peripheral nerve. In this review, new concepts concerning the clinical presentation, pathogenesis, diagnosis, and treatment of vasculitic neuropathy are discussed, particularly in relation to the syndrome of isolated peripheral nerve vasculitis.


Frequently asked questions about diabetic peripheral neuropathies. A number of issues pertaining to the diabetic peripheral neuropathies remain unanswered or controversial. The final solution to the disabling problem of diabetic neuropathy might well await the discovery of the exact pathogenesis and treatment of diabetes.


Quantitation of axon loss and conduction block in acute radial nerve palsies. Eleven acute radial nerve palsies were examined between 3 days and 14 weeks following the onset of the neuropathy. Our objective was to quantify the relative extent of axon loss and conduction block in radial motor fibers supplying the extensor and abductor pollicis longus (EPL/APL) muscles. In 10 of 11 cases, conduction block exceeded axon loss. Maximum motor and sensory conduction velocities were normal distal to the spiral groove, suggesting that the larger myelinated fibers were not selectively involved in this acute neuropathy. The localization of the conduction block and slowing was, in every case, across the spiral groove. This method provides a relatively simple way of assessing the approximate contributions of axon loss and conduction block, and fits well with the early and usually complete clinical recovery in these cases.


Muscle fiber conduction velocity and mean power spectrum frequency in neuromuscular disorders and in fatigue. This study investigated the relation of muscle fiber conduction velocity (MFCV) to difference power spectrum mean frequency (MF), their fatigue trends, and differences between their values and their fatigue trends in various neuromuscular disorders. Electromyographic interference pattern was recorded inside the biceps in continuous isometric maximal voluntary contractions. Each subject was encouraged to pull for as long as possible. Fatigue was calculated as percent of time to complete inability to sustain contraction. The MFCV was computed by cross-correlation. The MF was computed by differencing, windowing, FFT, squaring of coefficient, and repeat averaging. There were 33 healthy, 86 polyneuropathic, 28 myasthenic, 13 myotonic, and 32 myopathic patients. Both MFCV and MF changed significantly with fatigue--the MFCV linearly, while the MF in a markedly nonlinear fashion. Both were found to be insensitive to the end stages of muscle fatigue--the MFCV did not change its slope toward complete fatigue, and the MF did not change at all beyond the 40% fatigue point. A statistically sound fatigue regression equation was derived for each, and a nonlinear equation was found to best describe their relationship. Neither MFCV nor its fatigue changes were found to be significantly different across the neuromuscular disorders. The MF, however, was found to be significantly different in some neuromuscular disorders in both its average values and fatigue trends. This study showed, in contrast to the literature, a nonlinear relationship between MFCV and MF. It also shows that neither the MFCV nor the MF had reasonable diagnostic power on its own; however, the MF was very promising to serve as an adjunct to other variables.


Amiodarone-induced experimental acute neuropathy in rats. Amiodarone was injected endoneurially at increasing doses into the exposed tibial nerve of rats to study its electrophysiologic and pathologic effects on peripheral nerve fibers. Forty-five male Wistar rats were used, and each of the following concentrations was injected into 15 nerves: 25 micrograms/mL, 50 micrograms/mL, and 100 micrograms/mL. Microinjection of a 25 micrograms/mL concentration of amiodarone resulted in a subacute, incomplete conduction block evident at day 3 postinjection. This conduction block remained stable until day 10 and recovery was complete at day 35. Microinjection of a 50 micrograms/mL concentration of amiodarone produced a faster evolving conduction block, and significant axon degeneration (approximately 40% of fibers). Injection of a 100 micrograms/mL concentration resulted in severe acute motor axon degeneration followed by complete but delayed regeneration. Results of morphological studies closely correlated with electrophysiological findings. Amiodarone thus seems to have a direct toxic effect on axons at high concentrations in the peripheral nerve, and we suggest that different pathological changes described in human amiodarone neuropathy could be related to different concentrations of the drug in the nerve, perhaps due to variability of blood-nerve barrier efficacy.


Demyelinating polyneuropathy in eosinophilia-myalgia syndrome. Eosinophilia-myalgia syndrome (EMS) is a newly recognized disorder, characterized by myalgia, weakness, scleroderma-like changes, and eosinophilia. EMS is associated with lots of L-tryptophan allegedly contaminated with byproducts of the manufacturing process. We describe 3 patients with EMS who presented with a severe demyelinating sensorimotor polyneuropathy. Electrodiagnostic studies revealed multifocal conduction block, slowing and temporal dispersion of motor responses, and prolonged or absent F-responses. Despite plasmapheresis; corticosteroids; and, in 1 patient, cyclophosphamide, 2 patients died and the remaining patient experienced minimal recovery. Pathology revealed patchy perivascular infiltrates and fibrosis in the connective tissue of muscle and nerve. Autopsy of the central nervous system in 2 patients did not reveal changes unique to EMS. In addition to other organ involvement, EMS may manifest as a potentially fatal polyneuropathy, which initially appears to have prominent demyelinating features.


Effects of reduced joint mobility and training on Na,K-ATPase and Ca-ATPase in skeletal muscle. In guinea pigs, the ankle joint was partly immobilized in a position reducing dorsiflection to 105 degrees (as compared to the normal value of 30 degrees). When compared with the contralateral unrestrained leg, this led to a significant atrophy and a decrease in contractile force (-23%) of the gastrocnemius muscle. This was associated with a significant decrease in the total concentration of [3H]ouabain binding sites in gastrocnemius and plantaris muscle reaching minimum (-19% and -23%) after 3 weeks, but no evidence of degenerative changes. Total contents of Ca and Ca-ATPase were increased by 27% and 22%, respectively. After 4 to 5 weeks of reduced mobility, the concentration of [3H]ouabain binding sites in gastrocnemius muscle returned to control level. The lowest concentration of [3H]ouabain binding sites reached during reduced mobility was 258 +/- 13 pmol/g wet wt., and the maximum value attained following 3 weeks of reduced mobility and 3 weeks of training by running was 498 +/- 25 pmol/g wet wt., i.e, 93% higher. In soleus, training produced an increase of 25%. Clinically, it is important to realize that movable braces cannot prevent the development of muscular atrophy. The observed spontaneous recovery of the Na,K-pump concentration may partly explain why patients using movable casts show a better capacity for physical performance than those treated with complete immobilization. In conclusion, the total concentration of Na,K-pumps in guinea pig skeletal muscle undergoes downregulation and upregulation as a function of contractile activity as well as muscle length under conditions mimicking the constraints on mobility frequently used in the clinical treatment of lesions.


