Relaxin protein and gene expression in ovarian follicles of immature pigs

Abstract

Relaxin production by the ovarian follicle of gonadotropin-primed, prepubertal gilts is well documented. As far as we are aware, a source of relaxin in pig follicles, independent of gonadotropins, has not yet been reported. Therefore, the objective of this study was to determine whether relaxin is produced in porcine follicles in the absence of exogenous or cyclic gonadotropins. In immature pigs, immunoreactive relaxin was detected in fluids from small (1-3 mm), medium (4-5 mm) and large (>6 mm) follicles and localized to the theca interna of large follicles. Relaxin levels in follicular fluid significantly increased with follicle size (P<0·05). Relaxin mRNA was detected in whole small-and medium-sized follicles. In large follicles, the relaxin gene was expressed in thecal layers, but not granulosa cells. The abundance of relaxin transcript did not change with follicle size. In summary, relaxin protein and mRNA were detected in porcine follicles from immature animals, indicating that relaxin is produced in the porcine follicle in the absence of exogenous or cyclic gonadotropins. Relaxin's in vitro growth effects on porcine granulosa and theca cells support this follicular relaxin as a growth modulator during porcine follicular development.

Locally produced relaxin may affect testis and vas deferens function in rats

“…Its role in female reproduction is well known: relaxin is important during pregnancy and in preparation for parturition (Sherwood 2004), and it stimulates growth of the uterus in prepubertal animals (Bagnell et al 1993, Yan et al 2006. The main source of relaxin in females is the corpus luteum, but relaxin mRNA and/or protein have been detected also in preovulatory follicles (Bagnell et al 1993, Einspanier et al 1997, Ohleth et al 1998, and it has been suggested that relaxin exerts a paracrine or autocrine role in the ovary, promoting follicular growth and ovulation.…”

Relaxin Secretion and Gene Expression in Porcine Granulosa and Theca Cells Are Stimulated during In Vitro Luteinization1

“…Relaxin mRNA expression in cultured GC and TC was monitored using semi-quantitative RT-PCR [3]. Briefly, 1 g total RNA was incubated at 37ЊC for 30 min in singlestrength MMLV-RT reaction buffer; 1.5 g random primers; 125 M of deoxy (d) ATP, dCTP, dGTP, and dTTP; 1 U deoxyribonuclease (DNase) (ribonuclease [RNase]-free); and 20 U RNasin (RNase inhibitor).…”

“…Each set of PCR reactions included the appropriate volume of the RT blank. A pool of follicle RNA from pigs 60 and 72 h after treatment with eCG, which contains relaxin mRNA [6], and liver from neonatal pigs were used as positive and negative controls for relaxin transcript, respectively [3]. The post-eCG follicle pool was included in each set of RT-PCR reactions to account for variability between each set of reactions.…”

“…The post-eCG follicle pool was included in each set of RT-PCR reactions to account for variability between each set of reactions. To verify that relaxin and 18S cDNAs were amplified in the exponential range, an RT reaction of the post-eCG follicle pool was serially diluted, and each dilution was used as a template for relaxin and 18S PCR reactions [3].…”

“…Relaxin has been detected in media from follicle wall cultures, ovarian extracts from prepubertal gilts, and follicular fluid from prepubertal, cyclic and pregnant animals [1][2][3]. The presence of relaxin mRNA and protein in theca cells (TC) from pig follicles established the TC as the main site of relaxin production in the porcine follicle [4][5][6].…”