Incretin hormones, obesity and gut microbiota.

Over the past 40 years, the prevalence of obesity has risen dramatically, reaching epidemic proportions. By 2030 the number of people affected by obesity will reach 1.12 billion worldwide. Gastrointestinal hormones, namely incretins, play a vital role in the pathogenesis of obesity and its comorbidities. GIP (glucose-dependent insulinotropic polypeptide) and GLP-1 (glucagon-like peptide-1), which are secreted from the intestine after nutrient intake and stimulate insulin secretion from pancreatic ß cells, influence lipid metabolism, gastric empting, appetite and body weight. The gut microbiota plays an important role in various metabolic conditions, including obesity and type 2 diabetes and influences host metabolism through the interaction with enteroendocrine cells that modulate incretins secretion. Gut microbiota metabolites, such as short-chain fatty acids (SCFAs) and indole, directly stimulate the release of incretins from colonic enteroendocrine cells influencing host satiety and food intake. Moreover, bariatric surgery and incretin-based therapies are associated with increase gut bacterial richness and diversity. Understanding the role of incretins, gut microbiota, and their metabolites in regulating metabolic processes is crucial to develop effective strategies for the management of obesity and its associated comorbidities.

Intestinal heat shock proteins in metabolic syndrome: Novel mediators of obesity and its comorbidities resolution after metabolic surgery.

Over the past 40 years, the prevalence of obesity has risen dramatically, reaching epidemic proportions. Metabolic surgery has proven to be highly effective in treating obesity, leading to significant improvements or complete resolution of obesity-related comorbidities. Research conducted in both animals and humans suggests that the metabolic benefits achieved through metabolic surgery cannot be solely attributed to weight loss. Indeed, there has been an increasing recognition of intestinal inflammation as a novel factor influencing obesity. The gastrointestinal tract is continuously exposed to dietary components, particularly diets rich in saturated fats, which are known to contribute to obesity. It is now widely accepted that heat shock proteins can be released from various cells including intestinal epithelial cells and act as proinflammatory signals. Several studies have shown that circulating levels of glucose-regulated protein 78 (GRP78) are increased in subjects with obesity and correlate with the severity of the disease. Moreover, mice with a partial knockout of GRP78 are protected from diet-induced obesity. In this review, we discuss the role of GRP78 in the development of obesity. Several evidence suggests that GRP78 can influence adipogenesis, lipid droplets stabilization, insulin resistance, and liver steatosis. We also provide an update on GRP78 regulation following metabolic surgery, focusing on the bypass of the small intestine as a key factor for GRP78 secretion. Finally, we discuss the potential role of monoclonal antibodies against GRP78 as a treatment for obesity.

The first external validation of the Dallas steatosis index in biopsy-proven Non-alcoholic fatty liver Disease: A multicenter study.

AIMS: A new non-invasive tool (NIT) for non-alcoholic fatty liver disease (NAFLD) proposed in 2022 by the multi-ethnic Dallas Heart Study, i.e. the Dallas Steatosis Index (DSI), was validated herein using for the first time the gold standard i.e. liver biopsy-proven NAFLD. METHODS: This is a multicenter study based on samples and data from two Gastroenterology-Hepatology Clinics (Greece and Australia) and one Bariatric-Metabolic Surgery Clinic (Italy). Overall, n = 455 patients with biopsy-proven NAFLD (n = 374) and biopsy-proven controls (n = 81) were recruited. RESULTS: The ability of DSI to correctly classify participants as NAFLD or controls was very good, reaching an Area Under the Curve (AUC) = 0.887. The cut-off point that could best differentiate the presence vs. absence of NAFLD corresponded to DSI = 0.0 (risk threshold: 50% | Sensitivity: 0.88; Positive Predictive Value (PPV): 93.0%; F1-score = 0.91). DSI demonstrated significantly better performance characteristics than other liver steatosis indexes. Decision curve analysis revealed that the benefit of DSI as a marker to indicate the need for invasive liver assessment was confirmed only when higher DSI values, i.e. ≥ 1.4, were used as risk thresholds. DSI performance to differentiate disease progression was inadequate (all AUCs < 0.700). CONCLUSIONS: DSI is more useful for disease screening (NAFLD vs. controls) than to differentiate diseases stages or progression. The value of any inclusion of DSI to guidelines needs to be further studied.