Changes in the lateral diffusion of fluorescent lipid analogues in the surface membrane of adult male Schistosoma mansoni. The effect of serotonin on the fluidity of the tegumental membranes of adult male Schistosoma mansoni was assessed by the fluorescence recovery after photobleaching technique. It was demonstrated that the translational diffusion of 5-N'-octadecanoyl aminofluorescein is reduced by a mechanism involving G-protein coupled activation of adenylate cyclase and lowering of intracellular calcium concentration. Furthermore, the lateral diffusion coefficient and the mobile fraction appear to be controlled by calcium and cAMP dependent pathways respectively. No change in the diffusion of the fluorescent phospholipid N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-phosphatidyl choline was observed, suggesting the two probes used here partition into two different domains that are under independent control. An increase in the amount of protein associating with a membrane cytoskeleton is also demonstrated.


Homodimer formation of retinoid X receptor induced by 9-cis retinoic acid. Retinoid response pathways are mediated by two classes of receptors, the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs). A central question is whether distinct response pathways are regulated by these two classes of receptors. The observation that the stereoisomer 9-cis-retinoic acid binds with high affinity to RXRs suggested that this retinoid has a distinct role in controlling RXR activity, but it was almost simultaneously discovered that RXRs function as auxiliary receptors for RARs and related receptors, and are essential for DNA binding and function of those receptors. Hence, although RARs seem to operate effectively only as heterodimeric RAR/RXR complexes, RXRs themselves apparently function predominantly, if not exclusively, as auxiliary receptors. Here we report that 9-cis-retinoic acid induces RXR homodimer formation. Our results demonstrate a new mechanism for retinoid action by which a ligand-induced homodimer mediates a distinct retinoid response pathway.


[How can one achieve compliance?--On the after-care of psychiatric patients after clinic discharge]. For 384 of 558 hospitalized psychiatric patients who were discharged within six months and for whom it was possible to plan aftercare, compliance was controlled by a phone call three months after discharge at the institution or practice to which the patient had been directed. Eighty percent of the ambulatory patients complied (schizophrenic patients: 84%). After three months 70 percent of the compliers still were in treatment. The hospital's arranging an appointment for the patient had the strongest influence on compliance. The success of this strategy is confirmed by the literature.


Hepatitis C virus antibody response in acute and chronic non-A, non-B hepatitis. Patients with post-transfusion, community-acquired or hemodialysis-acquired non-A, non-B hepatitis (NANBH) were tested for antibody to hepatitis C virus (HCV) during acute-phase and resolving or chronicized illness. HCV appears to be involved in most cases of post-transfusion and hemodialysis-acquired NANBH, but only in 40% of community-acquired NANBH. Second generation HCV antibody assays are more specific and sensitive, favoring early detection of HCV seroconversion and identification of HCV-antibody-positive individuals years after exposure to the virus.


Behavior of antibody profile against hepatitis C virus in patients on maintenance hemodialysis. The prevalence of anti-hepatitis C virus (HCV) in dialysis setting is still a nonstandard datum. In particular, it is not known of the phenomenon is stable or increasing or decreasing, even in a given geographical area. We studied the behavior of anti-HCV prevalence during a 12-month follow-up in 415 hemodialysis patients treated at a single institution and belonging to a limited geographical area with standard HCV endemic. Point prevalence of anti-HCV has shown a tendency to growth linked in part of the incidence of infection, in part to new positivities in patients already on dialysis treatment. More than 50% of the new HCV-positive patients, had no history of classical parenteral transmission of the virus. These findings suggest that HCV infection is a phenomenon on the increase in dialysis units and that dialysis treatment emerges as an independent risk factor in contracting infection.


Antibodies against hepatitis C virus in hemodialysis patients in the central Italian region of Umbria: evaluation of some risk factors. The epidemiology of non-A, non-B hepatitis (NANBH) is still incomplete. To define the prevalence of antibodies against the main causative agent of NANBH, the hepatitis C virus (HCV) and the role of some risk factors, we tested sera from 269 patients on chronic dialysis at the hemodialysis units in our region in central Italy. We utilized the recently developed serological assay. Twenty-nine hemodialysis patients (13.3%) and 3 peritoneal dialysis patients (4.8%) were anti-HCV positive. Of these, 13 (40.6%) had antibodies to hepatitis B core antigen (anti-HBc) indicating prior hepatitis B infection. The anti-HCV seropositive patients had been on dialysis longer than the seronegative ones; they had received more transfusions than the others but without a significant difference. The prevalence rate of anti-HCV was statistically significantly higher among hemodialysis patients utilizing the same dialysis equipment for the previous 12 months.


Prevalence of hepatitis C virus antibodies in hemodialysis patients in the area of Milan. We studied the seroprevalence of antibodies to hepatitis C virus (HCV Ab) in a cohort of 229 chronic hemodialysis patients followed by 6 Dialysis Units in the Milan area. HCV Ab was present in 51 (22.3%) of 229 examined patients. Previous blood transfusions and surgery did not clearly influence HCV seroconversion, whereas duration of dialysis treatment seems to be strictly related to HCV Ab positivity.


Non-A, non-B hepatitis: clinical laboratory course in patients on hemodialysis and its correlation with the presence of anti-hepatitis C virus antibodies. 136 patients on hemodialysis, 89 males and 47 females, were studied; we evaluated the index of hepatic function (SGOT and SGPT) and antibodies against HCV. We observed 42 cases of increased transaminases classified as non-A, non-B (NANB) hepatitis. Antibodies against HCV were present in 40 patients. Among 42 patients with NANB hepatitis. 31 (73.8%) presented anti-HCV antibodies. No significant clinical or laboratory difference exists between anti-HCV-positive and -negative patients with NANB hepatitis. The distribution of patients who present anti-HCV antibodies is similar in post-transfusional and sporadic forms.


