Kobayashi Award 2019: The neuroendocrine regulation of the mammalian reproduction.

Mammalian reproductive function is a complex system of many players orchestrated by the hypothalamus-pituitary-gonadal (HPG) axis. The hypothalamic gonadotropin-releasing hormone (GnRH) and the consequent pituitary gonadotropin release show two modes of secretory patterns, namely the surge and pulse modes. The surge mode is triggered by the positive feedback action of estrogen secreted from the mature ovarian follicle to induce ovulation in females of most mammalian species. The pulse mode of GnRH release is required for stimulating tonic gonadotropin secretion to drive folliculogenesis, spermatogenesis and steroidogenesis and is negatively fine-tuned by the sex steroids. Accumulating evidence suggests that hypothalamic kisspeptin neurons are the master regulator for animal reproduction to govern the HPG axis. Specifically, kisspeptin neurons located in the anterior hypothalamus, such as the anteroventral periventricular nucleus (AVPV) in rodents and preoptic nucleus (POA) in ruminants, primates and others, and the neurons located in the arcuate nucleus (ARC) in posterior hypothalamus in most mammals are considered to play a key role in generating the surge and pulse modes of GnRH release, respectively. The present article focuses on the role of AVPV (or POA) kisspeptin neurons as a center for GnRH surge generation and of the ARC kisspeptin neurons as a center for GnRH pulse generation to mediate estrogen positive and negative feedback mechanisms, respectively, and discusses how the estrogen epigenetically regulates kisspeptin gene expression in these two populations of neurons. This article also provides the mechanism how malnutrition and lactation suppress GnRH/gonadotropin pulses through an inhibition of the ARC kisspeptin neurons. Further, the article discusses the programming effect of estrogen on kisspeptin neurons in the developmental brain to uncover the mechanism underlying the sex difference in GnRH/gonadotropin release as well as an irreversible infertility induced by supra-physiological estrogen exposure in rodent models.

Preeclampsia and gestational hypertension are associated to low maternal circulating kisspeptin levels: a systematic review and meta-analysis.

BACKGROUND: There are contradictory data concerning kisspeptin in gravids with preeclampsia and gestational hypertension (GH). OBJECTIVE: To conduct a meta-analysis of studies comparing maternal kisspeptin levels in gravids with and without preeclampsia or GH. MATERIAL AND METHODS: We searched PubMed, LILACS, and CNKI list of articles up to 20 August 2021, without language limitations, comparing circulating maternal kisspeptin levels, and maternal and neonatal outcomes in gravids with and without preeclampsia or GH. Meta-analyzed results are reported as standardized mean differences (SMD), and their 95% confidence interval (CI). RESULTS: Seven studies with a low-to-moderate risk of bias were eligible for meta-analysis. Gravids with preeclampsia or GH displayed significantly lower circulating kisspeptin levels (SMD, -0.68, 95% CI, -1.04 to -0.32), lower gestational ages at delivery (SMD, -2.22, 95% CI, -3.25 to -1.18), and birth weight (SMD, -2.16, 95% CI, -3.15 to -1.17), and significantly higher body mass indices (MD, 0.56, 95% CI, 0.24-0.88), systolic (SMD, 2.87, 95% CI, 2.22-3.53), and diastolic blood pressures (SMD, 2.57, 95% CI, 2.19-2.95). CONCLUSION: Gravids with preeclampsia or GH had lower kisspeptin levels as compared to normotensive controls.

Inducible Kiss1 knockdown in the hypothalamic arcuate nucleus suppressed pulsatile secretion of luteinizing hormone in male mice.

