The glucose-lowering effects of α-glucosidase inhibitor require a bile acid signal in mice.

AIMS/HYPOTHESIS: Bile-acid (BA) signalling is crucial in metabolism homeostasis and has recently been found to mediate the therapeutic effects of glucose-lowering treatments, including α-glucosidase inhibitor (AGI). However, the underlying mechanisms are yet to be clarified. We hypothesised that BA signalling may be required for the glucose-lowering effects and metabolic benefits of AGI. METHODS: Leptin receptor (Lepr)-knockout (KO) db/db mice and high-fat high-sucrose (HFHS)-fed Fxr (also known as Nr1h4)-KO mice were treated with AGI. Metabolic phenotypes and BA signalling in different compartments, including the liver, gut and endocrine pancreas, were evaluated. BA pool profiles were analysed by mass spectrometry. The islet transcription profile was assayed by RNA sequencing. The gut microbiome were assayed by 16S ribosomal RNA gene sequencing. RESULTS: AGI lowered microbial BA levels in BA pools of different compartments in the body, and increased gut BA reabsorption in both db/db and HFHS-fed mouse models via altering the gut microbiome. The AGI-induced changes in BA signalling (including increased activation of farnesoid X receptor [FXR] in the liver and inhibition of FXR in the ileum) echoed the alterations in BA pool size and composition in different organs. In Fxr-KO mice, the glucose- and lipid-lowering effects of AGI were partially abrogated, possibly due to the Fxr-dependent effects of AGI on decelerating beta cell replication, alleviating insulin hypersecretion and improving hepatic lipid and glucose metabolism. CONCLUSIONS/INTERPRETATION: By regulating microbial BA metabolism, AGI elicited diverse changes in BA pool composition in different host compartments to orchestrate BA signalling in the whole body. The AGI-induced changes in BA signalling may be partly required for its glucose-lowering effects. Our study, hence, sheds light on the promising potential of regulating microbial BA and host FXR signalling for the treatment of type 2 diabetes. DATA AVAILABILITY: Sequencing data are available from the BioProject Database (accession no. PRJNA600345; www.ncbi.nlm.nih.gov/bioproject/600345).

NPC1 is required for postnatal islet ß cell differentiation by maintaining mitochondria turnover.

Rationale: NPC1 is a protein localized on the lysosome membrane regulating intracellular cholesterol transportation and maintaining normal lysosome function. GWAS studies have found that NPC1 variants in T2D was a pancreatic islet expression quantitative trait locus, suggesting a potential role of NPC1 in T2D islet pathophysiology. Methods: Two-week-old Npc1-/- mice and wild type littermates were employed to examine pancreatic ß cell morphology and functional changes induced by loss of Npc1. Single cell RNA sequencing was conducted on primary islets. Npc1-/- Min6 cell line was generated using CRISPR/Cas9 gene editing. Seahorse XF24 was used to analyze primary islet and Min6 cell mitochondria respiration. Ultra-high-resolution cell imaging with Lattice SIM2 and electron microscope imaging were used to observe mitochondria and lysosome in primary islet ß and Min6 cells. Mitophagy Dye and mt-Keima were used to measure ß cell mitophagy. Results: In Npc1-/- mice, we found that ß cell survival and pancreatic ß cell mass expansion as well as islet glucose induced insulin secretion in 2-week-old mice were reduced. Npc1 loss retarded postnatal ß cell differentiation and growth as well as impaired mitochondria oxidative phosphorylation (OXPHOS) function to increase mitochondrial superoxide production, which might be attributed to impaired autophagy flux particularly mitochondria autophagy (mitophagy) induced by dysfunctional lysosome in Npc1 null ß cells. Conclusion: Our study revealed that NPC1 played an important role in maintaining normal lysosome function and mitochondria turnover, which ensured establishment of sufficient mitochondria OXPHOS for islet ß cells differentiation and maturation.

Dopamine-inhibited POMCDrd2+ neurons in the ARC acutely regulate feeding and body temperature.