Antibodies to hepatitis C virus in kidney transplantation. Ninety patients on dialysis, 241 cadaveric kidney donors and 27 cadaveric kidney recipients with a follow-up of 2 years, have been investigated as for anti-HCV positivity by means of 3 tests. As for patients on dialysis and cadaveric donors, the prevalence was 32 and 4%, respectively. As for transplanted patients, it must be noted that 4 negative recipients from positive donors seroconverted, but without any change in hepatic enzymes, while in 2 or 9 anti-HCV-positive recipients, hepatic enzymes increased after transplantation. Seroconversion in patients transplanted from a negative donor was not significantly different. We conclude that, according to their experience, anti-HCV positivity in the donors is not associated with a significant risk of infection in recipients of cadaveric grafts.


Histological features of non-A, non-B hepatitis in hemodialysis patients. In 12 hemodialysis (HD) patients with persistently raised serum alanine amino-transferase concentrations, 6 of which with anti-HCV antibodies, a liver biopsy was performed. The histological examination showed chronic persistent hepatitis or less significant changes in 11 patients and mild chronic active hepatitis in only 1. Non-A, non-B hepatitis seems to demonstrate in HD patients a low tendency to induce an active and progressive liver disease.


Epidemiology of hepatitis C in a population of hemodialysis patients. A search for antibodies against hepatitis C virus (HCV) was performed in 185 patients on chronic hemodialysis by means of 1st and 2nd generation ELISA tests. Immunoblot assays were performed on positive sera. This study shows a 38% prevalence of HCV-positive patients in our dialysis population according to the 2nd generation ELISA test which shows a higher specificity and sensitivity when compared to the 1st generation one (38 vs. 20%). A correlation was found between the prevalence of HCV-positive patients and how long they had been on dialysis and how many blood transfusions they had received.


HCV infection occupational hazard at dialysis units and virus spread, among relatives of dialyzed patients. It was the purpose of this study to evaluate the spread of HCV infection among the staff at Dialysis Units and among relatives of anti-HCV-positive hemodialysis patients. 122 health-care workers and 52 relatives were screened for presence of anti-HCV Ab. The control groups consisted of 100 health-care workers from other wards and 30 relatives of anti-HCV-negative subjects. 2.45% of the health-care staff and 5.8% of the relatives were found to be anti-HCV Ab carriers. None of the subjects in the control groups were positive. In conclusion, the results of this study indicate that there is a real possibility of HCV spread through occupational exposure although the corresponding percentage in less than that presumably due to within-the-family contagion by the virus.


Prevalence of HCV antibodies in a uraemic population undergoing maintenance dialysis therapy and in the staff members of the dialysis unit. We studied, by both 1st and 2nd generation assay, the prevalence rate of HCVAb in a population of 141 dialysis patients, 37 transplanted patients and 55 staff members. From this study emerges a higher sensitivity of the 2nd generation HCVAb test (15.38 versus 36.79% of positive responses, respectively), and a significant positive correlation between lengths of dialysis period. We have not found a significant difference between HCVAb-positive and -negative patients in relation to the blood transfusions. None of the 21 CAPD patients (home dialysis) resulted positive, even if transfused. Two nurses were positive. In our experience, the environmental factor seems more important. Since the isolation of the positive patients is an effective but not feasible measure, it is necessary to improve the operating management of the hemodialysis sessions, avoiding any contact between patients via material (instrumentation, monitors) and teaching the staff members to use severe preventive standards with all hemodialysis patients.


Hepatitis C virus in hemodialysis patients. Anti-HCV prevalence in 284 hemodialysis patients was assessed using the Abbott HCV-EIA test. Anti-HCV positivity was found in 19.4% of patients, was higher in males than in females and progressively increased with age. A correlation between time of dialysis treatment, transfusions and ALT course was found. These data suggest that HCV infection is a very important risk in dialysis units.


Hepatitis C virus in dialysis units: a multicenter study. Three hundred and eighty-seven chronic hemodialysis patients were evaluated, in a multicenter study, to investigate the epidemiology of hepatitis C virus. In anti-HCV seropositive patients, serum ALT values and blood transfusions were retrospectively compared; blood donors were studied for serum transaminases. In seropositive patients without previous blood transfusions, analysis of dialysis schedule was done. Eventually, the intrafamilial transmission of hepatitis C virus was studied in 104 family members. The prevalence of HCV infection in hemodialysis patients was 15.7%. The incidence of acute hepatitis was frequent, while chronic hepatitis incidence was less than expected (17.5%). Intrafamilial diffusion was low (1.9%). Blood-transfusion-related infections seem to be negligible, while cross-contamination in dialysis units seems to be very important.


Risk factors and clinical expression of HCV infection in hemodialysis patients. In our Dialytic Unit, the prevalence of antibodies to HCV (anti-HCV) was 16.4% in 1990 (73 hemodialyzed) and 21% in 1991 (105 hemodialyzed). The incidence of seroconversion was 13.1%. The transaminase behavior was similar in a group of anti-HCV-positive and a group of anti-HCV-negative patients and with previous HBV infection. Blood transfusions and dialytic age did not affect anti-HCV positivity.


Prospective and retrospective assessment of clinical and laboratory parameters in maintenance hemodialysis patients with and without HCV antibodies. In order to clear some aspects of HCV infection, we evaluated quarterly HCV markers by a RIBA 1 test (antigens c100-3 and 5.1.1) and monthly transaminases (ALT and AST) for 14 months in 89 HBsAg-free maintenance hemodialysis patients (MHP), and we retrospectively examined clinical records until the start of hemodialysis treatment. At the start of the study, 16 patients showed HCV antibodies (HCV+) and 73 were antibody-free (HCV-). 39 subjects of the staff were also examined. No HCV+ patient showed seroconversion, 10 showed irregular or persistent elevation of AST and ALT. In the retrospective evaluation 14 patients suffered from acute hepatitis (AH). Only 3 cases showed temporal relation with blood transfusions. In 1 case a 36-month temporary normalization of transaminases was noticed. 3 HCV-patients showed seroconversion (1 during AH), 13 showed severe or moderate elevations of transaminases. In the retrospective evaluation, 6 patients suffered from AH. All subjects of the staff were HCV- and showed no seroconversion or changes of transaminases. At the end of the study, we performed a RIBA 2 test containing the HCV antigens c100-3, 5.1.1, c22-3 and c33c. The 6 patients who suffered from AH showed at least 1 positivity for new proteins. Most of AH in MHP are likely due to HCV infection; besides transfusions, cross-infection during the dialytic procedure may be responsible for many cases of HCV infection; long-term normalization of transaminases may not secure against infectivity.