Accumulating evidence suggests that kisspeptin-GPR54 signaling is indispensable for gonadotropin-releasing hormone (GnRH)/gonadotropin secretion and consequent reproductive functions in mammals. Conventional Kiss1 knockout (KO) mice and rats are reported to be infertile. To date, however, no study has investigated the effect of inducible central Kiss1 KO/knockdown on pulsatile gonadotropin release in male mammals. Here we report an in vivo analysis of inducible conditional Kiss1 knockdown male mice. The mice were generated by a bilateral injections of either adeno-associated virus (AAV) vectors driving Cre recombinase (AAV-Cre) or AAV vectors driving GFP (AAV-GFP, control) into the hypothalamic arcuate nucleus (ARC) of Kiss1-floxed male mice, in which exon 3 of the Kiss1 gene were floxed with loxP sites. Four weeks after the AAV-Cre injection, the mice showed a profound decrease in the both number of ARC Kiss1-expressing cells and the luteinizing hormone (LH) pulse frequency. Interestingly, pulsatile LH secretion was apparent 8 weeks after the AAV-Cre injection despite the suppression of ARC Kiss1 expression. The control Kiss1-floxed mice infected with AAV-GFP showed apparent LH pulses and Kiss1 expression in the ARC at both 4 and 8 weeks after the AAV-GFP injection. These results with an inducible conditional Kiss1 knockdown in the ARC of male mice suggest that ARC kisspeptin neurons are responsible for pulsatile LH secretion in male mice, and indicate the possibility of a compensatory mechanism that restores GnRH/LH pulse generation.

Pharmacokinetic and pharmacodynamic modeling of the metastin/kisspeptin analog, TAK-448, for its anti-tumor efficacy in a rat xenograft model.

TAK-448 is the investigational metastin/kisspeptin analog, which is known to have an anti-tumor effect through suppression of androgen hormones (luteinizing hormone and testosterone) levels. This study developed pharmacokinetic-pharmacodynamic (PK/PD) models of TAK-448 and leuprorelin acetate (TAP-144) in a rat vertebral-cancer of the prostate (VCaP) androgen-sensitive prostate cancer xenograft model to quantitatively assess and compare the anti-tumor effects of both drugs. A potential contribution of the hormone-independent direct effects of TAK-448 to the tumor growth inhibition was also investigated in the in vivo rat xenograft model, because our in vitro experiments revealed that TAK-448 may also directly suppress VCaP cellular proliferation. The PK/PD model successfully described the time course of tumor growth inhibition after drug treatment as well as the development of resistance to the inhibition of androgen hormones, following drug treatment or castration. The EC50 of the hormone-dependent inhibitory effect of TAK-448 was much lower than that of TAP-144, and TAK-448 also has a faster onset of anti-tumor effect than TAP-144, demonstrating that TAK-448 has a stronger overall anti-tumor effect than TAP-144. In addition, model inference, by incorporating a hormone-independent inhibition pathway of TAK-448 into the PK-PD model, suggested that such a direct inhibition pathway for TAK-448 cannot be excluded, as also indicated by in vitro studies, but its EC50 would be approximately three orders of magnitude higher than that of the hormone-dependent pathway. This study helps to understand the potential and mechanism of TAK-448 as a prostate cancer treatment.

Circulatory metastin/kisspeptin-1 in polycystic ovary syndrome: a systematic review and meta-analysis with diagnostic test accuracy.

RESEARCH QUESTION: A close association between Kisspeptin-1 (KISS-1) and reproductive physiology has been reported, but the results on circulatory KISS-1 are ambiguous in patients with polycystic ovary syndrome (PCOS). A systematic review and meta-analysis were conducted to evaluate the association between KISS-1 and PCOS, and to test its diagnostic test accuracy (DTA) through DTA meta-analysis. DESIGN: Relevant studies were identified by searching PubMed and other databases in addition to manual searching of cross-references. Random-effects model was used to obtain standardized mean differences (SMD), pooled correlation coefficients and summary of DTA. Meta-regression and sub-group analyses were conducted to explore heterogeneity. The presence of publication bias was tested using funnel plot analysis. RESULTS: This meta-analysis finally included 12 studies. Compared with controls, women with PCOS showed significantly increased circulatory KISS-1 levels (SMD = 0.47; P = 0.002). Meta-analysis of correlations showed positive associations between KISS-1 and anti-Müllerian hormone (AMH) (P = 0.03), testosterone (P < 0.001) and dehydroepiandrosterone (P = 0.004). The pooled diagnostic odds ratio and area under curve were 13.71 and 0.835, respectively. A one-study leave-out sensitivity analysis indicated that no single study had a significant influence on the overall outcome, suggesting the robustness of this meta-analysis. CONCLUSIONS: This meta-analysis showed significantly increased KISS-1 level in PCOS, and its association with AMH reflects its role in reproductive physiology. In our DTA meta-analysis, KISS-1 showed good accuracy for PCOS detection. Further large-scale studies are required to establish its validity.

