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S Shubhada

Publications and source records attributed to S Shubhada.

11 recordsLinked to original sources
PubMed1993

Paracrine factors and the regulation of spermatogenesis.

MAIN PROBLEM: Although the gonadotropins and testosterone are required for normal spermatogenesis, it is believed that local control factors regulate spermatogenesis. For many years these regulatory factors had not been identified. Over the past five years, a number of growth factors have been identified in testis or isolated testicular cell types or secretions. Growth factors are key regulatory molecules which affect cell proliferation, meiosis, and differentiated function. These factors usually act in an autocrine (acting upon the cell which secreted it) or paracine (affecting another cell) manner and thus are involved in intercellular communications. METHODS: Growth factor secretion by testicular cell types or testis tissue has been analyzed using a variety of assays measuring cell proliferation in vitro, as well as assays using immunocytochemicals. Growth factor gene expression in testis has been analyzed by Northern blot analysis and in situ hybridization, which gives information concerning the stage and cell specific expression of the gene. Inbred strains of mice with mutations of deletions in a growth factor gene has been used to suggest the function of two specific factors in testicular development and growth. RESULTS: Among the growth factors expressed or secreted by testicular cell types, most are common to some other cell types in the body, such as transforming growth factors alpha and beta, epidermal growth factor, fibroblast-like growth factors, insulin-like growth factors, interleukins, endorphins, inhibin and activin, while others may be more testis specific such as mullerian inhibiting substance (anti-mullerian hormone) and Sertoli cell secreted growth factor.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Borate ion-assisted stabilization of beta-galactosidase from Aspergillus oryzae by polyhydroxy compounds in water-miscible organic solvents.

The stability of beta-galactosidase from Aspergillus oryzae in water-miscible organic solvents in different buffers at various pH values ranging from 4.6 to 8.0 was studied. The stability of the enzyme in all six organic solvents studied was dependent on pH and on the type of buffer ions present. At a given pH, destabilization by organic solvents was highest in sodium borate buffer. The destabilization of beta-galactosidase by these solvents could be reversed by addition of sugars or polyhydroxy compounds exclusively in sodium borate, suggesting a role of borate ions in stabilization. A similar effect of addition of mannitol was observed on deactivation of beta-galactosidase by N, N-dimethylformamide (DMF). Exclusively in sodium borate, at pH 8.0, the addition of mannitol (0.02 M) not only prevented the deactivation by DMF (8%, v/v) but increased the enzyme activity to the level at its optimum pH. Since beta-galactosidase from Aspergillus oryzae is a glycoprotein, complexation of the borate ions to the carbohydrate part may result in change in protein conformation, which, without leading to denaturation or inactivation of the enzyme, may facilitate interaction of the organic solvents with the enzyme leading to its denaturation. Such a denaturation is probably prevented by addition of polyhydroxy compounds, which appear to compete favorably with the carbohydrate moiety of the protein in complexing with borate ions. This should result in the enzyme regaining its native conformation.

Aspergillus oryzae↗
PubMed1991

Partial characterization of a unique mitogenic activity secreted by rat Sertoli cells.

Sertoli cell conditioned medium (SCCM) contains a potent mitogen, Sertoli cell secreted growth factor (SCSGF). A431 cells, derived from a human epidermoid carcinoma have provided an excellent model cell line for the study of this apparently unique activity secreted by rat Sertoli cells in vitro. Previously, it was shown that SCCM contained an epidermal growth factor (EGF)-like activity which was thought to be the mitogen for A431 cells. The present study showed that these two factors are distinct entities. The secretion of the EGF-like activity decreased with increasing number of culture days, while that of SCSGF and of another Sertoli cell specific protein, transferrin remained constant. The addition of SCCM stimulated whereas 2.5 ng/ml EGF inhibited the A431 cell growth. The proliferative response of A431 cells to a wide variety of growth factors and known Sertoli cell secretions was investigated. SCSGF was the only growth factor of known Sertoli cell secretions tested (transforming growth factors (TGF alpha, TGF beta), EGF, bombesin, fibroblast growth factor (FGF), platelet derived growth factor (PDGF), insulin-like growth factors 1 and 2 (IGF-1 and IGF-2), prostaglandins E-1 and E-2, insulin, transferrin and lactate) which stimulated A431 cell proliferation. SCSGF was mitogenic for A431 cells even in the presence of serum in the culture medium. The partially purified SCSGF was heat- and acid-stable, protease-sensitive with a molecular weight of 14,000. It did not bind to heparin or concanavalin A-Sepharose. The secretion of a mitogenic activity by the Sertoli cell which is different from other previously identified growth factors and which coincides with active spermatogenesis could have important implications in the regulation of spermatogenesis.

