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Abstract Background We previously reported that, among all the naturally occurring amino acids, l-valine is the most powerful luminal stimulator of glucagon-like peptide 1 (GLP-1) release from the upper part of the rat small intestine. This makes l-valine an interesting target for nutritional-based modulation of GLP-1 secretion. However, the molecular mechanism of l-valine-induced secretion remains unknown. Methods We aimed to investigate the effect of orally given l-valine in mice and to identify the molecular details of l-valine stimulated GLP-1 release using the isolated perfused rat small intestine and GLUTag cells. In addition, the effect of l-valine on hormone secretion from the distal intestine was investigated using a perfused rat colon. Results Orally given l-valine (1 g/kg) increased plasma levels of active GLP-1 comparably to orally given glucose (2 g/kg) in male mice, supporting that l-valine is a powerful stimulator of GLP-1 release in vivo (P > 0.05). Luminal l-valine (50 mM) strongly stimulated GLP-1 release from the perfused rat small intestine (P 0.05), suggesting that co-transport of l-valine and Na+ is not important for the depolarization necessary to activate the voltage-gated Ca2+-channels. Administration of the KATP-channel opener diazoxide (250 μM) completely blocked the l-valine induced GLP-1 response (P

Abstract Background Ten percent of the female population suffers from congenital abnormalities of the vagina, uterus, or oviducts, with severe consequences for reproductive and psychological health. Yet, the underlying causes of most of these malformations remain largely unknown. ADGRA3 (GPR125) is involved in WNT signaling and planar cell polarity, mechanisms vital to female reproductive tract development. Although ADGRA3 is a well-established spermatogonial stem cell marker, its role within the female urogenital system remains unclear. Results In this study, we found Adgra3 to be expressed throughout the murine female urogenital system, with higher expression pre-puberty than after sexual maturation. We generated a global Adgra3 −/− mouse line and observed imperforate vagina in 44% of Adgra3 −/− females, resulting in distension of the reproductive tract and infertility. Ovarian morphology, plasma estradiol, ovarian Cyp19a1, and vaginal estrogen receptor α (Esr1) expression were unaffected. However, compared to controls, a significantly lower bone mineral density was found in Adgra3 −/− mice. Whereas vaginal opening in mice is an estrogen-dependent process, 17β-estradiol treatment failed to induce vaginal canalization in Adgra3 −/− mice. Furthermore, a marked reduction in vaginal and ovarian progesterone receptor expression was observed concomitant with an upregulation of apoptotic regulators Bcl2, Bid, and Bmf in adult Adgra3 −/− females with a closed vagina. Conclusions Our collective results shed new insights into the complex mechanisms by which the adhesion receptor ADGRA3 regulates distal vaginal tissue remodeling during vaginal canalization via altered sex hormone responsiveness and balance in apoptotic regulators. This highlights the potential of ADGRA3 as a target in diagnostic screening and/or therapy for obstructive vaginal malformations in humans.

Toft-Bertelsen et al. describe repurposing of anti-influenza drug amantadine and its derivatives for the treatment of SARS-CoV-2. They show that Amantadine, Emodin and Xanthene show significant blockage of ionchannels formed by SARS-CoV-2 which are crucial for its assembly and pathophysiology.

The intestinal hormone, glucose-dependent insulinotropic polypeptide (GIP), is involved in important physiological functions, including postprandial blood glucose homeostasis, bone remodeling, and lipid metabolism. While mutations leading to physiological changes can be identified in large-scale sequencing, no systematic investigation of GIP missense variants has been performed. Here, we identified 168 naturally occurring missense variants in the human GIP genes from three independent cohorts comprising ~720,000 individuals. We examined amino acid changing variants scattered across the pre-pro-GIP peptide using in silico effect predictions, which revealed that the sequence of the fully processed GIP hormone is more protected against mutations than the rest of the precursor protein. Thus, we observed a highly species-orthologous and population-specific conservation of the GIP peptide sequence, suggestive of evolutionary constraints to preserve the GIP peptide sequence. Elucidating the mutational landscape of GIP variants and how they affect the structural and functional architecture of GIP can aid future biological characterization and clinical translation.