A new class of pentapeptide KISS1 receptor agonists with hypothalamic-pituitary-gonadal axis activation.

The kisspeptin (Kp, Kp-54, metastin)/KISS1R system plays crucial roles in regulating the secretion of gonadotropin-releasing hormone. Continuous administration of nonapeptide Kp analogs caused plasma testosterone depletion, whereas bolus administration caused strong plasma testosterone elevation in male rats. To develop a new class of small peptide drugs, we focused on stepwise N-terminal truncation of Kp analogs and discovered potent pentapeptide analogs. Benzoyl-Phe-azaGly-Leu-Arg(Me)-Trp-NH2 (16) exhibited high agonist activity for KISS1R and excellent metabolic stability in rat serum. A single injection of a 4-pyridyl analog (19) at the N-terminus of 16 into male Sprague Dawley rats caused a robust increase in plasma luteinizing hormone levels, but unlike continuous administration of nonapeptide Kp analogs, continuous administration of 19 maintained moderate testosterone levels in rats. These results indicated that small peptide drugs can be successfully developed for treating sex hormone deficiency.

The role of kisspeptin on aromatase expression in breast cancer.

AIM: Kisspeptin is a reproductive peptide hormone that has anti-metastatic roles in several cancer types including colon, lung, and brain cancer. However, in breast cancer, increasing of kisspeptin expression induces aggressiveness of tumors, which in turn exacerbates breast cancer prognosis. MATERIAL AND METHODS: Breast cancer cell lines MCF7 and SKBR3 were cultured in MEM (phenol red free) containing 10 % fetal bovine. Treatments were performed, at 70 % confluency, after 24-hour serum deprivation in serum free medium for 6, 24 and 48 hours. Aromatase (CYP19A1) and kisspeptin receptor (GPR54) mRNA expression were determined by real time Taqman Assay. RESULT: Kisspeptin induced aromatase (CYP19A1) and kisspeptin receptor (GPR54) mRNA expression, while this induction was abolished by kisspeptin receptor inhibitor in MCF7 cells. In SKBR3 cells, however, even though there was an increase in GPR54 mRNA expression with kisspeptin, the induction of CYP19A1 was not observed. CONCLUSION: The inducing effect of kisspeptin on aromatase expression is possibly mediated via kisspeptin receptor and estrogen receptor dependent mechanisms (Fig. 5, Ref. 27).

Aggressive interactions are associated with reductions in RFamide-related peptide, but not kisspeptin, neuronal activation in mice.

Aggressive interactions lead to changes in both future behavior and circulating testosterone (T) concentrations in animals across taxa. The specific neural circuitry and neurochemical systems by which these encounters alter neuroendocrine functioning are not well understood. Neurons expressing the inhibitory and stimulatory neuropeptides, RFamide-related peptide (RFRP) and kisspeptin, respectively, project to neural loci regulating aggression in addition to neuroendocrine cells controlling sex steroid production. Given these connections to both the reproductive axis and aggression circuitry, RFRP and kisspeptin are in unique positions to mediate post-encounter changes in both T and behavior. The present study examined the activational state of RFRP and kisspeptin neurons of male C57BL/6 mice following an aggressive encounter. Both winners and losers exhibited reduced RFRP/FOS co-localization relative to handling stress controls. Social exposure controls did not display reduced RFRP neuronal activation, indicating that this effect is due to aggressive interaction specifically rather than social interaction generally. RFRP neuronal activation positively correlated with latencies to display several offensive behaviors within winners. These effects were not observed in the anteroventral periventricular (AVPV) nucleus kisspeptin cell population. Together, these findings point to potential neuromodulatory role for RFRP in aggressive behavior and in disinhibiting the reproductive axis to facilitate an increase in T in response to social challenge.

Development and Validation of a Sensitive Enzymeimmunoassay for Determination of Plasma Metastin in Mithun (Bos frontalis).