Bariatric-metabolic surgery versus lifestyle intervention plus best medical care in non-alcoholic steatohepatitis (BRAVES): a multicentre, open-label, randomised trial.

BACKGROUND: Observational studies suggest that bariatric-metabolic surgery might greatly improve non-alcoholic steatohepatitis (NASH). However, the efficacy of surgery on NASH has not yet been compared with the effects of lifestyle interventions and medical therapy in a randomised trial. METHODS: We did a multicentre, open-label, randomised trial at three major hospitals in Rome, Italy. We included participants aged 25-70 years with obesity (BMI 30-55 kg/m2), with or without type 2 diabetes, with histologically confirmed NASH. We randomly assigned (1:1:1) participants to lifestyle modification plus best medical care, Roux-en-Y gastric bypass, or sleeve gastrectomy. The primary endpoint of the study was histological resolution of NASH without worsening of fibrosis at 1-year follow-up. This study is registered at ClinicalTrials.gov, NCT03524365. FINDINGS: Between April 15, 2019, and June 21, 2021, we biopsy screened 431 participants; of these, 103 (24%) did not have histological NASH and 40 (9%) declined to participate. We randomly assigned 288 (67%) participants with biopsy-proven NASH to lifestyle modification plus best medical care (n=96 [33%]), Roux-en-Y gastric bypass (n=96 [33%]), or sleeve gastrectomy (n=96 [33%]). In the intention-to-treat analysis, the percentage of participants who met the primary endpoint was significantly higher in the Roux-en-Y gastric bypass group (54 [56%]) and sleeve gastrectomy group (55 [57%]) compared with lifestyle modification (15 [16%]; p<0·0001). The calculated probability of NASH resolution was 3·60 times greater (95% CI 2·19-5·92; p<0·0001) in the Roux-en-Y gastric bypass group and 3·67 times greater (2·23-6·02; p<0·0001) in the sleeve gastrectomy group compared with in the lifestyle modification group. In the per protocol analysis (236 [82%] participants who completed the trial), the primary endpoint was met in 54 (70%) of 77 participants in the Roux-en-Y gastric bypass group and 55 (70%) of 79 participants in the sleeve gastrectomy group, compared with 15 (19%) of 80 in the lifestyle modification group (p<0·0001). No deaths or life-threatening complications were reported in this study. Severe adverse events occurred in ten (6%) participants who had bariatric-metabolic surgery, but these participants did not require re-operations and severe adverse events were resolved with medical or endoscopic management. INTERPRETATION: Bariatric-metabolic surgery is more effective than lifestyle interventions and optimised medical therapy in the treatment of NASH. FUNDING: Fondazione Policlinico Universitario A Gemelli, Policlinico Universitario Umberto I and S Camillo Hospital, Rome, Italy.

Long-term Outcomes of Transoral Outlet Reduction (TORe) for Dumping Syndrome and Weight Regain After Roux-en-Y Gastric Bypass.

BACKGROUND: Both weight regain and dumping syndrome (DS) after Roux-en-Y gastric bypass (RYGB) have been related to the dilation of gastro-jejunal anastomosis. The aim of this study is to assess the safety and long-term efficacy of endoscopic transoral outlet reduction (TORe) for DS and/or weight regain after RYBG. MATERIALS AND METHODS: A retrospective analysis was performed on a prospective database. Sigstad's score, early and late Arts Dumping Score (ADS) questionnaires, absolute weight loss (AWL), percentage of total body weight loss (%TBWL), and percentage of excess weight loss (%EWL) were assessed at baseline and at 6, 12, and 24 months after TORe. RESULTS: Eighty-seven patients (median age 46 years, 79% female) underwent TORe. The median baseline BMI was 36.2 kg/m2. Out of 87 patients, 58 were classified as "dumpers" due to Sigstad's score ≥ 7. The resolution rate of DS (Sigstad's score < 7) was 68.9%, 66.7%, and 57.2% at 6, 12, and 24 months after TORe, respectively. A significant decrease in Sigstad's score as well as in early and late ADS questionnaires was observed (p < 0.001). The median Sigstad's score dropped from 15 (11-8.5) pre-operatively to 2 (0-12) at 24 months. The %TBWL was 10.5%, 9.9%, and 8.1% at 6, 12, and 24 months, respectively. Further, "dumpers" with resolution of DS showed better weight loss results compared with those with persistent DS (p < 0.001). The only adverse event observed was a perigastric fluid collection successfully managed conservatively. CONCLUSION: TORe is a minimally invasive treatment for DS and/or weight regain after RYGB, with evidence of long-term efficacy.

