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Bariatric surgery improves both insulin sensitivity and secretion and can induce diabetes remission. However, the mechanisms and time courses of these changes, particularly the impact on β cell function, are difficult to monitor directly. In this study, we investigated the effect of Vertical Sleeve Gastrectomy (VSG) on β cell function in vivo by imaging Ca2+ dynamics in islets engrafted into the anterior eye chamber. Mirroring its clinical utility, VSG in mice results in significantly improved glucose tolerance, and enhanced insulin secretion. We reveal that these benefits are underpinned by augmented β cell function and coordinated activity across the islet. These effects involve changes in circulating GLP-1 levels which may act both directly and indirectly on the β cell, in the latter case through changes in body weight. Thus, bariatric surgery leads to time-dependent increases in β cell function and intra-islet connectivity which are likely to contribute to diabetes remission., Bariatric surgery procedures reduce weight and improve metabolic diseases. Here the authors investigate the effects of Vertical Sleeve Gastrectomy (VSG) on beta-cell function in mice using in vivo Ca2 + imaging and show that VSG increases beta-cell function and intra-islet connectivity.

BackgroundGlucocorticoids produced by the adrenal cortex are essential for the maintenance of metabolic homeostasis. Glucocorticoid activation is catalyzed by 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) and signalling is achieved through the glucocorticoid receptor (GR), a ligand-dependent transcription factor. Excess glucocorticoids are associated with insulin resistance and hyperglycaemia. A small number of studies have investigated the effects of bariatric surgery, a gastrointestinal procedure known to improve insulin sensitivity, on glucocorticoid metabolism, but the hypothesised mechanism is assumed to be via weight loss.AimTo investigate the effect of bariatric surgery on glucocorticoid metabolism in lean and obese mice.MethodsLean mice and HFD mice underwent Vertical Sleeve Gastrectomy (VSG) or sham surgery. Glucose and insulin tolerance tests were performed at four and ten weeks post operatively and circulating corticosterone was measured. Liver and adipose tissues were harvested from fed mice and 11β-HSD1 and GR levels were measured by quantitative RT-PCR or Western (immuno-) blotting, respectively.ResultsVSG did not cause excess weight loss in lean mice when compared to sham operated mice. However, both lean and HFD VSG mice displayed significantly improved glucose clearance and insulin sensitivity. Remarkably, VSG restores physiological corticosterone production in HFD mice and reduces11β-HSD1 levels at four and ten weeks post-surgery. Additionally, lean mice displayed significantly lowered mRNA levels of 11β-HSD1 in subcutaneous adipose tissue and GR in liver.ConclusionsBariatric surgery improves insulin sensitivity and reduces glucocorticoid activation at tissular level, under physiological and pathophysiological (obesity) conditions, irrespective of weight loss. The reduction of glucocorticoid exposure may represent an additional contribution to the health benefits of bariatric surgery. These findings point towards a physiologically relevant gut-adrenal axis.

Long non-coding RNAs (lncRNAs) are emerging as crucial regulators of {beta}-cell development and function. Consequently, the mis-expression of members of this group may contribute to the risk of type 2 diabetes (T2D). Here, we investigate roles for an antisense lncRNA expressed from the Pax6 locus (annotated as Pax6os1 in mice and PAX6-AS1 in humans) in {beta}-cell function. The transcription factor Pax6 is required for the development of pancreatic islets and maintenance of a fully differentiated {beta}-cell phenotype. Pax6os1/PAX6-AS1 expression was increased in pancreatic islets and {beta}-cell lines at high glucose concentrations, in islets from mice fed a high fat diet, and in those from patients with type 2 diabetes. Silencing or deletion of Pax6os1/PAX6-AS1 in MIN6 cells and EndoC-{beta}H1cells, respectively, upregulated {beta}-cell signature genes, including insulin. Moreover, shRNA-mediated silencing of PAX6-AS1 in human islets not only increased insulin mRNA, but also enhanced glucose-stimulated insulin secretion and calcium dynamics. In contrast, inactivation of Pax6os1 in mice was largely without effect on glucose homeostasis, though female Pax6os1 null mice on high fat diet (HFD) showed a tendency towards enhanced glucose clearance. Together, our results suggest that increased expression of PAX6-AS1 at high glucose levels may contribute to {beta}-cell dedifferentiation and failure in some forms of type 2 diabetes. Thus, targeting PAX6-AS1 may provide a promising strategy to enhance insulin secretion and improve glucose homeostasis in type 2 diabetes.