Metastin, also known as kisspeptin-10, is a potent stimulator of gonadotropin-releasing hormone (GnRH) neurons in the central nervous system. Recently, it has been emerged as a key player in the regulation of reproduction in mammals. Blood concentrations of metastin during different physiological stages in bovine species in general and mithun (Bos frontalis) in particular are not available. Lacking of such information may probably be due to non-availability of simple assay procedure to measure the peptide. Therefore, the objective of this study was to develop and validate a simple and sufficiently sensitive enzyme immunoassay (EIA) for metastin determination in mithun plasma using the biotin-streptavidin amplification system and second antibody coating technique. Biotin was coupled to metastin and used to bridge between streptavidin-peroxidase and the immobilized metastin antiserum in the competitive assay. The EIA was conducted directly in 150 µ L of unknown mithun plasma. Metastin standards ranging from 0.01-51.2 ng/150 µ L/well were prepared in hormone-free plasma. The lowest detection limit was 0.07 ng/mL plasma. Plasma volumes for the EIA, viz., 75, 150, and 200 µ L did not influence the shape of standard curve even though a drop in OD450 was seen with higher plasma volumes. A parallelism test was carried out to compare the endogenous mithun metastin with metastin standard used. It showed good parallelism with the metastin standard curve. For the biological validation of the assay, metastin was measured in (a) blood samples collected from 12 pregnant mithun cows during different stages of pregnancy, (b) in blood from seven early pregnant and 12 non-pregnant mithuns, and (c) in follicular fluid obtained from different types of follicle. It was found that the plasma metastin concentrations increased (P < 0.001) from first through last trimester of pregnancy. Plasma metastin levels were much higher (P < 0.001) in early pregnant than non-pregnant cows. Follicular fluid metastin concentrations were found to increase (P < 0.001) as the follicles grow and the highest levels were recorded in preovulatory follicles. In conclusion, a simple, sufficiently sensitive and direct EIA procedure has been developed for the first time to determine metastin levels in mithun. A wide range of metastin concentrations can be detected during different physiological stages in mithun using this metastin-EIA procedure.

Augmentation of Metastin/Kisspeptin mRNA Expression by the Proestrous Luteinizing Hormone Surge in Granulosa Cells of Rats: Implications for Luteinization.

Variations in mRNA levels and sources of metastin/kisspeptin, neurokinin B (NKB), dynorphin, and kisspeptin receptor GPR54 were examined in the ovaries of cycling rats. Kisspeptin and dynorphin mRNAs dramatically increased at 2000 h of the proestrous day. NKB mRNA also increased, but the peak was delayed by 6 h. GPR54 mRNA declined inversely with kisspeptin. Whole-ovary expressions of kisspeptin and dynorphin mRNAs, but not of NKB mRNA, were augmented by the administration of human chorionic gonadotropin (hCG). By means of laser-capture microdissection, kisspeptin mRNA was shown mostly in follicles at 2000 h of proestrus, whereas NKB and dynorphin were expressed mainly in interstitial tissues. GPR54 mRNA was detected equally in follicles, corpora lutea, and interstitial tissues. The hCG stimulated the follicular expression of kisspeptin and interstitial tissue expression of dynorphin mRNA. In primary cultures of granulosa cells prepared from equine chorionic gonadotropin-pretreated immature rats, hCG stimulated the expression of kisspeptin, dynorphin, and NKB mRNAs. Distortion of the corpus luteum and surrounding tissue borders was sometimes seen after intra-ovarian bursa administration of kisspeptin antagonist p234 for 3 days from proestrus. Progesterone production stimulated by hCG in granulosa cell culture was suppressed by p234. These data demonstrate that significant amounts of kisspeptin are synthesized in granulosa cells and dynorphin in interstitial tissues, in response to the proestrous luteinizing hormone surge, whereas granulosa cells also contain dynorphin and NKB, suggesting at least a role for kisspeptin in the luteinization of granulosa cells.

Determination of plasma kisspeptin concentrations during reproductive cycle and different phases of pregnancy in crossbred cows using bovine specific enzyme immunoassay.

