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Biomedical subjects

D Sinha

Publications and source records attributed to D Sinha.

At least 73 records · Page 4Linked to original sources
PubMed1985

Comparison of bleeding tendency, factor XI coagulant activity, and factor XI antigen in 25 factor XI-deficient kindreds.

The relationship of clinical bleeding tendency and factor XI antigen (XI:Ag) in factor XI deficiency was studied in 78 members of 25 factor XI-deficient kindreds. Factor XI:Ag was measured in a competitive radioimmunoassay, using monospecific, heterologous anti-factor XI antibody. 125I-labeled factor XI, and staphylococcal protein A as the precipitating agent. Deficiency of factor XI clotting activity (XI:C), less than 0.62 U/mL, occurred in 48 individuals, 22 of whom experienced postoperative or posttraumatic bleeding: Their mean factor XI:C was 0.21 +/- 0.04 U/mL (SEM), and factor XI:Ag was 0.23 +/- 0.04 U/mL. The remaining 26 had no clinical bleeding, many despite surgical challenge: Their mean factor XI:C was 0.30 +/- 0.04 U/mL, and factor XI:Ag was 0.34 +/- 0.05 U/mL. In all, 13 kindreds had between 1 and 11 members with bleeding; the other 12 had none with deficient hemostasis. Two heterozygous factor XI-deficient individuals appeared to be positive for cross-reacting material (CRM+). The slope of the regression line for factor XI:C and factor XI:Ag data points in the 78 individuals tested did not differ from control, and all points fell within 95% confidence limits derived from control. In conclusion, bleeding tendency appears to be consistent within a given kindred and is not determined exclusively by factor XI:C or factor XI:Ag levels.

Antigens↗
PubMed1984

Kinetics of the Factor XIa catalyzed activation of human blood coagulation Factor IX.

The kinetics of activation of human Factor IX by human Factor XIa was studied by measuring the release of a trichloroacetic acid-soluble tritium-labeled activation peptide from Factor IX by a modification of a method described for bovine Factor IX activation by Zur and Nemerson (Zur, M., and Y. Nemerson, 1980, J. Biol. Chem., 255:5703-5707). Initial rates of trichloroacetic acid-soluble 3H-release were linear over 10-30 min of incubation of Factor IX (88 nM) with CaCl2 (5 mM) and with pure (greater than 98%) Factor XIa (0.06-1.3 nM), which was prepared by incubating human Factor XI with bovine Factor XIIa. Release of 3H preceded the appearance of Factor IXa activity, and the percentage of 3H released remained constant when the mole fraction of 3H-labeled and unlabeled Factor IX was varied and the total Factor IX concentration remained constant. A linear correlation (r greater than 0.98, P less than 0.001) was observed between initial rates of 3H-release and the concentration of Factor XIa, measured by chromogenic assay and by radioimmunoassay and added at a Factor IX:Factor XIa molar ratio of 70-5,600. Kinetic parameters, determined by Lineweaver-Burk analysis, include Km (0.49 microM) of about five- to sixfold higher than the plasma Factor IX concentration, which could therefore regulate the reaction. The catalytic constant (kcat) (7.7/s) is approximately 20-50 times higher than that reported by Zur and Nemerson (Zur, M., and Y. Nemerson, 1980, J. Biol. Chem., 255:5703-5707) for Factor IX activation by Factor VIIa plus tissue factor. Therefore, depending on the relative amounts of Factor XIa and Factor VIIa generated in vivo and other factors which may influence reaction rates, these kinetic parameters provide part of the information required for assessing the relative contributions of the intrinsic and extrinsic pathways to Factor IX activation, and suggest that the Factor XIa catalyzed reaction is physiologically significant.

Factor IX↗
PubMed1984

Blood coagulation factor XIa binds specifically to a site on activated human platelets distinct from that for factor XI.

Binding of 125I-Factor XIa to platelets required the presence of high molecular weight kininogen, was enhanced when platelets were stimulated with thrombin, and reached a plateau after 4-6 min of incubation at 37 degrees C. Factor XIa binding was specific: 50- to 100-fold molar excesses of unlabeled Factor XIa prevented binding, whereas Factor XI, prekallikrein, Factor XIIa, and prothrombin did not. When washed erythrocytes, added at concentrations calculated to provide an equivalent surface area to platelets, were incubated with Factor XIa, only a low level of nonspecific, nonsaturable binding was detected. Factor XIa binding to platelets was partially reversible and was saturable at concentrations of added Factor XIa of 0.2-0.4 microgram/ml (1.25-2.5 microM). The number of Factor XIa binding sites on activated platelets was estimated to be 225 per platelet (range, 110-450). We conclude that specific, high affinity, saturable binding sites for Factor XIa are present on activated platelets, are distinct from those previously demonstrated for Factor XI, and require the presence of high molecular weight kininogen.