Accurate liquid biopsy for the diagnosis of non-alcoholic steatohepatitis and liver fibrosis.

OBJECTIVE: Clinical diagnosis and approval of new medications for non-alcoholic steatohepatitis (NASH) require invasive liver biopsies. The aim of our study was to identify non-invasive biomarkers of NASH and/or liver fibrosis. DESIGN: This multicentre study includes 250 patients (discovery cohort, n=100 subjects (Bariatric Surgery Versus Non-alcoholic Steato-hepatitis - BRAVES trial); validation cohort, n=150 (Liquid Biopsy for NASH and Liver Fibrosis - LIBRA trial)) with histologically proven non-alcoholic fatty liver (NAFL) or NASH with or without fibrosis. Proteomics was performed in monocytes and hepatic stellate cells (HSCs) with iTRAQ-nano- Liquid Chromatography - Mass Spectrometry/Mass Spectrometry (LC-MS/MS), while flow cytometry measured perilipin-2 (PLIN2) and RAB14 in peripheral blood CD14+CD16- monocytes. Neural network classifiers were used to predict presence/absence of NASH and NASH stages. Logistic bootstrap-based regression was used to measure the accuracy of predicting liver fibrosis. RESULTS: The algorithm for NASH using PLIN2 mean florescence intensity (MFI) combined with waist circumference, triglyceride, alanine aminotransferase (ALT) and presence/absence of diabetes as covariates had an accuracy of 93% in the discovery cohort and of 92% in the validation cohort. Sensitivity and specificity were 95% and 90% in the discovery cohort and 88% and 100% in the validation cohort, respectively.The area under the receiver operating characteristic (AUROC) for NAS level prediction ranged from 83.7% (CI 75.6% to 91.8%) in the discovery cohort to 97.8% (CI 95.8% to 99.8%) in the validation cohort.The algorithm including RAB14 MFI, age, waist circumference, high-density lipoprotein cholesterol, plasma glucose and ALT levels as covariates to predict the presence of liver fibrosis yielded an AUROC of 95.9% (CI 87.9% to 100%) in the discovery cohort and 99.3% (CI 98.1% to 100%) in the validation cohort, respectively. Accuracy was 99.25%, sensitivity 100% and specificity 95.8% in the discovery cohort and 97.6%, 99% and 89.6% in the validation cohort. This novel biomarker was superior to currently used FIB4, non-alcoholic fatty liver disease fibrosis score and aspartate aminotransferase (AST)-to-platelet ratio and was comparable to ultrasound two-dimensional shear wave elastography. CONCLUSIONS: The proposed novel liquid biopsy is accurate, sensitive and specific in diagnosing the presence and severity of NASH or liver fibrosis and is more reliable than currently used biomarkers. CLINICAL TRIALS: Discovery multicentre cohort: Bariatric Surgery versus Non-Alcoholic Steatohepatitis, BRAVES, ClinicalTrials.gov identifier: NCT03524365.Validation multicentre cohort: Liquid Biopsy for NASH and Fibrosis, LIBRA, ClinicalTrials.gov identifier: NCT04677101.

GLUT-1/PKM2 loop dysregulation in patients with non-ST-segment elevation myocardial infarction promotes metainflammation.