HCV infections in dialysis patients.

HCV infections in dialysis patients. HCV is seen as an emergent problem in Dialysis Units; in fact, the patients of these centers are at high risk for blood-transmitted infections, through coming microepidemies. For this reason and using the same method, we have studied the prevalence of anti-HCV and virus-derived hepatitis in a dialytic population of 105 patients. The prevalence of anti-HCV was 25.71%.


Hepatitis C virus-related acute and chronic hepatitis in hemodialysis patients. The incidence of HCV antibodies has been evaluated in 123 chronic hemodialysis (HD) patients (Group A; 55 M and 68 F) and in 37 consecutive HD patients (group B) admitted to our hospitals for acute hepatitis. In group A, HCV antibodies were present in 27% of the patients. 20 of 36 (55%) had previously received blood transfusions. 21 patients (58%) were also positive for HBV Ab. In 8 patients, ALT were significantly increased. In group B, the diagnosis of HCV-related acute hepatitis was made in 11 patients. 8 of them had previously received blood transfusions. Seroconversion occurred 2-3 months after onset of the disease.


Prevalence of hepatitis C infection in a hemodialysis unit. To define the prevalence of NANB hepatitis, anti-HCV antibodies were determined in 51 patients on renal replacement therapy, in 7 transplanted patients and 17 staff members of the hemodialysis unit. Anti-HCV antibodies were evaluated using immunoenzymatic methods (Ortho HCV ELISA Test System, 1st and 2nd generation). Among hemodialysis patients, seroconversion was respectively documented in 17.6% (9/51) and 52.9% (27/51); none of the transplanted patients were positive with the 1st generation test, while 3/7 were positive with the 2nd. No statistically significant difference was found in the prevalence antibodies between transfused and nontransfused patients. ALT levels were statistically greater in patients with anti-HCV antibodies (X2 2nd generation = 8.83; p less than 0.01). Our results suggest: (1) that hemodialysis represents a risk factor; (2) the validity of substitute markers and (3) more sensitivity of the 2nd than 1st generation test.


HCV infection in hemodialyzed patients: incidence and correlation with dialytic age. Hemodialyzed patients mean an high-risk population for hepatitis C infection. Our work was performed to evaluate the incidence of hepatitis C virus (HCV) infection in 51 hemodialyzed patients; the presence of anti-HCV antibodies was studied using the EIA and RIBA test of 1st and 2nd generation. 18 patients (35.29%) showed anti-HCV antibodies with the 1st test; 27 patients (52.94%) showed the presence of anti-HCV antibodies using 2nd generation test. The incidence of test positivity is not related to blood transfusions while it is strictly related with dialytic age. All HCV seropositive patients show antibodies against the c22-3 protein of "virus core". The presence in serum of anti-c22-3 antibodies means that in these patients, there is viral replication.


Prevalence of antibodies against hepatitis C virus in a dialysis unit. Anti-HCV was tested in 77 uremic patients, 48 on hemodialysis (HD), 29 on CAPD, by immunoenzymatic 1st and 2nd generation assays (ELISA I, II) and 4-antigen (4-RIBA) immublotting. The investigation was extended to the staff (n = 29) and to HCV-positive patients' families (n = 30). The prevalence using 2nd generation tests was double (21%) that in 1st generation tests (11%). A greater incidence in the HD than in the CAPD group (23 vs. 17%) and a highly significative correlation to dialytic age were observed. No one among the sanitary personnel and only 2 family members were found HCV positive, suggesting a low infectivity via the parenteral inevident route. Extracorporeal circulation and particularly the exposure time to the treatment seem to be the main risk factors.


On the positivity of the test for anti-HCV considerations in dialized patients. We have tested dialized patients with the ELISA test of 1st generation for anti-HCV. All patients who resulted positive were subjected to the RIBA test of 1st and 2nd generation. Since the positivity rate with this test is considerably more reliable, we suggest to confirm by RIBA test all anti-HCV-positive patients with ELISA in time.


Antibodies to hepatitis C virus (anti-HCV): prevalence in the same geographical area in dialysis patients, staff members, and blood donors. We studied the anti-HCV prevalence in the same period in all the dialysis patients, staff members and blood donors of the same geographical area. In our opinion, the low prevalence (10.26%) in dialysis patients is strictly correlated to the low prevalence (0.46%) in blood donors.


Confirmation of high prevalence of hepatitis C antibodies in hemodialysis patients by second generation immunoblot assay. Sera from 209 dialysis patients were tested for antibodies to hepatitis C virus (anti-HCV) by a 2nd generation enzyme-linked immunoassay (ELISA 2) using nonstructural and core antigens. Confirmation of reactivity was obtained by a 2nd generation immunoblot assay (RIBA 2) for antibodies to 4 separate antigens (5-1-1, c100-3, c33c, c22-3). ELISA 2 was positive in 99 sera, 95 of which were confirmed by RIBA 2, thus accounting for an anti-HCV prevalence of 45.5%. Anti-HCV positivity was correlated to longer duration of dialysis therapy (p less than 0.001), higher number of transfusions (p less than 0.001), history of kidney transplant (p less than 0.001) and of serum alanine/aspartate aminotransferase (AST/ALT; p less than 0.001) or gamma-glutamyltransferase (GGT) (p less than 0.001) increments. The most frequent RIBA 2 patterns were: reactivity to all 4 antigens (34 patients) and to c33c and c22-3 (45 patients). The former patients, compared to the latter, had higher values of AST (p less than 0.08), ALT (p less than 0.02), GGT (p less than 0.005), IgG (p less than 0.05). It is possible that the reactivity to all 4 antigens of RIBA 2 is a clue of a greater activity of viral hepatic disease.


