Several lines of evidence suggest that insulin-like growth factor 1 (IGF1) is involved in Sertoli cell (SC) proliferation and that its receptor (IGF1R) could mediate follicle-stimulating hormone (FSH) effects. To examine the role of the growth hormone (GH)-IGF1 axis on SC function, we evaluated the effects of GH and IGF1 on basal and FSH-modulated SC proliferation, as well as on anti-Müllerian hormone (AMH) and inhibin B expression and secretion in-vitro. SCs from neonatal pigs were incubated with (1) placebo, (2) 100 nM highly purified urofollitropin (hpFSH), (3) 100 nM recombinant GH (rGH), (4) 100 nM recombinant IGF1 (rIGF1), (5) 100 nM hpFSH plus 100 nM rGH, (6) 100 nM hpFSH plus 100 nM rIGF1, for 48 h. We found that IGF1, but not FSH nor GH, stimulated SC proliferation. Furthermore, an inhibitory effect of FSH, GH and IGF1 on AMH secretion, and a stimulatory role of FSH and IGF1, but not GH, on inhibin B secretion were found. These results suggest that the GH-IGF1 axis influences basal and FSH-modulated SC proliferation and function. We speculate that SC proliferation occurring in childhood might be supported by the increased serum IGF1 levels observed during this period of life.

Experimental evidence has shown that the IGF1 receptor (IGF1R) is involved in testicular development during embryogenesis. More recently, data gathered from mice granulosa cells and zebrafish spermatogonia suggest that IGF1R has a role in Follicle-stimulating hormone (FSH) signaling. No evidence has been reported on this matter in Sertoli cells (SCs) so far. The aim of the study was to evaluate the role, if any, of the IGF1R in FSH signaling in SCs. The effects of FSH exposure on myosin-phosphatase 1 (MYPT1), ERK 1/2, AKT308, AKT473, c-Jun N-terminal kinase (JNK) phosphorylation and on anti-Müllerian hormone (AMH), inhibin B and FSH receptor (FSHR) mRNA levels were assessed with and without the IGF1R inhibitor NVP-AEW541 in purified and functional porcine neonatal SCs. Pre-treatment with NVP-AEW541 inhibited the FSH-induced MYPT1 and ERK 1/2 phosphorylation, decreased the FSH-dependent Protein kinase B (AKT)308 phosphorylation, but did not affect the FSH-induced AKT473 and JNK phosphorylation rate. It also interfered with the FSH-induced AMH and FSHR down-regulation. No influence was observed on the FSH-stimulated Inhibin B gene expression. Conclusion. These findings support the role of theIGF1R in FSH signaling in porcine SCs. The possible influence of IGF1 stimulation on the FSH-mediated effects on SCs should be further explored.

There is ongoing debate as to whether the decline of sperm production in recent times may be related to a parallel increase in the rate of obesity and diabetes. Lower anti-Müllerian hormone (AMH) and inhibin B secretion have been observed in young hyperinsulinemic patients compared to healthy controls, suggesting a Sertoli cell (SC) dysfunction. The pathophysiological mechanisms underlying SC dysfunction in these patients are poorly understood. To the best of our knowledge, no evidence is available on the effects of insulin on SC function. Therefore, this study was undertaken to assess the effects of insulin on basal and follicle-stimulating hormone (FSH)-stimulated SC function in vitro. To accomplish this, we evaluated the expression of AMH, inhibin B and FSHR genes, the secretion of AMH and inhibin B and the phosphorylation of AKT473 and SC proliferation on neonatal porcine SC after incubation with FSH and/or insulin. We found that similar to FSH, the expression and secretion of AMH is suppressed by insulin. Co-incubation with FSH and insulin decreased AMH secretion significantly more than with FSH alone. Insulin had no effect on the expression and secretion of the inhibin B gene, but co-incubation with FSH and insulin had a lower effect on inhibin B secretion than that found with FSH alone. FSH and/or insulin increased AKT473 phosphorylation and SC proliferation. In conclusion, the results of this study showed that insulin modulates SC function. We hypothesize that hyperinsulinemia may therefore influence testicular function even before puberty begins. Therefore, particular care should be taken to avoid the onset of hyperinsulinemia in children to prevent a future deleterious effect on fertility.