Background: Bariatric surgery improves both insulin sensitivity and secretion in T2D. However, the impact on insulin secretion is difficult to monitor directly, requiring correction for insulin sensitivity. The degree to which surgery impacts β-cell function versus mass, and the time course over which these changes occur, also remain unclear. Aim: To investigate the effect of bariatric surgery on β-cell function in vivo, we imaged Ca2+ dynamics prospectively at the cellular level in islets engrafted into the anterior eye chamber. Methods: Ins1Cre mice were crossed to animals that express the genetic calcium indicator GCaMP6f behind a Lox PSTOPLoxP cassette. Isolated islets were engrafted into male C57BL6 mice maintained for 12 weeks on high fat/high sucrose diet (HFD). At pre- (-1) and post-operative weeks 4, 8 and 12, Vertical Sleeve Gastrectomy (VSG) (n=7) and sham (n=6) groups were anaesthetized, and islets imaged in the eye using a Nikon microscope equipped with a spinning disc, 488 nm laser and 20x/1.0 water immersion objective. Islets were scored as positive for wave activity when these were recurrent, had a defined origin and covered >75 % of the image plane. Glucose (3g/kg) tolerance and insulin secretion were assessed in parallel. Results: The VSG group demonstrated substantial weight loss during the first 4 weeks post-operatively but regained pre-operative weight by week 12. Nonetheless, VSG mice displayed significantly improved glucose tolerance and insulin secretion (p Conclusions: By imaging islet function in the eye in vivo we demonstrate that bariatric surgery leads to a time-dependent increase in glucose-induced Ca2+ dynamics of individual islets, likely to underlie increased insulin secretion. Disclosure E. Akalestou: None. K. Suba: None. L. Lopez Noriega: None. P.L. Chabosseau: None. I. Leclerc: Consultant; Spouse/Partner; Sun Pharmaceutical Industries Ltd. Research Support; Spouse/Partner; Servier. V. Salem: None. G.A. Rutter: Consultant; Self; Sun Pharmaceutical Industries Ltd. Research Support; Self; Servier, Sun Pharmaceutical Industries Ltd. Funding UK Wellcome Trust (212625/Z/18/Z); Rosetrees Trust (M825); UK Medical Research Council (MR/R022259/1); Harry Keen Diabetes UK Fellowship (BDA 15/0005317); European Union

Bariatric surgery improves both insulin sensitivity and secretion in type 2 diabetes. However, these changes are difficult to monitor directly and independently. In particular, the degree and the time course over which surgery impacts β cell function, versus mass, have been difficult to establish. In this study, we investigated the effect of bariatric surgery on β cell function in vivo by imaging Ca2+ dynamics prospectively and at the single cell level in islets engrafted into the anterior eye chamber. Islets expressing GCaMP6f selectively in the β cell were transplanted into obese male hyperglycaemic mice that were then subjected to either vertical sleeve gastrectomy (VSG) or sham surgery. Imaged in vivo in the eye, VSG improved coordinated Ca2+ activity, with 90% of islets observed exhibiting enhanced Ca2+ wave activity ten weeks post-surgery, while islet wave activity in sham animals fell to zero discernible coordinated islet Ca2+ activity at the same time point. Correspondingly, VSG mice displayed significantly improved glucose tolerance and insulin secretion. Circulating fasting levels of GLP-1 were also increased after surgery, potentially contributing to improved β cell performance. We thus demonstrate that bariatric surgery leads to time-dependent increases in individual β cell function and intra-islet connectivity, together driving increased insulin secretion and diabetes remission, in a weight-loss independent fashion.Significance StatementUsed widely to treat obesity, bariatric surgery also relieves the symptoms of type 2 diabetes. The mechanisms involved in diabetes remission are still contested, with increased insulin sensitivity and elevated insulin secretion from pancreatic β cells both implicated. Whilst the speed of remission – usually within a few days – argues for improvements in β cell function rather than increases in mass, a direct demonstration of changes at the level of individual β cells or islets has been elusive. Here, we combine vertical sleeve gastrectomy with intravital imaging of islets engrafted into the mouse anterior eye chamber to reveal that surgery causes a time-dependent improvement in glucose-induced Ca2+ dynamics and β cell - β cell connectivity, both of which likely underlie increased insulin release.