Dopamine acts on neurons in the arcuate nucleus (ARC) of the hypothalamus, which controls homeostatic feeding responses. Here we demonstrate a differential enrichment of dopamine receptor 1 (Drd1) expression in food intake-promoting agouti related peptide (AgRP)/neuropeptide Y (NPY) neurons and a large proportion of Drd2-expressing anorexigenic proopiomelanocortin (POMC) neurons. Owing to the nature of these receptors, this translates into a predominant activation of AgRP/NPY neurons upon dopamine stimulation and a larger proportion of dopamine-inhibited POMC neurons. Employing intersectional targeting of Drd2-expressing POMC neurons, we reveal that dopamine-mediated POMC neuron inhibition is Drd2 dependent and that POMCDrd2+ neurons exhibit differential expression of neuropeptide signaling mediators compared with the global POMC neuron population, which manifests in enhanced somatostatin responsiveness of POMCDrd2+ neurons. Selective chemogenetic activation of POMCDrd2+ neurons uncovered their ability to acutely suppress feeding and to preserve body temperature in fasted mice. Collectively, the present study provides the molecular and functional characterization of POMCDrd2+ neurons and aids our understanding of dopamine-dependent control of homeostatic energy-regulatory neurocircuits.

The Intestinal Effect of Atorvastatin: Akkermansia muciniphila and Barrier Function.

Studies have shown that the cholesterol-lowering medicine statins alter the gut microbiome, induce chronic metabolic inflammation, and disrupt glycemic homeostasis. In this study, we aimed to investigate whether effects of atorvastatin (Ator) on gut microbiome and metabolic inflammation could be causally correlated. Mice at 8-week age were fed with high-fat diet (HFD) or HFD with Ator (HFD+Ator) for 16 weeks. 16S rRNA sequencing of stool and RNA sequencing of colon tissue were employed to analyze the intestinal alterations that could be induced by Ator. A human colon carcinoma cell line (Caco2) was used for in vitro experiments on barrier function. Compared to HFD, HFD+Ator induced more weight gain, impaired glucose tolerance, and led to gut microbiota dysbiosis, such as suppressing Akkermansia muciniphila in mice. The expressions of tight junction (TJ) proteins were attenuated in the colon, and the serum LPS-binding-protein (LBP) level was elevated in HFD+Ator mice, so as to transcriptionally activate the intestinal nuclear factor-k-gene binding (NF-κB) signaling pathway. Consistently, Ator impaired the barrier function of Caco2, and treatment of supernatant of A. Muciniphila culture could decrease the intestinal permeability and recover the attenuated expression of TJ proteins induced by Ator. In conclusion, long-term use of Ator with HFD may alter gut microbiota, induce intestinal barrier dysfunction, and hence promote chronic inflammation that contributes to disrupted glycemic homeostasis.

Enhancing Acsl4 in absence of mTORC2/Rictor drove ß-cell dedifferentiation via inhibiting FoxO1 and promoting ROS production.

Rapamycin insensitive companion of mechanistic target of Rapamycin (Rictor), the key component of mTOR complex 2 (mTORC2), controls both ß-cell proliferation and function. We sought to study whether long chain acyl-CoA synthetase 4 (Acsl4) worked downstream of Rictor/mTORC2 to maintain ß-cell functional mass. We found Acsl4 was positively regulated by Rictor at transcriptional and posttranslational levels in mouse ß-cell. Infecting adenovirus expressing Acsl4 in ß-cell-specific-Rictor-knockout (ßRicKO) islets and Min6 cells knocking down Rictor with lentivirus-expressing siRNA-oligos targeting Rictor(siRic), recovered the ß-cell dysplasia but not dysfunction. Cell bioenergetic experiment performed with Seahorse XF showed that Acsl4 could not rescue the dampened glucose oxidation in Rictor-lacking ß-cell, but further promoted lipid oxidation. Transposase-Accessible Chromatin (ATAC) and H3K27Ac chromatin immunoprecipitation (ChIP) sequencing studies reflected the epigenetic elevated molecular signature for ß-cell dedifferentiation and mitigated oxidative defense/response. These results were confirmed by the observations of elevated acetylation and ubiquitination of FoxO1, increased protein levels of Gpx1 and Hif1an, excessive reactive oxygen species (ROS) production and diminished MafA in Acsl4 overexpressed Rictor-lacking ß-cells. In these cells, antioxidant treatment significantly recovered MafA level and insulin content. Inducing lipid oxidation alone could not mimic the effect of Acsl4 in Rictor lacking ß-cell. Our study suggested that Acsl4 function in ß-cell was context dependent and might facilitate ß-cell dedifferentiation with attenuated Rictor/mTORC2 activity or insulin signaling via posttranslational inhibiting FoxO1 and epigenetically enhancing ROS induced MafA degradation.

