The interplay between kisspeptin and endocannabinoid systems modulates male hypothalamic and gonadic control of reproduction in vivo.

Introduction: Male reproduction is under the control of the hypothalamus-pituitary-gonadal (HPG) axis. The endocannabinoid system (ECS) and the kisspeptin system (KS) are two major signaling systems in the central and peripheral control of reproduction, but their possible interaction has been poorly investigated in mammals. This manuscript analyzes their possible reciprocal modulation in the control of the HPG axis. Materials and methods: Adolescent male rats were treated with kisspeptin-10 (Kp10) and endocannabinoid anandamide (AEA), the latter alone or in combination with the type 1 cannabinoid receptor (CB1) antagonist rimonabant (SR141716A). The hypothalamic KS system and GnRH expression, circulating sex steroids and kisspeptin (Kiss1) levels, and intratesticular KS and ECS were evaluated by immunohistochemical and molecular methods. Non-coding RNAs (i.e., miR145-5p, miR-132-3p, let7a-5p, let7b-5p) were also considered. Results: Circulating hormonal values were not significantly affected by Kp10 or AEA; in the hypothalamus, Kp10 significantly increased GnRH mRNA and aromatase Cyp19, Kiss1, and Kiss1 receptor (Kiss1R) proteins. By contrast, AEA treatment affected the hypothalamic KS at the protein levels, with opposite effects on the ligand and receptor, and SR141716A was capable of attenuating the AEA effects. Among the considered non-coding RNA, only the expression of miR145-5p was positively affected by AEA but not by Kp10 treatment. Localization of Kiss1+/Kiss1R+ neurons in the arcuate nucleus revealed an increase of Kiss1R-expressing neurons in Kp10- and AEA-treated animals associated with enlargement of the lateral ventricles in Kp10-treated animals. In the brain and testis, the selected non-coding RNA was differently modulated by Kp10 or AEA. Lastly, in the testis, AEA treatment affected the KS at the protein levels, whereas Kp10 affected the intragonadal levels of CB1 and FAAH, the main modulator of the AEA tone. Changes in pubertal transition-related miRNAs and the intratesticular distribution of Kiss1, Kiss1R, CB1, and CB2 following KP and AEA treatment corroborate the KS-ECS crosstalk also showing that the CB1 receptor is involved in this interplay. Conclusion: For the first time in mammals, we report the modulation of the KS in both the hypothalamus and testis by AEA and revealed the KP-dependent modulation of CB1 and FAAH in the testis. KP involvement in the progression of spermatogenesis is also suggested.

Early intervention with breast milk mesenchymal stem cells attenuates the development of diabetic-induced testicular dysfunction via hypothalamic Kisspeptin/Kiss1r-GnRH/GnIH system in male rats.

Diabetic male infertility and sub fertility are major complications that may implicate both central and peripheral pathways as well as mechanisms controlling reproduction. This study was an attempt to explore the potential effect of breast milk mesenchymal stem cells (Br-MSCs) as a therapeutic tool for diabetic induced reproductive dysfunction at molecular level. Forty-five adult male Sprague Dawely rats were divided into 3 groups (n = 15); control group, diabetic group, and Br-MSCs treated diabetic group. The homing ability of Br-MSCs in diabetic treated rat testicles was confirmed via semi-quantitative RT- PCR analysis of a human specific Gapdh mRNA expression level. Our result showed that type1 diabetic rats exerted an elevation in blood glucose level and a reduction in body weight, fasting serum insulin, FSH, LH, and total testosterone levels, relative and absolute testicular weights, sperm count, motility, and live ratio. In addition, downregulation in the hypothalamic kisspeptin-GnRH system, HPG axis and testicular steroidogenesis compared to control group was noticed. Moreover, upregulation of testicular proinflammatory and apoptotic markers relative mRNA expression compared to control group was observed. Furthermore, a decrease in testicular tissue antioxidant activity (CAT, SOD, GSH) and an increase in lipid peroxidation (MDA) compared to control group was shown. However, Br-MSCs administration restored or even exceeded the normal physiological tone in most of these parameters to the point where a potential therapeutic role for Br-MSCs in type1diabetic induced infertility can be suggested.