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Biomedical subjects

Y I Shethna

Publications and source records attributed to Y I Shethna.

At least 19 recordsLinked to original sources

A modified bioassay for microbial mosquito larvicides.

The proposed bioassay-the grid assay-eliminates the influence of cannibalism while allowing recycling to take place. The LC50 values of the grid assay were intermediate to those of the group and the individual assay. The grid assay thus reflects the true toxicity values of the microbial pesticides being tested. Viable count studies of the microbial pesticides employed, viz., Bacillus thuringiensis H-14 and Bacillus sphaericus 2362, performed along with the assay support the results of the bioassay.

Animals↗

Isolation & characterization of angiogenic factors from cultured cells & conditioned media.

Angiogenic factors were isolated by ion exchange chromatography from three established cell lines viz., HEp 2, HeLa and CHO, a primary culture of mouse mammary adenocarcinoma and from conditioned media of HEp2 and cultured mammary adenocarcinoma cells. The angiogenic activity was assayed by chicken chorioallantoic membrane assay. The angiogenic factor eluted at 0.5 M, exhibited lambda max at 258 +/- 1 nm, contained protein and nucleic acid. The cellular angiogenic factor showed a ratio of 1:1 for protein and nucleic acid whereas the secreted angiogenic factor had 3-5 parts of protein to 1 part of nucleic acid. The angiogenic factor from HEp2 and CHO cells did not bind to heparin-agarose. Microheterogeneity of the angiogenic factors was established by SDS-PAGE. Antiserum raised against the cellular angiogenic factor from HEp2 cells, showed a titre of 1:1600 by ELISA. The angiogenic factor was directly localized on whole cells by ELISA. Cellular as well as secreted angiogenic factors crossreacted with the antibody. Neutralizing effect of the antiserum on induction of angiogenesis was detected on chicken chorioallantoic membrane.

Angiogenesis Inducing Agents↗

Differential cell behaviour on polylysine and gelatin in primary culture of mouse mammary adenocarcinoma.

Cells dissociated from spontaneous and transplanted tumours of C3HJax mammary gland have been cultured on polylysine and gelatin substrates. The isolated cells proliferated to form monolayers with high degree of organoid structure as indicated by formation of alveolar cavities. Differences were observed in the cell attachment, growth pattern, number and size of alveolar cavities, cells which lined the cavity and cell morphology on polylysine and gelatin substrates as compared to conventional cell culture plastic surface. On polylysine more than 90% cells attached rapidly, within 15-45 min after plating, with or without serum and formed confluent monolayers marked by presence of large and small alveolar cavities. Multiple interacting cell types took part in organization of the cavity. Cells lining the cavity constantly proliferated and rearranged to expand it. On gelatin, 60-70% cells attached over a period of 6-24 hr in presence of serum and formed confluent monolayers dominated by small alveolar cavities. Cells forming the cavities were epithelial in nature and cavities once formed did not increase in size. Upon subculture, the cell morphology on these substrates was strikingly different. On polylysine, the predominant cell type had numerous irregular microvilli whereas on gelatin, cells had smoother boundaries with a few stunted cytoplasmic extensions. The cell attachment on conventional surface was low, 40-50%. When seeded at high cell density, formation of alveolar cavities was suppressed and at low cell density, cultures were marked by contact inhibition of cells and failure to attain confluence. These results suggest differential behaviour and interaction of mammary tumour epithelium with the substrates used.

Adenocarcinoma↗

Production & formulation of Bacillus thuringiensis var. israelensis & B. sphaericus 1593.

Three fermentation media each for bulk growth of B. thuringiensis var. israelensis and B. sphaericus 1593 were formulated using defatted groundnut cake (Arachis hypogea) as the first nitrogen source and gram flour (Cicer arientinum), soy bean (Glycine max) and defatted milk powder as the second nitrogen source. Medium containing gram flour showed highest toxicity (14.45 micrograms/l) in case of B. thuringiensis var. israelensis whereas medium containing milk powder was found to be highly toxic with B. sphaericus 1593 (51.39 micrograms/l). Sustained release floating pellet formulations of B. thuringiensis var. israelensis and B. sphaericus 1593 exhibited toxicity of 77 per cent and above for 42 days at a dose of 500 micrograms/l for 4th instar larvae of Culex pipiens quinquefasciatus Say.

Animals↗

Isolation & characterization of tumour angiogenesis factor from solid tumours & body fluids from cancer patients.

Tumour angiogenesis factor (TAF) was isolated from malignant solid tumours (10) and from pleural and peritoneal fluids (10) collected from cancer patients. Normal tissues and body fluids from individuals with no clinical history of cancer did not show any detectable levels of TAF. Also, no angiogenic activity was detectable in the benign tumour samples studied (2). The TAFs isolated were all ribonucleoproteins. Molecular weight determination by SDS-PAGE (9%) of the TAFs isolated by DEAE cellulose chromatography of tumour extracts showed them to be 18,000 dalton (D) ribonucleoproteins, while the TAFs isolated by immunoaffinity chromatography (using immobilized anti-TAF IgG) of solid tumour extracts and body fluids had a molecular weight of 38,000 D. The TAFs isolated by both the methods were found to be angiogenic by the chick chorioallantoic membrane and the mouse intradermal assays. Immunoaffinity chromatography could be used for the one-step purification of TAF from solid tumour extracts as well as from body fluids.

Angiogenesis Inducing Agents↗

Growth & toxicity of Bacillus thuringiensis var israelensis.

