Expression of HER-2/neu oncoprotein in human breast cancer: a comparison of immunohistochemical and western blot techniques. Three hundred and one primary breast cancers from patients with tumor infiltrated lymph nodes were analyzed for the presence of HER-2/neu oncoprotein by two procedures: Western blot (WB) and immunohistochemistry (IHC). Overexpression of this protein was found by WB in 16.6% of the tumors, and by IHC in 16.3%. Concordance between the two methods was found in 95% of tumors (286/301). In 7 cases we found HER-2/neu by IHC but not by WB, while the opposite was found in the remaining 8 patients. This discrepancy was found mainly in samples with HER-2/neu values just above the cut points and were therefore close to the sensitivity limits of the procedures used here. This study helps to define the parameters that should be considered to evaluate the immunostaining for HER-2/neu as positive (i.e., membrane staining, IHC score of 2 or more). The results obtained by both techniques were correlated with several currently used prognostic factors. Higher HER-2/neu protein expression was found in tumors lacking estrogen or progesterone receptors, in tumors with high S-phase fraction and in patients with more than 3 positive lymph nodes. In contrast, no relationship was found between overexpression of this protein and tumor size, ploidy, or age of the patient. Patients with elevated HER-2/neu expression showed a significantly worse overall survival by both methods, IHC (p = 0.05) and WB (p = 0.001). In conclusion, there is very high agreement between IHC and WB when measuring expression of HER-2/neu and both techniques showed prognostic significance.


Immunoenzymatic assay of erbB2 protein in cancer and non-malignant breast tissue. Relationships with clinical and biochemical parameters. The erbB2-encoded protein p185 was determined in 130 breast cancer specimens and in 29 non-malignant breast tissues using a recently available ELISA method. The assay showed good characteristics of precision and accuracy. In the non-malignant tissue p185 concentrations were normally distributed and directly correlated with estrogen receptors (ER) and progesterone receptors (PR). In cancer tissue p185 showed higher concentrations than in non-malignant tissue. No relationships were found between p185 and the main clinical and pathological parameters, except that a direct association with nuclear grade was found. We have categorized the breast cancer samples according to p185 concentrations as p185-negative (concentrations lower than the higher non-malignant tissue) and p185-positive. Our data suggested a different behaviour of p185 in samples with low or high p185 concentrations. Indeed, in p185-negative samples the concentrations were directly correlated with both ER and PR. Conversely, p185-positive samples were directly associated with node status and pT, and inversely associated with ER and PR.


Presence of p53 mutation in human cervical carcinomas associated with HPV-33 infection. In this present study, we report the mutation of the p53 gene in vivo in human primary carcinomas of cervix and cervical intraepithelial neoplasia (CIN). The association of the HPV subtypes with the tumors was determined by multiplex primer polymerase chain reaction (PCR) amplification. The mutation of the p53 gene was detected using PCR amplification of the p53 exons followed by SSCP (single strand conformation polymorphism) and DNA sequencing analysis. The p53 mutation was detected in two out of two HPV-33 positive carcinomas but was absent in the HPV-16/-18 positive carcinomas (0 out of 8 cases). The p53 mutation was also detected in one out of four HPV-negative cervical carcinomas. No mutation of the p53 gene was detected in the CIN specimens (0 out of 7 cases). The two mutations in the HPV-33 associated cervical carcinoma were detected at codon 273 (CGT to TGT; arginine to cysteine) and intron 5 (24 base pair downstream of the 3' end of exon 5). The p53 mutation at codon 273 has been previously reported in one of the HPV-negative cervical carcinoma cell line (C33A). Our results indicate that mutation of the p53 gene is not a common event in human cervical cancers (3/14), and may be related to the infection of HPV-16/18 in the tumor. However, mutation of the p53 gene was detected in cervical carcinomas associated with HPV-33 and may be an important genetic event in this subgroup of carcinomas.


Expression of p53 gene in B-CLL peripheral lymphocytes. The p53 tumor suppressor gene expression has been studied in 23 B-CLL cases at different clinical stages. The analysis failed to show a direct correlation with each stage, but the significantly lower frequency of a BglII RFLP in the pathologic population suggests a role of this gene in B-CLL. Northern Blot analysis showed the expression of p53 mRNA in all the B-CLL cases. A protocol for the RT-PCR methods was set up to study a very small amount of materials which should be better used for sequence analysis.


Molecular cloning and characterization of gravity specific cDNA in rice (Oryza sativa L.) suspension callus. Rice (Oryza sativa L. var. Nipponbare) suspension callus was exposed to gravity stress at 450,000 g for 2 hours, after which poly(A)+RNA was isolated and a cDNA library was constructed. Three different gravity specific cDNAs, namely, GSC 128, GSC 233 and GSC 381 of 0.67, 0.60 and 0.68 kilobase pairs and transcripts of 1.9, 1.6 and 2.0 kb, respectively, were isolated by differential screening and Northern hybridization. The maximum level of transcript was achieved after 4 hours of exposure to gravity at 450,000 g for GSC 128, 2 hours for GSC 233 and 8 hours for GSC 381 followed by a gradual decrease to undetectable levels with the extension of gravitation time. Callus (GSC 128), shoot and callus (GSC 381) and root and callus (GSC 233) specific expression of transcripts was identified. Although the protection of callus by treatment with ABA, kinetin and sucrose extended the period of expression of mRNA in suspension callus after gravity exposure, the expression of gravity-inducible mRNA was exclusively regulated by the degree of callus viability or survival after the stress. In addition, we demonstrated that the level of GSC 381 transcript was markedly increased by exposing the cell to periodical gravity stress, suggesting that this mRNA is expressed and translated into special proteins which are closely related to the survival of the cell against gravity stress. The sequence of GSC 233 and GSC 381, consisting of 417 and 531 base pairs of the longest open reading frames, encode polypeptides with calculated molecular weights of 15.29 and 19.47 kDa, respectively. A sequence homology search against a data bank revealed that GSC 233 and GSC 381 differed from other stress inducible genes in terms of the coding sequence and expression characteristics.


Nerve-dependent expression of c-myc protein during forelimb regeneration of Xenopus laevis froglets. The consequences of denervation on the expression of c-myc protein have been analyzed on the regenerating forelimb of young froglets of Xenopus laevis. The level of c-myc expression, low in control limbs and enhanced in the regenerate, is transiently increased after a three-hour total denervation. For this protein, the level of expression is not a function of the quantity of nerve in the regenerate. Four days after denervation, c-myc signal is back to its base level observed in the regenerate. A different pattern of expression is obtained for an S phase marker (PCNA protein) taken as a control in the same experimental conditions. The data presented here show that the nervous system normally exerts a negative control on the expression of c-myc and PCNA proteins in the limb regenerate of Xenopus.


Expression of c-erbB-2 gene in human head and neck carcinoma. The c-erbB-2 oncoprotein is a transmembrane protein the presence of which has been associated with poor prognosis in several human neoplasms. However, there has been no comprehensive assessment of its value as a potential prognostic marker in head and neck squamous cell carcinoma. Archival specimens from 93 patients, treated surgically for squamous cell carcinoma of the head and neck between 1981 and 1989, were analyzed by immunohistochemistry using an anti-c-erbB-2 monoclonal antibody; of these, 43 (46%) were positive for c-erbB-2 staining. The majority of stained specimens (41%) displayed staining predominantly at the cell surface, while mixed membrane and cytoplasmic staining was less common (9%). Only 4% shared exclusively cytoplasmic staining. Since the specimens were archival, the cytoplasmic staining is probably a consequence of variable handling and/or fixation at the time of tissue removal. Therefore, only cases exhibiting distinct cell surface membrane staining in more than 10% of tumor cells were regarded as positive. There is a definite association between immunohistochemical detection of c-erbB-2 and head and neck squamous cell carcinoma, since almost half of the tumor specimens manifested detectable c-erbB-2 protein. However, this association could not be extended to a predicted disease progression or outcome, since there was no significant correlation between c-erbB-2 staining and tumor size, stage of disease, histologic differentiation, lymph node status or patient survival.


Differential expression of the mouse and human Thy-1 gene in embryonal carcinoma cells. Mouse P19 embryonal carcinoma (EC) cells express on their surfaces a Thy-1 glycoprotein. The expression of Thy-1 at the mRNA and protein levels is down-regulated during differentiation induced by retinoic acid (RA). Thy-1 is also expressed in human NTERA-2 EC cells, but its expression is not down-regulated during RA-induced differentiation. As a first step towards understanding differential regulation of the mouse and human Thy-1 gene in EC cells, we have introduced genomic DNA fragments encompassing the mouse or human Thy-1 gene into NTERA-2 and P19-derived cells and analyzed surface properties of the transfectants. In the transient transfection assay, both mouse and human Thy-1 genes were expressed on cell surfaces at comparable levels. P19-derived stable transfectants exhibited great clonal variations in the expressions of the transfected Thy-1 gene products, which in part reflected copy numbers. There was no simple correlation between the expression of the transfected Thy-1 gene and two stem cell surface markers, TEC-1 and TEC-4. In the course of differentiation induced by RA several clones with a surface phenotype of EC cells exhibited a significant decrease in the expression of the transfected mouse Thy-1, whereas expression of the human Thy-1 was less efficiently down-regulated. The results suggest the presence of multiple cis- and trans-acting elements controlling expression of the mouse and human Thy-1 genes in P19 EC cells and their differentiated derivatives.


Increase of vinblastine accumulation by inhibitors of calmodulin-dependent cell functions in rat ascites hepatoma AH66 cells. ML-9, an inhibitor for myosin light chain kinase, and W-7, a calmodulin inhibitor, suppressed the efflux of vinblastine and increased the intracellular accumulation of vinblastine, but W-5, an inactive compound for calmodulin, did not so in rat ascites hepatoma AH66 cells, which have a multidrug-resistant phenotype. In sensitive counterpart AH66F cells, W-7 and ML-9 were less effective. W-7 and ML-9 did not interfere with [3H]azidopine photolabeling of P-glycoprotein in the plasma membrane from AH66 cells. While P-glycoprotein is reported to be superphosphorylated by protein kinases, W-7 did not influence the phosphorylation of the P-glycoprotein in AH66 cells. There may be an unknown Ca(2+)-calmodulin-dependent mechanism in the extrusion of vinblastine from AH66 cells.