Animals↗
PubMedDate not supplied

Inhibition of ornithine decarboxylase activity by follicle stimulating hormone in primary culture of rat Sertoli cells.

The effect of follicle stimulating hormone on the activity of ornithine decarboxylase (ODC) was determined in primary culture of rat Sertoli cells. Three different FSH preparations (NIH oFSH-S-15, S-16, and eFSH) inhibited ODC activity in rat Sertoli cells under different media conditions. The inhibition was both time- and dose-dependent. The mechanism of the FSH inhibitory effect was studied using dibutyryl cyclic adenosine monophosphate (dbcAMP), 1-methyl-3-isobutylxanthine (MIX), forskolin, and isoproterenol. All of these agents, known to elevate cellular cAMP levels, inhibited ODC activity in cultured rat Sertoli cells. The combined effect of each of these substances plus FSH was either greater than, or equal to, that of FSH alone, and was not additive. Dibutyryl cyclic guanosine monophosphate had no effect on the ODC activity. These findings suggest that FSH inhibition of ODC activity in the rat Sertoli cell may be mediated by cAMP.

1-Methyl-3-isobutylxanthine↗
PubMedDate not supplied

Polyamine profiles in rat testis, germ cells and Sertoli cells during testicular maturation.

Polyamine cellular concentrations (putrescine, spermidine and spermine) in the rat testis and testicular cell types were determined by fluorescence spectroscopy of their dansyl derivatives. A method is described to separate dansylated polyamines by high performance liquid chromatography in less than 12 minutes. In rat Sertoli cells, polyamine concentrations (per mg DNA) were greater than those in germ cells and the testis. The concentrations of all three polyamines increased with age. Concentrations of spermidine and spermine in germ cells also increased with age and leveled off after 27 to 35 days. On the other hand, higher putrescine levels were found in the testis of young rats (13 to 22 days) while the greatest spermidine and spermine contents were observed in the testis from rats of 31 to 35 days old. Of great interest, Sertoli cells from all age groups studied released a relatively large quantity of putrescine and a smaller amount of spermidine, but no spermine, into culture media. The amount of polyamine released by Sertoli cells varied with the age of the animal. Sertoli cells from 27-day-old rats released the greatest quantity of putrescine on a per mg DNA basis. The release of putrescine increased after hypotonic treatment that removed contaminating germ cells from the remaining Sertoli cells. It is concluded that cellular polyamine levels in the rat testis, germ cells and cultured Sertoli cells and the amount of polyamines released by Sertoli cells were age-dependent during the first wave of spermatogenesis.

Aging↗
PubMedDate not supplied

The changing profiles of L-ornithine decarboxylase and S-adenosyl-L-methionine decarboxylase activities in testicular cell types during sexual maturation of male rats.

To understand the involvement of polyamines in testicular maturation and spermatogenesis, the activity of two enzymes involved in the polyamine synthetic pathway, L-ornithine decarboxylase (ODC) and S-adenosyl-L-methionine decarboxylase (AdoMetDC) were determined in whole testis and isolated testicular cell types during the sexual maturation of male rats. ODC activities per mg protein in whole testis and cultured Sertoli cells from 12- to 14-day-old rats were very high but declined as the age of the animals increased. ODC activities in both germ cells and interstitial cells increased as the age of the rats increased, reached a maximum at 22 days and then declined rapidly. On the other hand, AdoMetDC activity in whole testis, Sertoli cells, germ cells and interstitial cells was highest from 27 to 35 days, 18 to 22 days, 22 to 27 or 35 days, and 27 days of age, respectively, then decreased and remained at a constant lower level after 45 days. After 35 or 45 days of age, cellular AdoMetDC activities were relatively higher than ODC activities in the whole testis and germ cells, while the activity of both enzymes was comparable in the Sertoli cells. Correlation of these data with the development of germ cells during rat testicular maturation suggests that the high ODC activity in both Sertoli and germ cells is associated mainly with cell proliferation while the increase in AdoMetDC activity is most likely associated with the maturation of Sertoli cells and the meiosis of germ cells, particularly the formation of pachytene spermatocytes and the meiotic cell division of spermatocytes into spermatids.

Adenosylmethionine Decarboxylase↗
PubMedDate not supplied

Differential effects of FSH on the activities of S-adenosyl-L-methionine decarboxylase and ornithine decarboxylase in Rat sertoli cells.

