Advantages of pulsatile hormone treatment for assessing hormone-induced gene expression by cultured rat Sertoli cells

Abstract

In response to various hormonal (follicle-stimulating hormone [FSH] and testosterone [T]) and biochemical inputs, testicular Sertoli cells (Sc) produce factors that regulate spermatogenesis. A number of FSH- and T-responsive Sc-specific genes, necessary for spermatogenesis, have been identified to date. However, the hormone-induced in vitro expression pattern of most of these genes is reported to be inconsistent at various time points in primary rat Sc cultures. As a matter of convenience, cultured Sc are constantly exposed to hormones for a few hours to days in the reported literature, although Sc are exposed to pulsatile FSH and T in vivo. The major aim of the present study is to evaluate the advantage, if any, of the in vitro administration of pulsatile hormone (FSH and T in combination) treatment on gene expression of cultured Sc as compared with that of constant hormone treatment. Pulsatile treatment (a 30-min hormonal exposure every 3 h) mimicking the in vivo condition reveals a more prominent effect of hormones in augmenting gene expression as compared with constant treatment. Our results indicate that the expressions of Stem cell factor (Scf, only responsive to FSH), Claudin11 (only responsive to T) and Transferrin (both FSH- and T-responsive) mRNAs are significantly higher at 12 h upon pulsatile treatment than upon constant hormonal treatment. Maximal expression of relevant genes because of pulsatile treatment with hormones suggests that this protocol provides a more suitable premise for assessing hormone-induced gene expression in isolated Sc than one involving constant exposure to hormones.

Follicle-Stimulating Hormone (FSH) Action on Spermatogenesis: A Focus on Physiological and Therapeutic Roles

“…As expected, in immature Sertoli cells, genes involved in cell growth and proliferation, metabolism, and the MAPK and Wingless-related integration site (Wnt) pathways were upregulated, whereas in mature cells, genes relevant to the differentiated function of Sertoli cells, involved in phagocytosis, cytoskeleton remodeling, glucose metabolism, and insulin signaling, were expressed. In this study, FSH was delivered with testosterone in a pulsatile fashion, expected to amplify the gene responses, as previously described [60]. This observation is very interesting because it suggests that hormone target cells decode a pulsatile signal vs. a monotonous signal in a quantitatively different manner.…”

“…For example, the FSHR couples not only to Gα s but also to Gα i , which is involved in the post-natal mitotic response of Sertoli cells to FSH via extracellular-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK)-dependent signaling [59]. These data are supported by recent evidence that FSH limits the expression of Gα s , which fails to downregulate Gα i activity in infant primates [60]. In this line, a dynamic computational model predicted that PKA inhibition does not compromise phosphatidylinositide(3,4,5) trisphosphate (PIP3) production in pre-pubertal rats [61].…”

Hormone induced differential transcriptome analysis of Sertoli cells during postnatal maturation of rat testes

“…For example, pulsatile LH-releasing hormonal-therapy was found to be effective over constant GnRH in restoring the normal gonadotropin secretion inducing fertility in men with hypogonadotropic eunuchoidism [ 47 ]. We have recently demonstrated that for hormone induced gene expression study, pulsatile treatment of hormones (both FSH and T in combination) to cultured Sc from 18 days-old pre-pubertal rats’ shows better readouts as compare to that of the conventional, constant stimulation with hormones [ 48 ]. Although the establishment of hypothalamo-hypophyseal testicular axis steadily improves with age in rodents, GnRH pulse generator activity initiates during fetal development [ 49 ].…”

Claudins: New Players in Human Fertility and Reproductive System Cancers

“…Claudin-11 is known to be regulated hormonally, and its knockout models result in infertility [87]. In rat primary cultures of testicular Sertoli cells, claudin-11 expression was reported to be modulated both by testosterone as well as follicle-stimulating hormone (FSH) [88]. Additionally, in mice models, suppressed levels of gonadotropins affected the localization of claudin-11 in Sertoli cells [25].…”