[A preliminary study of immuno-electron microscopy of endothelium in cardiovascular system]. The immunocytochemical localizations of the von Willebrand factor (vWF), endothelin-1 (ET-1) and the beta 2-adrenergic receptor (beta 2R) in cultured human umbilical vein endothelial cells (HUVEC) after thrombin treatment have been demonstrated. Competitive ELISA analysis of these treated cells indicate an increase in vWF deposition on the subendothelial matrix (1.5 folds vs control) and vWF release (3.8 folds vs control) in the culture medium. The vWF-peroxidase, immunogold labeling and in situ hybridization of ET-1 were demonstrated on HUVEC. Double labelling with different sized gold particles for the ET-1 and beta 2R locations were performed on the hearts of young adult WEY, SHR, and SHRsp rats. The results were as follows. (1) Positive vWF deposits were detected on the extracellular matrix, Weibel-Palade bodies and secreting vesicles in the cytoplasm. It is suggested that both the constituted and regulated pathways are involved in the secretion of vWF stimulated by thrombin. (2) The gene expression of ET-1 was detected by light microscopic in situ hybridization in HUVEC. (3) ET-1 was detected by immunoelectron microscopy in the lysosome, cell surface, and extracellular matrix, suggesting that lysosome possibly plays an important role in the processing of ET-1 in HUVEC. (4) The presence of both ET-1 and beta 2R have been successfully detected in adult heart tissues and the endothelial cells of rats. These observations suggest the functional importance of ET-1 and beta 2R.


Lack of reversal of daunorubicin resistance in HL60/AR cells by cyclosporin A. Cyclosporin A and verapamil are substrates for P-glycoprotein. Both agents are known to reverse multidrug resistance in cells overexpressing P-glycoprotein. In this investigation, we have examined the effects of cyclosporin A and verapamil on multidrug resistance in HL60/AR cells that lack P-glycoprotein. In addition, a correlation was sought between an alteration in plasma membrane potential as measured with cationic dye DIOC5 and overexpression of P-glycoprotein. HL60/AR cells accumulated 3 fold less daunorubicin than HL60 cells. The drug accumulation defect and drug resistance in HL60/AR cells were partially corrected by verapamil and buthionine sulfoximine. However, cyclosporin A had no detectable effect on daunorubicin accumulation or drug resistance in HL60/AR cells. The multidrug resistant P338/ADR cell line overexpressed P-glycoprotein and exhibited depolarization of plasma membrane when compared to its corresponding drug sensitive parental cell line. In contrast, HL60/AR cells lacked P-glycoprotein and plasma membrane potentials were similar to those of drug sensitive HL60 cells. These results suggest that [1] verapamil modulates daunorubicin transport by a mechanism independent of P-glycoprotein, [2] the mechanisms of reversal of multidrug resistance by verapamil and cyclosporin A are distinct, and [3] the plasma membrane depolarization in multidrug resistant cell lines that overexpress P-glycoprotein, as determined by DIOC5, may be due to an increased efflux of cationic dye by P-glycoprotein, rather than a true measurement of plasma membrane potential in multidrug resistant cells.


[Using 131I-MIBG and 99mTc-MDP bone scan for localization of rare extra-adrenal pheochromocytomas: report of 2 cases]. It is difficult, but important, to diagnose extra-adrenal pheochromocytomas before surgery. Failure to recognize the existence and nature of an extra-adrenal pheochromocytoma can cause life-threatening problems even in a minor surgical operation. We present two rare cases of extra-adrenal pheochromocytomas which were detected by 131I-MIBG scintigraphy. One of them was intrapericardial, and the other was a vesical pheochromocytoma. We used a combined 99mTc-MDP bone scan and 131I-MIBG scintigraphy to locate the pheochromocytomas. Both cases of extra-adrenal pheochromocytoma were correctly diagnosed preoperatively and successfully resected in the subsequent operations.


Human lung cancers growing on extracellular matrix: expression of oncogenes and growth factors. The goal of this study was to evaluate the extracellular matrix (ECM) as a model for growing human lung cancers and to study the feasibility of its application for cellular and molecular studies of tumor biology. Bovine corneal endothelial cell ECM coated dishes were evaluated as a growth substrate for tumor cultures. Growth success, morphology and oncoprotein/growth factor expression for 74 different lung cancers (adenocarcinoma, epidermoid carcinoma and small cell carcinoma) were compared after seeding fresh surgical explants onto bovine corneal endothelial cell ECM and plastic culture substrate. Nineteen out of 74 tumors (26%) plated on ECM demonstrated measurable growth. Growth on ECM was superior to growth on plastic for the lung tumors. All 19 tumor cultures showed malignant morphology and functions. They were examined under the light microscope, and in all cases pre- and post-cytology confirmed malignancy. Tumor cells seeded on ECM retained their malignant phenotype in comparison to tumors grown on plastic. Several oncoproteins (c-myc, c-Ha-ras, c-erbB-2) and growth factors/receptors (EGF, EGF-R, TGF alpha) were immunostained. These analyses were performed immediately after disaggregation of tumor cells obtained surgically and after seeding on ECM or plastic. Strong expression of oncoproteins/growth factors was detected in tumor cells immediately after surgery or when the cells were plated on ECM. On the other hand, moderate or no expression was observed in the same type of cells on plastic.


[Blood pressure and metabolic response to converting enzyme inhibitor in hypertensive patients: comparison between delapril and captopril]. The antihypertensive and metabolic responses to delapril and captopril in hypertensive patients were studied. Forty-six hypertensive patients entered the study and were divided into two groups. The delapril group included 21 essential hypertensive and five renoparenchymal hypertensive patients; while the captopril group included 11 essential hypertensive and nine renoparenchymal hypertensive patients. The patients in the delapril group took delapril 7.5 mg twice a day for 2 weeks. If the antihypertensive effect was inadequate, the dose was increased to 15mg twice a day, and then to 30mg twice a day. The period of delapril treatment was 12 weeks. The patients in the captopril group took captopril 25 mg twice a day or three times a day for 12 weeks. After delapril treatment, there were significant decreases in the systolic (from 163 +/- 17 to 141 +/- 15 mmHg) and diastolic blood pressure (from 105 +/- 13 to 91 +/- 10 mmHg). There were also significant decreases in the systolic (from 161 +/- 18 to 141 +/- 24 mmHg) and diastolic blood pressure (from 100 +/- 10 to 90 +/- 12 mmHg) after captopril treatment (p < 0.001). The pulse rates in both groups showed no significant changes after treatment. The laboratory data in both groups showed few changes after treatment. Plasma renin activity increased after delapril treatment. A cough was the side effect most commonly seen. We conclude that hypertensive patients have the same blood pressure and metabolic responses to delapril as captopril.


Expression of protein kinase C in human renal cell carcinoma cells with inherent resistance to doxorubicin. The expression of protein kinase C (PKC) was analyzed in 18 primary cell cultures of human renal cell carcinomas by means of immunocytochemistry. We found that a high PKC expression significantly correlates with both resistance to doxorubicin and high P-glycoprotein expression. These data support the hypothesis that PKC is involved in inherent drug-resistance by phosphorylation and regulation of P-glycoprotein.


Energy metabolism and effects of energy depletion or exposure to glutamate. The entire program of the first day of the IBRO satellite meeting entitled Ions, Water, and Energy in Brain Cells was devoted to the subject of energy. There were three sessions on the topics of energy metabolism, activation, and development and pathological conditions, followed by a final general discussion on the contents of the day's topics. During this general discussion there were spirited exchanges on the role of glycogen in the energy metabolism of the brain, on the metabolic source of the energy consumed by functional activity, e.g., glycolytic or oxidative energy metabolism, and on the sources of the acid-equivalents that are responsible for the tissue acidosis accompanying cerebral hypoxia. Despite the arguments pro and con presented on all of the issues that were discussed, it is doubtful that a consensus was achieved on most of the issues.


Ca2+ and filopodial responses to glutamate in cultured astrocytes and neurons. Neurons and glia exhibit complex homeostatic interactions via shared extracellular space which can involve metabolites, inorganic ions, and neurotransmitters. Focal application of glutamate to both human and rat central nervous system astrocytes in primary culture produced a rapid, transient increase in both cytoplasmic and nuclear Ca2+. These Ca2+ waves can propagate at up to 15-20 micron/s for long distances (millimetres) through the astrocyte syncitium. Oscillatory Ca2+ signals were frequently observed under control conditions and were enhanced by glutamate application. These Ca2+ signals were paralleled by rapid extensions of filopodia from the astrocyte cell margin and apical surface near the point of glutamate application. Focal application of glutamate to rat hippocampal neurons also elicited rapid, transient increases in intracellular Ca2+. Levels of Ca2+ signals were consistently two- to three-fold greater in pyramidal neurons cultured from CA1 than in those from CA3. Filopodial extension was extensive in CA1 neurons, but rare in CA3 neurons, and in either case observable only during the first few days of primary culture. Diversity of glial and neuronal responses to binding the glutamate receptors may reflect their roles in homeostatic interactions.


The regulation of pH in the central nervous system. The pHi regulation from intracellular acidosis in the central nervous system appears to be mediated by mechanisms driven by the large inwardly directed Na+ gradient. The involvement of these mechanisms in pHi regulation of neurones and glial cells has been investigated in the leech central nervous system using ion-selective microelectrodes. For recovery from acidification, there appear to be three separate mechanisms: Na+/H+ exchange, Na(+)-dependent Cl-/HCO3- exchange, and Na+-HCO3- cotransport. All three mechanisms have a profound effect on the maintenance of pHi homeostasis in glial cells; whereas in leech neurones, as in other neuronal cells studied previously, the predominant mechanisms are Na+/H+ and Na(+)-dependent Cl-/HCO3- exchange. In addition to acid extrusion mechanisms we also found evidence for Na(+)-independent Cl-/HCO3- exchange. At alkaline pHi this exchanger may mediate some of the pHi recovery from intracellular alkalinization.


Alkaline extracellular pH shifts generated by two transmitter-dependent mechanisms. Recent studies of the effect of gamma-aminobutyric acid (GABA) on brain extracellular pH are reviewed. Experiments were performed on isolated turtle cerebellum, using double-barrelled pH-sensitive microelectrodes. Superfusion of GABA (1 mM) caused a rapid extracellular alkaline shift accompanied by a rise in extracellular K+. Washout of GABA was often associated with an acid rebound, concomitant with an undershoot of extracellular K+. The GABA-evoked alkaline shift was blocked by picrotoxin and mimicked by the GABA-A agonists isoguvacine and muscimol. The response persisted in the nominal absence of extracellular calcium, but it was reversibly abolished in nominally bicarbonate free media. In contrast, extracellular alkaline shifts evoked by repetitive stimulation of the parallel fibers were amplified in bicarbonate-free media and were insensitive to picrotoxin. These results indicate the existence of separate, transmitter-dependent mechanisms of extracellular alkalinization: (i) a GABA-A receptor mediated process, most likely associated with efflux of bicarbonate ions across GABA-A anion channels and (ii) a bicarbonate-independent process associated with excitatory synaptic transmission.


