**7. 11-hydroxysteroid dehydrogenase in the adrenal gland - expression profile under conditions of testosterone withdrawal**

As mentioned above, the enzyme11-HSD catalyzes the interconversion of glucocorticoids to inert metabolites in man and rodents and plays a crucial role in regulating the action of corticosteroids. Inhibition of 11-HSD allows access of cortisol or corticosterone to the mineralcorticoid receptors where they act as mineralcorticoids [148]. Northern blot analyses revealed expression of mRNAs encoding both 11-HSD1 and 11-HSD2 in the whole rat

adrenal gland. *In situ* hybridization of rat adrenal cortex and medulla demonstrated specific localization of 11 HSD1 mRNA predominantly to the cells at the corticomedullary junction, within the inner cortex, suggesting that the oxoreductase enzyme may serve to maintain high medullary glucocorticoid concentrations required for catecholamine biosynthesis. In contrast, 11-HSD2 mRNA was more uniformly distributed in the cortex and was low/absent in the medulla [149, 150]. The expression of 11-HSD2 has been demonstrated in rat adrenal gland by immunohistochemical and molecular analyses and the 11-HSD2 antigen was confined to the zona fasciculata and zona reticularis, but not in the zona glomerulosa or medulla [149-151]. The ubiquitous presence of 11-HSD2 in sodiumtransporting epithelia revealed that mineralcorticosteroid action is facilitated by this enzyme which metabolizes glucocorticoids and allows aldosterone to bind to the nonselective mineralcorticoid receptor [151].

Hydrohysteroid Dehydrogenases – Biological Role and Clinical Importance – Review 133

**Figure 7.** 11-HSD2 immunoreactivity in rat adrenal gland zones. 35 days after EDS (a, b); 7 days after EDS (c, d); 21 days after EDS (e, f). 11-HSD2- immunoreactivity in the zona fasciculata (ZF) and zona

**localization/distribution and relation to Polycistic Ovaries Syndrome and** 

Glucocorticoids exert their effects in all parts of the body and they are involved in a number of physiological processes, including female reproduction. The ovary is also affected by the glucocorticoids and it is well known that the reproductive function may be impaired in cases of adrenal hyperactivity. The ovaries express glucocorticoid receptors and one of the prominent glucocorticoids affecting ovarian function is the cortisol [157]. Ovaries lack the necessary enzymes for cortisol synthesis and cortisol is not produced *de novo* [158] but it was delivered by the circulation. The 11β-HSD enzymes play a crucial role in controlling the

reticularis (ZR), and the adipocytes of adrenal capsula adipose (A). Less sensitive were the adrenocorticocytes of zona glomerulosa (ZG). No positive signals in the medulla (M). x 200.

**8. 11β –hydroxysteroid dehydrogenase in the ovary – cellular** 

**obesity in women** 

Using EDS experimental model in adult rats [152] we found that the dynamic of 11-HSD2 expression correlated with the changes of serum T levels following the exposure after EDS [153]. The lowest 11-HSD2 staining intensity was found 7 days after EDS followed by progressive increase in the immunoreactivity on day 14 and 21 after EDS [152]. Moreover, the restoration of 11-HSD2 activity on day 14 after EDS corresponded with unchanged glandular and serum corticosterone levels in treated rats on day 15 reported by Plecas et al. [154]. Enzymatic assays on tissue homogenates showed extensive conversion of corticosterone to its 11-dehydro product in an NAD+-dependent manner in adrenal gland [151]. Using enzymehistochemistry a strong reduction was found in the activity of NADH2 cytochrome-C-reductase that is involved in NAD+-synthesis as a cofactor in the adrenal gland after EDS treatment of adult rats [155]. Immunohistochemical analysis revealed that the 11-HSD2 expression pattern in adrenal gland of EDS treated rats [152] is very similar to the enzymehistochemical profile of NADH2- cytochrome-C-reductase [155], supporting the view that 11-HSD2 acts as high-affinity NAD+-dependent dehydrogenase in the rat adrenal gland [151]. On the other hand, the increase in the expression of 11β-HSD2 in rat adrenal gland on day 14 after EDS treatment [152] coincided with the appearance of the repopulation of testosterone-producing Leydig cells in the testis [135]. These data suggested a possible role of the gonadal steroids, especially of testosterone, as modulators of the adrenal gland functional activity and they are consistent with previously reported results related to the direct impact of testosterone on the key steps in the adrenal gland steroidogenesis [156]. The above mentioned findings characterized 11β-HSD2 (high-affinity NAD+- dependent unidirectional dehydrogenase) as a potential target of testosterone action in rat adrenal cortex. Our data from EDS experimental model provided new evidence for expression of 11-HSD2 in the adrenal gland under conditions of testosterone withdrawal. The EDS results bring additional elucidation on the functional significance of 11β-HSD system in rat adrenal gland and the regulatory role of testosterone in its activity [152]. Together with our previous studies [135,153], these data suggested the relationship between 11-HSD2 expression in adrenal gland and kinetics of restoration of testosterone production during renewal of testicular adult LCs population after EDS treatment. (Figure 7)