Kisspeptin, a decapeptide and potent secretagogue of GnRH has been emerged recently as a master player in the regulation of reproduction in animals. Determination of kisspeptin in peripheral circulation is, therefore, very important for studying the control of its secretion and its role on reproduction in bovine species, the information on which is not available during any physiological state in this species, may probably be due to non-availability of simple assay procedure to measure the hormone. Therefore, the objective of this study was to develop and validate a simple and sufficiently sensitive enzyme immunoassay (EIA) for kisspeptin determination in bovine plasma using the biotin-streptavidin amplification system and second antibody coating technique. Biotin was coupled to kisspeptin and used to bridge between streptavidin-peroxidase and the immobilized kisspeptin antiserum in the competitive assay. The EIA was conducted directly in 100 µl of unknown bovine plasma. Kisspeptin standards ranging from 0.01 to 25.6 ng/100 µl/well were prepared in hormone-free plasma. The lowest detection limit was 0.1 ng/ml plasma. Plasma volumes for the EIA, viz., 50, 100 and 200 µl did not influence the shape of standard curve even though a drop in OD450 was seen with higher plasma volumes. A parallelism test was carried out to compare the endogenous bovine kisspeptin with kisspeptin standard used. It showed good parallelism with the kisspeptin standard curve. For the biological validation of the assay, plasma kisspeptin was measured in blood samples collected from six non-lactating cyclic cows during entire estrous cycle and from 18 pregnant cows during different stages of pregnancy. The mean plasma kisspeptin concentration during different days of the estrous cycle was different (P<0.001). Three peaks of kisspeptin were recorded, one on a day before appearance of preovulatory LH surge, second at day 6 and third one at day 18 of the estrous cycle. Plasma kisspeptin concentrations increased (P<0.001) from first through last trimester of pregnancy. Kisspeptin concentrations were also measured in different follicular, luteal and placental tissues. Follicular and placental kisspeptin levels increased (P<0.01) during follicular development and with the advancement of pregnancy, respectively. On the other hand, luteal concentrations of kisspeptin decreased (P<0.01) with its developmental process. In conclusion, a simple, sufficiently sensitive and direct EIA procedure has been developed for the first time to determine plasma kisspeptin levels in bovine. A wide range of kisspeptin concentrations can be detected during different physiological stages in bovine using this kisspeptin-EIA procedure.

Kisspeptin and its Current Clinical Status- A Systematic Review.

BACKGROUND: Kisspeptin was initially known as metastin for its role in suppressing metastasis in melanoma and breast cancer. Later, based on its ability to stimulate GPR54, its importance in maintaining an intact hypothalamic-pituitary-ovarian axis was recognised, which is the basis for the widespread application of the drug in several conditions such as secondary amenorrhea, regulation of puberty onset, ovarian function, trophoblast invasion, fertility regulation, parturition, and lactation. This systematic study aims to evaluate the current status of kisspentin in clinical trials. METHODS: The keywords 'kisspeptin' or 'metastin' were used in the clinicaltrials.gov website and Clinical Trial Registry of India (CTRI) to find eligible clinical trials or records carried out without time constraints until February 26, 2023. RESULTS: A total of 33 records were identified through clinical trial databases. All records were screened, and four trials were rejected as they failed to meet the inclusion criteria. Finally, 29 (87.9%) reports of interventional clinical trials with kisspeptin were reviewed. CONCLUSION: Kisspeptin can be viewed as a multipurpose drug with considerably fewer side effects due to its effects simulating normal physiological processes in our body.

Temperature differentially regulates the two kisspeptin systems in the brain of zebrafish.

Kisspeptins encoded by the kiss1 and kiss2 genes play an important role in reproduction through the stimulation of gonadotropin-releasing hormone (GnRH) secretion by activating their receptors (KissR1 EU047918 and KissR2 EU047917). To understand the mechanism through which temperature affects reproduction, we examined kiss1 and kiss2 and their respective receptor (kissr1 and kissr2) gene expression in the brain of male zebrafish exposed to a low temperature (15°C), normal temperature (27°C), and high temperature (35°C) for 7-days. kiss1 mRNA levels in the brain were significantly increased (2.9-fold) in the low temperature compared to the control (27°C), while no noticeable change was observed in the high temperature conditions. Similarly, kissr1 mRNA levels were significantly increased (1.5-2.2-folds) in the low temperature conditions in the habenula, the nucleus of the medial longitudinal fascicle, oculomotor nucleus, and the interpeduncular nucleus. kiss2 mRNA levels were significantly decreased (0.5-fold) in the low and high temperature conditions, concomitant with kissr2 mRNA levels (0.5-fold) in the caudal zone of the periventricular hypothalamus and the posterior tuberal nucleus. gnrh3 but not gnrh2 mRNA levels were also decreased (0.5-fold) in the low and high temperature conditions. These findings suggest that while the kiss1/kissr1 system is sensitive to low temperature, the kiss2/kissr2 system is sensitive to both extremes of temperature, which leads to failure in reproduction.

