Biomedical subjects
Publications and source records attributed to R K Kulkarni.
The calmodulin content of the yeast and mycelial phases of Ceratocystis ulmi was determined by radioimmunoassay. Calmodulin levels increased at the G1-S boundary of the cell cycle, coinciding with the first visible appearance of buds or germ tubes. However, in both phases the cellular calmodulin levels were equivalent. No differential synthesis was observed.
The addition of the tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA) to serum-starved quiescent Reuber H35 hepatoma cells results in a rapid 5- to 11-fold increase in the incorporation of 32Pi into a Mr = 32,000 ribosomal protein. The Mr = 32,000 protein was the major phosphorylated protein extracted from isolated 80 S ribosomes and was identified as the 40 S ribosomal protein S6 based upon its migration in two-dimensional gels. Insulin, which has been demonstrated to increase the phosphorylation of S6 in a number of cell lines, caused a 10- to 20-fold increase in the incorporation of 32Pi into this Mr = 32,000 ribosomal protein. S6 phosphorylation was dose- and time-dependent being detected as early as 5 min following the addition of 1.6 microM TPA. Maximal phosphorylation of ribosomal protein S6 was achieved by 60 min and remained elevated for at least 90 min in the presence of TPA. The 50% effective dose for TPA was estimated to be 0.14 microM. Based upon the altered migration of S6 in pH 8.5 urea-polyacrylamide gels, it was demonstrated that the increased 32Pi labeling of S6 by TPA was due to a net increase in the incorporation of phosphates into the S6 molecule. Non-tumor-promoting phorbol esters were ineffective in increasing the phosphorylation of S6. In whole cells, exogenously added 1 mM 8-bromoadenosine 3':5'-monophosphate failed to substantially increase phosphorylation of S6 suggesting that the TPA-induced phosphorylation of S6 occurs via a cyclic AMP-independent mechanism. The S6 amino acid residue phosphorylated in response to TPA was phosphoserine. A possible role for protein kinase C in the phosphorylation of ribosomal protein S6 is discussed.
The possible role of insulin-mediated phosphorylation of ribosomal protein S6 in the control of cell proliferation was examined in insulin-unresponsive mouse melanoma calls (PG19) and insulin-responsive melanoma x fibroblast clone 100A. In the hybrid cells, under conditions of growth arrest in medium with low serum, ribosomal protein S6 was rapidly phosphorylated in response to insulin or serum. The phosphorylation of the S6 protein increased over a wide range of insulin concentrations, suggesting that insulin stimulated the phosphorylation by interacting with both high- and low-affinity receptors. In contrast, in growth-arrested melanoma cells, an intermediate level of S6 phosphorylation was observed. Insulin caused only a marginal increase and serum caused a small but consistent increase in the level of S6 phosphorylation in the melanoma cells. Cell cycle analysis revealed that both cell lines arrested growth to a similar degree in the G1 phase of the cell cycle; thus, the higher baseline level of S6 phosphorylation observed in the melanoma cells was not attributable to less complete growth arrest of these cells in medium with low serum. The S6 phosphorylation results correlate well with previous results suggesting that the hybrid cells, but not the parental melanoma cells, can become growth-limited for processes regulated by insulin.
Clinical isolates of C. albicans were grown in defined media with various carbon sources then examined by scanning electron microscopy in an attempt to note changes in cell surface features. The mode of budding, bud scar morphology, surface topography, and intercellular matrix varied according to the carbon source.
An in vitro method has been developed for screening of candidate biomaterials in an early phase of their development. The test is based on L-929 mouse fibroblast cultures and their response to powdered polymer samples. It applies microscopic observation for the detection of morphological changes, uses dye exclusion testing for cell viability determination, and utilizes estimation of population doublings as an end point. The test is shown to be reliable and reproducible and is compared to in vivo implantation studies in rats, previously reported.
An in vivo method is described for screening polymeric materials for biocompatibility. The test is based on grading acute and subacute tissue reactions at 7 and 28 days, respectively, following implantation in rats. The methods is reproducible and reliable. It is designed to provide uniform test criteria for biocompatibility assessment in the early phases of the development of surgical implant materials.
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