Oxidized cholesterols have emerged as important signaling molecules of immune function, but little is known about the role of these oxysterols during mycobacterial infections. We found that expression of the oxysterol-receptor GPR183 was reduced in blood from patients with tuberculosis (TB) and type 2 diabetes (T2D) compared to TB patients without T2D and was associated with TB disease severity on chest x-ray. GPR183 activation by 7α,25-dihydroxycholesterol (7α,25-OHC) reduced growth of Mycobacterium tuberculosis (Mtb) and Mycobacterium bovis BCG in primary human monocytes, an effect abrogated by the GPR183 antagonist GSK682753. Growth inhibition was associated with reduced IFN-β and IL-10 expression and enhanced autophagy. Mice lacking GPR183 had significantly increased lung Mtb burden and dysregulated IFNs during early infection. Together, our data demonstrate that GPR183 is an important regulator of intracellular mycobacterial growth and interferons during mycobacterial infection.

The choroid plexus (CP) plays a major role in controlling the entry of substances and immune cells into the brain as it forms the blood-cerebrospinal fluid barrier (BCSFB) in the brain ventricles. Dysregulated immune cell trafficking through the epithelial cell (EC) layer of CP is central for the pathogenesis of infectious diseases in the brain and many neurodegenerative disorders. In vitro studies elucidating the function of the CP have so far been limited to the monolayer culture of CP ECs. To mimic immune cell migration across the CP barrier, a three-dimensional model would be advantageous. Here, we present an in vitro platform for studies of the immune cell trafficking based on CP explants/organoids. The explants were generated from fragments of mouse CPs in Matrigel, where the cells formed luminal spaces and could be maintained in culture for at least 8 weeks. We demonstrate expression of the major CP markers in the explants, including transthyretin and aquaporin 1 as well as ZO1 and ICAM-1, indicating a capacity for secretion of cerebrospinal fluid (CSF) and presence of tight junctions. CP explants displayed CP-like cell polarization and formed an intact EC barrier. We also show that the expression of transthyretin, transferrin, occludin and other genes associated with various functions of CP was maintained in the explants at similar levels as in native CP. By using dendritic cells and neutrophils, we show that the migration activity of immune cells and their interactions with CP epithelium can be monitored by microscopy. Thereby, the three-dimensional CP explant model can be used to study the cellular and molecular mechanisms mediating immune cell migration through CP epithelium and other functions of choroid EC. We propose this platform can potentially be used in the search for therapeutic targets and intervention strategies to improve control of (drug) substances and (immune) cell entry into the central nervous system.

B1 cells constitute a specialized subset of B cells, best characterized in mice, which is abundant in body cavities, including the peritoneal cavity. Through natural and antigen-induced antibody production, B1 cells participate in the early defense against bacteria. The G protein-coupled receptor 183 (GPR183), also known as Epstein-Barr virus-induced gene 2 (EBI2), is an oxysterol-activated chemotactic receptor that regulates migration of B cells. We investigated the role of GPR183 in B1 cells in the peritoneal cavity and omentum. B1 cells expressed GPR183 at the mRNA level and migrated towards the GPR183 ligand 7α,25-dihydroxycholesterol (7α,25-OHC). GPR183 knock-out (KO) mice had smaller omenta, but with normal numbers of B1 cells, whereas they had fewer B2 cells in the omentum and peritoneal cavity than wildtype (WT) mice. GPR183 was not responsible for B1 cell accumulation in the omentum in response to i.p. lipopolysaccharide (LPS)-injection, in spite of a massive increase in 7α,25-OHC levels. Lack of GPR183 also did not affect B1a- or B1b cell-specific antibody responses after vaccination. In conclusion, we found that GPR183 is non-essential for the accumulation and function of B1 cells in the omentum and peritoneal cavity, but that it influences the abundance of B2 cells in these compartments.

