Quantitative histochemical assessment of the heterogeneity of glycogen phosphorylase activity in liver parenchyma of fasted rats using the semipermeable membrane technique and the PAS reaction

Summary Glycogen phosphorylase (EC 2.4.1.1) has been demonstrated in sections of liver from rats starved for 24 h. The method is based on the measurement of the amount of glycogen formed after incubation in a gelled medium containing glucose 1-phosphate as substrate, using the semipermeable membrane technique. Glycogen was demonstrated with the periodic acid-Schiff (PAS) reaction.Phosphorylase activity appeared to be highest in periportal areas. The optimum substrate concentration for revealing activity of the enzyme was 60–120mm. After incubation in the absence of substrate, the staining intensity, as measured cytophotometrically as the mean integrated absorbance at 560 nm, was similar to that of an unincubated section.p-Chloromercuribenzoate, a non-specific inhibitor of glycogen phosphorylase activity, reduced the formation of final reaction product attributable to phosphorylase activity completely. The Michaelis constants (K _m ) of the enzyme in periportal and pericentral areas differed. This was probably due to the presence of thea form only in periportal areas and of thea andb forms in pericentral areas. The mean integrated absorbances in both the periportal and pericentral areas increased linearly with incubation time (4–16 min). A linear relationship was also found with section thickness (4–10 µm). The total activity of glycogen phosphorylase in the periportal areas was double the pericentral activity.It is concluded that the semipermeable membrane technique, combined with the PAS reaction for glycogen, can be used as a valid method for the demonstration and quantification of glycogen phosphorylase activity in livers from starved rats.     

白纹伊蚊幼虫龄期的发育历期

在实验室内20±1℃, 23±1℃, 25±1℃, 28±1℃和30±1℃恒温条件下, 观察了白纹伊蚊(Aedes albopictus)幼虫(广州株)的发育历期.幼虫的总发育历期和各龄幼虫的发育历期随温度的上升而缩短, 各龄幼虫历期在总发育历期中占有恒定的百分比, 据此计算出其捕获机率.各龄幼虫的发育历期与温度呈直线回归关系.通过积温公式, 求出各龄幼虫的积温常数.此外还探索了营养对幼虫发育历期的影响, 从而影响各龄幼虫捕获机率的计算.     

Diurnal variation in 5′-nucleotidase activity in rat liver

Summary The diurnal variation of 5-nucleotidase activity in periportal and pericentral areas of rat liver parenchyma has been determined with quantitative histochemical means. 5-Nucleotidase activity was estimated using microdensitometry in cryostat sections after being incubated with a medium according to Wachstein and Meisel (1957). It appeared that 5-nucleotidase activity was significantly higher in pericentral areas than in periportal areas throughout the daily cycle and showed a maximum at the end of the light period. It was concluded that 5-nucleotidase activity may be related with the capacity to diminish messenger RNA resulting in protein breakdown.     

Protofibrin clots induced by calcium and zinc

During the course of studies with fibrin protofibrils, produced by adding hirudin to thrombin-activated fibrinogen prior to the onset of gelation, turbid clots were observed to be generated merely by adding Ca(II) or Zn(II) to protofibrils. The rate of gelation (CT) and turbidity of the “protofibrin” clots increases with cation levels in a concentration-dependent manner, with Zn(II) much more potent than Ca(II). For example, 50 μM Zn(II) generated a more turbid protofibrin clot than 0.5 mM Ca(II). In combination, levels of Zn(II) and Ca(II), which individually have no effect, induce protofibril gelation. The generation of protofibrin clots by Zn(II) is decreased at increasing ionic strength. Apparently, the underlying electrostatic forces that bind the monomers in fibrin and protofibrin gels are similar. SEM micrographs show that Ca(II)- or Zn(II)-induced protofibrin clots (600–1500Å thick) are essentially indistinguishable from those formed directly from fibrinogen and thrombin with divalent cation. The protofibrin fibers induced by the cations are thicker than the fibers formed directly from fibrinogen and thrombin in the absence of divalent cation. Branching appears brought about the the divalent cation-sensitive lateral association of different protofibril strands. These findings describe simple experimental methods for separately studying the early and late stages of fibrin gelation.     

Changes in the acinar distribution of some enzymes involved in carbohydrate metabolism in rat liver parenchyma after experimentally induced cholestasis

Extrahepatic cholestasis induced by ligation and transsection of the common bile duct caused a change in the parenchyma/stroma relationship in rat liver. Two weeks after ligation, the periportal zones of the parenchyma were progressively invaded by expanding bile ductules with surrounding connective tissue diverging from the portal areas. Parenchymal disarray developed and small clumps of hepatocytes or isolated hepatocytes were scattered within the expanded portal areas. These cells showed normal activity of lactate, succinate and glutamate dehydrogenase and may, therefore, be considered to be functionally active. After cholestasis the remainder of the liver parenchyma showed adaptational changes with respect to glucose homeostasis, as demonstrated by histochemical means. Glycogen stores disappeared completely whereas glycogen phosphorylase activity increased about ten fold. The increased glycogen phosphorylase activity and glycogen depletion indicate a greater glycogenolytic capacity in liver parenchyma after bile duct ligation to maintain as far as possible a normal plasma glucose concentration. The parenchymal distribution pattern of glucose-6-phosphatase activity did not change significantly after bile duct ligation. The isolated hepatocytes within the expanded portal tracts showed a high activity of this enzyme whereas the pericentral parenchyma was only moderately active. The distribution patterns of glucose-6-phosphate dehydrogenase and lactate dehydrogenase activity in the liver parenchyma were also largely unchanged after bile duct ligation, but the histochemical reaction for glucose-6-phosphate dehydrogenase activity demonstrated infiltration of the remainder of the parenchyma by non-parenchymal cells, possibly Küpffer cells and leucocytes as part of an inflammatory reaction. Under normal conditions the mitochondrial enzymes succinate and glutamate dehydrogenase show an opposite heterogenous distribution pattern in liver parenchyma. Following cholestasis both enzymes became uniformly distributed. The underlying regulatory mechanism for these different changes in distribution patterns of enzyme activities is not yet understood.     