Mechanisms of glial swelling induced by glutamate. The mechanisms of glutamate-induced glial swelling have been studied using an in vitro model that permits detection of cell volume changes with high accuracy. The model allows for a close control of the extracellular environment to study in isolation the effect of defined extracellular alterations occurring in brain under pathophysiologic conditions. Glutamate was applied in concentrations between 50 microM and 10 mM to either C6 glioma cells or astrocytes from primary culture. Glutamate uptake was assessed by HPLC measurements of amino acids in the extracellular medium. Glutamate at all concentrations tested caused glial swelling, which, however, was moderate, with maximal average volume increases between 5.0 +/- 1.92 and 18.38 +/- 1.6% of control at 50 microM and 5 mM glutamate, respectively. Swelling was concentration dependent and correlated with glutamate uptake. After removal of all extracellular glutamate by glial uptake, cell volume spontaneously normalized. Pretreatment of the cells for 90 min with ouabain (1 mM) to abolish the extracellular/intracellular Na+ gradient, prevented glutamate-induced swelling. It is concluded that while glial cells readily accumulate glutamate from the extracellular environment to protect neurons from excitotoxic effects, swelling results from the increase of intracellular osmotic activity due to the uptake of Na+ and glutamate.


Use of cell cultures to differentiate among effects of various ischemia factors on astrocytic cell volume. Mouse astrocytes were subjected to in vitro models of ischemia (hypoxia with or without substrate deprivation, excess potassium, or elevated glutamate). Three hours of hypoxia alone or with substrate deprivation had little effect upon the morphology of astrocytes but did cause disaggregation of polyribosomes. Excess (12-50 mM) potassium added (as KCl) to a normal isotonic medium also caused no swelling; it did, however, cause a shrinkage of cell volume. When 50 mM potassium was substituted for a similar amount of sodium, marked swelling occurred. Swelling of astrocytes was also seen after addition of glutamate (50 microM to 1 mM) to the culture medium. These results show that ischemia per se does not result in astrocytic swelling; rather, microenvironmental alterations such as rising glutamate levels and changes in the sodium/potassium ratios result in astrocytic swelling. We conclude that one can use astrocytes in culture to dissect out the mechanisms that cause postischemic alterations in astrocytes in vivo.


Drug therapy for chronic urticaria.

Drug therapy for chronic urticaria. Given the variability of patient problems, it is difficult to construct a single drug therapy regimen for treatment of chronic urticaria. However, the following regimen should prove to be a useful outline to follow for most cases. The first line of therapy will usually be antihistamines. In general, antihistamines should be always used on a regular basis and not only after hives occur. If drowsiness or anticholinergic adverse symptoms limit the use of one drug in effective doses, other H1-blockers should be tried. For day-time use, the newer, less sedating antihistamines are preferred. If antihistamines fail to control symptoms when used at full doses, addition of glucocorticosteroids can be tried for short periods. Most patients respond to doses equivalent to 40 mg of prednisone daily. The end point of use of corticosteroids is to reach quickly an effective low, alternate-day dose followed by their discontinuation.


Effect of calcium concentration on survival, proliferation and activities of alkaline phosphatase, 5'-nucleotidase, gamma-glutamyltransferase and lactate dehydrogenase of adult rat hepatocytes cultured in serum-free medium. Mature rat hepatocytes were cultured on collagen coated dishes in serum-free alpha-modified Eagle's minimum essential medium containing 0.1 microM insulin, 0.1 microM dexamethasone, 10 mM pyruvate and Ca2+ at concentrations of 0-2 mM. Survival of nondivided cells was best in medium containing 2 mM Ca2+. Proliferation during 5-day culture was greatest with 0.4 mM Ca2+, but DNA synthesis was scarcely affected by the concentration of Ca2+. Both the activities of alkaline phosphatase, 5'-nucleotidase, gamma-glutamyltransferase and lactate dehydrogenase and the number of cell nuclei of cultures in 0.1 mM and 2 mM Ca2+ media were assayed over a 5-day period, and their activities were calculated as enzyme activities per unit number of cell nuclei. Alkaline phosphatase activity increased rapidly during the first day of culture in both media, and its activity in 0.1 mM medium was higher than that in 2 mM medium after culture for 3 days. The activity of 5'-nucleotidase became higher in 0.1 mM medium than in 2 mM medium from day 2 and was maximal on day 3 in both media. gamma-Glutamyltransferase activity increased and lactate dehydrogenase activity decreased with time in culture, both activities showing no appreciable difference in the two media.


The influence of vasodilating beta-blockers on cardiac function and vascular resistance in essential hypertension. In nearly all forms of established hypertension, the cardinal hemodynamic disturbance is an increased total peripheral resistance, while cardiac output is abnormally low, particularly during exercise. When left untreated, total peripheral resistance increases, cardiac output falls, and blood pressure increases over time. The coronary reserve is reduced, and renal as well as cerebral resistance increases and blood flow falls. Antihypertensive agents effect central hemodynamics differently. Ordinary beta-blockers do usually not reduce total peripheral resistance much below pretreatment level, and cardiac output is chronically depressed, particularly during exercise. However, the beta-blockers greatly reduce the workload on the heart by decreasing the heart rate-pressure product. Modern beta-blockers with vasodilating activity--like carvedilol--are based on a combination of beta-blockade and vasodilatation. Such beta-blockers also induce a marked decrease in the pressure-heart rate product, and some reduction in total peripheral resistance. They cause less depression of exercise cardiac output than ordinary beta-blockers. Blood flow to the kidneys and the brain is maintained. From a theoretical point of view, this type of antihypertensive treatment should maintain good blood pressure control, reduce cardiac workload and be associated with less side-effects than ordinary beta-blockers.


Coordinate embryonic expression of three zebrafish engrailed genes. We have identified three genes, expressed in zebrafish embryos, that are members of the engrailed gene family. On the basis of sequence comparisons and analyses of their expression patterns, we suggest that two of these genes, eng2 and eng3, are closely related to the En-2 gene of other vertebrates. The third gene, eng1, is probably the zebrafish homolog of En-1. Subsets of cells at the developing junction between the midbrain and hindbrain express three different combinations of these genes, revealing a previously unknown complexity of this region of the CNS. Other cells, for example, jaw and myotomal muscle precursors, express two of the three genes in combinations which, in the myotomal muscles, change during development. Cells in the developing hindbrain and fins express only a single engrailed gene. We propose that the fates and patterning of these cells may be regulated by the coordinate expression of particular combinations of these closely related homeoproteins.


Autocatalysis and phenotypic expression of Drosophila homeotic gene Deformed: its dependence on polarity and homeotic gene function. Previously published experiments have shown that the endogenous Dfd gene can be ectopically activated by its own (heat-shock-driven) product in a subset of cells of different segments. This results in the differentiation of maxillary structures like cirri and mouth hooks in places where they normally do not appear, and represents a phenomenon of autocatalysis of homeotic gene function that differs from the normal activation process. We show that this out-of-context activation occurs in cells belonging to the anterior compartments of the three thoracic and the A1 to A8 abdominal segments and that it requires the normal function of the polarity genes wingless (wg) and engrailed (en). The wg product, in addition to that of Dfd, appears to be sufficient to activate the endogenous Dfd gene in many embryonic cells. We have studied the effect of several homeotic genes on Dfd activation and phenotypic expression: Scr, Antp, Ubx and Abd-B repress Dfd both transcriptionally and at the phenotypic level, if their products are in sufficient amounts. The endogenous abd-A gene does not have a noticeable effect, but when it is replaced by an hsp70-abd-A gene, which produces a high and uniform level of expression, the phenotypic expression of Dfd is suppressed. Our results also suggest that the differentiation of cirri is induced by Dfd-expressing cells in non-expressing neighboring cells, and that this interaction occurs across the parasegmental border.


Mox-1 and Mox-2 define a novel homeobox gene subfamily and are differentially expressed during early mesodermal patterning in mouse embryos. We have isolated two mouse genes, Mox-1 and Mox-2 that, by sequence, genomic structure and expression pattern, define a novel homeobox gene family probably involved in mesodermal regionalization and somitic differentiation. Mox-1 is genetically linked to the keratin and Hox-2 genes of chromosome 11, while Mox-2 maps to chromosome 12. At primitive streak stages (approximately 7.0 days post coitum), Mox-1 is expressed in mesoderm lying posterior of the future primordial head and heart. It is not expressed in neural tissue, ectoderm, or endoderm. Mox-1 expression may therefore define an extensive 'posterior' domain of embryonic mesoderm before, or at the earliest stages of, patterning of the mesoderm and neuroectoderm by the Hox cluster genes. Between 7.5 and 9.5 days post coitum, Mox-1 is expressed in presomitic mesoderm, epithelial and differentiating somites (dermatome, myotome and sclerotome) and in lateral plate mesoderm. In the body of midgestation embryos, Mox-1 signal is restricted to loose undifferentiated mesenchyme. Mox-1 signal is also prominent over the mesenchyme of the heart cushions and truncus arteriosus, which arises from epithelial-mesenchymal transformation and over a limited number of craniofacial foci of neural crest-derived mesenchyme that are associated with muscle attachment sites. The expression profile of Mox-2 is similar to, but different from, that of Mox-1. For example, Mox-2 is apparently not expressed before somites form, is then expressed over the entire epithelial somite, but during somitic differentiation, Mox-2 signal rapidly becomes restricted to sclerotomal derivatives. The expression patterns of these genes suggest regulatory roles for Mox-1 and Mox-2 in the initial anterior-posterior regionalization of vertebrate embryonic mesoderm and, in addition, in somite specification and differentiation.


Connectin, a target of homeotic gene control in Drosophila. The homeotic genes of Drosophila encode transcription factors that specify morphological differences between segments. To identify the genes that they control, we developed a chromatin immunopurification approach designed to isolate in vivo binding sites for the products of the homeotic gene Ultrabithorax. Here, we report the analysis of one immunopurified binding site. This 110 bp fragment maps within a regulatory region of a gene under homeotic control, connectin. A 4 kb DNA fragment, including the immunopurified binding site, is sufficient to reproduce the appropriate homeotic control within a subset of the full tissue distribution of connectin. Analysis of the role of the 110 bp binding site indicates that it mediates transcriptional controls by Ultrabithorax and other homeotic genes. This is the first report of a functional in vivo binding site isolated using the chromatin immunopurification method. We also show that the protein product of the connectin gene is predicted to be a cell-surface molecule containing leucine-rich repeats. The protein, connectin, can mediate cell-cell adhesion thus suggesting a direct link between homeotic gene function and processes of cell-cell recognition.