mineralcorticoid receptor [151].

adrenal gland. *In situ* hybridization of rat adrenal cortex and medulla demonstrated specific localization of 11 HSD1 mRNA predominantly to the cells at the corticomedullary junction, within the inner cortex, suggesting that the oxoreductase enzyme may serve to maintain high medullary glucocorticoid concentrations required for catecholamine biosynthesis. In contrast, 11-HSD2 mRNA was more uniformly distributed in the cortex and was low/absent in the medulla [149, 150]. The expression of 11-HSD2 has been demonstrated in rat adrenal gland by immunohistochemical and molecular analyses and the 11-HSD2 antigen was confined to the zona fasciculata and zona reticularis, but not in the zona glomerulosa or medulla [149-151]. The ubiquitous presence of 11-HSD2 in sodiumtransporting epithelia revealed that mineralcorticosteroid action is facilitated by this enzyme which metabolizes glucocorticoids and allows aldosterone to bind to the nonselective

Using EDS experimental model in adult rats [152] we found that the dynamic of 11-HSD2 expression correlated with the changes of serum T levels following the exposure after EDS [153]. The lowest 11-HSD2 staining intensity was found 7 days after EDS followed by progressive increase in the immunoreactivity on day 14 and 21 after EDS [152]. Moreover, the restoration of 11-HSD2 activity on day 14 after EDS corresponded with unchanged glandular and serum corticosterone levels in treated rats on day 15 reported by Plecas et al. [154]. Enzymatic assays on tissue homogenates showed extensive conversion of corticosterone to its 11-dehydro product in an NAD+-dependent manner in adrenal gland [151]. Using enzymehistochemistry a strong reduction was found in the activity of NADH2 cytochrome-C-reductase that is involved in NAD+-synthesis as a cofactor in the adrenal gland after EDS treatment of adult rats [155]. Immunohistochemical analysis revealed that the 11-HSD2 expression pattern in adrenal gland of EDS treated rats [152] is very similar to the enzymehistochemical profile of NADH2- cytochrome-C-reductase [155], supporting the view that 11-HSD2 acts as high-affinity NAD+-dependent dehydrogenase in the rat adrenal gland [151]. On the other hand, the increase in the expression of 11β-HSD2 in rat adrenal gland on day 14 after EDS treatment [152] coincided with the appearance of the repopulation of testosterone-producing Leydig cells in the testis [135]. These data suggested a possible role of the gonadal steroids, especially of testosterone, as modulators of the adrenal gland functional activity and they are consistent with previously reported results related to the direct impact of testosterone on the key steps in the adrenal gland steroidogenesis [156]. The above mentioned findings characterized 11β-HSD2 (high-affinity NAD+- dependent unidirectional dehydrogenase) as a potential target of testosterone action in rat adrenal cortex. Our data from EDS experimental model provided new evidence for expression of 11-HSD2 in the adrenal gland under conditions of testosterone withdrawal. The EDS results bring additional elucidation on the functional significance of 11β-HSD system in rat adrenal gland and the regulatory role of testosterone in its activity [152]. Together with our previous studies [135,153], these data suggested the relationship between 11-HSD2 expression in adrenal gland and kinetics of restoration of testosterone production

during renewal of testicular adult LCs population after EDS treatment. (Figure 7)

**Figure 7.** 11-HSD2 immunoreactivity in rat adrenal gland zones. 35 days after EDS (a, b); 7 days after EDS (c, d); 21 days after EDS (e, f). 11-HSD2- immunoreactivity in the zona fasciculata (ZF) and zona reticularis (ZR), and the adipocytes of adrenal capsula adipose (A). Less sensitive were the adrenocorticocytes of zona glomerulosa (ZG). No positive signals in the medulla (M). x 200.