Kisspeptin and kisspeptin receptor immunoreactivity in euploid and aneuploid choriodecidual tissues of recurrent pregnancy losses.

OBJECTIVE: To study choriodecidual immunoreactivity of kisspeptin (KISS1) and its receptor (KISS1R) in recurrent pregnancy loss (RPL) due to aneuploidy (AnE) and unexplained (UE) RPL in comparison to control elective abortions (EAbs). DESIGN: This is a case-control study. SETTING: Tertiary care facility and affiliated research institute. PATIENT(S): Patients with either UE RPL (n = 10) or RPL due to AnE (n = 10) vs. a control group of patients who underwent EAb (n = 10). INTERVENTION(S): Immunohistochemistry of archived choriodecidual tissue samples. MAIN OUTCOME MEASURE(S): Histoscores of KISS1 and KISS1R immunoreactivity in the syncytiotrophoblast (SyT), cytotrophoblast (CyT), decidual glands (DeGs), and decidual stroma (DeS) across the 3 study groups. RESULT(S): There was no difference in both maternal and gestational ages among the 3 groups. Kisspeptin immunoreactivity was similar in the SyT, CyT, DeGs, and DeS of all groups. Similarly, KISS1R expression was not different in the DeGs or DeS among all study groups. In addition, there was no difference in KISS1R immunoreactivity in the SyTs and CyTs between patients with RPL due to AnE and those with UE RPL. However, KISS1R was significantly lower in the SyT and CyT of patients with RPL due to AnE and UE RPL than in those who underwent EAb. CONCLUSION(S): The expression of KISS1R is lower in the chorionic tissues of euploid (unexplained) and aneuploid RPLs than in the control group. The current results broaden our understanding of the role played by KISS1 and KISS1R in early placentation. Further investigation is necessary to determine whether KISS1 activity is the cause or a sequel of defective placentation.

Short-term depletion of plasma estrogen affects hypothalamic kisspeptin-neurokinin B-dynorphin A neurons, gonadotrophs, and pulsatile luteinizing hormone secretion in female rats.

Kisspeptin-neurokinin B-dynorphin A (KNDy) neurons in the arcuate nucleus (ARC) regulate pulsatile luteinizing hormone (LH) secretion. These neurons express estrogen receptors and are negatively regulated by estrogen. This study aimed to determine whether estrogen supplementation after short-term ovariectomy-induced estrogen depletion has different effects on KNDy neurons depending on the timing of the supplementation. To decrease endogenous estradiol (E2) for a short time, adult female rats received a tube filled with E2 one week after ovariectomy and utilized it one week later (O1w + E). From the results of immunohistochemistry, the response to E2 was attenuated in KNDy neurons of O1w + E rats. Enlarged LH-secreting cells in the anterior pituitary were found in O1w + E rats; however, such enlarged LH cells were not found in ones without previous short-term E2 depletion. From the analysis of LH pulses, plasma LH levels were increased in O1w + E rats relative to ones without previous short-term E2 depletion. These results suggested that once endogenous sex steroids were depleted, the response to E2 in hypothalamic KNDy neurons did not fully recover in one week. Thus, short-term sex steroid depletion due to gonadectomy could alter the response to the sex steroids in KNDy neurons even though the period without sex steroids is only one week, and the alteration is likely to affect plasma hormone levels.

Sex-specific association between urinary kisspeptin and pubertal development.