High prevalence of antibodies to hepatitis C virus in hemodialysis units using a second generation assay. We studied the prevalence and significance of antibodies to hepatitis C virus (HCV) in patients and staff from 3 dialysis units, using a 2nd generation assay (2nd g.a.; Ortho HCV). Of 277 patients, 151 (55%) were positive by 2nd and 85 (31%) by 1st g.a. Significant associations with the anti-HCV carrier status were: blood transfusions, retrospective finding of elevated ALT and duration of dialysis treatment, independently of transfusions. Of the 74 staff members, 5 were positive by 2nd and 3 by 1st g.a. Our data suggest that the 2nd g.a. is more sensitive in detecting HCV exposure in dialysis units and that duration of dialysis is a significant factor in acquiring HCV infection.


Comparison between first and second generation tests to determine the frequency of anti-HCV antibodies in uremic patients in replacement dialytic therapy. Research was carried out on 74 hemodialysis patients to determine the frequency of anti-HCV antibodies; this was done by means of the 1st and 2nd generation screening and control tests. The 1st generation tests showed 9 HCV-seropositive patients, while the 2nd generation tests demonstrated 15 HCV-seropositive patients. It seems evident that the 2nd generation tests are more sensitive than those of the 1st.


Anti-HCV antibody presence in hemodialyzed patients: assessment after one year regarding some preventive measures chosen. The authors report on the presence of anti-HCV antibodies in a uremic chronic (UC) patient population after 6 months and 1 year with regard to certain selective preventive measures. The nonincreased number of HCV-positive subjects among the UC population could lead to the conclusion that the risk of environmental spreading of HCV is less than the risk of HBV diffusion.


HCV in a group of chronic hemodialysis patients. Since HCV appears to be the major cause of post-transfusion non-A, non-B hepatitis in Italy, this study determines the presence of anti-HCV in a risk group. Among 26 patients, 9 were anti-HCV in a risk group. Among 26 patients, 9 were anti-HCV positive with the ELISA test and all of them were confirmed with the RIBA test of 2nd generation. Only 1 had a poussés movement of ALT levels. Hemodialyzed patients are reactive for HCV probably for the transfusional therapy.


Hepatitis C virus antibodies in dialysis pediatric patients. A new 4-antigen recombinant immunoblot assay (4-RIBA) for confirmation of hepatitis C virus (HCV) C-100 enzyme-linked immunosorbent assay (ELISA) reactivity was tested in serum samples of 11 pediatric patients on dialysis. Of 6 HCV C-100 ELISA-positive samples, all were 4-RIBA positive. The 2nd generation ELISA picked up 1 additional case confirmed by 4-RIBA. The 2nd generation tests increased the prevalence of HCV-positive cases.


Comparison between first- and second-generation test for anti-hepatitis C virus antibodies in hemodialysis patients. The serum of 387 hemodialysis patients from 9 dialysis units was checked for anti-hepatitis C virus antibodies with a 1st-generation ELISA (Ortho) test: 61 patients were repeatedly positive. In order to avoid false-positive results, these sera were tested with a 1st-generation confirmatory RIBA test, 2nd-generation screening ELISA test and 2nd-generation confirmatory RIBA test. The 2nd-generation ELISA test confirmed data obtained with 1st-generation ELISA, however, the 1st-generation confirmatory RIBA test underestimated the number of anti-HCV-positive patients.


Hepatitis C: reality of a renal unit.

Hepatitis C: reality of a renal unit. Hepatitis C virus (HCV) seems to be the most important agent of non-A, non-B hepatitis. This study was undertaken to assess the prevalence of hepatitis C in our renal unit. Twelve patients (29%) had antibodies against HCV (anti-HCV). Seropositive patients were on hemodialysis for a longer period than seronegatives. Statistically significant associations with anti-HCV were: blood transfusions, at least 1 episode of elevated value of transaminases (2-fold) and fluctuations of transaminases. Our findings confirm the high prevalence of anti-HCV in hemodialyzed patients, the importance of parenteral transmission and the high probability of liver disease in anti-HCV-positive patients.


Serum epidemiological trial on the prevalence of the anti-cytomegalovirus antibodies in patients under substitutive treatment with hemodialysis and CAPD. The detection of IgG and IgM anti-cytomegalovirus is very important when movements of serum transaminases are noted in absence of positivity for virus of hepatitis in uremic patients on substitutive treatment; in our patients, we have noted a positivity of 67% for the IgG class. Cytomegalic infection must be taken into consideration for the high parenteral transmissibility in addition to the high grade of immunodepression of uremic patients.


HCV incidence in a dialysis center: preliminary reports. Incidence of seroconversion to anti-HCV from December 1989 to May 1991 was evaluated in patients and in their sexual partners. In December '89, 13 of 66 and in May '91, 26 of 75 patients were HCV positive. 9 of 13 new, seroconverted patients had been transfused. Seroconverted mean dialytic age was lower than that of previous HCV-positive patients. All the sexual partners were HCV negative. Blood transfusion seems to be the main risk factor for HCV infection, while environmental contamination is still debatable.


