Multifunctional host-defense peptides isolated from skin secretions of the banana tree dwelling frog Boana platanera (Hylidae; Hylinae).

Five host-defense peptides (figainin 2PL, hylin PL, raniseptin PL, plasticin PL, and peptide YL) were isolated from norepinephrine-stimulated skin secretions of the banana tree dwelling frog Boana platanera (Hylidae; Hylinae) collected in Trinidad. Raniseptin PL (GVFDTVKKIGKAVGKFALGVAKNYLNS.NH2) and figainin 2PL (FLGTVLKLGKAIAKTVVPMLTNAMQPKQ. NH2) showed potent and rapid bactericidal activity against a range of clinically relevant Gram-positive and Gram-negative ESKAPE + pathogens and Clostridioides difficile. The peptides also showed potent cytotoxic activity (LC50 values < 30 µM) against A549, MDA-MB-231 and HT29 human tumor-derived cell lines but appreciably lower hemolytic activity against mouse erythrocytes (LC50 = 262 ± 14 µM for raniseptin PL and 157 ± 16 µM for figainin 2PL). Hylin PL (FLGLIPALAGAIGNLIK.NH2) showed relatively weak activity against microorganisms but was more hemolytic. The glycine-leucine-rich peptide with structural similarity to the plasticins (GLLSTVGGLVGGLLNNLGL.NH2) and the non-cytotoxic peptide YL (YVPGVIESLL.NH2) lacked antimicrobial and cytotoxic activities. Hylin PL, raniseptinPL and peptide YL stimulated the rate of release of insulin from BRIN-BD11 clonal ß-cells at concentrations ≥100 nM. Peptide YL was the most effective (2.3-fold increase compared with basal rate at 1 µM concentration) and may represent a template for the design of a new class of incretin-based anti-diabetic drugs.

Dopamine signalling in pancreatic islet cells and role in adaptations to metabolic stress.

OBJECTIVES: Dopamine and related receptors are evidenced in pancreatic endocrine tissue, but the impact on islet ß-cell stimulus-secretion as well as (patho)physiological role are unclear. METHODS: The present study has evaluated islet cell signalling pathways and biological effects of dopamine, as well as alterations of islet dopamine in rodent models of diabetes of different aetiology. KEY FINDINGS: The dopamine precursor L-DOPA partially impaired glucose tolerance in mice and attenuated glucose-, exendin-4, and alanine-induced insulin secretion. The latter effect was echoed by the attenuation of glucose-induced [Ca2+]i dynamics and elevation of ATP levels in individual mouse islet cells. L-DOPA significantly decreased ß-cell proliferation rates, acting predominantly via the D2 receptor, which was most abundant at the mRNA level. The administration of streptozotocin (STZ) or high-fat diet (HFD) in mice significantly elevated numbers of dopamine-positive islet cells, with HFD also increasing colocalization of dopamine with insulin. At the same time, colocalization of dopamine with glucagon was increased in STZ-treated and pregnant mice, but unaffected by HFD. CONCLUSION: These findings highlight a role for dopamine receptor signalling in islet cell biology adaptations to various forms of metabolic stress.

The impact of diabetes and obesity on fertility and the potential role of gut hormones as treatment.

AIMS: Alongside its metabolic implications, obesity and associated diabetes impair female reproductive function, causing infertility and polycystic ovarian syndrome (PCOS). Recently, gut hormones and their receptors have been identified in various reproductive organs indicating their potential regulatory effects on reproductive function. This review aims to give an overview of their potential effects. METHODS: This review focuses on literature that outlines modifications during obesity, diabetes and related infertility with an emphasis on gut hormones and their therapeutic potential. RESULTS: Evidence suggests that bariatric surgery has positive effects on fertility and PCOS where major alterations in metabolism occurs through restoration of gut hormone levels. This is thought to be due to the indirect effect weight loss and regulation of blood glucose has on the hypothalamic-pituitary-ovarian and hypothalamic-pituitary-adrenal axis influencing reproduction. CONCLUSIONS: Further research is required to elucidate the cellular mechanisms involved in the direct effects of gut hormone receptor activation on reproductive tissues. Current observations suggest a therapeutic role for gut hormones in infertility/PCOS associated with metabolic pathophysiology.

GLP-1 receptor agonism and GIP receptor antagonism induce substantial alterations in enteroendocrine and islet cell populations in obese high fat fed mice.