Kisspeptins play a crucial role during pubertal development, but little is known about how their peripheral concentrations relate to sexual maturation. This is partly due to the lack of non-invasive, quick, and reliable peripheral kisspeptin measures, which limit widespread testing. Here, we investigated the relationship between kisspeptin concentrations measured from midstream urine samples with 2-h retention periods and developmental markers (age, self-reported pubertal status, and saliva concentrations of testosterone and DHEA sulphate ) in 209 typically developing 9- to 15-year-old males and females. As a result of the study, we found marked sex differences. Kisspeptin concentrations were similar between sexes until around 12 years of age, but, thereafter, kisspeptin concentrations in females did not change significantly, whereas, in males, there was a clear positive correlation with developmental measures. Our results replicate previous findings regarding kisspeptin concentration changes across the pubertal transition obtained from blood samples, suggesting that measuring these peptides in urine has the potential for exploring kisspeptins' peripheral effects and their associations with pubertal status.

Hypothalamic KNDy neuron expression in streptozotocin-induced diabetic female rats.

Kisspeptin neurons, i.e. KNDy neurons, in the arcuate nucleus (ARC) coexpress neurokinin B and dynorphin and regulate gonadotropin-releasing hormone/luteinizing hormone (LH) pulses. Because it remains unclear whether these neurons are associated with reproductive dysfunction in diabetic females, we examined the expression of KNDy neurons detected by histochemistry in streptozotocin (STZ)-induced diabetic female rats 8 weeks after STZ injection. We also evaluated relevant metabolic parameters - glucose, 3-hydroxybutyrate, and non-esterified fatty acids - as indicators of diabetes progression. Severe diabetes with hyperglycemia and severe ketosis suppressed the mRNA expression of KNDy neurons, resulting in low plasma LH levels and persistent diestrus. In moderate diabetes with hyperglycemia and moderate ketosis, kisspeptin-immunoreactive cells and plasma LH levels were decreased, while the mRNA expression of KNDy neurons remained unchanged. Mild diabetes with hyperglycemia and slight ketosis did not affect KNDy neurons and plasma LH levels. The number of KNDy cells was strongly and negatively correlated with plasma 3-hydroxybutyrate levels. The vaginal smear analysis showed unclear proestrus in diabetic rats 3-5 days after STZ injection, and the mRNA expression of kisspeptin in the ARC was decreased 2 weeks after STZ injection in severely diabetic rats. Kisspeptin neurons in the anteroventral periventricular nucleus (AVPV), which induce an LH surge, were unaffected at 2 and 8 weeks after STZ injection regardless of the diabetes severity. These results suggest that diabetes mellitus progression in females may negatively affect ARC kisspeptin neurons but not AVPV kisspeptin neurons, implicating a potential role of ARC kisspeptin neurons in menstrual disorder and infertility.

Circulating kisspeptin and anti-müllerian hormone levels, and insulin resistance in women with polycystic ovary syndrome: A systematic review, meta-analysis, and meta-regression.

OBJECTIVE: This systematic review and meta-analysis aimed to summarize the available evidence regarding circulating kisspeptin and anti-müllerian hormone (AMH) and the homeostasis model assessment of insulin resistance (HOMA-IR) index in adolescents and women with and without polycystic ovary syndrome (PCOS). METHOD: We performed a comprehensive literature search in Medline, Embase, Cochrane, Scopus, and Web of Science for studies evaluating circulating kisspeptin levels in women with and without PCOS published until September 24th, 2020. Co-primary outcomes were the HOMA-IR index and AMH. The quality of included studies was assessed using the Newcastle-Ottawa Scale. Random-effects models were used to estimate outcomes, and effects reported as mean difference (MD) or standardized MD (SMD) and their 95 % confidence interval (CI). The systematic review and meta-analysis was registered in the International Prospective Register of Systematic Reviews (PROSPERO) as number CRD42020205030. RESULTS: We evaluated 18 studies including, 1282 PCOS cases and 977 controls. Participants with PCOS were younger (MD = -2.38 years, 95 %CI -4.32 to -0.44), with higher BMI (MD = 1.16, 95 % CI 0.54-1.78), waist-to-hip ratio (MD = 0.04, 95 %CI 0.02 to 0.05), circulating kisspeptin (SMD = 1.15, 95 %CI 0.68-1.62), luteinizing hormone (SMD = 1.29, 95 %CI 0.76-1.83), AMH (SMD = 0.97, 95 %CI 0.60-1,34), total testosterone (SMD = 2.48, 95 %CI 1.73-3.23), free testosterone (SMD = 1.37, 95 %CI 0.56-2.17), and dehydroepiandrosterone sulphate (SMD = 0.72, 95 %CI 0.32-1.13) levels, and Ferriman-Gallwey score (SMD = 5.08, 95 %CI 2.76-7.39), and lower sex hormone-binding globulin level (SMD = -1.34, 95 %CI -2.15 to -0.52). Besides, participants with PCOS had higher HOMA-IR index (SMD = 0.76, 95 %CI 0.35-1.17), and circulating insulin (SMD = 0.75, 95 %CI 0.30-1.19), leptin (SMD = 2.82, 95 %CI 1.35-4.29), and triglycerides (SMD = 2.15, 95 %CI 1.08-3.23) levels than participants without the syndrome. The meta-regression did not identify significant factors influencing circulating kisspeptin. CONCLUSION: Patients with PCOS showed higher kisspeptin, LH, insulin, AMH, and androgen levels and HOMA-IR index, and lower sex hormone-binding globulin levels than those without the syndrome.