ABSTRACT Bones have been suggested to be a target for glucagon‐like peptide ‐1 (GLP‐1); however, studies of the effects on human bones so far have given diverging results. We hypothesized that GLP‐1, together with glucagon‐like peptide‐2 and glucose‐dependent insulinotropic polypeptide, plays a role in the gut–bone axis. We examined the acute effect of three GLP‐1 receptor ligands [GLP‐1 (7‐36)amide, GLP‐1 (9‐36)amide, and exenatide] on markers of bone remodeling. Eight healthy, normal‐weight participants, with a mean age of 24.3 years, were studied for 4 days in a double‐blinded, randomized clinical trial. Blood was collected before and after s.c. injection of GLP‐1 (7‐36)amide (1.5 nmol/kg), GLP‐1 (9‐36)amide (1.5 nmol/kg), exenatide (2.4 nmol/subject), or saline. Plasma was analyzed for bone markers and for osteoprotegerin (OPG), PTH, and IGF‐1 levels. All ligands were tested in vitro for their cAMP‐inducing activity on the human GLP‐1 receptor. GLP‐1 (7‐36)amide decreased CTX‐levels, compared with placebo (area under the curve [AUC] ±SD 0 to 120 min = –2143 ± 1294 % × min versus –883 ± 1557 % × min; p < 0.05). No difference was observed between placebo and GLP‐1 (9‐36)amide, or between placebo and exenatide, although exenatide had a similar potency as GLP‐1 (7‐36)amide for cAMP formation in vitro (EC50 of 0.093 and 0.054 nmol/L). However, exenatide reached maximum plasma concentration at 90 min versus 15 min for GLP‐1 (7‐36)amide, and plasma CTX was significantly decreased during the second hour of the study after exenatide injections compared with placebo (AUC ±SD –463.1 ± 218 % × min and –136 ± 91 % × min; p < 0.05). There was no effect of the injections on bone formation markers (P1NP and osteocalcin) or on OPG, PTH and IGF‐1 levels. In conclusion, we show that GLP‐1 receptor agonists, but not the primary metabolite GLP‐1 (9‐36)amide, decrease bone resorption, and suggest that GLP‐1 may be part of the gut–bone axis. © 2019 The Authors. JBMR Plus is published by Wiley Periodicals, Inc. on behalf of the American Society for Bone and Mineral Research.

Glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-2 (GLP-2) are hormones secreted from the enteroendocrine cells after a meal. They exert their actions through activation of G protein-coupled receptors (R), the GIPR and GLP-2R, respectively. Both have been reported to influence metabolism. The purpose of the study was to investigate the role of the hormones in the regulation of lipid and bone homeostasis by subchronic treatment with novel GIPR and GLP-2R antagonists. Rats were injected once daily with vehicle, GIPR, or GLP-2R antagonists for 3 weeks. Body weight, food intake, body composition, plasma lipoprotein lipase (LPL), adipokines, triglycerides and the marker of bone resorption carboxy-terminal collagen crosslinks (CTX), were examined. In rats, subchronic treatment with GIPR antagonist, rat GIP (3-30)NH2, did not modify food intake and bone resorption, but significantly increased body weight, body fat mass, triglycerides, LPL, and leptin levels compared with vehicle treated rats. Subchronic (Pro3)GIP (a partial GIPR agonist), GLP-2(11-33), and GLP-2(3-33) (GLP-2R antagonists) treatment did not affect any parameter. The present results would be consistent with a role for GIP, but not GLP-2, in the maintenance of lipid homeostasis in rats, while neither GIPR nor GLP-2R antagonism appeared to influence bone resorption in rats.

Bone homeostasis displays a circadian rhythm with increased resorption during the night time as compared to day time, a difference that seems—at least partly—to be caused by food intake during the day. Thus, ingestion of a meal results in a decrease in bone resorption, but people suffering from short bowel syndrome lack this response. Gut hormones, released in response to a meal, contribute to this link between the gut and bone metabolism. The responsible hormones appear to include glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1), known as incretin hormones due to their role in regulating glucose homeostasis by enhancing insulin release in response to food intake. They interact with their cognate receptors (GIPR and GLP-1R), which are both members of the class B G protein-coupled receptors (GPCRs), and already recognized as targets for treatment of metabolic diseases, such as type 2 diabetes mellitus (T2DM) and obesity. Glucagon-like peptide-2 (GLP-2), secreted concomitantly with GLP-1, acting via another class B receptor (GLP-2R), is also part of this gut-bone axis. Several studies, including human studies, have indicated that these three hormones inhibit bone resorption and, moreover, that GIP increases bone formation. Another hormone, peptide YY (PYY), is also secreted from the enteroendocrine L-cells (together with GLP-1 and GLP-2), and acts mainly via interaction with the class A GPCR NPY-R2. PYY is best known for its effect on appetite regulation, but recent studies have also shown an effect of PYY on bone metabolism. The aim of this review is to summarize the current knowledge of the actions of GIP, GLP-1, GLP-2, and PYY on bone metabolism, and to discuss future therapies targeting these receptors for the treatment of osteoporosis.