Effects of sustained activation of glucagon-like peptide-1 (GLP-1) receptors (GLP-1R) as well as antagonism of receptors for glucose-dependent insulinotropic peptide (GIP) on intestinal morphology and related gut hormone populations have not been fully investigated. The present study assesses the impact of 21-days twice daily treatment with the GLP-1R agonist exendin-4 (Ex-4), or the GIP receptor (GIPR) antagonist mGIP(3-30), on these features in obese mice fed a high fat diet (HFD). HFD mice presented with reduced crypt depth when compared to normal diet (ND) controls, which was reversed by Ex-4 treatment. Both regimens lead to an enlargement of villi length in HFD mice. HFD mice had increased numbers of GIP and PYY positive ileal cells, with both treatment interventions reversing the effect on PYY positive cells, but only Ex-4 restoring GIP ileal cell populations to ND levels. Ex-4 and mGIP (3-30) marginally decreased GLP-1 villi immunoreactivity and countered the reduction of ileal GLP-1 content caused by HFD. As expected, HFD mice presented with elevated pancreatic islet area. Interestingly, mGIP(3-30), but not Ex-4, enhanced islet and beta-cell areas in HFD mice despite lack of effect of beta-cell turnover, whilst Ex-4 increased delta-cell area. Co-localisation of islet PYY or GLP-1 with glucagon was increased by Ex-4, whilst islet PYY co-immunoreactivity with somatostatin was enhanced by mGIP(3-30) treatment. These observations highlight potential new mechanisms linked to the metabolic benefits of GLP-1R agonism and GIPR antagonism in obesity.

Disturbed ovarian morphology, oestrous cycling and fertility of high fat fed rats are linked to alterations of incretin receptor expression.

Obesity is a major cause of infertility in females with a direct correlation between energy intake and reproductive dysfunction. To explore underlying mechanisms, disturbances in reproductive health and incretin/reproductive hormone receptor expression were studied in female Wistar rats fed a high-fat-diet for 20-weeks. Metabolic parameters and ovarian/adrenal gene expression were monitored along with estrous cycling and fertility upon mating. High-fat-feeding significantly increased body weight, plasma insulin and HOMA-IR, indicative of obesity and insulin resistance. Estrous cycles were prolonged compared to normal chow-fed rats, with 50 % having an average cycle length ≥ 7days. Reproductive outcomes revealed high-fat-diet reduced litter size by 48 %, with 16 % rats unable to achieve pregnancy. Furthermore, 80 % of the high-fat group took > 35 days to become pregnant compared to 33 % fed a normal-diet. Also, 35 % of pups born to high-fat-fed rats were eaten by mothers or born dead which was not observed with control rats. These changes were associated with downregulation of Amh, Npy2R and GcgR gene expression in ovaries with upregulation of InsR and Glp-1R genes. In adrenals, Glp-1R, GipR, Npy2R, InsR, GcgR, GshR and Esr-1 genes were upregulated. Histological analysis of high-fat-diet ovaries and adrenals revealed changes in morphology with significantly increased number of cysts and reduced adrenal capsule thickness. Circulating levels of insulin, testosterone and progesterone was significantly higher in high-fat group with reduced FSH levels in plasma. These data demonstrate that high-fat feeding disrupts female reproductive function and suggest important interactions between gut and reproductive hormones in ovaries and adrenals which merit further investigation.

RYGB surgery has modest effects on intestinal morphology and gut hormone populations in the bypassed biliopancreatic limb but causes reciprocal changes in GLP-2 and PYY in the alimentary limb.

Roux-en-Y gastric-bypass (RYGB) induced alterations in intestinal morphology and gut-cell hormone expression profile in the bypassed biliopancreatic-limb (BPL) versus the alimentary-limbs (AL) are poorly characterised. This pilot study has therefore explored effects following RYGB in high-fat-diet (HFD) and normal-diet (ND) rats. Female Wistar rats (4-week-old) were fed HFD or ND for 23-weeks prior to RYGB or sham surgeries. Immunohistochemical analysis of excised tissue was conducted three-weeks post-surgery. After RYGB, intestinal morphology of the BPL in both HFD and ND groups was unchanged with exception of a small decrease in villi width in the ND-RYGB and crypt depth in the HFD-RYGB group. However, in the AL, villi width was decreased in ND-RYGB rats but increased in the HFD-RYGB group. In addition, crypt depth decreased after RYGB in the AL of HFD rats. GIP positive cells in either limb of both groups of rats were unchanged by RYGB. Similarly, there was little change in GLP-1 positive cells, apart from a small decrease of numbers in the villi of the BPL in HFD rats. RYGB increased GLP-2 cell numbers in the AL of ND-RYGB rats, including in both crypts and villi. This was associated with decreased numbers of cells expressing PYY in the AL of ND-RYGB rats. The BPL appears to maintain normal morphology and unchanged enteroendocrine cell populations despite being bypassed in RYGB-surgery. In contrast, in the AL, villi area is generally enhanced post-RYGB in ND rats with increased numbers of GLP-2 positive cells and decreased expression of PYY.

