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• 1. [Article] Cellular dynamics of the intestinal epithelium of coho salmon, Oncorhynchus kisutch, as influenced by temperature and x-rays

  The cellular dynamics of the intestinal epithelium in the coho salmon, Oncorhynchus kisutch, has been studied to determine the effects of temperature and whole-body X-irradiation. In vivo experiments utilizing ...

  Citation × Citation Citation Abstract Copy Title: Cellular dynamics of the intestinal epithelium of coho salmon, Oncorhynchus kisutch, as influenced by temperature and x-rays Author: Johnson, Tod Schaefer The cellular dynamics of the intestinal epithelium in the coho salmon, Oncorhynchus kisutch, has been studied to determine the effects of temperature and whole-body X-irradiation. In vivo experiments utilizing colchicine mitotic inhibiting technique and tritiated thymidine (³H-TdR) radioautography were employed in order to evaluate the kinetics of the cell population. In vitro radiotracer enzyme assays were used to measure the conversion of ¹⁴C-2-thymidine to the precursors of DNA synthesis, dTMP, dTDP and dTTP. Colchicine solution at 0.04 mg/100 gm body weight was injected and fish sacrificed from 1 to 18 hours. Arrested mitotic figures occurred only in the basal region of the intestinal folds for all time periods. A series of ³H-TdR experiments were carried out using salmon thermally acclimated at 5°, 10°, or 18° C. Fish were injected intraperitoneally with ³H-TdR solution (1.5 μ C[subscript i]/gm body weight), then sacrificed sequentially from 10 minutes to 37 days after injection. Samples of the intestine were processed either for radioautography or for liquid scintillation studies. Radioautographs indicated initial ³H-TdR cell labeling occurred in the basal region of the intestinal folds, with subsequent cell migration up the folds, and sloughing of cells into the lumen from the tips of the folds. Renewal times were temperature dependent, with estimates of 13-15, 23-25, and in excess of 35 days for 18°, 10° and 5°C respectively. Whole-body X-irradiation at 10° C produced considerable cell damage by 21-22 days after exposure at 1,000 R level. The gastro-intestinal syndrome was manifested by 24 to 26 days after exposures of 4,000 and 8,000 R. ³H-TdR incorporation into the intestinal nucleoprotein fraction was dose dependent through 160 minutes after injection. A series of invitro radiotracer enzyme assay experiments were carried out using ¹⁴C-2-thymidine and the intestinal post-microsomal fraction (> 104,000 x G for 1 hr) from thermally acclimated coho salmon. Thymidine kinase exhibited pH optimum at pH 7.9 ± 0.1 while thymidylate kinase showed maximum activity at pH 6.8 ± 0.1. Formation of dTMP, dTDP, and dTTP exhibited saturation type substrate dependency curves, while dTMP activities measured at 5°, 100, and 18° C resulted in non-competitive inhibition at the lower temperatures. Relative thymidine kinase activities estimated for the different temperature levels were 3.6, 2.5, and 2.0 for 5° -5° C vs 10° -10° C, 5° -5° C vs 18° -18° C, and 10° -10° C vs 18° -18° C, respectively. Lowry protein assays indicated there was an inverse relationship between total protein per unit volume intestinal microsomal supernatant fraction and fish acclimation temperature. Title: Cellular dynamics of the intestinal epithelium of coho salmon, Oncorhynchus kisutch, as influenced by temperature and x-rays Author: Johnson, Tod Schaefer The cellular dynamics of the intestinal epithelium in the coho salmon, Oncorhynchus kisutch, has been studied to determine the effects of temperature and whole-body X-irradiation. In vivo experiments utilizing colchicine mitotic inhibiting technique and tritiated thymidine (³H-TdR) radioautography were employed in order to evaluate the kinetics of the cell population. In vitro radiotracer enzyme assays were used to measure the conversion of ¹⁴C-2-thymidine to the precursors of DNA synthesis, dTMP, dTDP and dTTP. Colchicine solution at 0.04 mg/100 gm body weight was injected and fish sacrificed from 1 to 18 hours. Arrested mitotic figures occurred only in the basal region of the intestinal folds for all time periods. A series of ³H-TdR experiments were carried out using salmon thermally acclimated at 5°, 10°, or 18° C. Fish were injected intraperitoneally with ³H-TdR solution (1.5 μ C[subscript i]/gm body weight), then sacrificed sequentially from 10 minutes to 37 days after injection. Samples of the intestine were processed either for radioautography or for liquid scintillation studies. Radioautographs indicated initial ³H-TdR cell labeling occurred in the basal region of the intestinal folds, with subsequent cell migration up the folds, and sloughing of cells into the lumen from the tips of the folds. Renewal times were temperature dependent, with estimates of 13-15, 23-25, and in excess of 35 days for 18°, 10° and 5°C respectively. Whole-body X-irradiation at 10° C produced considerable cell damage by 21-22 days after exposure at 1,000 R level. The gastro-intestinal syndrome was manifested by 24 to 26 days after exposures of 4,000 and 8,000 R. ³H-TdR incorporation into the intestinal nucleoprotein fraction was dose dependent through 160 minutes after injection. A series of invitro radiotracer enzyme assay experiments were carried out using ¹⁴C-2-thymidine and the intestinal post-microsomal fraction (> 104,000 x G for 1 hr) from thermally acclimated coho salmon. Thymidine kinase exhibited pH optimum at pH 7.9 ± 0.1 while thymidylate kinase showed maximum activity at pH 6.8 ± 0.1. Formation of dTMP, dTDP, and dTTP exhibited saturation type substrate dependency curves, while dTMP activities measured at 5°, 100, and 18° C resulted in non-competitive inhibition at the lower temperatures. Relative thymidine kinase activities estimated for the different temperature levels were 3.6, 2.5, and 2.0 for 5° -5° C vs 10° -10° C, 5° -5° C vs 18° -18° C, and 10° -10° C vs 18° -18° C, respectively. Lowry protein assays indicated there was an inverse relationship between total protein per unit volume intestinal microsomal supernatant fraction and fish acclimation temperature. Close