Endocytosis is a fundamental process involved in trafficking of various extracellular and transmembrane molecules from the cell surface to its interior. This enables cells to communicate and respond to external environments, maintain cellular homeostasis, and transduce signals. G protein-coupled receptors (GPCRs) constitute a family of receptors with seven transmembrane alpha-helical domains (7TM receptors) expressed at the cell surface, where they regulate physiological and pathological cellular processes. Several herpesviruses encode receptors (vGPCRs) which benefits the virus by avoiding host immune surveillance, supporting viral dissemination, and thereby establishing widespread and lifelong infection, processes where receptor signaling and/or endocytosis seem central. vGPCRs are rising as potential drug targets as exemplified by the cytomegalovirus-encoded receptor US28, where its constitutive internalization has been exploited for selective drug delivery in virus infected cells. Therefore, studying GPCR trafficking is of great importance. This review provides an overview of the current knowledge of endocytic and cell localization properties of vGPCRs and methodological approaches used for studying receptor internalization. Using such novel approaches, we show constitutive internalization of the BILF1 receptor from human and porcine γ-1 herpesviruses and present motifs from the eukaryotic linear motif (ELM) resources with importance for vGPCR endocytosis.

Insulin is a vital part of diabetes treatment, whereas glucagon is primarily used to treat insulin-induced hypoglycemia. However, glucagon is suggested to have a central role in the regulation of body weight, which would be beneficial for diabetic patients. Since the glucagon effect on blood glucose is known to be transient, it is relevant to investigate the pharmacodynamics of glucagon after repeated dosing. In the present study, we used telemetry to continuously measure blood glucose in streptozotocin induced diabetic Sprague-Dawley rats. This allowed for a more detailed analysis of glucose regulation compared to intermittent blood sampling. In particular, we evaluated the blood glucose-lowering effect of different insulin doses alone, and in combination with a long acting glucagon analog (LAG). We showed how the effect of the LAG accumulated and persisted over time. Furthermore, we found that addition of the LAG decreased body weight without affecting food intake. In a subsequent study, we focused on the glucagon effect on body weight and food intake during equal glycemic control. In order to obtain comparable maximum blood glucose lowering effect to insulin alone, the insulin dose had to be increased four times in combination with 1 nmol/kg of the LAG. In this set-up the LAG prevented further increase in body weight despite the four times higher insulin-dose. However, the body composition was changed. The insulin group increased both lean and fat mass, whereas the group receiving four times insulin in combination with the LAG only significantly increased the fat mass. No differences were observed in food intake, suggesting a direct effect on energy expenditure by glucagon. Surprisingly, we observed decreased levels of FGF21 in plasma compared to insulin treatment alone. With the combination of insulin and the LAG the blood glucose-lowering effect of insulin was prolonged, which could potentially be beneficial in diabetes treatment.

Several herpes- and poxviruses have captured chemokine receptors from their hosts and modified these to their own benefit. The human and viral chemokine receptors belong to class A 7 transmembrane (TM) receptors which are characterized by several structural motifs like the DRY-motif in TM3 and the C-terminal tail. In the DRY-motif, the arginine residue serves important purposes by being directly involved in G protein coupling. Interestingly, among the viral receptors there is a greater diversity in the DRY-motif compared to their endogenous receptor homologous. The C-terminal receptor tail constitutes another regulatory region that through a number of phosphorylation sites is involved in signaling, desensitization, and internalization. Also this region is more variable among virus-encoded 7TM receptors compared to human class A receptors. In this review we will focus on these two structural motifs and discuss their role in viral 7TM receptor signaling compared to their endogenous counterparts.