Immunoreactivity of Kisspeptin and Kisspeptin Receptor in Eutopic and Ectopic Endometrial Tissue of Women With and Without Endometriosis.

Endometriosis is characterized by the presence of ectopic endometrial tissues. Mechanisms of tissue dissemination in endometriosis may be similar to those involved in tumor metastasis. We hypothesize that dysregulation of kisspeptin (KISS1), a metastasis suppressor in endometrial carcinoma, may contribute to the pathogenesis of endometriosis. In this study, we characterized the immunoreactivity of kisspeptin and its receptor, KISS1R, in eutopic and ectopic endometrial tissue of women with and without endometriosis, in proliferative and secretory menstrual cycle phases. Immunohistochemistry was performed using KISS1 and KISS1R antibodies on samples from women with (n = 35) and without (n = 14) endometriosis. Samples from women with endometriosis included eutopic endometrium (n = 20) samples, superficial endometriotic implants (SUP, n = 10) deep infiltrating endometriotic implants (DIE, n = 15), and ovarian endometriomas (OMA, n = 15). Immunoreactivity was quantified using histoscores. KISS1 and KISS1R immunoreactivity was significantly lower in eutopic endometrial stroma of women with versus without endometriosis, regardless of the menstrual cycle phase (P = 0.001 and P = 0.015 respectively). In endometriotic implants, KISS1 levels were significantly lower in both glandular and stromal components of DIE (P < 0.01) and OMA (P < 0.01) compared to SUP. KISS1R immunoreactivity was lower in the glandular component of OMA (P = 0.035) compared to SUP. KISS1 and KISS1R levels are lower in eutopic endometrial stroma from women with versus without endometriosis, consistent with a role for decreased KISS1 expression in the pathogenesis of endometriosis. As deeply invasive lesions showed lower KISS1 levels than superficial lesions, downregulation of KISS1 levels may contribute to implant invasiveness.

Distribution of GnRH and Kisspeptin Immunoreactivity in the Female Llama Hypothalamus.

Llamas are induced non-reflex ovulators, which ovulate in response to the hormonal stimulus of the male protein beta-nerve growth factor (ß-NGF) that is present in the seminal plasma; this response is dependent on the preovulatory gonadotrophin-releasing hormone (GnRH) release from the hypothalamus. GnRH neurones are vital for reproduction, as these provide the input that controls the release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) from the pituitary gland. However, in spontaneous ovulators, the activity of GnRH cells is regulated by kisspeptin neurones that relay the oestrogen signal arising from the periphery. Here, we investigated the organisation of GnRH and kisspeptin systems in the hypothalamus of receptive adult female llamas. We found that GnRH cells exhibiting different shapes were distributed throughout the ventral forebrain and some of these were located in proximity to blood vessels; sections of the mediobasal hypothalamus (MBH) displayed the highest number of cells. GnRH fibres were observed in both the organum vasculosum laminae terminalis (OVLT) and median eminence (ME). We also detected abundant kisspeptin fibres in the MBH and ME; kisspeptin cells were found in the arcuate nucleus (ARC), but not in rostral areas of the hypothalamus. Quantitative analysis of GnRH and kisspeptin fibres in the ME revealed a higher innervation density of kisspeptin than of GnRH fibres. The physiological significance of the anatomical findings reported here for the ovulatory mechanism in llamas is still to be determined.