Sertoli Cell Morphology Research Articles

Effect of some phthalate esters and other testicular toxins on primary cultures of testicular cells

Mixed cultures of Sertoli and germ cells were prepared from rat testes and their response to some model testicular toxins was studied. Cultures consisted of a monolayer of Sertoli cells to which clusters of spermatocytes and spermatogonia adhered. With time in culture, germ cells progressively detached from the Sertoli cells into the medium. Addition of mono-(2-ethylhexyl) phthalate (MEHP) to the culture medium resulted in a concentration-dependent increase in the rate of germ-cell detachment over the range 10 −7–10 −4 m. No such effect was produced by di-(2-ethylhexyl) phthalate or 2-ethylhexanol. An increased rate of germ-cell detachment was also produced by other phthalate monoesters known to cause testicular damage in vivo, whereas similar concentrations of a number of monophthalates not known to affect the testis in vivo had no such effect on the cultures. Known age and species differences in the testicular toxicity of di-(2-ethylhexyl) phthalate could be reproduced in cultures treated with MEHP. There was little effect on the viability of either germ cells or Sertoli cells at concentrations of MEHP that caused marked germ-cell detachment, but there were changes in Sertoli-cell morphology. Increased germ-cell detachment was also observed in cultures treated with 10 −7–10 −4 m-AF1312/TS, a compound that affects Sertoli cells in vivo, but was not seen in cultures exposed to a range of other testicular toxins with different target cells in the testis. Thus, the effects produced by phthalate monoesters in vitro may reflect damage to the Sertoli cells. Testicular cell cultures could be of value both for screening compounds and for studying underlying mechanisms of testicular toxicity.

Effects of gossypol on rat Sertoli and Leydig cells in primary culture and established cell lines.

In order to evaluate the direct effect of gossypol on testicular cells, we used primary cultures of rat Leydig and Sertoli cells. No alteration in Leydig cell survival, morphology, or testosterone production was seen during three days of culture with up to 3 microgram/ml gossypol. With higher concentrations (3 to 7 microgram/ml) of gossypol, there was a reduction in cell survival but no change in androgen secretion. In contrast, there was a marked change in Sertoli cell morphology after five days of gossypol treatment. Large vacuoles and electron dense granules appeared in the cytoplasm, but these effects were reversed within six days of removing gossypol from the medium. There was a significant decrease in androgen binding protein (ABP) secretion by Sertoli cells in the presence of gossypol. We also tested the effect of gossypol on the growth of three established cell lines. Two Sertoli-derived cell lines, TM4 and TR-ST, were more sensitive than a Leydig-derived cell line (TM3). These results suggest that, of the somatic cell types in the testis, the Sertoli cells are most sensitive to gossypol.

Fine structure and FSH binding of Sertoli cells in the blue fox (Alopex lagopus) in different stages of reproductive activity.

A clearly different Sertoli cell morphology was found in the immature blue fox and in the adults in and out of season. Immature cells had small, basally, located nuclei rich in peripheral heterochromatin, and few cytoplasmic organelles. Sertoli cells from adults in season had basally located, large, convoluted nuclei with homogenous nucleoplasma and prominent nucleoli. The basal and intermediate parts of the cytoplasm contained an extensively developed ER, numerous mitochondria, free ribosomes, lipid droplets and residual bodies, while the apical cytoplasm showed few distinct structures. In Sertoli cells from adults out of season the nuclei were dislocated towards the lumen, and apart from numerous dilated cisternae of ER, there were generally fewer organelles, but more glycogen particles. A marked rise in FSH binding was found towards the breeding season.

Effects of serum components on the morphology of Sertoli cells in culture.

Studies of Sertoli cell structure, maturation, and function have been aided by the use of in vitro systems. Although numerous papers have appeared that utilize the Sertoli cell culture model, few papers have dealt with the characterization of these cells under various culture environments. Recently, it has been reported that the addition of serum to the culture medium prevents induction of long cytoplasmic appendages in cultured Sertoli cells that have been treated with FSH, TSH, or c-AMP. The purpose of this investigation was to determine which serum components, obtained by gel filtration, are capable of inhibiting the morphological response induced by FSH, TSH, or c-AMP. Sertoli cell-enriched cultures were prepared using collagenase and trypsin digestion, each followed by gravity sedimentation. Untreated cells grown on plastic or glass substrates assumed an epithelioid appearance after several days. Cells treated with FSH, TSH, or c-AMP formed long cytoplasmic appendages after 1-2 days. This response was prevented or reversed by the addition of fetal calf serum (10%), crystallized bovine serum albumin (0.25%-2%), or purified albumin obtained by gel filtration of whole serum (0.25%). It was also found that fractions that elute between the void volume and the initial albumin fractions (molecular weights of approximately 50,000 and greater) mimic the hormone-induced response after only 10-12 hours. The results of this investigation indicate that albumin is the primary serum component responsible for inhibiting morphological alterations induced by FSH, TSH, and c-AMP. Furthermore, it is apparent that the production of long filamentous cytoplasmic appendages in Sertoli cells can be induced by a wide variety of substances.