Selective venous sampling in the differential diagnosis of ACTH-dependent Cushing's syndrome. Bilateral simultaneous inferior petrosal sinus sampling, associated with the oCRH stimulation test (100 micrograms i.v. as a bolus) was performed in 22 patients with Cushing's syndrome and no signs of pituitary abnormalities. Catheters were inserted into both femoral veins. More than one site in the superior and inferior vena cava was sampled before reaching the inferior petrosal sinuses. Blood samples for ACTH and beta-endorphin were gently aspirated from both petrosal sinuses and from a peripheral vein simultaneously. Blood was drawn at 0, 5, 10 and 15 min after oCRH injection. Seventeen of 22 patients showed an ipsilateral to peripheral vein ratio higher than 1.5, and 12 patients showed a lateralization of ACTH levels after oCRH stimulation. Seventeen patients underwent transsphenoidal pituitary surgery. Nine patients had a pituitary adenoma at the expected side; 1 at the contralateral side, while in 2 it was central. Three of 4 patients in whom the ipsilateral/peripheral ratio was less than 1.5 had the highest ACTH levels at the superior or inferior vena cava, not responsive to oCRH stimulation. One of these had a mediastinal and one a pulmonary mass. The third one, with an occult ectopic source, is still under investigation. At immunohistochemical and biological in vitro studies, both tumors were shown to secrete ACTH. In 13 patients in whom both beta-endorphin and ACTH measurements were performed, these hormones showed similar patterns of response. In conclusion, simultaneous bilateral petrosal sinus catheterization is a useful tool in the differential diagnosis of Cushing's syndrome as concerning pituitary and ectopic forms.(ABSTRACT TRUNCATED AT 250 WORDS)


Sex differences in the regulation of oxytocin receptors by ovarian steroids in the ventromedial hypothalamus of the rat. The facilitation of sexual receptivity by oxytocin (OT) in female rats is related to the regulation of oxytocin receptors (OTR) by ovarian steroids in the ventromedial nuclei (VMN) of the hypothalamus. In a previous study, we have shown that estradiol benzoate (EB) causes a twofold increase in OTR binding in the VMN. Progesterone (P) then modulates levels of the estrogen-induced OTR and increases the area occupied by the receptors by acting on the neuronal membrane. In the present study, we compared the effects of EB and P on OTR binding between males and females. In both sexes, EB increased the density of OTR and the area covered by the receptors at the level of the medial and caudal VMN. In estrogen-primed females, P further increased OTR levels in the medial VMN and the area covered by OTR at the level of the caudal VMN. By contrast, P did not modulate OTR binding in estrogen-primed males. Thus, the behavioral insensitivity of male rats to ovarian hormones, in particular to P, may be related to sex differences affecting the modulation of OTR binding.


Evidence that prolactin mediates the stimulatory effects of estrogen on tuberoinfundibular dopamine neurons in female rats. The effects of ovariectomy and estrogen on prolactin secretion and/or the activity of tuberoinfundibular dopamine (TIDA) neurons were examined by either concurrently measuring concentrations of prolactin in plasma and 3,4-dihydroxyphenylacetic acid (DOPAC) in the median eminence of female rats or by determining the rate of DA synthesis (accumulation of 3,4-dihydroxyphenylalanine (DOPA) after the administration of a decarboxylase inhibitor) in the median eminence. For comparison, concentrations of alpha-melanocyte-stimulating hormone (alpha MSH) in plasma and DOPAC in the intermediate lobe of the pituitary (an index of the activity of tuberohypophysial DA neurons) were also determined. Ovariectomy produced a time-dependent decrease in the accumulation of DOPA and the concentrations of DOPAC in the median eminence and prolactin in plasma with maximal effects occurring by 7 days. Estrogen administration to ovariectomized rats increased plasma prolactin and median eminence DOPAC concentrations to levels comparable to those in diestrous controls. In contrast, neither ovariectomy nor estrogen replacement altered the concentrations of alpha MSH in plasma or DOPAC in the intermediate lobe. Administration of the DA agonist bromocriptine blocked the ability of estrogen to increase plasma prolactin and median eminence DOPAC concentrations. Also, administration of antiserum to rat prolactin blocked the stimulatory action of estrogen on median eminence DOPAC concentrations. Taken together, these results indicate that the stimulatory effect of estrogen on the activity of TIDA neurons is mediated by prolactin.


Sexual differences in kappa opioid receptor-mediated regulation of tuberoinfundibular dopaminergic neurons. The purpose of the present study was to examine the acute effects of kappa opioid receptor blockade or activation on the activity of tuberoinfundibular dopaminergic (TIDA) neurons in gonadally-intact or castrated male and female rats. In the absence of drug treatment, the basal activity of TIDA neurons (accumulation of 3,4-dihydroxyphenylalanine, DOPA, in the median eminence after administration of a decarboxylase inhibitor) in male rats was approximately one third of that in diestrous females. In male rats, blockade of kappa opioid receptors following administration of the kappa antagonist norbinaltorphimine (NOR-BNI) increased the activity of TIDA neurons suggesting that these neurons are tonically inhibited by endogenous kappa opioids. By contrast, NOR-BNI had no effect on TIDA neuronal activity in gonadally-intact diestrous female rats, but increased the activity of these neurons in ovariectomized female rats. These results suggest that ovarian hormones block the inhibitory effects of endogenous kappa opioids on the activity of TIDA neurons. Activation of kappa opioid receptors following administration of the kappa agonist U-50,488 caused a dose-related decrease in TIDA neuronal activity in diestrous female rats. U-50,488 had no effect on TIDA neuronal activity in gonadally-intact male rats, but decreased the activity of these neurons in orchidectomized male rats. Taken together, these results reveal a sexual difference in the responsiveness of TIDA neurons to kappa opioid receptor agonists and antagonists, and suggest that gonadal steroid-induced gender differences in the basal activity of TIDA neurons may be due, in part, to differences in tonic inhibitory regulation of these neurons by endogenous kappa opioids.


Coordinative mineralocorticoid and glucocorticoid receptor-mediated control of responses to serotonin in rat hippocampus. In a previous study we showed that selective occupation of the mineralocorticoid receptor (MR) in hippocampal slices from adrenalectomized (ADX) rats attenuates the membrane hyperpolarization and resistance decrease induced in CA1 pyramidal neurons by serotonin (5HT). In the present study we established responses to 5HT in the hippocampal slice when not only MRs but also glucocorticoid receptors (GRs) were occupied, using either a combination of selective MR and GR ligands or different concentrations of the endogenous mixed agonist corticosterone. We observed that the GR agonist RU 28362 blocks the attenuating action of the MR agonist aldosterone on responses to 3, 10 and 30 microM 5HT; RU 28362 by itself did not affect 5HT responses. If a low concentration of the mixed agonist corticosterone (0.5 nM, close to the Kd for the MR) was continuously perfused in vitro, 5HT responses were steadily depressed with a delay of 2 h, while high levels of corticosterone (5 nM, around Kd for GR) only temporarily reduced 5HT responses. Finally, 5HT responses in slices from sham-operated rats (with relatively high plasma corticosterone levels) were similar to the responses obtained in slices from ADX rats. These data suggest that the previously reported MR-mediated attenuation of 5HT responses may be limited to conditions of low adrenocortical activity or pathophysiological conditions where the balance of MR- and GR-mediated effects is disturbed.


