Hormonal regulation of activities of 4-ene-5 beta- and 5 alpha-reductases and 17 beta-hydroxy-dehydrogenase in immature golden hamster testis

We have reported [1-3] in immature golden hamster testis that 5 beta-reductase is localized in the tubular nongerm cells, while 5 alpha-reductase is present in the interstitial tissue and that the 17 beta-hydroxy-dehydrogenase activity is found predominantly in the tubular nongerm cells. Hormonal regulation of these enzyme activities was examined in the present study. Male golden hamsters were hypophysectomized on day 22 after birth. The hypophysectomized hamsters in groups of 3-8 were injected daily with 10 micrograms NIH-LH-S19, 50 micrograms NIAMD-Rat-FSH-B-1, 8 or 16 micrograms NIAMD-oFSH-13, 8 micrograms NIAMD-oFSH-13 plus 5 or 10 micrograms NIH-LH-S19, 1 mg testosterone propionate or saline for 5 days starting from day 23. Testicular homogenates of the treated hamsters and intact hamsters on day 28 were incubated with [14C]4-androstene-3,17-dione and NADPH, and enzyme activity (nmol/testes/h) was estimated. The activities of 5 beta- and 5 alpha-reductases and 17 beta-hydroxy-dehydrogenase decreased significantly 6 days after hypophysectomy. In the hypophysectomized hamster testis, a distinct response to FSH but not to LH in the activities of 5 beta-reductase and 17 beta-hydroxy-dehydrogenase was found. The injection of LH in addition to FSH showed no significant additive effects on these enzyme activities. The 5 alpha-reductase activity was stimulated significantly by LH plus FSH but not by LH alone, FSH alone or androgen. These results show that 5 beta-reduction of 4-ene-3-ketosteroids takes place in the Sertoli cells under the influence of FSH while 5 alpha-reduction occurs in the interstitial cells under the influence of LH and FSH in immature hamster testis.     

Ontogenesis of gastrin cells in the pyloric antrum and duodenum of the mouse

Summary Ontogenesis of gastrin cells was studied in the pyloroduodenal mucosa of the mouse using anti-human G17 serum, R-1301, and anti-human G34(1–15) serum, R-2703. R-1301-immunostained cells first appeared in the pyloric mucosa of 14-day-old fetuses. Cells stained with both R-1301 and R-2703 appeared immediately after birth, and gradually increased in number to the adult level. Most R-1301-reactive cells were also reactive to R-2703, whereas some cells that reacted with R-1301 exhibited very weak or no reaction with R-2703. The discrepancy between these two immunoreactivities is discussed.In the duodenum, a considerable number of R-1301-reactive cells were present from the perinatal stage and through out adult development. A few R-2703-reactive cells were seen in the duodenum of young mice but not of the adult.     

GQ1b, a bioactive ganglioside that exhibits novel nerve growth factor (NGF)-like activities in the two neuroblastoma cell lines

To clarify the role of gangliosides in the morphological and biochemical differentiation of neuronal cell cultures, the model cell culture system represented by two neuroblastoma cell lines, GOTO and NB-1, which were established from adrenal gland and metastatic neck lymph node, respectively, was examined. We found that the total ganglioside fraction from human brain had two remarkable effects on these cell lines, which are similar to those of nerve growth factor (NGF): (a) an increase in the cell number, and (b) an increase in the neurite number and the total length of neurites. In these cases, the genuine effector in total gangliosides could not be ascribed to a possibly contaminating NGF-like protein, but rather to a particular molecular species of the gangliosides, GQ1b, which could completely replace the effector function not only qualitatively but also quantitatively. Our results provide direct evidence for the participation of gangliosides in such functions.     

Correlation between the presence of T-antigen and the reinitiation of host DNA synthesis in senescent human diploid fibroblasts after SV40 infection

Senescent human diploid fibroblasts, TIG-1, had labelling indices of about 0.5-3% when labelled with [3H]thymidine for 3 days in fresh medium containing 10% fetal bovine serum. When these cells were infected with SV40, the percentage of nuclei incorporating [3H]thymidine increased by about 10-fold. The frequency of T-antigen-positive cells and that of [3H]thymidine-incorporating cells were almost the same. About 80% of T-antigen-positive cells were also positive to incorporation of [3H]thymidine, and the same result was obtained in infected young cells. These results indicated that senescent human diploid cells which are brought to synthesize T-antigen always initiate DNA synthesis as young cells do. The characteristics of senescent cells as compared with younger cells was low incidence of T-antigen-positive cells after infection. The basis of low susceptibility of senescent cells to initiate DNA synthesis by SV40 infection thus seems to be concerned with an event after the adsorption of virus, but before the synthesis of a detectable amount of T-antigen.     