Atorvastatin Targets the Islet Mevalonate Pathway to Dysregulate mTOR Signaling and Reduce ß-Cell Functional Mass.

Statins are cholesterol-lowering agents that increase the incidence of diabetes and impair glucose tolerance via their detrimental effects on nonhepatic tissues, such as pancreatic islets, but the underlying mechanism has not been determined. In atorvastatin (ator)-treated high-fat diet-fed mice, we found reduced pancreatic ß-cell size and ß-cell mass, fewer mature insulin granules, and reduced insulin secretion and glucose tolerance. Transcriptome profiling of primary pancreatic islets showed that ator inhibited the expression of pancreatic transcription factor, mechanistic target of rapamycin (mTOR) signaling, and small G protein (sGP) genes. Supplementation of the mevalonate pathway intermediate geranylgeranyl pyrophosphate (GGPP), which is produced by 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase, significantly restored the attenuated mTOR activity, v-maf musculoaponeurotic fibrosarcoma oncogene homolog A (MafA) expression, and ß-cell function after ator, lovastatin, rosuvastatin, and fluvastatin treatment; this effect was potentially mediated by sGP prenylation. Rab5a, the sGP in pancreatic islets most affected by ator treatment, was found to positively regulate mTOR signaling and ß-cell function. Rab5a knockdown mimicked the effect of ator treatment on ß-cells. Thus, ator impairs ß-cell function by regulating sGPs, for example, Rab5a, which subsequently attenuates islet mTOR signaling and reduces functional ß-cell mass. GGPP supplementation could constitute a new approach for preventing statin-induced hyperglycemia.

The delayed effects of antibiotics in type 2 diabetes, friend or foe?

An increasing amount of evidence suggests that the delayed effect of antibiotics (abx) on gut microbiota after its cessation is not as favorable as its immediate effect on host metabolism. However, it is not known how the diverse abx-dependent metabolic effects influence diabetic subjects and how gut microbiota is involved. Here, we treated db/db mice with abx cocktail for 12 days and discontinued for 24 days. We found that db/db mice showed decreased body weight and blood glucose after abx treatment, which rapidly caught up after abx cessation. Twenty-four days after abx withdrawal, db/db mice exhibit increased plasma, hepatic total cholesterol (TC) levels and liver weight. The gut microbiota composition at that time showed decreased relative abundances (RAs) of Desulfovibrionaceae and Rikenellaceae, increased RA of Erysipelotrichaceae and Mogibacteriaceae, which were correlating with the reduced short-chain fatty acids (SCFAs) in gut content, such as propionic acid and valeric acid and with the elevated fecal taurine-conjugated bile acids (BAs) levels. The molecular biology studies showed inhibited hepatic BA synthesis from cholesterol, impeded intracellular transportation and biliary excretion of cholesterol that all conferred to liver TC accumulation. The associations among alterations of gut microbiota composition, microbial metabolite profiles and host phenotypes suggested the existence of gut microbiota-linked mechanisms that mediate the unfavorable delayed effects of abx on db/db mice cholesterol metabolism. Thus, we call upon the caution of applying abx in diabetic animal models for studying microbiota-host interaction and in type 2 diabetes subjects for preventing chronic cardiovascular consequences.