The 412 retrotransposon and the development of gonadal mesoderm in Drosophila. We have shown that the expression of the 412 retrotransposon provides a useful early marker for the development of the gonadal mesoderm in Drosophila embryos. 412 is initially expressed in a set of parasegmentally repeated stripes from parasegments (PS) 2-14 in the mesoderm at the extended germ band stage. During germ band retraction the bulk of 412 expression declines except in dorsolateral clusters of cells in PS10, 11 and 12, where high levels of 412 expression remain. These mesodermal cell clusters are associated with germ cells and subsequently they coalesce, rounding up to form the gonads. The gonadal mesoderm thus appears to originate specifically from three abdominal parasegments, PS10, 11 and 12. We show that the maintenance of high levels of 412 expression in gonadal mesoderm is not induced by contact with germ cells, but rather depends on genetic control by the homeotic genes abdominal-A and Abdominal-B.


Ectopic expression of UBX and ABD-B proteins during Drosophila embryogenesis: competition, not a functional hierarchy, explains phenotypic suppression. The Abdominal-B (Abd-B) gene, a member of the bithorax complex (BX-C), specifies the identities of parasegments (PS) 10-14 in Drosophila. Abd-B codes for two structurally related homeodomain proteins, ABD-B m and ABD-B r, that are expressed in PS10-13 and PS14-15, respectively. Although ABD-B m and r proteins have distinct developmental functions, ectopic expression of either protein during embryogenesis induces the development of filzkörper and associated spiracular hairs, structures normally located in PS13, at ectopic sites in the larval thorax and abdomen. These results suggest that other parasegmental differences contribute to the phenotype specified by ABD-B r activity in PS14. Both ABD-B m and r repress the expression of other homeotic genes, such as Ubx and abd-A, in PS10-14. However, the importance of these and other cross-regulatory interactions among homeotic genes has been questioned. Since ectopic UBX protein apparently failed to transform abdominal segments, González-Reyes et al. (González-Reyes, A., Urquía, N., Gehring, W.J., Struhl, G. and Morata, G. (1990). Nature 344, 78-80) proposed a functional hierarchy in which ABD-A and ABD-B activities override UBX activity. We tested this model by expressing UBX and ABD-B m proteins ectopically in wild-type and BX-C-deficient embryos. Ectopic ABD-B m does not prevent transformations induced by ectopic UBX. Instead, ectopic UBX and ABD-B m proteins compete for the specification of segmental identities in a dose-dependent fashion. Our results support a quantitative competition among the homeotic proteins rather than the existence of a strict functional hierarchy. Therefore, we suggest that cross-regulatory interactions are not irrelevant but are important for repressing the expression of competing homeotic proteins. To explain the apparent failure of ectopic UBX to transform the abdominal segments, we expressed UBX at different times during embryonic development. Our results show that ectopic UBX affects abdominal cuticular identities if expressed during early stages of embryogenesis. In later embryonic stages, abdominal segments become resistant to transformation by ectopic UBX while thoracic segments remain susceptible. Head segments also show a similar stage-dependent susceptibility to transformation by ectopic UBX in early embryogenesis but become resistant in later stages. These results suggest that abdominal and head identities are determined earlier than are thoracic identities.


Virulence factors in anaerobic bacteria. Various surface structures can be expressed in Bacteroides fragilis, but little is known about capsular structures in other non-spore-forming anaerobes. Fimbriae have been isolated from Bacteroides fragilis and Porphyromonas gingivalis. The importance of iron-repressible outer membrane proteins as virulence factors in Bacteroides fragilis is under study. The low endotoxic activity of Bacteroides fragilis lipopolysaccharide can be attributed to the chemical composition of this organism's lipid A. A tissue culture system for the demonstration of Bacteroides fragilis enterotoxin has recently been described. The toxins A and B of Clostridium difficile are immunologically distinct. The importance of IgA proteases and other enzymes as virulence factors in anaerobic bacteria remains unclear.


Inactivation of Bacillus subtilis glutamine synthetase by metal-catalyzed oxidation. Instability of Bacillus subtilis glutamine synthetase in crude extracts was attributed to site-specific oxidation by a mixed-function oxidation, and not to limited proteolysis by intracellular serine proteases (ISP). The crude extract from B. subtilis KN2, which is deficient in three intracellular proteases, inactivated glutamine synthetase similarly to the wild-type strain extract. To understand the structural basis of the functional change, oxidative modification of B. subtilis glutamine synthetase was studied utilizing a model system consisting of ascorbate, oxygen, and iron salts. The inactivation reaction appeared to be first order with respect to the concentration of unmodified enzyme. The loss of catalytic activity was proportional to the weakening of subunit interactions. B. subtilis glutamine synthetase was protected from oxidative modification by either 5 mM Mn2+ or 5 mM Mn2+ plus 5 mM ATP, but not by Mg2+. The CD-spectra and electron microscopic data showed that oxidative modification induced relatively subtle changes in the dodecameric enzyme molecules, but did not denature the protein. These limited changes are consistent with a site-specific free radical mechanism occurring at the metal binding site of the enzyme. Analytical data of the inactivated enzyme showed that loss of catalytic activity occurred faster than the appearance of carbonyl groups in amino acid side chains of the protein. In B. subtilis glutamine synthetase, the catalytic activity was highly sensitive to minute deviations of conformation in the dodecameric molecules and these subtle changes in the molecules could be regarded as markers for susceptibility to proteolysis.


Physiological roles of acetoacetyl-CoA thiolase in n-alkane-utilizable yeast, Candida tropicalis: possible contribution to alkane degradation and sterol biosynthesis. The presence of two types of thiolases, acetoacetyl-CoA thiolase and 3-ketoacyl-CoA thiolase, was demonstrated in peroxisomes of n-alkane-grown Candida tropicalis [Kurihara, T., Ueda, M., & Tanaka, A. (1989) J. Biochem. 106, 474-478], while acetoacetyl-CoA thiolase was also shown to be present in cytosol. The activity of the enzyme in cytosol was constant irrespective of culture conditions, while the peroxisomal enzyme was inducibly synthesized in the alkane-grown yeast cells. These results indicate that peroxisomal acetoacetyl-CoA thiolase participates in alkane degradation, while the cytosolic enzyme is associated with other fundamental metabolic processes, probably sterol biosynthesis, because this enzyme can catalyze the first step of the sterol biosynthesis. 3-Hydroxy-3-methylglutaryl (HMG)-CoA reductase, a key regulatory enzyme of sterol biosynthesis, was found to be localized exclusively in microsomes of the alkane-grown yeast cells. These results suggest that yeast peroxisomes do not contribute to sterol biosynthesis, unlike the case of mammalian cells.


Endothelium-derived relaxing factor contributes to the regulation of endothelial permeability. To determine whether endothelium-derived relaxing factor (EDRF) contributes to the regulation of endothelial permeability, the transendothelial flux of 14C-sucrose, a marker for the paracellular pathway across endothelial monolayers (Oliver, J. Cell. Physiol. 145:536-548, 1990), was examined in monolayers of bovine aortic endothelial cells grown on collagen-coated filters. The permeability coefficient of 14C-sucrose was significantly decreased by 10(-3) M 8-Bromoguanosine 3',5'-cyclic monophosphate or by 5 x 10(-6) M glyceryl trinitrate, an activator of soluble guanylate cyclase. Depletion of L-arginine from endothelial monolayers increased 14C-sucrose permeability from 3.21 +/- 0.59 to 3.88 +/- 0.50 x 10(-5) cm.sec-1 (mean +/- SEM; n = 6; P < 0.05). The acute administration of 5 x 10(-4) M L-arginine to monolayers depleted of this amino acid decreased 14C-sucrose permeability from 2.91 +/- 0.27 to 2.52 +/- 0.26 x 10(-5) cm.sec-1 (n = 11; P < 0.05). 14C-sucrose permeability was increased by 10(-7) M bradykinin and this effect was enhanced by the presence of each one of the following compounds: 10(-5) M methylene blue, 4 x 10(-6) M oxyhemoglobin, 5 x 10(-4) M NG-methyl-L-arginine or 5 x 10(-4) M N omega-nitro-L-arginine. These results suggest that EDRF contributes to the sealing of the endothelial monolayer and that EDRF released by bradykinin acts as a feedback inhibitor attenuating the increase in endothelial permeability induced by this peptide. Because endothelial cells have the ability to contract and relax and possess guanylate cyclase responsive to nitric oxide, our results suggest that EDRF decreases 14C-sucrose permeability by relaxing endothelial cells, thereby narrowing the width of endothelial junctions.


Lanthanum influx into cultured human keratinocytes: effect on calcium flux and terminal differentiation. Trivalent cation lanthanum (La) binds to calcium binding sites of cells and either mimics the properties of calcium or inhibits the effects of calcium by displacing calcium from its binding sites. Extracellular calcium induces differentiation of human epidermal keratinocytes in culture, in part by increasing the intracellular calcium levels (Cai). Therefore, in this study we determined the effect of La on differentiation and intracellular calcium levels of keratinocytes. We observed that La inhibited the production of cornified envelopes, a marker for terminal differentiation of keratinocytes. La inhibited the calcium requiring envelope cross-linking enzyme, transglutaminase, in a direct manner, presumably, by displacing calcium from its binding site on the enzyme. La inhibited the influx and the efflux of 45Ca from keratinocytes. Paradoxically, extracellular La appeared to increase the Cai levels of keratinocytes as measured by the fluorescent probe indo-1. However, subsequent experiments revealed that indo-1 bound La with a higher affinity than Ca and emitted fluorescence in the same wavelength as the Ca bound form. Using this probe, we observed that La enters keratinocytes in a dose-dependent fashion and achieves concentrations exceeding 80 nM when the external La concentration is raised to 300 microM. This fully accounted for the apparent increase in Cai when La was added to the cells. Treatment of cells with ionomycin increased indo-1 fluorescence maximally in the presence of La indicating influx of La via this Ca specific ionophore. Our results indicate that La enters cells and inhibits calcium mediated keratinocyte differentiation both by blocking Ca influx and by blocking calcium regulated intracellular processes such as transglutaminase directed cornified envelope formation.


[Surgery for pseudarthrosis of the scaphoid bone performed in Hungary 1988]. Authors analyse, based on an all-round inquiry, the operations performed for pseudarthrosis of the scaphoid in 1988. It was found that 344 of 472 operations were reconstructive, 128 palliative. The single types of operation are briefly described. They hint at the fact that the operative treatment of scaphoid pseudarthrosis in Hungary is carried out mostly in Centers.