Prostaglandin stimulation of adenosine 3',5'-monophosphate accumulation in cultured chondrocytes in the presence or absence of parathyroid hormone

The effect of prostaglandin analogues on the cyclic AMP level in cultured chondrocytes were examined. Prostaglandin E1 at 0.4 to 30 microM, increased the intracellular concentration of cyclic AMP in chondrocytes. Its effect was rapid, being evident within 1 min and reaching a maximum in 10 to 20 min. The maximum level was sustained until 30 min after its addition and then decreased gradually. Prostaglandin D2 and E2 also increased the cyclic AMP level in chondrocytes, but they had less effect than prostaglandin E1. Prostaglandin A1 had no effect on the nucleotide level in chondrocytes, although they markedly increased the level in fibroblasts. The time course of stimulation of cyclic AMP accumulation in chondrocytes by prostaglandin E1, D2 or E2 was quite different from that by parathyroid hormone (PTH): the effect of prostaglandin was slower and more sustained than that of PTH. PTH potentiated the effect of prostaglandin E1, E2, or D2 on the cyclic AMP level in chondrocytes and that the combined effects of prostaglandin and PTH were more than additive. Addition of an inhibitor of cyclic nucleotide phosphodiesterase with prostaglandin, PTH or both produced a synergistic effect on the accumulation of cyclic AMP in the chondrocytes. These findings suggest that prostaglandin E1, E2, and D2 increase the synthesis of cyclic AMP and that the combined effect of the prostaglandins and PTH on the cyclic AMP level in chondrocytes is partly attributed to the synergistic synthesis of cyclic AMP in the cells.     

Membrane Potentials of Heterotrophically Cultured Tobacco Cells

The electrophysiological properties of the membrane of Nicotianatabacum var. Sarnsun cultured cells were determined using amicroelectrode technique in standard medium containing 1 mMKC1, 1 mM NaCl and 1 mu CaCl₂ at pH 7. Tobacco callus was derivedfrom the pith (E_m=–104.4%16.2 mV). The membrane potentialsof the callus cells did not show a symmetrical Gaussian distributionbut were scattered over a wide range. The percentage of highmembrane potential cells increased as the subculture was continueduntil about 11 months and then decreased. The response of themembrane potential to electric stimulus, ionic composition,metabolic inhibitors, sugars and amino acids was characteristicof high (E_m=–{small tilde}–160 220 mV; H-cells)and low (E_m=–80{small tilde}–90 mV; L-cells) membranepotential cells. The membrane potential of H-cells was largelydepolarized by addition of CN^–, carbonium cyanide m-chlorophenylhydrazone,decyclohexylcarbodiimide, and triphenyltin chloride and transientlydepolarized by addition of glucose, galactose, mannose or sucrose,and D-alanine, L-alanine or Llysine, but the membrane potentialof L-cells was not. (Received December 3, 1982; Accepted March 16, 1983)     

Formation of Chlorophyll-Protein Complexes in Greening Cucumber Cotyledons in Light and then in Darkness

The changes in chlorophyll-protein complexes (CPs) in cucumbercotyledons during illumination and subsequent dark incubationwere studied by SDS-polyacrylamide gel electrophoresis. Whenetiolated cucumber seedlings were illuminated, chlorophyll wassynthesized and CPs were formed. In the early phase of greening(6 h of illumination), light-harvesting chlorophyll a/b-proteincomplex (LHCP) was the main GP. As the greening proceeded, P700chlorophyll a-protein complex (CP1) accumulated. When 6-h illuminatedseedlings were transferred to darkness, CP1 accumulated concomitantlywith a decrease in LHCP without new chlorophyll synthesis. Thechanges in the amounts of CPs in the dark became smaller withthe progress of greening and were not observed after 72 h ofillumination. These changes were confirmed by examining thechlorophyll/P700 ratio and the low temperature absorption spectrumof cotyledons. These results suggest that in the early phaseof greening, CPs were unstable and their chlorophyll moleculeseasily exchanged with those of other kinds of CPs. (Received October 14, 1982; Accepted December 1, 1982)     

Proliferative kinetics and differentiation of murine blast cell colonies in culture: Evidence for variable G0 periods and constant doubling rates of early pluripotent hemopoietic progenitors

Several investigators have described hemopoietic colonies expressing multilineage differentiation in culture. We recently identified a class of murine hemopoletic progenitors which form blast cell colonies with very high replating efficiencies. In order to clarify further the relationship between progenitors for blast cell colonies and progenitors for the multilineage hemopoietic colonies in culture, we carried out analyses of kinetic and differentiation properties of murine blast cell colonies. Serial observations of the development of blast cell colonies into multilineage (and single lineage) colonies in cultures of spleen cells obtained from 5-fluorouracil (5-FU)-treated mice confirmed the transitional nature of the murine blast cell colonies. The data also suggested that the early pluripotent progenitors are in G₀ for variable periods, and that when triggered into cell cycle, they proliferate at relatively constant doubling rates during the early stages of differentiation. The notion that some of the pluripotent progenitors are in G₀ was also supported by long-term thymidine suicide studies in which spleen cells were exposed to ³H-thymidine with high specific activity for 5 days in culture, washed, and assayed for surviving progenitors. Comparison of replating abilities of day-7 and day-16 blast cell colonies from normal as well as 5-FU-treated mice indicated that some of the day-7 blast cell colonies are derived from maturer populations of progenitors which are sensitive to 5-FU. In contrast, progenitors for the day-16 blast cell colonies are dormant in cell cycle and were not affected by 5-FU treatment. Previously we reported that progenitors for day-16 blast cell colonies have a significant capacity for self-renewal. These observations suggest the hypothesis that the capability for self-renewal is accompanied by long periods of G₀, and that once commitment to differentiation takes place, then active cell division occurs.