AIM: The functional capacity of the immune cells is strongly dependent on their metabolic state and inflammatory responses are characterized by a greater use of glucose in immune cells. This study is aimed to establish the role of glucose metabolism and its players [glucose transporter-1 (GLUT-1) and pyruvate kinase isozyme M2 (PKM2)] in the dysregulation of adaptive immunity and inflammation observed in patients with non-ST-segment elevation myocardial infarction (NSTEMI). METHODS AND RESULTS: We enrolled 248 patients allocated to three groups: NSTEMI patients, chronic coronary syndromes (CCS) patients, healthy subjects (HS). NSTEMI patients showed higher expression of GLUT-1 and an enhanced glucose uptake in T cells as compared to CCS patients (p < 0.0001; p = 0.0101, respectively) and HS (p = 0.0071; p = 0.0122, respectively). PKM2 had a prevalent nuclear localization in T lymphocytes in NSTEMI (p = 0.0005 for nuclear versus cytoplasm localization), while in CCS and HS was equally distributed in both compartments. In addition, the nuclear fraction of PKM2 was significantly higher in NSTEMI compared to HS (p = 0.0023). In NSTEMI patients, treatment with Shikonin and Fasentin, which inhibits PKM2 enzyme activity and GLUT-1 mediated glucose internalization, respectively, led to a significant reduction in GLUT-1 expression along with the downregulation of pro-inflammatory cytokine expression. CONCLUSIONS: NSTEMI patients exhibit dysregulation of the GLUT-1/PKM2 metabolic loop characterized by nuclear translocation of PKM2, where it acts as a transcription regulator of pro-inflammatory genes. This detrimental loop might represent a new therapeutic target for personalized medicine.

Upper gut heat shock proteins HSP70 and GRP78 promote insulin resistance, hyperglycemia, and non-alcoholic steatohepatitis.

A high-fat diet increases the risk of insulin resistance, type-2 diabetes, and non-alcoholic steato-hepatitis. Here we identified two heat-shock proteins, Heat-Shock-Protein70 and Glucose-Regulated Protein78, which are increased in the jejunum of rats on a high-fat diet. We demonstrated a causal link between these proteins and hepatic and whole-body insulin-resistance, as well as the metabolic response to bariatric/metabolic surgery. Long-term continuous infusion of Heat-Shock-Protein70 and Glucose-Regulated Protein78 caused insulin-resistance, hyperglycemia, and non-alcoholic steato-hepatitis in rats on a chow diet, while in rats on a high-fat diet continuous infusion of monoclonal antibodies reversed these phenotypes, mimicking metabolic surgery. Infusion of these proteins or their antibodies was also associated with shifts in fecal microbiota composition. Serum levels of Heat-Shock-Protein70 and Glucose-Regulated Protein78were elevated in patients with non-alcoholic steato-hepatitis, but decreased following metabolic surgery. Understanding the intestinal regulation of metabolism may provide options to reverse metabolic diseases.

Prevalence and predictors of non-alcoholic steatohepatitis in subjects with morbid obesity and with or without type 2 diabetes.

AIM: To investigate the prevalence of biopsy-proven non-alcoholic steatohepatitis (NASH) in a cohort of patients with morbid obesity and with or without type 2 diabetes (T2D) and to find non-invasive predictors of NASH severity. METHODS: We evaluated a cohort of 412 subjects (age 19-67 years, body mass index-BMI: 44.98 kg/m2), who underwent fine-needle liver biopsy during bariatric surgery. Thirty-six percent of the subjects were affected by T2D. Liver biopsies were classified according to the Kleiner's NAFLD Activity Score (NAS). NAFLD Fibrosis Score (NFS), AST/ALT ratio, AST to Platelet ratio (APRI), fibrosis-4 score (FIB4) were calculated. A neural network analysis (NNA) was run to predict NASH severity. RESULTS: The prevalence of biopsy-proven NASH was 63% and 78% in subjects with obesity and without or with T2D, respectively. T2D doubled the risk of NASH [OR 2.079 (95% IC=1.31-3.29)]. The prevalence of NAFL increased with the increase of BMI, while there was an inverse correlation between BMI and NASH (r=-0.145 p=0.003). Only mild liver fibrosis was observed. HOMA-IR was positively associated with hepatocyte ballooning (r=0.208, p<0.0001) and fibrosis (r=0.159, p=0.008). The NNA highlighted a specificity of 77.3% using HDL-cholesterol, BMI, and HOMA-IR as main determinants of NASH. CONCLUSIONS: Our data show a higher prevalence of NASH in patients with morbid obesity than reported in the literature and the pivotal role of T2D among the risk factors for NASH development. However, the inverse correlation observed between BMI and biopsy-proven NASH suggests that over a certain threshold adiposity can be somewhat protective against liver damage. Our model predicts NASH presence with high specificity, thus helping identifying subjects who should promptly undergo liver biopsy.