[Anatomical bases of ultrasonic examination of the shoulder joint]. Authors present the parts of the shoulder that can be represented by ultrasound. Beside the rotator cuff of accentuated significance (within this, first of all the tendon of the supraspinatus muscle) the subacromial bursa, the tendon of the long head of the biceps, the muscle-tendon junction of the infraspinatus muscle are analysed. It is stated that the ultrasonography is suitable for a reliable representation of the soft tissues, surrounding the shoulder joint, further in possession of literary data they see possibility for a very exact judgement of the pathological alterations, too.


[Experience with the surgical management of childhood fractures and epiphyseal injuries]. Authors analyse 1792 fractures in children, treated during 10 years in their Department and distributed in age groups. They take the part of conservative treatment of the majority, about two third, of fractures in childhood. They consider as operative indication certain forms of epiphyseal injuries, the dislocated intraarticular fractures, the unstable fractures, certain cases of polytrauma and the open fractures.


[Stabilization of the injured acromioclavicular joint using a new type of fixation plate]. The opinions in the question of transitory fixation of the joint during the treatment of the acromioclavicular articulation are rather different. The complications of the widely used methods--break of the implant, its tear from the bone, the wandering and the disturbances of wound healing--are well known. To decrease the number of complications authors have developed a new type of hooked plate. The plate is fitted to the anterior surface of the acromial end of the clavicle and its hook is fixed from below in the bore hole of the acromion. To choose the optimal place of the bore hole a drill guide was prepared. The operative method is described and the advantages of the method are summarized. The "Debrecen-plate" was used during 2 years in 39 injuries. 35 patients were controlled 6-24 months after the operation. Based on the results of the treatment a wider use of the method is suggested.


[Overhead extension in delayed diagnosis of congenital hip dislocations in children, following failure of treatment with the Pavlik harness]. Authors report, based on a retrospective examination of 103 dislocated hips of 85 children, on experiences with the overhead treatment of congenital hip dislocations, resistant to Pavlik harness or lately recognized. The results are compared with those of 23 hips of 20 children treated with Pavlik harness and plaster casts. Follow-up time was 3-15, in average 7 years. It is stated that the overhead extension treatment not followed by stiff fixation resulted in 92 per cent reduction in resistant and lately recognized hip dislocations. This procedure has caused in only 4 per cent severe necrosis of the femoral head with lasting consequences. Their experience with Pavlik harness combined with plaster cast are unfavourable as in 4 of 23 hips treated subtotal, in another 7 partial femoral head necrosis has developed.


[Value of osteotomy of the proximal end of the metatarsal base in Köhler II disease]. Authors briefly describe the etiology of Köhler II. disease and the possibilities of the operative treatment. The osteotomy of the proximal end of the metatarsus, performed by them, the biomechanical basis of the operation, the indication are detailed and the favourable experiences gained with this procedure are reported.


[Measuring the looseness of the knee joint by using the Kt 1000 arthrometer]. Authors measured the stability in the sagittal plane of the intact knee joints of 50 young men with the MEDmetric KT 1000 apparatus. The greatest movement during the measurements on 100 knees was observed in 25 degrees flexion and during the application of force 89N. In 70 degrees flexion the movement was only half of that observed in 25 degrees flexion. The difference of the average movements measured bilaterally was neither in active nor in passive experiments more than 0.5 mm. The difference between the average values of the compliance indexes on the left and right side was only 0.01 mm. In all measurements the frequency distribution of the differences between the two sides was essentially the same. The difference between the movements on the two sides did not exceed 2 mm-s in 90-100 percent of the cases.


[Experience with the "Oxford" type uni-condylar meniscus- and surface replacing knee prosthesis. Preliminary report]. Authors report on the first implantation of a new type of an unicondylar meniscus and surface replacing knee prosthesis in this country. After a brief literary review the prosthesis, the instruments developed for the implantation, the indications and the early favourable results of their two operations are described.


[Enhancement of the ossification of decalcified bone]. Authors have proven with previous experiments that the decalcinated bone can be used as transplant. In this paper their experiments to increase the osteogene capacity of this kind of bone for replacement are described. It is stated that with the use of electric after treatment both the speed of the development of new bone after bone replacement both the rebuilding on the recipient bone ends may be advantageously influenced. With their method the bone defect in the rabbit's radius could be healed in four weeks.


[Experience with the management of scapula fractures]. Authors have treated 37 fractures of the scapula during 3 years. The questions of the aetiology, classification, diagnosis and therapy of this injury are reviewed. It is stated that the majority of the scapula fractures heal on conservative therapy; in consequence the functional treatment has a leading role. The fractures of the articular surfaces with step formation, the fractures of the scapular neck process dislocated in a high degree, or unstable fractures of the scapular neck combined with the comminutive fractures of the operative therapy are assessed. Attention is called to the most frequent operative complications and to the possibility of their prevention.


[Experience with the management of childhood diaphyseal fractures]. Author reports on experiences gained during 10 years the diaphyseal fractures in childhood. Whereas certain types of fracture near to the joints have a generally accepted absolute operative indication, the surgery in diaphyseal fractures is rarely held to be indicated. Osteosynthesis was performed in 18 per cent of all cases. Surgery is thought to be indicated in open fractures, in fractures that cannot be reduced or that are unstable, in polytrauma and in bilateral cases. In closed fractures plate osteosynthesis, in severe open fractures the fixateur externe was found to be the best solution. In certain cases Kuntscher nailing of the femur, medullary splinting of the lower arm or diafixation of the lower leg are performed in older children.


[Examination of infection and infectivity of open injuries]. Authors examined the circumstances of bacteriological infection of open fractures and large size injuries of the soft tissues. Bacteriological examinations were made from the ichor before the primary wound care and immediately after its finishing. It is stated that in open injuries there is only a potential possibility of infection and with an early wound care its danger can be substantially decreased. In the purulent infection after open injuries one has always to think on the possibility of secondary hospital infection.


[Experience with the Bristow procedure]. Authors assess the results of 17 Bristow's operations performed in the therapy recurrent anterior dislocation of the shoulder. The literature of the subject is reviewed, the technical details of the operations are described and the possibilities of complications are pointed out. On the basis of literary data and their own favourable experiences the wider use of this method is suggested.


[Fracture of the lower thoracic spine in ankylosing spondylitis]. Authors report on experiences gained during the treatment of the injury of the lower thoracal spine in a 64 year old male patient and review the relating literature. As a consequence of the basic disease fractures may occur following rather trivial traumas. As all ligaments are calcified a particular unstable state will develop following the fracture.


[Management of agricultural injuries at our department]. Authors want to call attention with the presentation of 4 cases to the significance of the agricultural injuries and to the difficulties of the therapy. The importance of the primary wound care is stressed. An adequate excision of the wound, debridement and an open wound care are essential in severely crushed, soiled cases. Therapy of severely crushed injuries, complicated with multiple fractures needs individual weighing and a reconstruction in more sittings.


[Experience with modular system tumor prostheses]. Authors elaborated a modular tumor prosthesis by the use of which the bone segment lost after the resection of hip, femur, knee and proximal tibial tumors can be supplemented in the desired measure. The components of the prosthesis set and the field of its use are described. In the last one and half year modular tumor prostheses were inserted in 10 patients: in 6 primary bone tumors and 4 solitary metastases. One shoulder, 5 hip, 3 stiff knee and 1 tibia diaphysis supplementing modular prostheses were inserted. In spite of 2 complications the initial experiences are encouraging, and in adequate indication the possibility of retaining limb and articular function beside good functional results is given.


[Management of upper arm fractures using a thermoplastic functional brace]. Authors report on the functional bracing of the fractured humerus. The advantage of the method is the preserved muscle and joint function with the use of a brace. This means ease for the patient in respect of the personal hygiene and living conditions. In elderly patients all other type of fixation around the chest means a substantial danger to the patients life because of the breath's movements depression.


[Results of using bone cement and phenol lavage in the surgical management of giant cell bone tumors]. Authors report on experiences gained with phenol lavage used as adjuvant and filling with bone cement in the surgical treatment of giant cell tumors of the bone. The effect of the adjuvant therapy was a considerable decrease in the rate of relapses: in 11 patients 1 relapse was found only in contrary to the 15 relapses in the control material (41 per cent) after 36 excochleations. They think that with adequate indications both the phenol lavage and filling with bone cement are useful supplements to the surgical therapy.


[Distally pedicled fasciocutaneous flaps]. Authors used for the supplementation of the soft parts of the lower extremity reversed fasciocutaneous flaps in 8 cases; 6 of them were burned patients. In the Hungarian literature no paper on the use of distally pedicled flaps was published until now.


[Chondromatosis in the metacarpophalangeal joint]. The case of a 57 year old male patient is described, operated for synovial chondromatosis of the third metacarpophalangeal joint of the left hand. The pathogenesis and the opinions on the therapy of this disease, described rarely because of its localization even in the world literature, are discussed.


[Incidental scaphoideo-trapezoid arthrodesis instead of Matti operation]. Author reports on a technical fault during Matti--Russe's operation. The corticospongious graft was inserted instead of the pseudarthrosis, in the scaphoid-trapezial joint. Because of the scaphoid-trapezial ankylosis developed, the persistent pseudarthrosis and the degeneration in the carpal bones the freeing of complaints could be reached only by intercarpal arthrodesis and wrist denervation, according to Wilhelm.


[Ventricular hyperkinetic arrhythmias: a current therapeutic problem. The possible greater use of beta blockers]. The therapeutic approach to cardiac arrhythmias is constantly evolving due to our improved understanding of their mechanisms and clinico-prognostic implications, even if uncertainties and controversies continue to be a marked feature of this sector, perhaps more than in any other field of medicine. The frequent finding of cardiac arrhythmias in the healthy and cardiopathic population justifies the importance which the question of the diagnosis and treatment of cardiac rhythm disorders has now assumed, even if, as far as the latter is concerned, the aggressive approach has been considerably modified over the past years. This has occurred in view of the still unproven value of indiscriminate anti-arrhythmic treatment for the purposes of prolonging life. This treatment has only been demonstrated to be of value in a few studies in selected subgroups of high-risk patients. In addition, it should be underlined that it has been reported that anti-arrhythmic drugs may possible aggravate or induce new arrhythmia. This potential pro-arrhythmic effect has become increasingly recurrent due to the widespread use and diffusion of this category of drugs. Such considerations should therefore encourage greater caution in the use of these drugs. Cardiac arrhythmias may be benign or life-threatening, symptomatic or asymptomatic; they may be a warning sign of sudden death, or be the cause or effect of heart failure, be the expression of an acute or chronic heart disease, or the clinical manifestation, at a cardiac level, of an extracardiac pathology. Within this broad-ranging clinical context, arrhythmia often gives rise to therapeutic dilemmas which must be resolved with extreme rationality, taking into account the results of all available clinical trials. The results of the Cardiac Arrhythmias Suppression Trial (CAST) showed that clinical judgements of therapeutic efficacy, made in the absence of carefully controlled studies, are often incorrect. On the basis of these findings beta-blocking drugs may find increasing use, since while they are not anti-arrhythmic drugs in the strict sense of the term, they are safer due to their negligible pro-arrhythmic effect, the lower incidence of collateral effects and their proven efficacy in post-infarction. The role of beta-blockers in the treatment of manifest heart failure should not be over-looked, since by countering the deleterious effect of increased catecholamines they may improve the prognosis, thus reducing the incidence of sudden death.