Chemically defined media containing glutamic acid, glutamine and aspartic acid at a 20 mM concentration individually supported abundant growth and sporulation of Bacillus thuringiensis var israelensis. The parasporal crystals produced in these media were toxic to fourth instar larvae of Culex pipiens quinquefasciatus Say. The medium containing 20 mM of glutamine induced early sporulation of B. thuringiensis var israelensis. An LC50 value of 3.92 micrograms/l was obtained. Calcium alginate and agar agar immobilized formulations of Bacillus thuringiensis var israelensis exhibited toxicity of 90 per cent and above to C. pipiens quinquefasciatus Say larvae, over a period of 90 days.

Animals↗

Purification and Some Biological Properties of Asparaginase from Azotobacter vinelandii.

Asparaginase was found in the soluble fraction of cells of Azotobacter vinelandii, and its activity remained the same during growth of the organism in a nitrogen-free medium. The specific activity and the yield of A. vinelandii increased twofold in the presence of ammonium sulfate. Within limits, the temperature (30 to 37 degrees C) and pH (6.5 to 8.0) of the medium showed little effect on the levels of enzyme activity. The enzyme was purified to near homogeneity by standard methods of enzyme purification, including affinity chromatography, and had optimum activity at pH 8.6 and 48 degrees C. The approximate molecular weight was 84,000. The apparent K(m) value for the substrate was 1.1 x 10 M. Metal ions or sulfhydryl reagents were not required for enzyme activity. Cu, Zn, and Hg showed concentration-dependent inhibition, whereas amino and keto acids had no effect on the enzyme activity. Asparaginase was stable when incubated with rat serum and ascites fluid. The enzyme had no effect on the membrane of sheep erythrocytes and did not inhibit the incorporation of radioactive precursors into deoxyribonucleic acid, ribonucleic acid, and protein in Yoshida ascites sarcoma cells. Asparaginase activity was not detected in the tumor cells.

Journal Article↗

Oxalate, formate, formamide, and methanol metabolism in Thiobacillus novellus.

Thiobacillus novellus was able to grow with oxalate, formate, formamide, and methanol as sole sources of carbon and energy. Extensive growth on methanol required yeast extract or vitamins. Glyoxylate carboligase was detected in extracts of oxalate-grown cells. Ribulose bisphosphate carboxylase was found in extracts of cells grown on formate, formamide, and thiosulfate. These data indicate that oxalate is utilized heterotrophically in the glycerate pathway, and formate and formamide are utilized autotrophically in the ribulose bisphosphate pathway. Nicotinamide adenine dinucleotide-linked formate dehydrogenase was present in extracts of oxalate-, formate-, formamide-, and methanol-grown cells but was absent in thiosulfate- and acetate-grown cells.

Alcohol Oxidoreductases↗

Purification and properties of an NADP+-specific isocitrate dehydrogenase from Rhizobium meliloti.

An NADP+-specific isocitrate dehydrogenase has been purified and characterized from Rhizobium meliloti. The enzyme showed Mn++ or Mg++ requirement. The apparent Km values were 2.00 x 10(-5) M and 1.51 x 10(-5) M for DL-isocitrate and NADP+, respectively. The enzyme was inhibited by ATP, to a lesser extent by ADP and AMP. alpha-Ketoglutarate also inhibited the enzyme activity. Oxalacetate and glyoxylate together inhibited the enzyme activity. The inhibition was competitive. Studies with thiol inhibitors suggested that the enzyme contained a sulfhydryl group at or near the active site. The enzyme has an approximate molecular weight of 60,000. Fluorescence studies suggested that the enzyme contained tryptophan.

Adenine Nucleotides↗

Mode of action of a purified antitumor protein from the proteinaceous crystal of Baccillus thuringiensis subsp. thuringiensis on Yoshida ascites sarcoma cells.

A purified antitumor protein from the proteinaceous crystal of Bacillus thuringiensis subsp. thuringiensis inhibits the growth of Yoshida ascites sarcoma both in vivo and in vitro. Exogenous respiration of the tumor cells was unaffected by the protein at a concentration as high as 500 mug/ml. The antitumor protein inhibits the uptake and incorporation of labeled precursors into macromolecules. However, the ratio of incorporation over uptake is not affected by the protein. Further, the protein brings about the leakage of 260-nm-absorbing material, proteins, and (32)P-labeled cellular constituents from the Yoshida ascites sarcoma cells. The results show that the action of the antitumor protein appears to alter the cellular permeability of the tumor cells.

Animals↗

Isolation and characterization of some new oxalate-decomposing bacteria.

Forty-one cultures degrading and assimilating oxalate were isolated from chicken dung. Characterizarion indicated six different types. One of these belonged to thhe genus Alcaligenes hitherto never reported to degrade oxalte. Three groups of Pseudomonas strains differed physiologically from strains already known.

Alcaligenes↗

Oxalate and formate in Alcaligenes and Pseudomonas species.

Oxalate is metabolized by the glycerate pathway involving glyoxylate carboligase in Alcaligenes LOx and Pseudomonas KOx, and by the serine pathway involving hydroxypyruvate reductase in Ps.MOx and Ps.AM1 (var. 470). Although A.LOx does not grow on formate, stimulation of growth was observed in the presence of amino acids and a few Kreb's cycle intermediates. A.LOx possesses two different mechanisms for the oxidation of formate: (1) the constitutive formate oxidase which is present in the particulate fraction of oxalate-grown and succinate-plus-formate-grown cells; (2) the inducible NAD-linked formate dehydrogenase present in the 100 000 x g supernatant fraction of the cell-free extracts of oxalate-grown cells alone. The two systems occur simultaneously in oxalate-grown cells. The effect of inhibitors on formate oxidase activity and the other enzyme activities of the particulate formate-oxidizing fraction indicate that the oxidation of formate is linked to the respiratory chain.

Alcaligenes↗