Aims/hypothesis Mitochondrial oxidative metabolism is central to glucose-stimulated insulin secretion (GSIS). Whether Ca2+ uptake into pancreatic beta cell mitochondria potentiates or antagonises this process is still a matter of debate. Although the mitochondrial Ca2+ importer (MCU) complex is thought to represent the main route for Ca2+ transport across the inner mitochondrial membrane, its role in beta cells has not previously been examined in vivo. Methods Here, we inactivated the pore-forming subunit of the MCU, encoded by Mcu, selectively in mouse beta cells using Ins1Cre-mediated recombination. Whole or dissociated pancreatic islets were isolated and used for live beta cell fluorescence imaging of cytosolic or mitochondrial Ca2+ concentration and ATP production in response to increasing glucose concentrations. Electrophysiological recordings were also performed on whole islets. Serum and blood samples were collected to examine oral and i.p. glucose tolerance. Results Glucose-stimulated mitochondrial Ca2+ accumulation (pppMcu-null (βMcu-KO) animals, in vitro, as compared with wild-type (WT) mice. Interestingly, cytosolic Ca2+ concentrations increased (pMcu-KO animals. βMcu-KO mice displayed impaired in vivo insulin secretion at 5 min (ppMcu-KO ( Conclusions/interpretation MCU is crucial for mitochondrial Ca2+ uptake in pancreatic beta cells and is required for normal GSIS. The apparent compensatory mechanisms that maintain glucose tolerance in βMcu-KO mice remain to be established.

BackgroundBariatric surgery has been established to improve insulin sensitivity and glucose clearance, but also increases insulin and glucagon secretion. Each of the above effects have also been observed following treatment with sodium glucose co-transporter 2 (SGLT2) inhibitors.AimTo determine whether there is an effect of bariatric surgery (Vertical Sleeve Gastrectomy; VSG) on renal SGLT2 expression in mice.MethodsEighteen lean mice underwent VSG (n=8) or sham (n=9) surgery. Glucose tolerance tests with or without treatment with the SGLT2 inhibitor dapagliflozin were performed four weeks post operatively, in order to assess if pharmacological SGLT2 inhibition has the same euglycemic effects after bariatric surgery. Kidneys were harvested from fed mice and SGLT2 expression was analysed using Quantitative reverse-transcription PCR and immunofluorescence.ResultsVSG mice displayed significantly improved glucose tolerance (AUC=103±6.8; AUC=66.6±2.9 in control and VSG mice, respectively; pConclusionsVertical sleeve gastrectomy in lean animals causes a significant inhibition of SGLT2 expression in the kidney cortex. These findings are in line with our previous results on the effects of Duodenal Jejunal Bypass in lean rats, and point towards a physiologically-relevant gut-kidney axis. SGLT2 inhibition may thus be an important mechanism through which bariatric surgery improves glucose tolerance in man.

Background: Bariatric surgery is known to improve glucose tolerance and weight loss, but can also cause a postprandial increase in renal glucose excretion following oral glucose tolerance test. Both effects have also been observed following treatment with sodium glucose co-transporter 2 (SGLT2) inhibitors. Aim: To investigate whether there is an effect of bariatric surgery (Vertical Sleeve Gastrectomy; VSG) on renal SGLT2 expression in mice. Methods: Fourteen lean mice underwent VSG (n=8) or sham (n=6) surgery. Glucose Tolerance tests with or without treatment with the SGLT2 inhibitor dapagliflozin were performed four weeks post operatively, in order to assess if exogenous SGLT2 inhibition has the same euglycemic effects as bariatric surgery. Kidneys were harvested from fed mice and analysed using Quantitative real-time PCR (qRT-PCR) and immunofluorescence. Results: VSG did not cause any weight loss when compared to sham operated mice (p=0.37, Mann-Whitney test). However, VSG mice displayed significantly improved glucose tolerance (AUC=103 ±6.8; AUC =66.6 ±2.9 in control and VSG mice, respectively; p0.05). In contrast, treatment of sham mice with dapagliflozin increased glucose tolerance, though to a lesser extent than VSG. Nevertheless, qPCR and immunofluorescence analysis on mouse kidneys demonstrated a significant lowering of SGLT2 expression at both the protein (n=5, p=0.0007) and mRNA (n=7, 0 Conclusions: Both gastric and intestinal surgery in lean animals causes a significant inhibition of SGLT2 in the kidney cortex. These findings are in line with our previous results on the effects of Duodenal Jejunal Bypass in lean rats, and point towards a physiologically-relevant gut-kidney axis. SGLT2 inhibition may thus be an important mechanism through which bariatric surgery improves glucose tolerance in man. Disclosure E. Akalestou: None. L. Lopez Noriega: None. P.L. Chabosseau: None. I. Leclerc: None. G.A. Rutter: Consultant; Self; Sun Pharma. Funding UK Wellcome Trust