Autoradiographic evidence of opioid binding sites in rat growth plate chondrocytes. After 125I-beta-endorphin was intravenously injected in rats, an autoradiographic study of distal femur growth plates was performed. Results show a specific binding of the radiolabelled peptide to chondrocytes suggesting the possible existence of an opiate modulation of growth plate physiology.


Alteration in the immunoreactivity of the calcineurin subunits after ischemic hippocampal damage. Dephosphorylation processes of target proteins are critical to the reversible regulation of intracellular signal transduction systems. Further, brain damage such as ischemic insult induces marked changes in protein kinase activity. To study these changes more thoroughly, specific monoclonal antibodies of the A and B subunits of calcineurin (protein phosphatase 2B) were raised, and regional alterations in the immunoreactivity of calcineurin in the rat hippocampus were investigated after a transient forebrain ischemic insult causing selective and delayed hippocampal CA1 pyramidal cell damage. In normal rats it was found that both the calcineurin A and the B subunits showed high immunoreactivity in the dendritic fields of the hippocampal formation. The immunoreactivity of subunit A in the strata oriens, the radiatum of the CA1 subfield and in the stratum lucidum of the CA3 subfield was most intense, whereas the immunoreactivity in the other CA3 subfields and in the dentate gyrus was relatively low. In contrast, the dendritic fields of the hippocampal formation were equally immunoreactive to calcineurin subunit B, although the stratum lucidum of the CA3, where the mossy fibers from the dentate granule cells terminate, showed a very high immunoreactivity of the B subunit. After transient forebrain ischemia in the CA1 subfield, where selective pyramidal cell death occurred two days after this ischemia, a marked loss of immunoreactivity in both subunits was observed, along with morphological pyramidal cell damage. A recovery of the immunoreactivity of A and B subunits in the strata oriens and radiatum was later noted 30 days after ischemia. In the stratum lucidum of the CA3, the immunoreactivity of both the A and B subunits was transiently depressed from 6 to 24 h, followed by a marked immunoreactivity enhancement from four to 30 days after ischemia. Further, in the histologically intact dentate gyrus, both the immunoreactivity of the A and B subunits in the molecular layer were transiently enhanced from four to 14 days after ischemia, particularly in the supragranular layer. The results clearly indicate that the protein dephosphorylation systems were markedly altered in the whole hippocampal formation during the recirculation period following ischemia. Further, the transient depression in the calcineurin immunoreactivity seen in the mossy fiber terminals may reflect modulated synaptic activity of the dentate granule cells, which may play a pivotal role in the delayed and selective death of the CA1 pyramidal cells. Thus, calcineurin appears to be an excellent marker enzyme for the detection of neuronal activity and synaptic plasticity after brain damage, such as an ischemic insult.


Involvement of tyrosyl residue(s) in binding of endothelin and sarafotoxin to their receptors in rat brain and heart. The possible involvement of tyrosyl residue(s) in the binding of endothelins and sarafotoxins to their receptors was examined by the use of tetranitromethane. Incubation of rat cerebellar, hypothalamic, caudate putamen and atrial membranes with the reagent (100 microM, pH 8.1, 20 min) was accompanied by a decrease in their capacity to bind endothelins and sarafotoxin. Experiments employing nitration at different pH values (6.0 vs. 8.1) and studies on the effect of dithiothreitol on the nitrated preparations indicated that the modified residue(s) is most probably a tyrosyl and not a cysteinyl residue, and that there is no oligomerization of the receptors. These data are discussed in relation to the structural data recently obtained by cloning and expression of the endothelin/sarafotoxin receptors.


Carboplatin/etoposide in small cell lung cancer. Carboplatin/etoposide is an active combination in small cell lung cancer. In phase II studies, it produces results that appear equivalent to cisplatin/etoposide, but it has not been compared in a randomized study. It has a better toxicity profile than cisplatin/etoposide when compared in non-small cell lung cancer. The combination of carboplatin/etoposide is very well tolerated by elderly patients. Carboplatin/etoposide lacks important nonhematologic side effects, which has led to its assessment in dose escalation studies with colony-stimulating factors and autologous bone marrow transplantation.


Combined modality therapy for small cell lung cancer. Small cell carcinoma is one of the solid cancers for which chemotherapy is effective and can prolong the survival of patients. There is a general agreement that the standard regimen is PVP (cisplatin/etoposide) or alternating PVP-CAV (cyclophosphamide/doxorubicin/vincristine) therapy, replacing CAV, which had been considered standard. For the improvement of the treatment results of small cell carcinoma combined modality therapy including radiotherapy and/or surgery is essential, especially against limited disease. The effect of prophylactic cranial irradiation is obscure, because of the relapse to other sites of organs and the late complications of brain radiation. The present status of combined modality therapy for small cell carcinoma is reviewed and the future essential trials are discussed.


Carboplatin/etoposide/vincristine therapy in small cell lung cancer. Carboplatin is one of the most active agents in untreated small cell lung cancer (SCLC; 14% complete response, CR; and 61% CR + partial response, PR). The combination carboplatin/etoposide/vincristine (CEV) (phase II trial) led to an overall remission rate of 84% in patients with limited disease, with 52% CR. The median survival time with this combination was 13 months in patients with limited disease and 9.5 months in those with extensive disease. The 4-year survival rates are 26% in limited disease and 8% in extensive disease, with a plateau of the survival curve. This regimen is highly effective and exhibits low toxicity in SCLC. To evaluate the role of carboplatin in combination chemotherapy in patients with extensive SCLC, a phase III trial was performed. In this ongoing trial comparing CEV and etoposide/vincristine in SCLC patients with extensive disease, CR and overall response rates are higher in the CEV arm (CR 32 vs. 17%, CR + PR 80 vs. 60%), with statistically significant difference. In summary, chemotherapy regimens containing platinum compounds are among the most active in the treatment of SCLC. The use of the new compound carboplatin instead of cisplatin has led to similar or increased remission rates and is preferable because it has fewer side effects. Preliminary results from this ongoing, prospective, randomized phase III trial will be presented.