Binding, Competitive↗
PubMed1984

Immunological studies of human placentae: complement components in pre-eclamptic chorionic villi.

Forty human placentae were studied by immunofluorescence for the presence of complement (C) components C1q, C4, C3d, C6 and C9 with the use of characterized antisera. The tissues were grouped as control placentae from 20 normal pregnancies and 20 from cases of pre-eclampsia (PE): the PE samples were sub-grouped as being obtained from patients with mild or severe PE. All of the C components studied were found in the same distribution for test and control samples, but statistical analysis of each pattern of distribution revealed that the deposition of C1q, C3d and C9 were increased in PE as compared to normal tissues. This impression was strengthened by the finding that the deposition of these C components was further increased when chorionic villus immunopathology was compared between mild and severe PE. These data indicate that immunological mechanisms are operating in PE chorionic villi, and they suggest that, among other mechanisms, immune processes may be operative in the pathophysiology of this clinical disease, and that more studies along these lines are in order to rule either in or out this possibility.

Adolescent↗
PubMed1984

Amidolytic assay of human factor XI in plasma: comparison with a coagulant assay and a new rapid radioimmunoassay.

The traditional coagulant assay for plasma factor XI suffers from a relatively high coefficient of variation, the need for rare congenitally deficient plasma, and a poor correlation between precision and sensitivity. We have developed a simple functional amidolytic assay for factor XI in plasma using the chromogenic substrate PyrGlu-Pro-Arg-p-nitroanilide (S-2366). After inactivation of alpha 1-antitrypsin, CI inhibitor, and other plasma protease inhibitors with CHCI3, plasma was incubated with kaolin, in the absence of added calcium, which limited the enzymes formed to those dependent on contact activation. Soybean trypsin inhibitor was used to minimize the action of kallikrein on the substrate. Once the reaction was complete, corn trypsin inhibitor was used to inactive factor XIIa, the enzyme generated by exposure of plasma to negatively charged surfaces, which had activated the factor XI. The assay is highly specific for factor XI, since plasma totally deficient in that zymogen yielded only 1%-3% of the enzymatic activity in normal plasma under identical conditions. The requirements for complete conversion of factor XI to XIa in plasma within 60 min were, respectively, factor XII, 0.6 U/ml, and high molecular weight kininogen, 0.2 U/ml. Prekallikrein was not an absolute requirement for complete activation but did accelerate the reaction. The intraassay coefficient of variation was 3.4%, and the mean of 35 normal plasmas was 1.00 U +/- 0.24 SD. In addition, a new rapid radioimmunoassay was devised using staphylococcal protein A as the precipitating agent for a complex of factor XI antigen with monospecific rabbit antibody. The mean was 1.01 U +/- 0.30 SD. The correlation coefficients for amidolytic versus coagulant and amidolytic versus radioimmunoassay were r = 0.95 for the former and 0.96 for the latter. Thus, a simple, accurate amidolytic assay and a radioimmunoassay have been devised for measuring factor XI in plasma that correlate well with the coagulant activity of factor XI, as determined in our laboratory.

Animals↗
PubMed1982

Specific reactivity of lipid vesicles conjugated with oriented anti-lactose antibody fragments.

The method previously described (Sinha, D. and Karush, F. (1979) Biochem. Biophys. Res. Commun. 90, 554--560) for the oriented attachment of immunoglobulins to lipid vesicles has been used to confer specific reactivity on liposomes by their conjugation with anti-lactose Fab' fragments derived from rabbit IgG antibody. It is estimated that one-third of the Fab' fragments was irreversibly attached to liposomal membrane, resulting in a membrane concentration of 2 mmol of Fab' per mol of total lipid. The specific reactivity of the modified liposomes was demonstrated by agglutination with a multivalent, lactose-containing diheteroglycan. The availability of virtually all of the binding sites of the attached antibody for reaction with ligand was established by a fluorescence quenching titration with N-(N epsilon-Dnp-L-lysyl)-p-aminophenyl-beta-lactoside. An intrinsic association constant of 8.9 x 10(6) M-1 was found for the attached Fab' compared to a value of 2.8 x 10(6) M-1 for free anti-lactose Fab'. In addition the maximum values for the quenching by bound ligand of the fluorescence of free and attached antibody were the same. It can be concluded that the chemical procedures used to effect attachment of the antibody to the lipid vesicles allow retention of the original structure of the antibody site and its accessibility to external components.