Follicle stimulating hormone (FSH) modulates testicular function via Sertoli cells. The effect of FSH on S-adenosyl-L-methionine decarboxylase (AdoMetDC) activity was investigated in cultured Sertoli cells isolated from 18-day-old rats. In contrast to our previous finding that FSH inhibited Sertoli cell ornithine decarboxylase (ODC) activity, FSH stimulated AdoMetDC activity 160 to 300% above the level of the control cells during the initial 2 to 6 hours of treatment. The stimulated enzyme activity declined to 20 to 30% below the control values by 12 hours of exposure to FSH, and then rebounded to 120 to 125% of control values. To determine whether or not the initial opposite effects of FSH on AdoMetDC and on ODC activities are mediated by the same mechanism, various agents which increase intracellular cAMP level were used. All agents studied stimulated Sertoli cell AdoMetDC activity at 5 hours after their inclusion while they significantly inhibited ODC activity. In the presence of FSH, the stimulatory effect of these agents on AdoMetDC was either equal to or slightly greater than that caused by FSH or either agent alone. The combination of dbcGMP with FSH or with dbcAMP resulted in a synergistic or additive effect on AdoMetDC, as well as ODC, activity. The data suggest that the action of FSH on AdoMetDC activity in Sertoli cells is also mediated through cAMP, as concluded previously for ODC activity. Regardless of the effects of FSH on both enzymes, ODC always exhibited a relatively greater activity than did AdoMetDC in cultured rat Sertoli cells.

Adenosylmethionine Decarboxylase↗
PubMedDate not supplied

Sertoli cell secretion of a factor that inhibits the incorporation of 3H-thymidine into cells in vitro.

Rat Sertoli cells secrete a low molecular weight factor in vitro that inhibits the incorporation of 3H-thymidine into cells. Although the addition of Sertoli cell-conditioned medium (rSCCM) resulted in nearly a threefold stimulation of cell growth, the incorporation of 3H-thymidine was decreased in a dose-dependent manner and did not reflect the increase in cell number. Peritubular cell- and germ cell-conditioned medium did not contain this inhibitory activity. Nor did the conditioned medium from fibroblasts and a variety of cell lines tested. A low molecular weight filtrate of rSCCM (< 1,000 Da) contained virtually all of the 3H-thymidine inhibiting activity, as well as about 50% of the mitogenic activity in the rSCCM. The inhibitory activity was eliminated upon removal of the rSCCM and was not due to either growth inhibition or a toxic effect on cell proliferation.

Animals↗
PubMedDate not supplied

Sertoli cell-conditioned medium affects nucleoside utilization in vitro.

Rat Sertoli cell-conditioned medium (SCCM) or low molecular weight filtrate of SCCM (< 1,000 Da molecular weight) inhibited the uptake of [3H]thymidine into cells in culture; [3H]thymidine is incorporated into DNA by means of a salvage pathway. The incorporation of radioactivity into DNA from [14C]N5,N10-methylene-tetrahydrofolate, required for the thymidylate synthetase reaction, was not inhibited by SCCM and reflected the increase in cell number. SCCM specifically inhibited the incorporation of pyrimidines ([3H]uracil, [3H]thymidine) with no effect on the transport or incorporation of [3H]adenosine. Inhibition of thymidine uptake by SCCM could have occurred by a direct competition, i.e., the secretion of thymidine into the medium by the Sertoli cells, or by an indirect mechanism that would result in an inhibition of transport. The activity was partially purified by membrane ultrafiltration, ion exchange chromatography, and sequential extraction. Addition of SCCM filtrate (< 1,000 Da molecular weight) to growth-arrested cells cultured in the presence of inhibitors of dihydrofolate reductase resulted in cell proliferation, suggesting that the factor is involved in thymidine biosynthesis. This activity may play a role in the regulation of nucleoside biosynthesis and/or utilization in the testis.

3T3 Cells↗
PubMedDate not supplied

Sertoli cell secreted growth factor. Cellular origin, paracrine and endocrine regulation of secretion.

Rat and human Sertoli cells in culture secrete a growth factor, Sertoli cell secreted growth factor (SCSGF). The aims of the present study were (1) to evaluate other testicular cell types as additional sources of SCSGF, as well as their paracrine effect, and (2) to study the hormonal regulation of SCSGF secretion using an A431 cell growth assay. The Sertoli cell was the only testicular cell type tested that secreted SCSGF activity in vitro. Peritubular cells enhanced Sertoli cell attachment and SCSGF secretion. Spermatogenic cells had no effect. The secretion of SCSGF was specifically stimulated by follicle-stimulating hormone (FSH) and testosterone. Treatment with agents that increase intracellular cAMP levels and adenosine stimulated the secretion of mitogenic activity into Sertoli cell-conditioned medium by three- to fivefold. This growth factor, secreted by the Sertoli cell and regulated by FSH and testosterone, may play a critical role in the regulation of spermatogenesis.

Adenosine↗