Prolonged administration of oral etoposide plus cisplatin in extensive stage small cell lung cancer. Etoposide is a highly schedule-dependent agent. We previously reported that a 21-day schedule of oral etoposide had good activity in small cell lung cancer (SCLC). The current phase II study was designed to test the combination of 21-day oral etoposide with cisplatin in hopes of capitalizing on etoposide's schedule dependency. Sixteen extensive stage SCLC patients were treated with cisplatin 100 mg/m2 day 1 plus 21 days of low-dose oral etoposide 50 mg/m2/day. Chemotherapy was repeated every 28 days for 4 cycles. Blood counts were monitored weekly, and etoposide was discontinued if the leukocyte or platelet count dropped below 2.0 x 10(9)/l or 75 x 10(9)/l, respectively. Fifteen of 16 patients were evaluable for response; 13 achieved either a complete (13%) or partial response (73%), for an overall response rate of 86% (95% confidence interval, 62-93%). Median response duration was approximately 7 months; median survival was not reached. Thirteen patients (81%) received all the planned cycles of chemotherapy. In cycle 1 of chemotherapy, the median leukocyte nadir was 2.8 x 10(9)/l (range, 0.1-6.3 x 10(9)/l; median platelet nadir was 180 x 10(9)/l (range, 51-397 x 10(9)/l). Life-threatening leukopenia (less than 1.0 x 10(9)/l) was unusual (2 of 58 cycles). There was 1 treatment-related death. One patient developed mild renal insufficiency that resolved after therapy. Nonhematologic toxicities were uncommon, but alopecia occurred in all patients. These data do not suggest that a major survival benefit will be derived for patients with extensive stage SCLC by increasing the duration of etoposide administration when used in combination with cisplatin. A randomized study is needed to determine if this long-term schedule of etoposide plus cisplatin is superior to the standard schedule of etoposide plus cisplatin.


Platinum/etoposide therapy in non-small cell lung cancer. Non-small cell lung cancer (NSCLC) is a major health problem in the United States and in many other parts of the world. Identification of systemic therapy that can maintain or improve quality of life and prolong survival is essential because locally advanced disease or distant metastasis is found in the majority of these patients. A variety of combination regimens have produced response rates of 20% or greater in stage IV NSCLC patients in cooperative group trials, including studies conducted within the Eastern Cooperative Oncology Group (ECOG). Although no regimen has emerged as clearly superior, the highest 1-year survival rate (25%) in ECOG trials was observed in EP(etoposide/cisplatin)-treated patients. Subsequently, a randomized trial in which EP was compared to etoposide/carboplatin revealed no significant differences in survival or response rate, and toxicity was slightly less with etoposide/carboplatin. Results from recent phase II trials in which stage III NSCLC patients received preoperative EP and concurrent thoracic irradiation have shown response rates of 65-85%, as well as acceptable toxicity and surgical morbidity and mortality. Future stage IV NSCLC trials should include studies of platinum/etoposide plus new agents. Also a study comparing platinum/etoposide versus supportive care should be considered. The objectives of this study would be to compare survival, quality of life, and cost-effectiveness ratio. The results of recent phase II studies in stage III NSCLC patients suggest that preoperative cisplatin/etoposide and concurrent thoracic irradiation should be considered for evaluation in a phase III trial in stage IIIa NSCLC patients and that this regimen should also be evaluated in stage IIIb patients.


Cisplatin/carboplatin therapy in non-small cell lung cancer. Based on in vitro experiments suggesting cytotoxic activity of at least a supra-additive effect, a phase I clinical study of carboplatin/cisplatin sequential combination treatment was carried out in 17 patients with non-small cell lung cancer (NSCLC). At 300 mg/m2 carboplatin and 80 mg/m2 cisplatin, the median leukocyte nadir was 2.6 x 10(9)/l; half the patients exhibited grade 3 or higher leukopenia. When cisplatin was increased to 100 mg/m2 the leukocyte nadir fell to 0.7 x 10(9)/l and grade 3 or 4 leukopenia was universal. A phase II pilot study of carboplatin 300 mg/m2 followed by 80 mg/m2 cisplatin was subsequently initiated in patients with NSCLC. Currently, 11 patients have entered the study and 8 have been added from the phase I study. Interim analysis shows that 8 patients have had at least a partial response to cisplatin/carboplatin, for a response rate of 42%. The response rate in previously untreated patients was 47%. At this time, 12 patients are alive but median survival has not yet been reached. No particularly serious adverse effects were observed. Four to five repeat administrations are planned in responders. Future studies with the cisplatin/carboplatin combination in NSCLC are warranted.


Carboplatin plus oral etoposide in the management of unresectable non-small cell lung cancer. Preliminary results of a Vanderbilt trial. Forty-two patients with unresectable non-small cell lung cancer (NSCLC) were treated with carboplatin (300, 350 or 400 mg/m2) and 21 days of oral etoposide (50 mg/m2/day). Thirty-three patients were evaluable for response (too early to assess the remaining patients) and 9 patients (27%) achieved a partial remission. Median survival was 29 weeks (range, 4+ to 71+ weeks). The primary toxicity was myelosuppression. Leukocyte and platelet count nadirs occurred between days 22 and 29. In cycle 1, median leukocyte nadir was 2.8 x 10(9)/l and the platelet nadir was 142 x 10(9)/l. Nonhematologic toxicities were modest and included alopecia in all patients, mild nausea, mild to moderate diarrhea, and an occasional increase in serum creatinine. Carboplatin plus oral etoposide is a tolerable outpatient regimen with modest activity against unresectable NSCLC.