Agglutination↗
PubMed1981

Neoplastic growth of carcinogen-treated mammary transplants as influenced by fat intake of donor and host.

Female Wistar-Furth rats were fed either a high-fat (HF) or a low fat (LF) diet from weaning. The HF and LF diets contained 20% and 0.5% corn oil, respectively. Mammary gland explants from 50-day-old rats of both dietary groups were exposed to dimethylbenz[a]anthracene in organ culture before grafting to HF- or LF-hosts. A total of 4 groups were involved: LF leads to LF, HF leads to LF, LF leads to HF, and HF leads to HF, where the designations before and after the arrow describe the dietary treatment of the donor and host, respectively. Final tumor incidences were as follows: 28%, 20%, 72% and 76%, presented in the order of the above 4 groups. Tumors also appeared earlier in the HF-hosts. Results of this experiment indicate that regardless of the nutritional status of the donor, it was the fat intake of the host that governed the subsequent neoplastic growth of transformed cells, suggesting that the action of fat is primarily exerted at the promotional stage of carcinogenesis.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1981

Anticarcinogenic effect of selenium in rats treated with dimethylbenz[a]anthracene and fed different levels and types of fat.

The present investigation reports the effect of selenium supplementation on 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary carcinogenesis in rats fed either a 5% or a 25% corn oil diet. A reduction in tumorigenesis in both groups was observed with 2.5 p.p.m. of dietary selenium. Selenium supplementation also inhibited the development of hyperplastic alveolar nodules in the mammary gland subsequent to DMBA treatment. In addition, the appearance of mammary neoplasia was reduced by selenium in rats fed a high-saturated fat diet (coconut oil), indicating that the type of fat consumed did not influence the antitumorigenic effectiveness of selenium. The lack of a correlation between the anticarciongenic efficacy of selenium and its ability to suppress lipid peroxidation in the mammary tissue of rats fed either a high-saturated fat or a high-unsaturated fat diet suggests that the inhibitory action of selenium is probably not mediated by its antioxidant function in lipid metabolism

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1977

Hyperplastic alveolar nodules of the rat mammary gland: tumor-producing capability in vivo and in vitro.

Carcinogen-induced hyperplastic alveolar nodules failed to form tumors after transplantation into isologous female rats; instead they developed ductal or ductal-alveolar outgrowths. Mammary glands from mid-pregnant female rats showed similar outgrowths following transplantation to the isologous hosts. In vitro and in vivo studies failed to demonstrate that subcarcinogenic doses of 7,12-dimethylbenz(a)anthracene can induce mammary tumorigenesis in the hyperplastic alveolar nodules. Our observations suggest that carcinogen-induced hyperplastic alveolar nodules are not 'preneoplastic' lesions in mammary carcinogenesis in the rat.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1976

Neonatal modification of endocrine functions and mammary carcinogenesis in the rat.

Effects of neonatal androgenization or neonatal ovariectomy in female rats on endocrine functions and mammary tumourigenesis are examined. Pituitary gonadotrophin contents (both LH and FSH) are significantly lower in neonatally androgenized rats (TT) and significantly increased in neonatally ovariectomized rats (NO) when compared with controls of the same age. Plasma and pituitary prolactin levels are higher in TT rats than in the control rats of the same age, but the difference is not significant. Mammary tumours developing in TT rats after DMBA treatment are predominantly fibroadenomata, and lactogenesis in TT rats occurs almost entirely in those receiving DMBA treatment. Neonatal ovariectomy in female rats protects against subsequent induction of mammary cnacer by DMBA. The relationship between neonatal modification of endocrine functions and mammary tumourigenesis is discussed.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1975

Site of origin of mammary tumors induced by 7,12-dimethylbenz(a)anthracene in the rat.

Serial sections of mammary glands from female Sprague-Dawley rats were examined at different intervals after rats were treated with 7,12-dimethylbenz[a]anthracene (CMBA). DMBA was given either by intravenous (iv) injection or by local application. Lesions with histologic characteristics typical of mammary adenocarcinomas were observed in the mammary glands as early as 20 days after DMBA treatment, when no other lesions were detectable. Whereas hyperplastic alveolar nodules in the mammary glands began to appear 35 days after iv injection of DMBA, they were never observed after local application of the carcinogen. All mammary tumors were adenocarcinomas of ductal origin.

9,10-Dimethyl-1,2-benzanthracene↗