Production of 25-Hydroxycholesterol by Testicular Macrophages and Its Effects on Leydig Cells1

Abstract

Testicular macrophages secrete 25-hydroxycholesterol, which can be converted to testosterone by neighboring Leydig cells. The purposes of the present studies were to determine the mode of production of this oxysterol and its long-term effects on Leydig cells. Because oxysterols are produced both enzymatically and by auto-oxidation, we first determined if testicular macrophages possess cholesterol 25-hydroxylase mRNA and/or if macrophage-secreted products oxidize cholesterol extracellularly. Rat testicular macrophages had 25-hydroxylase mRNA and converted 14C-cholesterol to 14C-25-hydroxycholesterol; however, radiolabeled cholesterol was not converted to 25-hydroxycholesterol when incubated with medium previously exposed to testicular macrophages. Exposure of Leydig cells to 10 microg/ml of 25-hydroxycholesterol, a dose within the range known to result in high basal production of testosterone when tested from 1 to 6 h, completely abolished LH responsiveness after 2 days of treatment. Because 25-hydroxycholesterol is toxic to many cell types at 1-5 microg/ml, we also studied its influence on Leydig cells during 4 days in culture using a wide range of doses. Leydig cells were highly resistant to the cytotoxic effects of 25-hydroxycholesterol, with no cells dying at 10 microg/ml and only 50% of cells affected at 100 microg/ml after 2 days of treatment. Similar conditions resulted in 100% death of a control lymphocyte cell line. These results demonstrate that 1) testicular macrophages have mRNA for cholesterol 25-hydroxylase and can convert cholesterol into 25-hydroxycholesterol, 2) macrophage-conditioned medium is not capable of auto-oxidation of cholesterol, 3) Leydig cells are highly resistant to the cytotoxic influences of 25-hydroxycholesterol, and 4) long-term treatment with high doses of 25-hydroxycholesterol results in loss of LH responsiveness. These results support the concept that testicular macrophages enzymatically produce 25-hydroxycholesterol that not only is metabolized to testosterone by Leydig cells when present at putative physiological levels but also may exert inhibitory influences on Leydig cells when present for extended periods at very high concentrations that may occur under pathological conditions.

Oxysterols regulate expression of the steroidogenic acute regulatory protein

“…Although the fold increase seen in oxysterol-treated MA-10 cells was greater than that seen in Y1 cells, this appears to be due at least in part to a higher basal level of StAR protein expression in Y1 cells. Consistent with previous results with Leydig cells (Lukyanenko et al 2001), MA-10 cells were resistant to cytotoxicity by oxysterols.…”

“…The higher sensitivity of MA-10 cells to 25 OHC may reflect just such a physiological situation in the testis. There, comparable levels of 25 OHC produced by testicular macrophages can induce differentiation of primary rat progenitor and immature Leydig cells and long-lasting increases in 'basal' steroidogenesis in adult Leydig cells; 25 OHC further elicits the generation of up to 3-fold higher concentrations of testosterone than of the oxysterol itself, indicating that the induced production of steroid is not simply a result of direct conversion of 25 OHC to testosterone (Yee & Hutson 1985, Lukyanenko et al 2001, Chen et al 2002. Thus, the loss of oxysterol production in testes depleted of macrophages may account for the observed reduction in steroidogenic enzyme and testosterone levels (Bergh et al 1993, Cohen et al 1997.…”

Gonadal macrophage infiltration in congenital lipoid adrenal hyperplasia

“…This is partly explained by the notion that hyperplasia of Leydig cells in infants is less manifested than that of the theca cells in adult patients (10,12). Intriguingly, testicular macrophages have the potential to provide 25-hydroxycholesterol as a direct substrate for P450 side-chain cleavage enzyme in an alternate pathway for steroidogenesis, which bypasses the STAR-dependent delivery of cholesterol into the inner mitochondrial membrane (15,16). Thus, potential steroidogenic cooperation may also occur between the macrophages and the ovarian theca cells that, similar to the testicular Leydig cells, make androgens as well.…”

Uropathogenic E. coli induce different immune response in testicular and peritoneal macrophages: implications for testicular immune privilege

“…Alternatively/additionally TM are known producers of 25-hydroxycholestrol, an oxysterol that can negatively impact luteinizing hormone stimulated Leydig cell testosterone production [37]. Vice versa testosterone suppresses production of 25-hydroxycholestrol in TM pointing to an interesting control loop between TM and Leydig cells involving two factors with established potent inhibitory function on innate immune responses and likely candidates in establishment testicular immune privilege [38][39][40]. A further layer of complexity is added as TM secreted 25-hydroxycholestrol can be converted to testosterone in Leydig cells.…”