Lately, nickel oxide nanoparticles (NiO NPs) have been employed in different industrial and biomedical fields. Several studies have reported that NiO NPs may affect the development of reproductive organs inducing oxidative stress and, resulting in male infertility. We investigated the in vitro effects of NiO NPs on porcine pre-pubertal Sertoli cells (SCs) which undergone acute (24 h) and chronic (from 1 up to 3 weeks) exposure at two subtoxic doses of NiO NPs of 1 μg/ml and 5 μg/ml. After NiO NPs exposure we performed the following analysis: (a) SCs morphological analysis (Light Microscopy); (b) ROS production and oxidative DNA damage, gene expression of antioxidant enzymes (c) SCs functionality (AMH, inhibin B Real-time PCR analysis and ELISA test); (d) apoptosis (WB analysis); (e) pro-inflammatory cytokines (Real-time PCR analysis), and (f) MAPK kinase signaling pathway (WB analysis). We found that the SCs exposed to both subtoxic doses of NiO NPs didn’t sustain substantial morphological changes. NiO NPs exposure, at each concentration, reported a marked increase of intracellular ROS at the third week of treatment and DNA damage at all exposure times. We demonstrated, un up-regulation of SOD and HO-1 gene expression, at both concentrations tested. The both subtoxic doses of NiO NPs detected a down-regulation of AMH and inhibin B gene expression and secreted proteins. Only the 5 μg/ml dose induced the activation of caspase-3 at the third week. At the two subtoxic doses of NiO NPs a clear pro-inflammatory response was resulted in an up-regulation of TNF-α and IL-6 in terms of mRNA. Finally, an increased phosphorylation ratio of p-ERK1/2, p-38 and p-AKT was observed up to the third week, at both concentrations. Our results show the negative impact of subtoxic doses NiO NPs chronic exposure on porcine SCs functionality and viability.

Duchenne muscular dystrophy (DMD) is an X-linked disease caused by mutations in DMD gene translating in lack of functional dystrophin and resulting in susceptibility of myofibers to rupture during contraction. Inflammation and fibrosis are critical hallmarks of DMD muscles, which undergo progressive degeneration leading to loss of independent ambulation in childhood and death by early adulthood. We reported that intraperitoneal injection of microencapsulated Sertoli cells (SeC) in dystrophic mice translates into recovery of muscle morphology and performance thanks to anti-inflammatory effects and induction of the dystrophin paralogue, utrophin at the muscle level, opening new avenues in the treatment of DMD. The aim of this study is to obtain information about the direct effects of SeC on myoblasts/myotubes, as a necessary step in view of a translational application of SeC-based approaches to DMD. We show that (i) SeC-derived factors stimulate cell proliferation in the early phase of differentiation in C2C12, and human healthy and DMD myoblasts; (ii) SeC delay the expression of differentiation markers in the early phase nevertheless stimulating terminal differentiation in DMD myoblasts; (iii) SeC restrain the fibrogenic potential of fibroblasts, and inhibit myoblast-myofibroblast transdifferentiation; and, (iv) SeC provide functional replacement of dystrophin in preformed DMD myotubes regardless of the mutation by inducing heregulin β1/ErbB2/ERK1/2-dependent utrophin expression. Altogether, these results show that SeC are endowed with promyogenic and antifibrotic effects on dystrophic myoblasts, further supporting their potential use in the treatment of DMD patients. Our data also suggest that SeC-based approaches might be useful in improving the early phase of muscle regeneration, during which myoblasts have to adequately proliferate to replace the damaged muscle mass.