Antipsychotic medication effects on neuropsychological functions. The investigator of antipsychotic drug effects on neuropsychological functions faces a range of conceptual, methodological, and technical obstacles. Some of these hurdles are pointed out in a brief review of existing literature, and a more detailed consideration of instrumentation issues is presented. The need for hypothesis-driven assessment strategies is highlighted. Interpretive difficulties are posed by the polyfactorial nature of the most widely used neuropsychological tests, and emphasis is placed on the use and development of methods that enable isolation of specific neuropsychological constructs. Examples of this strategy are drawn from ongoing work in the Neuropsychology Unit of the Clinical Research Center for the Study of Schizophrenia at Hillside Hospital, where hypotheses about the behavioral site of action of dopamine agonists and antagonists are used to guide the construction of novel computerized tests.


Lysosomal enzyme activities in frozen, non-cultured chorionic villi for prenatal diagnosis of enzymopathies. Normal reference values of lysosomal enzyme activities (alpha-glucosidase, mannosidase, fucosidase and arylsulfatase-A) were determined in chorionic villi obtained from artificial abortion in the first trimester of normal pregnancies (gestational weeks 6 to 11). Villi were homogenized comparatively either in saline or in Triton X-100 detergent. The alpha-glucosidase, mannosidase and arylsulfatase-A enzyme activities significantly diminished if homogenization was done in saline instead of Triton-X while the difference in fucosidase activity was not significant. Significant correlation was detected between alpha-glucosidase activity and week of gestation. It is suggested that Triton X-100-homogenization should be used for the lysosomal enzyme determinations in chorionic villi because the solubilization of enzymes from the lysosomes is complete in this case than with homogenization in saline.


[Activity of gamma-glutamyltranspeptidase in serum and urine of patients with polycystic renal disease and hepatic cystic disease]. The determination of gamma-glutamyltranspeptidase activity was carried out by the Rosalka et al. method in the serum and urine in the control group of healthy persons (20 cases), in the group of polycystic renal disease and hepatic cystic disease (18 patients), and in the group of polycystic renal disease (in 32 patients in the serum, and in 28 patients in the urine). No significant differences were found in the values of the enzyme activity in the studied patients in relation to the control group.


Somatostatin immunopositive neurons in the small intestine of the bullfrog (Rana catesbeiana). Somatostatin immunopositive neurons in the small intestine of the bullfrog (Rana catesbeiana) were studied using immunohistochemistry and surgical denervation of the mesenteric nerve. Immunopositive nerve elements were distributed throughout the small intestine, including nerve fibers in the myenteric plexus, circular muscle layer, submucosal layer, and mucosa. Somatostatin immunopositive nerve cell bodies occurred in the myenteric plexus but not in the submucosal layer. These cell bodies were surrounded by immunopositive nerve fibers forming basket-like terminals, and thus some of these cells may be interneurons. After denervation of the mesenteric nerve, adrenaline immunopositive nerve fibers disappeared almost completely from the small intestine, but no changes occurred in the distribution of somatostatin immunopositive neurons. Neurons in the coeliac ganglion projecting into the small intestine were adrenaline immunopositive but somatostatin immunonegative. The results indicate that somatostatin immunopositive neurons in the small intestine of the bullfrog are primarily intrinsic in origin.


[State of the pancreatic insular apparatus in experimental diabetes mellitus with varying degrees of severity in the rat]. The interrelationship between endocrine cells of the Langerhans islands in experimental diabetes provoked in female Wistar rats are studied. The content of the insulin in B-cells, glucagon in A-cells and somatostatin in D-cells was determined by method of indirect immunofluorescence with the use of monoclonal antibodies and antiserum. It is established that an decrease of the insulin content in B-cells is followed by an increase of glucagon in A-cells and that of somatostatin in D-cells. The increase of the glucagon content is proportional to glycemia level and to the decrease of the insulin in B-cells.


Restriction fragment length polymorphism (RFLP) analysis provides evidence for a high degree of homology of mitochondrial DNAs from rat hepatomas versus normal rat livers. Where differences have been reported between tumor and normal mitochondrial DNA (mtDNA), they have generally involved limited modifications of the genome (Taira et al., Nucleic Acids Res. 11:1635, 1983; Shay and Werbin, Mutat. Res. 186:149, 1987). However, Corral et al. (Nucleic Acids Res. 16:10935, 1988; 17:5191, 1989) observed recombination between cytochrome oxidase subunit I (COI) and NADH dehydrogenase subunit 6 (ND6), two genes normally on opposite sides of the circular mitochondrial genome. In rat hepatoma mtDNA COI and ND6 were reported to be separated by only 230 base pairs (Corral et al., 1988, 1989). We have performed RFLP analysis on mtDNA from normal rat livers and rat hepatomas, using COI and ND6 probes. Additional experiments compared end-labeled DNA fragments produced by EcoRI and HindIII digestion of mtDNA. These studies failed to provide any evidence for genetic recombination in rat hepatoma mtDNA, even in the same cell line used by Corral et al. Rather, they support the conclusion that mtDNA from tumor and normal tissues exhibits a low degree of heterogeneity.


The aminoquinazoline group as a replacement for the salicylamide group: the design and synthesis of a novel highly selective beta 1 adrenoceptor partial agonist. The high potency at beta 1 receptors, excellent selectivity (beta 1/beta 2) and high degree of agonism displayed by compounds such as 1 is believed to be due in part to the salicylamide side chain. Two conformations of salicylamide are known to exist in the crystal state (2 and 3), but ab initio calculations suggest that in the absence of crystal packing forces 2 should be more stable. The aminoquinazoline group was judged to be a good replacement for salicylamide in 1, and consequently the oxypropanolamine derivative 4 was prepared. 4 shows extremely high potency at the beta 1 receptor, and excellent beta 1/beta 2 selectivity. It has comparable in vitro activity to 1, although it displays a lower degree of agonism. These results suggest that in this system aminoquinazoline appears to be an excellent mimic of the salicylamide group.


Molecular characterization of the smallest secondary constriction region (qh) of human chromosome 16. We report the smallest secondary constriction region (h) in human chromosome 16. The cytochemical, cytogenetic, and molecular techniques revealed the complex heterogeneity of heterochromatin observed in this region. The heteromorphisms can be found due to the variation in centromeric (c) region alone or in combination with the h region. Routine selective staining techniques fail to differentiate the C region from the h region. However, the fluorescence in situ hybridization technique clearly demonstrated that the centromere of chromosome 16, which is composed of 340-base-pair dimers arranged in a tandem array of 1.7-kb higher-order repeat units, is not heteromorphic in the present case, but other molecular cytogenetic techniques demonstrated the presence of a very small h region. The evolution of heterochromatin of this region is discussed.


Very low calorie diets and recently developed anti-obesity drugs for treating overweight in non-insulin dependent diabetics. Weight reduction in non-insulin dependent diabetes mellitus (NIDDM) patients improves metabolic control, reduces cardiovascular risk factors, has blood pressure lowering effects and improves the well-being of the patient. This paper describes the role of very low calorie diets (VLCD), exercise, beta-adrenergic drugs and serotoninergic agents in the treatment of overweight in NIDDM. VLCD reduce body weight and improve glucose metabolism. Physical exercise programmes in addition to dietary restriction substantially contribute to weight loss and metabolic control in NIDDM. New specific beta-adrenergic agents, exhibiting virtually no beta 1 or beta 2 activity, increase energy expenditure and weight loss probably by enhancement of the basal metabolic rate. The target tissue in humans of this beta-adrenergic effect is as yet unknown. These drugs seem to enhance weight loss when used in combination with (very) low calorie diets compared to dietary restriction alone. Serotoninergic drugs reduce body weight by decreasing appetite, in particular for carbohydrates. Furthermore these drugs seem to improve insulin receptor sensitivity.


Production of phytochelatins in Polygonum cuspidatum on exposure to copper but not to zinc. We studied cellular resistance to copper of plant cells Polygonum cuspidatum. When callus of P. cuspidatum was incubated on medium containing 100 microM cupric sulfate, the callus grew as well as the control callus did. The copper content of the callus, however, was elevated to a similar level of the medium. When cell extracts of callus exposed to 100 microM cupric sulfate were fractionated by gel filtration chromatography, a specific copper peak was eluted at the region of molecular weights between 4000 and 1000. Since an appearance of the copper-containing materials was inhibited by buthionine sulfoximine and the partially purified copper-containing materials contained only three amino acids: glutamic acid, glycine and cystine, the materials were supposed to be gamma-glutamyl peptides phytochelatins. Callus of P. cuspidatum synthesized phytochelatins from 50 microM cupric sulfate and maximally at 100-150 microM cupric sulfate. When induction of phytochelatins by another heavy metal, zinc, was analyzed, the callus, however, did not synthesize phytochelatins on exposure to zinc sulfate up to 1 mM. These findings suggested that phytochelatins were required for resistance to copper but probably not to zinc in the plant cells.


[New information about the biomechanics of fracture fixation]. The better knowledge of the biomechanics of bone tissue and fracture healing resulted in the development of the AO principles and its methods of treatment. I have demonstrated on the femora of rabbits that the consequence of progressive dynamization versus the current unchanging stability of fracture fixation was a more rapid healing. According to our knowledge the more elastic, dynamizating or dynamizable fixation of the fracture are the methods of future. The new biodegradive implantates make not only their removal superfluous but help the fracture healing, too.


[Double dislocation on the same finger (based on author's own cases as well as data from the literature)]. Author describes now the third time this very rare injury. New cases are demonstrated and the knowledge originating partly from all other authors, partly developed by himself is summarized.