Evidence for Involvement of GIP and GLP-1 Receptors and the Gut-Gonadal Axis in Regulating Female Reproductive Function in Mice.

Substantial evidence suggests crosstalk between reproductive and gut-axis but mechanisms linking metabolism and reproduction are still unclear. The present study evaluated the possible role of glucose-dependent-insulinotropic-polypeptide (GIP) and glucagon-like-peptide-1 (GLP-1) in reproductive function by examining receptor distribution and the effects of global GIPR and GLP-1R deletion on estrous cycling and reproductive outcomes in mice. GIPR and GLP-1R gene expression were readily detected by PCR in female reproductive tissues including pituitary, ovaries and uterine horn. Protein expression was confirmed with histological visualisation of incretin receptors using GIPR-Cre and GLP1R-Cre mice in which the incretin receptor expressing cells were fluorescently tagged. Functional studies revealed that female GIPR-/- and GLP-1R-/- null mice exhibited significantly (p < 0.05 and p < 0.01) deranged estrous cycling compared to wild-type controls, indicative of reduced fertility. Furthermore, only 50% and 16% of female GIPR-/- and GLP-1R-/- mice, respectively produced litters with wild-type males across three breeding cycles. Consistent with a physiological role of incretin receptors in pregnancy outcome, litter size was significantly (p < 0.001-p < 0.05) decreased in GIPR-/- and GLP-1R-/- mice. Treatment with oral metformin (300 mg/kg body-weight), an agent used clinically for treatment of PCOS, for a further two breeding periods showed no amelioration of pregnancy outcome except that litter size in the GIPR-/- group was approximately 2 times greater in the second breeding cycle. These data highlight the significance of incretin receptors in modulation of female reproductive function which may provide future targets for pharmacological intervention in reproductive disorders.

Differential effects of RYGB surgery and best medical treatment for obesity-diabetes on intestinal and islet adaptations in obese-diabetic ZDSD rats.

Modification of gut-islet secretions after Roux-En-Y gastric bypass (RYBG) surgery contributes to its metabolic and anti-diabetic benefits. However, there is limited knowledge on tissue-specific hormone distribution post-RYGB surgery and how this compares with best medical treatment (BMT). In the present study, pancreatic and ileal tissues were excised from male Zucker-Diabetic Sprague Dawley (ZDSD) rats 8-weeks after RYGB, BMT (daily oral dosing with metformin 300mg/kg, fenofibrate 100mg/kg, ramipril 1mg/kg, rosuvastatin 10mg/kg and subcutaneous liraglutide 0.2mg/kg) or sham operation (laparotomy). Insulin, glucagon, somatostatin, PYY, GLP-1 and GIP expression patterns were assessed using immunocytochemistry and analyzed using ImageJ. After RYGB and BMT, body weight and plasma glucose were decreased. Intestinal morphometry was unaltered by RYGB, but crypt depth was decreased by BMT. Intestinal PYY cells were increased by both interventions. GLP-1- and GIP-cell counts were unchanged by RYGB but BMT increased ileal GLP-1-cells and decreased those expressing GIP. The intestinal contents of PYY and GLP-1 were significantly enhanced by RYGB, whereas BMT decreased ileal GLP-1. No changes of islet and beta-cell area or proliferation were observed, but the extent of beta-cell apoptosis and islet integrity calculated using circularity index were improved by both treatments. Significantly decreased islet alpha-cell areas were observed in both groups, while beta- and PYY-cell areas were unchanged. RYGB also induced a decrease in islet delta-cell area. PYY and GLP-1 colocalization with glucagon in islets was significantly decreased in both groups, while co-staining of PYY with glucagon was decreased and that with somatostatin increased. These data characterize significant cellular islet and intestinal adaptations following RYGB and BMT associated with amelioration of obesity-diabetes in ZDSD rats. The differential responses observed and particularly those within islets, may provide important clues to the unique ability of RYGB to cause diabetes remission.