Cyclooxygenase 2, Prostaglandins and TGFb1 on Testicular Function

CALANDRA, RICARDO S; MATZKIN MARIA EUGENIA; GONZALEZ-CALVAR SILVIA I; GONZALEZ CANDELA; MAYERHOFER ARTUR; FRUNGIERI MONICA B

Sao Paulo, SP, Brasil.

Workshop; II Workshop on Male Reproductive Biology; 2009

FeSBE e das Sociedades Brasileiras de Farmacologia e Terapêutica Experimental (SBFTE), de Fisiologia (SBFis), de Biologia Celular (SBBC) e de Endocrinologia e Metabologia - Departamento de Endocrinologia Básica (SBEM)

      Cyclooxygenase (COX) is the main regulatory enzyme in the synthesis of prostaglandins (PGs). This enzyme is present in two isoforms named COX1 and COX2. In most cell types COX1 is present as a contitutive isoform whereas COX2, the inducible form, is expressed only in the early stages in the processes of differentiation and cellular division. Induction or presence of COX-2 has been reported not only in isolated cells, but also in cells in various tissues such as female reproductive organs. COX-1- or COX-2-knockout mice may provide many clues in establishing the role of both isoforms in reproduction. On this regards, in the female, the role of COX 2 and PGs has shown a direct participation in the reproductive process such as ovulation, fertilization, implantation and parturation (Lim et al, 1997; Sirois et al, 2004).         Concerning to the male reproductive system, fertility is not affected in COX-deficient mice (Loftin et al, 2002). However, early and more recent studies suggest that PGs decrease plasma testosterone in male rats and rhesus (Saksena et al, 1973 ;  Kimball et al, 1979; Romanelli et al, 1995). Evenmore, a recent study suggests a novel mechanism in male aging involving COX2 as a potential mechanism to explain the delay age-related decline in testosterone biosynthesis (Wang et al, 2005). On the other hand, recently has been detected COX2 expression and production of PGF2á in spermatogenic and somatic cells from testes of adult rats (Winnall et al, 2007).          The golden (Syrian) hamster is a seasonal breeder and consequently the function of the pituitary-testicular axis undergoes annual cyclic variations (Steger et al,1985). Thus, the golden hamster represents a versatile experimental model where changes in the photoperiod can greatly alter male gonadal function. It is well known that when is exposed to short photoperiods (SP< than 12.5h light/day) undergoes a morphological and physiological testicular regression followed by a spontaneous recrudescence. This SP-induced gonadal regression is accompanied by a marked decrease in serum levels of FSH, LH and prolactin (Bex et al.,1978; Frungieri et al,1996) and in blood and intratesticular concentrations of testosterone, its precursors and  metabolites (Chandrashekar et al,1989; Frungieri et al, 1996).             On this context, our Group has shown :1) by immunohistochemical assays the presence of COX2 in testes from reproductive active Syrian hamster (LD) Leydig cells but not in testes from other adult species (mice, rats, monkeys and pigs) (Frungieri et al, 2006) and 2) expression of COX2 in hamsters testes occurred in peripubertal,  pubertal and adult reproductively active hamsters along with high serum levels of testosterone, LH and prolactin. On the other hand, prepubertal and maximal photoperiodically regressed adult hamsters with rather low serum levels of these hormones, did not express testicular COX2 (Frungieri et al, 2006) and these results were confirmed by RT-PCR. In addition, was investigated the effects of various PGs on testicular testosterone production and found that PGF2á  within a range of 100 pM and 1 ìM, in the presence of 100 mIU/ml hCG, was significantly inhibited whereas on the other hand, we described production of PGD2 and a significant increase in basal testosterone production in LD adult hamster Leydig cells in the presence of 1 ìM of PGD2 (Schell et al, 2007).Testicular concentration and content of PGF2á in reproductively active hamsters as well as its production from isolated Leydig cells were also determined. These cells were also incubated with an inhibitor of COX2 (Meloxican) and a 2.5-fold decrease in PGF2á  production was observed and PGF2á  receptors were also revealed in adult LD hamster testis. The inhibitory mechanism of PGF2á on testosterone production involves in hCG-stimulated Leydig cells a decreased in gene expression of testicular StAR and 17â-hydroxysteroid dehydrogenase enzymes. Furthermore, other studies observed that treatment with PGF2á decrease StAR expression in ovaries from different especies (Chung et al, 1998 ; Diaz et al, 2005).       The mechanisms involves in the influence of COX2 / PGs on testicular steroidogenesis hamster Leydig cells was also studied and in brief, among others,were found : 1) testosterone, LH and prolactin, by RT-PCR, Western blot and immunoassay, stimulated mRNA and protein COX2 expression and PGs production; 2) testosterone and LH effecys was abolished by the specific antiandrogen bicalutamide (Bi); 3) testosterone also stimulated phosphorylation of the extracellular signal-regulated kinase isoforms ½ (erk ½); 4) Bi and  an inhibitor of the mitogen-activated protein kinase ½ (MEK ½) abolished testosterone actions on erk ½ and COX2 (Matzkin et al, 2009).       Moreover, TGF-âs are involved during testicular development and expressed by Leydig, peritubular and Sertoli cells (Mullaney et al, 1993). TGF-â1 and 3 influence steroidogenesis in fetal and neonatal rat testis (Gautier et al, 1997; Olaso et al, 1999). TGF-â1 exerts is influence via TGFB-RII, and TGFB-RI (ALK-1 and ALK-5). Recently, it has been reported that TGF-b1 induces COX expression and PGs synthesis through MAPK and PI3K (Rodriguez-Barbero et al., 2006). Moreover, it was suggest that endoglin, a TFG-b1 co-receptor, is involved in the COX regulation (Jerkic et al., 2006).   We analyze TGF-b1 system in Leydig cells from adult active and maximal regressed hamsters in culture, and we detected the presence of ALK-1 and 5 and TGF-â1. Also, this cytokine increases the expression of p15 through ALK-5. This effect involves MAPK: p38 and erk 1/2 (Gonzalez et al, 2008). These preliminary results suggest that TGF-â1 may regulate Leydig cell function through the modulation of specific genes.      In summary, PGs, particularly PGF2á, influence in hamsters testicular androgen production and the androgen milieu appears to be crucial for the regulation of the link between steroidogenesis-PGs during sexual development as well as photoperiodics seasonal changes.