Effects of interleukin-8 on suppression of human lymphocyte polarization and migration by anti-LFA-1 antibody. We investigated the role of lymphocyte function-associated antigen 1 (LFA-1) in human T cell polarization and migration assay by using monoclonal antibody specific to beta chain (CD18) and alpha chain (CD11a). T cell polarization in response to fetal calf serum (FCS) and colchicine was suppressed by the addition of CD18 and CD11a antibodies. Furthermore, T cell migration in response to lymphocyte chemotactic factor (LCF) and casein was markedly depressed by the addition of CD18 and CD11a antibodies. Additional studies to evaluate effects of interleukin 8 (IL-8) on polarization and migration of T cells preincubated with CD18 or CD11a antibody showed that IL-8 restored the capability of migration of T cells, whereas did not restore polarization activity of such cells. These studies indicate that LFA-1 plays a role in the polarization and migration of T cells and that IL-8 may positively interfer with LFA-1-adhesion molecules.


[Three-dimensional CT studies of pelvic fractures]. Authors use in the diagnosis of fractures of the pelvis a three dimension programme developed by them to the tomographic apparatus of the Siemens Somatom CR computer. With the demonstration of a few pictures of their own they call attention to the possibility of diagnosis given by the 3D-CT programme. On the basis of their experiences the advantages and disadvantages of this modern imaging method are summarized.


[The regulation of human cardiac activity during the cyclical change of barometric pressure under sealed-cabin conditions]. A cyclic change of barometric pressure from 790 to 720 mm Hg once a day to twice within 3 days under sealed conditions results in a functional rearrangement of the mechanisms to control cardiac activity which cause the predominance of vagotonic responses determined by a decrease of the body reserves and its asthenization. At the beginning of staying under pressurized conditions (2nd week) it appears as an occurrence of meteorotropic responses (in 13% of subjects tested) and at the end of living in pressurized conditions (9th week) as a significant decline with a change ("removal") in: pressure, index of strain, vegetative index of rhythm, index of regulation processes, index of vegetative equilibrium and as an increase in the number of individuals responding to a cyclic change of barometric pressure up to 44% which is indicative of a moderate relationship between manifestation rate of these sensations and an effect duration of a given factor.


[Diagnosis and management of hand injuries caused by foreign bodies]. The authors survey the possibilities of diagnosis of the frequent injuries of the hand caused by foreign bodies. It is proven by them that almost all foreign bodies can be demonstrated using radiological methods (native picture, ultrasound examination, xeroradiography, CT, MRI). They support the immediate surgical care in the therapy of injuries caused by foreign body, they deal however with the problems of the later therapy, too. Their opinion is backed up by the demonstration of case reports.


[Surgical treatment of hallux valgus by peg osteotomy]. Authors describe a method of therapy of hallux valgus not yet published in this country. Its advantage is that as a consequence of the shortening of the proximal phalanx the tension of the soft tissues is decreased and as a final result the movement occurs between the original cartilaginous surfaces.


[Management of metatarsalgia using Helal's method for metatarsal osteotomy]. Authors report on the result of Helal's metatarsal osteotomies performed on 62 feet of 48 patients for metatarsalgia. In 85 per cent the result proved to be excellent, in 15 per cent there was no improvement. Analysing the causes of the postoperative complaints they call attention to the overload of the marginal arches and to the metatarsalgia developing on the non osteotomized neighbouring arch. These complaints appeared in 1/3 of their material and could be generally well influenced with conservative treatment.


[Experience with the AGC-2000 type bicondylar surface-substituting knee prosthesis]. Authors report on the first use of bicondylar surface substitution knee prosthesis of a new type. The prosthesis and the instruments developed for implantation are demonstrated. In the first five operations the appropriateness of the implantation instruments, the exactness of their handling, the assortment of the series of prosthesis have met the plus which, compared to the types known until now, can secure the exactness of the operation and improve the ratio of the lasting good results.


[Osteosynthesis of femoral neck fractures by double-cannulated screws (preliminary report)]. Authors report on a new osteosynthesis based on the double nailing of femoral neck fracture and screwing, and developed from the "Uppsala" double screwing. The preliminaries of the double cannulated screwing method, its advantages, the experiences gained in 30 cases operated in one year and the early results are described.


[Problems of rehabilitation after femoral neck fractures in Hungary and possible solutions]. Authors examined the results of the 4 months rehabilitation of 753 cases of hip fractures treated in one year in their Institute. It was stated that a significant part of the patients had to be discharged too early. It is contributed to the inadequacy of the rehabilitation possibilities of the patients that 3/4 of those living after 4 months have complaints, only half of them can leave their flat, only 1/5 walks without a stick and 1/3 with one stick. On the basis of Swedish experiences they see the possibility of improving their results in a closer cooperation with the primary health care.


[Posterior dislocation of the median end of the clavicle]. Authors discuss, based on the experiences gained in the operative treatment of posterior sternoclavicular dislocation in two patients, the frequency, the symptoms, the classification, the diagnosis and treatment of this rare injury. One of the two injuries was fresh the other inveterated. No description of the operative treatment of similar injuries was found in the literature of this country. Finally the importance of the early recognition and the definitive treatment of this rare dislocation are stressed.


[Acromioclavicular dislocation associated with fracture of the coracoid process]. The joining of the acromioclavicular dislocation and the fracture of the coracoid process is rather rare. The number of cases described in the world literature is under 20, in this country no such case was described yet. Author reports on a case treated with operation, that healed with good result. The viewpoints of the diagnosis and treatment of this injury are reviewed. The results of the conservative and operative treatment are hitherto equally favourable. Author thinks the operative treatment of injuries with complete dislocation reasonable.


[Arthrosis following knee joint instability and its surgical prevention]. Authors describe the more important injuries that must be corrected by all means during the operative treatment of the acute instability of the knee joint to prevent later arthrosis. They call attention to the fact that in the treatment of the acute instability of the knee joint the prevention of the arthrosis can be realized only with a reconstructive operation, performed in time and with modern technique. The biomechanical causes of the arthrosis, developing during the persistent instability are analysed. The extraarticular operative methods used by them in chronic cases are described.


[Femoral neck fracture--selection of treatment method]. Authors followed for 4 months the life of 753 patients with femoral fractures of the hip and state, that with an extension of the operative indication the hospital mortality was decreased to the half during 15 years in spite of the further rise of the age. It is contributed first of all to the inadequacy of the rehabilitation that the later mortality is high and it is the double of that in the hospital: not more than 3 of 4 injured reaches the 4 months control.


Neurotransmitters and the pharmacology of the suprachiasmatic nuclei. The suprachiasmatic nucleus is the major component of the biological clock responsible for the generation and the regulation of circadian rhythms in behavioral and physiological functions. The SCN acts as an endogenous circadian pacemaker that becomes entrained to the light/dark cycle. Although many neuropeptides and neurotransmitters have been localized within and around the SCN, their role in the generation of the circadian signal still has to be elucidated. There are some data about the transmitters that are involved in the mechanism of entrainment. The way the SCN modulates the homeostatic control systems that are involved in the control of behaviour, also needs more clarification.


Simultaneous occurrence of monoclonal gammopathy and acute secondary leukemia with overexpression of P-glycoprotein. A 52-year-old woman, previously treated with chemo- and radiotherapy for Hodgkin's disease, developed an acute non-lymphoid leukemia and, contemporarily, an IgG-kappa paraproteinemia. Cytogenetic analysis showed a major clone, representing 90% of observed metaphases, with monosomy of chromosomes 5 and 14. In addition, leukemic cells exhibited a high expression of the P-glycoprotein, a transmembrane glycoprotein involved in the multidrug-resistance mechanism. Possible explanations for this cluster of findings are provided.


Modulation of the 5-HT1C receptor-mediated behavior by 5-HT2, but not 5-HT1A, receptor activation. The effects of 5-HT1A-receptor agonists 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) and gepirone, a 5-HT1A/5-HT2-receptor agonist 5-methoxy-N,N-dimethyltryptamine (5-MeODMT) and a 5-HT2-receptor agonist (+-)1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane ((+/)DOI) on the 5-HT1C-receptor-mediated exploratory hypoactivity in rats, induced by m-trifluoromethylphenylpiperazine (TFMPP) or m-chlorophenylpiperazine (m-CPP), were studied in the open field test. (+/-)DOI attenuated the effects of TFMPP and abolished those of m-CPP (not dose-dependently). 5-MeODMT showed a weak antagonistic action only at one, intermediate dose. The effects of TFMPP or m-CPP were not changed by 8-OH-DPAT or gepirone. At the same time, 8-OH-DPAT, gepirone, 5-MeODMT and (+/-)DOI themselves practically did not change the exploratory activity of rats. The obtained results permit an assumption that a functional interaction exists between 5-HT1C- and 5-HT2-receptors, but not between 5-HT1C- and 5-HT1A-ones.


Relationship of protease-resistant protein, scrapie-associated fibrils and tubulofilamentous particles to the agent of spongiform encephalopathies. Tubulofilamentous particles and scrapie-associated fibrils (SAF) are ultrastructural markers, while protease-resistant protein (PrP) is a molecular biological marker for all spongiform encephalopathies. Review of all published work has suggested that PrP molecules aggregate to form a three-dimensional SAF. Further reports have suggested that a single-stranded DNA wraps round SAF and acquires an outer protein coat to form tubulofilamentous particles. As incubation period increases in the infected animals, larger amounts of PrP molecules are committed to form SAF, interfering with the normal supply of PrP to cell membranes which become disrupted and eventually fragment, resulting in vacuoles typical of those found in spongiform encephalopathies.


Isolation and characterization of a cDNA encoding a synaptonemal complex protein. A gene encoding a 65-kilodalton antigen of the rat synaptonemal complex, SC65, has been cloned by screening rat testis lambda gt11 and lambda ZAPII cDNA expression libraries using polyclonal antibodies against rat synaptonemal complex proteins. The longest open reading frame, initiating at an ATG codon in the cDNA, encodes a protein of 431 amino acids, with a relative molecular mass of 50,000. Immunological analysis locates the SC65 gene product on the synaptonemal complex between the pairing faces of the parallel aligned cores of homologous chromosomes in spermatocytes. Of the rat tissues examined, the SC65 gene is transcribed in testis, brain, and heart at similar levels, and in the liver at a much lower level. The DNA sequence extending about 80 base pairs downstream of the translation termination codon has 93% similarity to the identifier sequence present in the rat genome in 1 x 10(5)-1.5 x 10(5) copies and in cDNA clones of precursors of brain-specific mRNAs. The amino acid sequence encoded by the SC65 gene contains an acidic region in the C-terminal domain of the protein, potential glycosylation sites, and at least one possible phosphorylation site. The protein shows no overall similarity to proteins of known function, nor is there similarity to protein sequences present in GenBank or EMBL data bases.


