A Metabolite of Methoxychlor, 2,2-Bis(p-Hydroxyphenyl)-1,1,1-Trichloroethane, Reduces Testosterone Biosynthesis in Rat Leydig Cells Through Suppression of Steady-State Messenger Ribonucleic Acid Levels of the Cholesterol Side-Chain Cleavage Enzyme1

Abstract

Postnatal development of Leydig cells involves transformation through three stages: progenitor, immature, and adult Leydig cells. The process of differentiation is accompanied by a progressive increase in the capacity of Leydig cells to produce testosterone (T). T promotes the male phenotype in the prepubertal period and maintains sexual function in adulthood; therefore, disruption of T biosynthesis in Leydig cells can adversely affect male fertility. The present study was designed to evaluate the ability of a xenoestrogen, methoxychlor (the methoxylated isomer of DDT [1,1, 1-trichloro-2,2-bis(p-chlorophenyl)ethane]), to alter Leydig cell steroidogenic function. Purified progenitor, immature, and adult Leydig cells were obtained from, respectively, 21-, 35-, and 90-day-old Sprague-Dawley rats treated with graded concentrations of the biologically active metabolite of methoxychlor, 2, 2-bis(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE), and assessed for T production. HPTE caused a dose-dependent inhibition of basal and LH-stimulated T production by Leydig cells. Compared to the control value, reduced T production by progenitor and immature Leydig cells was apparent after 10 h of HPTE treatment in culture; the equivalent time for adult Leydig cells was 18 h. The reversibility of HPTE-induced inhibition was evaluated by incubating Leydig cells for 3, 6, 10, 14, or 18 h and measuring T production after allowing time for recovery. After treatment with HPTE for 3 h, T production by immature and adult Leydig cells for the 18-h posttreatment period was similar to the control value, but that of progenitor Leydig cells was significantly lower. The onset of HPTE action and the reversibility of its effect showed that Leydig cells are more sensitive to this compound during pubertal differentiation than in adulthood. T production was comparable when control and HPTE-treated immature Leydig cells were incubated with pregnenolone, progesterone, and androstenedione, but HPTE-treated Leydig cells produced significantly reduced amounts of T when incubations were conducted with 22R-hydroxycholesterol (P < 0.01). This finding suggested that HPTE-induced inhibition of T production is related to a decrease in the activity of cytochrome P450 cholesterol side-chain cleavage enzyme (P450(scc)) and cholesterol utilization. The reduced steady-state mRNA level for P450(scc) in HPTE-treated Leydig cells was demonstrated by reverse transcription-polymerase chain reaction and densitometry. In conclusion, this study showed that HPTE causes a direct inhibition of T biosynthesis by Leydig cells at all stages of development. This effect suggests that reduced T production could be a contributory factor in male infertility associated with methoxychlor and, possibly, other DDT-related compounds.

Influence of long-term dietary administration of procymidone, a fungicide with anti-androgenic effects, or the phytoestrogen genistein to rats on the pituitary–gonadal axis and Leydig cell steroidogenesis

“…The mechanism remains unknown, but may involve activation of the estrogen receptor by genistein. This suggestion is supported by the recent report that the xenoestrogen methoxychlor (a derivative of DDT) alters steroidogenesis by immature rat Leydig cells by suppressing the expression of P450 scc and that the antiestrogen ICI 182 780 prevents this effect (Akingbemi et al 2000). In addition, genistein and its derivatives were found to be potent inhibitors of P450c21 and human adrenocortical 3 -hydroxysteroid dehydrogenase in vitro (Ohno et al 2002).…”

The effect of environmental contaminants on testicular function

“…73 Exposing Leydig cells (progenitor, immature and adult cells) isolated from 21-, 35-and 9-day-old rats to MXC and 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane decreased the mRNA levels of P-450scc, decreased testosterone production and decreased cholesterol utilisation by Leydig cells, thereby inhibiting testosterone biosynthesis at all stages of development. 74 It has also been shown that MXC and 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane inhibit the activities of 3b-HSD and 17b-HSD in both human and rat testis. The mode of action has been suggested to be by competition with the cofactors and not the substrate.…”

Influence of pesticides on male fertility

“…By blocking the expression of this protein, Leydig cells produce less testosterone in vitro. Another mechanism of action is the reduction of P450 cholesterol side-chain cleavage enzyme (P450 scc ) activity by, for example, dimethoate, roundup, 2,2-bis(p-hydroxyphenyl)-1,1,1-trichloroethane, a metabolite of methoxychlor, and ketoconazole (26,31,33,34). P450 scc is an enzyme that catalyzes the first reaction in the testosterone biosynthesis pathway: the conversion of cholesterol to pregnenolone.…”