The chemistry of 9 alpha-hydroxy steroids. 2. Epimerization and functionalization of 17 alpha-ethynylated 9 alpha-hydroxy steroids

Practical routes to 9 alpha-hydroxypregnenes were developed by epimerization and hydration of 17 alpha-ethynyl-9 alpha,17 beta-dihydroxyandrost-4-en-3-one. In the three different methods of epimerization which were used, the C-9 alpha hydroxy group was not susceptible to rearrangement or other side reactions. C-21 functionalized 9 alpha-hydroxypregnenes were obtained by introducing a 17 alpha-halogenated ethynyl group into 9 alpha-hydroxyandrost-4-ene-3,17-dione. Epimerization and hydration by the 17 beta-nitrooxy method produced 21-halogenated 9 alpha-hydroxypregnenes, which were further converted into 21-acetoxy-9 alpha-hydroxypregn-4-ene-3,20-dione.     

Mucosal epithelial cells and Langerhans cells are targets for infection by the immunosuppressive type D retrovirus simian AIDS retrovirus serotype 1

Simian acquired immune deficiency syndrome (SAIDS) caused by the type D retrovirus SRV-1 results in opportunistic infections and a spectrum of oral lesions similar to those seen in humans with AIDS. To better understand the pathogenesis of these oral lesions we have retrospectively examined the oral mucosa from ten rhesus monkeys that died with SAIDS and prospectively examined the oral mucosa of ten additional animals inoculated with SRV-1 to determine at what time, and in what cells SRV-1 infection of the oral mucosa occurs. Using single and double label immunohistologic techniques, and electron microscopy we detected SRV-1 in clusters of oral epithelial cells and rare Langerhans cells as early as 1 month postinoculation.     

Binding of the transition metal ion [(H20)(NH3)5Ru(II)]2+ to nucleosomal core and internucleosomal DNA

The goal of this study is to establish the nature of pentammineruthenium(III) binding to DNA in intact mouse liver nuclei. Also, we wish to determine whether the nucleosomal organization of mouse chromatin has a substantial effect on the relative Ru(III) binding levels of internucleosomal and nucleosomal core DNA. These questions are important because ammineruthenium compounds share chemical and biological properties with the cis-dichlorodiammineplatinum(II) or cisplatin chemotherapeutic agent. Therefore, they represent a potential class of new chemotherapeutic agents. We find that in intact nuclei the predominant DNA binding site for pentammineruthenium(II), followed by air oxidation to pentammineruthenium(III), is N-7 guanine, as is the case with cisplatin. Also, the Ru(III) distribution between internucleosomal and nucleosomal core DNA was found to be nearly identical as probed with three non-specific deoxyribonucleases.     

Absence of the Epstein-Barr virus genome in the normal thymus, thymic epithelial tumors, thymic lymphoid hyperplasia in a European population

It has previously been shown that the Epstein-Barr virus (EBV) genome may be detected in some thymic tumors. We have investigated specimens of normal thymus, thymitis with lymphoid hyperplasia and a large spectrum of thymic epithelial tumors obtained from european patients for the presence of EBV genome by in situ hybridization and DNA-blotting methods. Cell lines established from seven of the thymic tumors were also tested for EBV. No EBV genome was demonstrated in any of the tumors examined, which included various types of thymoma and thymic carcinomas, nor in the non-neoplastic thymic specimens. However, unlike previous reports, no examples of lymphoepithelial-like thymic carcinoma, nor specimen from Asian patients were included in this study. We suggest that EBV is linked to a specific epithelial tumor type, namely the lymphoepithelial-like carcinoma, regardless of its site, and not to thymic tumors in general.     

Absence of the Epstein-Barr virus genome in the normal thymus,thymic epithelial tumors,thymic lymphoid hyperplasia in a European population

It has previously been shown that the EpsteinBarr virus (EBV) genome may be detected in some thymic tumors. We have investigated specimens of normal thymus, thymitis with lymphoid hyperplasia and a large spectrum of thymic epithelial tumors obtained from european patients for the presence of EBV genome by in situ hybridization and DNA-blotting methods. Cell lines established from seven of the thymic tumors were also tested for EBV. No EBV genome was demonstrated in any of the tumors examined, which included various types of thymoma and thymic carcinomas, nor in the non-neoplastic thymic specimens. However, unlike previous reports, no examples of lymphoepithelial-like thymic carcinoma, nor specimen from Asian patients were included in this study. We suggest that EBV is linked to a specific epithelial tumor type, namely the lymphoepithelial-like carcinoma, regardless of its site, and not to thymic tumors in general. Supported by the SFB 172, C8, grant to BB and by the Deutsche Forschungsgemeinschaft, grant Ki 370/1-1