Epidemio-entomological survey on malarial vector mosquitoes in Kyongbuk, Korea. In order to determine population dynamics of Anopheles sinensis, a survey based on average number of female mosquitoes per trap-night was carried out during the period of 5 years from 1987 to 1991. A. sinensis first appeared between the 2nd and 20th April, and were trapped in large number between the 5th and 12th July. The number of trapped mosquitoes began to decrease from mid-August, and a few were collected until mid-November, each year. The average number of A. sinensis in July was 542.6 per trap-night in 1987, but in 1989 increased abruptly to 1,331.4, and then decreased to considerably lower levels, 271.9 in 1990 and 372.1 in 1991. The nocturnal activity of A. sinensis to attack humans was found to become active in the early night, and it was gradually decreasing at mid-night, however, then slightly increasing toward dawn. The immature stage of A. sinensis in the rice paddies was first found in the correlation pattern with peak adult densities in early July. The highest larval density of A. sinensis in the study area was 21,226 x 10(3) in early July 1990. The larval A. sinensis showed high resistance level and resistance ratios against 3 kinds of organo-phosphorous compounds, diazinon, malathion, and fenitrothion, but low resistance against fenthion. The present results indicated that the population density of A. sinensis in Kyongbuk area is decreasing over the five-year from 1987 to 1991.


Scrapie-associated tubulofilamentous particles in human Creutzfeldt-Jakob disease. Scrapie-associated fibrils (SAF) were demonstrated by a simple negative staining method for electron microscopy from fresh and frozen brains with naturally occurring human Creutzfeldt-Jakob disease (CJD). The findings confirm that SAF occur as an internal part of a larger three-layer particle. The two outer coats of SAF can be disrupted by detergent alone or can be digested in two stages by a combination of proteolytic enzymes and subsequent treatment with DNase and mung bean nuclease. Examination of thin sections from the cerebral cortex of brains from patients with CJD revealed the presence of 26-30-nm diameter tubulofilamentous particles, identical to those previously described in natural scrapie of sheep and bovine spongiform encephalopathy and also in experimentally induced scrapie in mice and hamsters and CJD-infected mice and chimpanzees. Thus, it would appear that the particles are not contaminants passaged in experimental animals.


Interrelationships of endoplasmic reticulum, mitochondria, intermediate filaments, and microtubules--a quadruple fluorescence labeling study. To study the interrelationships of endoplasmic reticulum, mitochondria, intermediate filaments, and microtubules, we have developed a quadruple fluorescence labeling procedure to visualize all four structures in the same cell. We applied this approach to study cellular organization in control cells and in cells treated with the microtubule drugs vinblastine or taxol. Endoplasmic reticulum was visualized by staining glutaraldehyde-fixed cells with the dye 3,3'-dihexyloxacarbocyanine iodide. After detergent permeabilization, triple immunofluorescence was carried out to specifically visualize mitochondria, vimentin intermediate filaments, and microtubules. Mitochondria in human fibroblasts were found to be highly elongated tubular structures (lengths up to greater than 50 microns), which in many cases were apparently fused to each other. Mitochondria were always observed to be associated with endoplasmic reticulum, although endoplasmic reticulum also existed independently. Intermediate filament distribution could not completely account for endoplasmic reticulum or mitochondrial distributions. Microtubules, however, always codistributed with these organelles. Microtubule depolymerization in vinblastine treated cells resulted in coaggregation of endoplasmic reticulum and mitochondria, and in the collapse of intermediate filaments. The spatial distributions of organelles compared with intermediate filaments were not identical, indicating that attachment of organelles to intermediate filaments was not responsible for organelle aggregation. Mitochondrial associations with endoplasmic reticulum, on the other hand, were retained, indicating this association was stable regardless of endoplasmic reticulum form or microtubules. In taxol-treated cells, endoplasmic reticulum, mitochondria, and intermediate filaments were all associated with taxol-stabilized microtubule bundles.


Inhalant allergens in asthmatic children in Taiwan: comparison evaluation of skin testing, radioallergosorbent test and multiple allergosorbent chemiluminescent assay for specific IgE. The multiple allergosorbent chemiluminescent assay (MAST-CLA) is a method for measuring total and allergen-specific IgE in human serum by a chemiluminescent immuno-enzymatic system. To compare the results of MAST tests with those of radioallergosorbent (RAST) and skin tests as an adjunct to the diagnosis of inhalant allergens in Taiwan, 195 asthmatic children, aged 5 to 15 years, were studied. All MAST tests had valid positive and negative control threads. The most important allergens in our patients were the two mite species: Dermatophagoid pteronyssinus and Dermatophagoid farinae (87.5% and 82.1%, respectively). There were also large responses (MAST-CLA class > or = 2) to the cockroach mix (28.45%), Alternaria (24.3%), and Eucalyptus (32.6%). The individual efficiency percentage between MAST-CLA and skin tests was D. pteronyssinus 91%, D. farinae 88%, cockroach mix 85%, feather mix 75%, dog dander 64%, Candida 75%, Aspergillus 79%, Alternaria 88%, ragweed mix 62% and Eucalyptus 76%. Comparison of the results of allergen-specific IgE measured by MAST-CLA and RAST were also significantly correlated for all four allergens (D. farinae, Candida, Alternaria and grass mix, r = 0.79-0.92). MAST-CLA was randomly duplicated and proven reproducible in 89% of the tests. Changes between positive and negative results occurred in only 3.6% of the tests. MAST-CLA is a simple in vitro test for specific IgE to 35 allergens, which compares favorably with RAST. It is concluded that MAST-CLA and RAST are similar in their ability to measure allergen-specific IgE, and correlate equally well with skin tests and clinical history in asthmatic children.


Body axis determination during early development in amphibians. The specification of the main axes of the body is a phenomenon based on cell communication and is among the early crucial events of embryonic development. Upon fertilization, the amphibian egg reorganizes its cytoplasmic content, leading to the establishment of the future dorsal-ventral axis of the body. Heterogeneous distribution of maternal components confers cellular regionalization after only a few mitoses. Development up to the 4000-cell stage proceeds almost entirely on maternal materials, and during this period there is remodeling of the chromatin to set up specific gene expression in various regions of the embryo. The zygote at this stage has already undertaken cellular interactions leading to mesoderm formation and regionalization. Dorsal mesodermal components then induce the formation of the Spemann's organizer, a structure directly involved in the specification of the anterior-posterior axis of the embryo (head to tail). Molecular analysis of these phenomena has allowed the identification of growth-factor-like and transcription-factor-like proteins that have characteristics typical of specification factors. We will review the recent advances on these molecules and will also discuss the putative role of retinoic acid as a posteriorizing agent.


The effect of alpha-2 agonists and antagonists on the upper urinary tract of the rat. We examined the effect of the selective alpha-2 agonist dexmedetomidine and antagonist atipamizole on the upper urinary tract, renal pelvic pressure and ureteral peristalsis. Experiments were performed on twelve Sprague-Dawley female rats weighing 275-323 grams, with administration of urethane (1.2 micrograms/kg). Ventilatory support was provided through a tracheotomy. A continuous normal saline infusion was maintained through the left iliac vein at a rate of 2.5 ml/hr. Arterial pressure was measured at the left iliac artery, which was cannulated with a PE-100 tube connected to a pressure transducer. A mid-line incision was then made from the xyphoid to the symphysis to expose the left kidney, both ureters, and the bladder. The bladder was intubated and drained to avoid bladder pressure increase. Measurements of urine output rate were made from the right ureter and renal pelvic or ureteral pressure was measured using a nephrostomy placed into the pelvis. A ureterostomy was produced by introducing another catheter, into the upper segment of the left ureter for ureteral pressure measurements. The rats were divided into two groups as follows: 1) dexmedetomidine group (n = 6); injected intravenously with 2 micrograms/kg of dexmedetomidine dissolved in 0.5 ml saline. 2) atipamizole group (n = 6); injected intravenously with 2 micrograms/kg of atipamizole dissolved in 0.5 ml saline. Ureteral peristaltic frequency, baseline pressure, and contraction amplitude were compared before, after, and between the bolus injections of 2 micrograms/kg dexmedetomidine (n = 6) or 2 micrograms/kg atipamizole (n = 6) in 0.5 ml saline. The results showed that dexmedetomidine at 2 micrograms/kg produced a significant decrease in arterial pressure and an increase in urine output from 1.2 + 0.8 to 3.6 + 1.2 ml/min. There was no effect on the baseline pelvic pressure of 6.8 + 1.2 cmH2O or amplitude of the renal pelvic contractions: 3.5 + 0.6 cmH2O. The frequency of pelvic contractions was reduced from 0.37 + 0.03 to 0.27 + 0.02 Hz. Atipamizole at 2 micrograms/kg produced a significant reduction in urine flow rate of 1.1 + 0.8 to 0.6 + 0.2 ml/min. Atipamizole also showed no significant effects on baseline pelvic pressure or frequency, but increased the amplitude of pelvic contractions from control values of 3.0 + 0.9 to 3.4 + 0.9 cmH2O. Dexmedetomidine reduced both the baseline ureteral pressure of 8.5 + 2.4 and peristaltic contraction pressure of 11.5 + 2.3 cmH2O in 4/6 rats. Atipamizole reduced base-line ureteral pressure and increased peristaltic rate.(ABSTRACT TRUNCATED AT 400 WORDS)


Blood levels of platelet-activating factor in endotoxin-sensitive and endotoxin-resistant mice during endotoxemia. Platelet-activating factor (PAF) is a phosphoglyceride secreted by a variety of cells and has been implicated in endotoxin toxicities. To further confirm its role in endotoxin-induced tissue injuries and death, we conducted an experiment on endotoxin-resistant (C3H/HeJ strain) and endotoxin-sensitive (C3H/HeN strain) mice. The experiment consisted of three parts: 1) the LD50 of endotoxin from E. coli 0127:B8 cells was quantitated in C3H/HeN mice; 2) the lethality of PAF in C3H/HeJ mice at a dose lethal to C3H/HeN mice was determined; and 3) the blood levels of PAF in C3H/HeJ and C3H/HeN mice were measured after a dose of endotoxin lethal to the C3H/HeN strain was injected. PAF contained in the blood samples was extracted by a solid phase procedure and assayed by a radioimmunoassay method. The results showed that endotoxin-resistant and endotoxin-sensitive mice were equally susceptible to death induced by the same lethal dose of PAF. After injection with endotoxin, the blood PAF levels in C3H/HeN mice increased significantly (p < 0.01) at 60 minutes and 90 minutes, with a peak level three times that of the control group. The blood PAF levels in C3H/HeJ mice, however, remained unelevated throughout the experiment. The timing of the occurrence of the peak blood PAF level in the C3H/HeN mice corresponded with the emergence of their illness from the endotoxin injection. These findings shed new light on our understanding of the resistant mechanisms of C3H/HeJ mice to bacterial endotoxin and affirm the possible role of PAF in mediating endotoxin toxicities.