Heparin binding domain of human antithrombin III inferred from the sequential reduction of its three disulfide linkages. An efficient method for structural analysis of partially reduced proteins

Human antithrombin III (AT-III) was partially reduced under mild conditions in the absence or presence of low molecular weight heparin. Quantitation of reduced disulfide bonds was facilitated by the application of a water-soluble color reagent, 4-N,N-dimethylaminoazobenzene-4'-iodoacetamido-2'-sulfonic acid (S-DABIA). The study shows that the three disulfide linkages of AT-III can be sequentially reduced, with Cys8-Cys128 being the most sensitive, followed by Cys21-Cys95, while Cys247-Cys430 is the most resistant to the mild reduction conditions. The rate of reduction of Cys8-Cys128 and Cys21-Cys95 was significantly decreased in the presence of heparin. The reduction of Cys8-Cys128 was also found to correlate quantitatively with the loss of heparin-accelerated antithrombin activity, heparin binding affinity, and heparin-induced fluorescence enhancement. These results suggest that Cys8-Cys128 is required for the integrity of the heparin binding domain of AT-III and support previous findings that lysyl residues surrounding Cys128 (Lys107, Lys114, Lys125, and Lys136) constitute an important part of the heparin binding site in AT-III.     

Heparan sulfate-mediated binding of epithelial cell surface proteoglycan to thrombospondin

Purified NMuMG mouse mammary epithelial cell surface proteoglycan (PG), a membrane-intercalated core protein bearing both heparan sulfate and chondroitin sulfate glycosaminoglycan (GAG) chains, binds to a thrombospondin (TSP) affinity column and is eluted by a salt gradient. Double immunofluorescence microscopy demonstrates extensive co-localization of bound exogenous TSP and cells bearing exposed cell surface PG at their apical surface. The binding, as assayed by both methods, is heparitinase-sensitive, but not chondroitinase-sensitive. Alkali-released heparan sulfate chains bind to a TSP affinity column, similarly to native PG, whereas the chrondroitin sulfate chains do not. Core protein does not bind to TSP. These results indicate that NMuMG cells bind TSP via their surface PG and that the binding is mediated by the heparan sulfate chains.     

Association of a basic 25K protein with membrane coating granules of human epidermis

Keratinocytes of the upper granular layers contain unique round-to-oval granules, 100-500 nm in diameter, in their peripheral cytoplasm. These granules (known as membrane coating granules [MCG], or lamellar granules) fuse with the apical cell surface of uppermost granular cells and discharge their contents into the intercellular space, where they are believed to play a role in establishing the permeability barrier of the epidermis and possibly in regulating the orderly desquamation of terminally differentiated keratinocytes. Using two monoclonal antibodies originally prepared against hair follicle antigens, we have identified a 25K epidermal protein in association with both MCG-like granules in the peripheral cytoplasm of granular cells as well as MCG-derived intercellular material. This protein is relatively basic (pI greater than 8), largely aqueous soluble, methionine deficient, and is relatively abundant in epidermis (comprising up to approximately 0.1% of soluble proteins). Its distribution is restricted to the granular layer of keratinized (cornified) stratified squamous epithelia. The identification of this protein component opens new avenues for studying the molecular mechanisms underlying the establishment of permeability barrier and/or regulation of desquamation.     

Inhibition of HIV replication by naphthalenemonosulfonic acid derivatives and a bis naphthalenedisulfonic acid compound

Several naphthalenemonosulfonic acid analogs and a bis naphthalenedisulfonic acid have been evaluated for anti-HIV activity in assays using H9 and MOLT-3 cells. Among the naphthalenemonosulfonic acids, a 4-amino-5-hydroxy compound and a 4,5-diamino compound showed low anti-HIV activity (upto 50% inhibition) at non-toxic doses. The bis naphthalenedisulfonic acid compound demonstrated significant suppression of HIV-1 antigen expression as measured by monoclonal antibodies to p17 (95%), p24 (94%) and syncytia inhibition (82%) at a dose of 20 micrograms/ml that was non-toxic to the host cells. The bis naphthalenedisulfonic acid analog represents a new class of compounds which may be effective in the treatment of HIV infected patients. The structure activity relationship and a probable mode of action of these compounds is discussed.     

Label-retaining cells reside in the bulge area of pilosebaceous unit: implications for follicular stem cells, hair cycle, and skin carcinogenesis

Inconsistent with the view that hair follicle stem cells reside in the matrix area of the hair bulb, we found that label-retaining cells exist exclusively in the bulge area of the mouse hair follicle. The bulge consists of a subpopulation of outer root sheath cells located in the midportion of the follicle at the arrector pili muscle attachment site. Keratinocytes in the bulge area are relatively undifferentiated ultrastructurally. They are normally slow cycling, but can be stimulated to proliferate transiently by TPA. Located in a well-protected and nourished environment, these cells mark the lower end of the "permanent" portion of the follicle. Our findings, plus a reevaluation of the literature, suggest that follicular stem cells reside in the bulge region, instead of the lower bulb. This new view provides insights into hair cycle control and the possible involvement of hair follicle stem cells in skin carcinogenesis.     

