Follicle-stimulating hormone, testosterone, and hypoxia differentially regulate UDP-glucuronosyltransferase 1 isoforms expression in rat Sertoli and peritubular myoid cells

Abstract

Uridine diphosphoglucuronosyltransferases (UGTs) are detoxifying enzymes responsible for the metabolism of endogenous and xenobiotics compounds. UGT isoforms are widely distributed in rat tissues showing a constitutive and inducible gene expression. However, little information is available concerning UGTs expression in testis. The UGT1A1, UGT1A2, and UGT1B1 mRNAs expression in whole rat testis, in Sertoli and peritubular myoid cells in basal conditions, and after hormonal and hypoxic stimulation were investigated by reverse transcriptase-polymerase chain reaction (RT-PCR). Constitutive expression of each UGT1 isoform was present in rat testis with higher levels of UGT1A2. UGT transcripts were also detected in Sertoli and peritubular myoid cells. After FSH stimulation, Sertoli cells showed an increase in UGT1B1 mRNA expression, whereas the levels of UGT1A1 and UGT1A2 resulted unmodified. The main effect induced by testosterone was a decrease of UGT1B1 mRNA expression in peritubular myoid cells, whereas in Sertoli cells an increase in UGT1A1 and UGT1B1 was observed. In hypoxic conditions, a reduction in UGTs mRNA levels was detected in both cell types. These findings suggest that rat UGT1 isoforms are regulated in testis by hormonal and environmental factors. Thus, it was speculated that alterations in UGTs expression and/or activity may be involved in the pathogenesis of testis injury.

Introduction

Uridine diphosphoglucuronosyltransferases (UGTs) are a family of isoenzymes located in the endoplasmic reticulum that catalyse the transfer of the glucuronic acid from uridine diphosphoglucuronic acid to a wide variety of endogenous compounds including bilirubin, steroid hormones, fat-soluble vitamins, and thyroid hormones [1]. UGTs are also involved in the metabolism and inactivation of various xenobiotic compounds such as drugs, teratogens, and carcinogens [2]. Based on their aminoacid sequence similarities, two families of UGTs have been characterized, UGT1 and UGT2, consisting of drug-glucuronidating forms and steroid-glucuronidating forms, respectively. However, it has been demonstrated that UGT1 family is also able to glucoronidate steroids [3], [4]. Furthermore, UGT1 family was divided in two subgroups, UGT1A and UGT1B, according to their preferential substrate specificity. The members of UGT1 gene complex differ in their first exon but share a single set of commonly used exons (2–5), which encode the common carboxyl terminal of UGT1 isoforms. The variable first exon codes for the N terminus of the enzyme, which determines substrate specificity. In rats, multiple UGT1 isoforms have been identified showing a different tissue distribution and a specific gene expression regulation by xenobiotics and endobiotics. The liver is the main site of expression of most of these enzymes, except for UGT1A7, UGT1A8, UGT1A10, which are expressed only in extrahepatic tissues [5], [6]. Furthermore, a direct evidence indicates a widely constitutive and inducible UGT1 expression in several tissues such as brain, lung, intestine, skin, and gonads. Testis has been demonstrated a considerable site of expression of both UGT1 [7] and UGT2 [8], [9] families.

In the last decades the rates of testis cancer, cryptorchidism, and hypospadias are increasing [10] while reduction of the sperm counts and male fertility are observed [11]. The reasons of the increased frequency of testicular disorders have been discussed in an intriguing debate. A dozen of synthetic and natural chemicals interfering with the reproductive system, directly or through disruption of endocrine functions, have been discovered. UGTs play a central role in the cellular detoxification of several compounds, preventing accumulation of potentially dangerous xenobiotics or metabolites [12], [13] thus avoiding their subsequent bioactivation to even more toxic reactive intermediates. The production of such reactive species can be enhanced by hypoxic conditions, leading to an increased susceptibility to cell injury [14]. The glucuronidation prevents the covalent binding of these compounds to cellular proteins, lipids, and DNA thus hampering cellular damage leading to neoplastic transformation, developmental defects or specific organ diseases.

Although the function and regulation of UGTs are well characterized in several tissues such as liver and kidney, little information is available concerning the relationship between testis and the UGT system.

In the present work, the expression of UGT1A1, UGT1A2, and UGT1B1 isoforms in whole rat testis was tested. Moreover, primary cultures of Sertoli and peritubular myoid cells in basal conditions and after hormonal stimulation were analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR). Finally, the influence of hypoxia on UGT isoforms expression was evaluated.