Effect of testosterone on serum immunoactive inhibin concentrations in intact and hypophysectomized male rats

Intact and hypophysectomized rats were treated with graded doses of testosterone via subcutaneous Silastic implants over a 13-week period. Serum inhibin concentrations fell 50% (P less than 0.001) after 2 weeks of hypophysectomy, remaining suppressed at this level for 13 weeks. The administration of testosterone to hypophysectomized rats (serum testosterone values 2-12 ng/ml; control values 5.5 ng/ml) was without effect on serum inhibin values. In contrast, administration of testosterone to intact animals for 7 weeks resulted in an initial fall (P less than 0.05) in inhibin levels to 50-70% of controls then increasing to reach control levels at higher doses. Serum FSH concentrations were similarly biphasic with increasing dose of testosterone and values for these two hormones were significantly correlated (r = 0.44, P less than 0.01). Segments of seminiferous tubules in culture from rats after various times of hypophysectomy showed a partly suppressed secretion of inhibin. The administration of testosterone did not modify inhibin production although inhibin production was sensitive to FSH. It is concluded that (1) serum inhibin concentrations are partly suppressed after hypophysectomy and testosterone has no effect on serum inhibin values; and (2) the suppression of serum inhibin in intact rats treated with increasing doses of testosterone is attributable to the concomitant fall in serum FSH concentrations.     

兔的一种新病毒：Ⅱ.一株兔出血症...

In this paper a strain of Rabbit Hemorrhagic Disease Virus (RHDV) was isolated and purified from the diseased rabbit livers with a method of using chloroform, two-phase of polyethylene-glycol-dextran sulfate sodium and sucrose density gradient centrifugation. Purified virus was nonenveloped, icosahedeal symmetry with a triangulation number of 3, and 33-37 nm in diameter. The capsid was composed of 32 capsomeres with central holes in an outer diameter of about 9nm. Two types of viral particles having different sedimentation coefficient, 130s and 166s could be identified after sucrose density gradient centrifugation. Probably no less than four virion proteins with molecular weight of 66.4, 65.0, 63.5, 41.0 x 10(3) dalton were detected by SDS-polyacrylamide gel electrophoresis. Viral nucleic acid was extracted from purified virus by using SDS-proteinase K-phenol. Tests with diphenylamine, formaldehyde, and staining with acridine orange as well as the curves of thermal denaturation showed that this kind of virus had a single-stranded DNA. The molecular weight of the ssDNA was approx 2.1 x 10(6) dalton as determined by electron microscopy. Data indicate that the RHDV may like the parvovirus of the family Parvoviridae.     

新疆温泉煤田早、中侏罗世孢粉组合及其地层意义*

新疆温泉煤田艾肯拜尔段和柯克它乌组的孢粉化石,计有42属59种.根据孢粉类型和含量变化等特征,建立2个孢粉组合(自下而上):1. Dictyophyllidites-Chordasporites-Jugasporites 组合;2. Cyathidites-Neoraistrickia-Pseudopicea 组合.第一组合产于艾肯拜尔段,时代可能为早侏罗世,第二组合产于柯克它乌组,时代暂定为早、中侏罗世.根据孢粉组合反映的植物群面貌是:裸子植物非常茂盛,其中以松柏纲为主,蕨类植物较少(包括真蕨纲植物);反映的古气候属于温暖湿润的亚热带型.     

黑龙江绥滨地区晚侏罗世东荣组的沟鞭藻类

本文对黑龙江绥滨86-11孔的沟鞭藻类进行了初步研究,共描述化石10属16种(包括2新种,6未定种);其中不少是在我国首次描述的。这些化石分成两个组合,其优势分子分别为Gonyaulacysta jurassica和Amphorula delicata。根据组合内各分子的时代分布,两化石组合的时代分别为Late Oxfordian—Early Kimmeridgian和Portlandian期,并对其产出层位东荣组的时代作了讨论,认为属于晚侏罗世。     

Hepatitis B virus DNA integration and expression of an erb B-like gene in human hepatocellular carcinoma.

Southern blot studies on Hepatitis B Virus (HBV) DNA integration in 13 human hepatocellular carcinomas (HCCs) patients revealed the presence of several distinct HBV integration sites in different human liver disease patients. In one HCC patient the DNA fragment containing the HBV integration also hybridized to an erb B probe. The erb B/HBV co-migrating DNA fragment was cloned and sequenced, and showed that HBV DNA is integrated next to a cellular DNA fragment which is homologous to the tyrosine protein kinase domain of the human epidermal growth factor receptor gene and other cell surface receptor genes. The virus-integrated cellular DNA sequence is expressed in this HCC patient, suggesting a possible role for this gene in hepatocarcinogenesis.