Machine Learning-Based Plasma Metabolomics in Liraglutide-Treated Type 2 Diabetes Mellitus Patients and Diet-Induced Obese Mice.

Liraglutide, a glucagon-like peptide-1 receptor agonist, is effective in the treatment of type 2 diabetes mellitus (T2DM) and obesity. Despite its benefits, including improved glycemic control and weight loss, the common metabolic changes induced by liraglutide and correlations between those in rodents and humans remain unknown. Here, we used advanced machine learning techniques to analyze the plasma metabolomic data in diet-induced obese (DIO) mice and patients with T2DM treated with liraglutide. Among the machine learning models, Support Vector Machine was the most suitable for DIO mice, and Gradient Boosting was the most suitable for patients with T2DM. Through the cross-evaluation of machine learning models, we found that liraglutide promotes metabolic shifts and interspecies correlations in these shifts between DIO mice and patients with T2DM. Our comparative analysis helped identify metabolic correlations influenced by liraglutide between humans and rodents and may guide future therapeutic strategies for T2DM and obesity.

HMBA ameliorates obesity by MYH9- and ACTG1-dependent regulation of hypothalamic neuropeptides.

The global epidemic of obesity remains a daunting problem. Here, we report hexamethylene bisacetamide (HMBA) as a potent anti-obesity compound. Peripheral and central administration of HMBA to diet-induced obese mice regulated the expression of hypothalamic neuropeptides critical for energy balance, leading to beneficial metabolic effects such as anorexia and weight loss. We found that HMBA bound to MYH9 and ACTG1, which were required for the anti-obesity effects of HMBA in both NPY-expressing and POMC-expressing neurons. The binding of HMBA to MYH9 and ACTG1 elevated the expression of HEXIM1 and enhanced its interaction with MDM2, resulting in the dissociation of the HEXIM1-p53 complex in hypothalamic cells. Subsequently, the free HEXIM1 and p53 translocated to the nucleus, where they downregulated the transcription of orexigenic NPY, but p53 and acetylated histone 3 upregulated that of anorexigenic POMC. Our study points to a previously unappreciated efficacy of HMBA and reveals its mechanism of action in metabolic regulation, which may propose HMBA as a potential therapeutic strategy for obesity.

Glucagon-like peptide-1 analog liraglutide leads to multiple metabolic alterations in diet-induced obese mice.

Liraglutide, a glucagon-like peptide-1 analog, has beneficial metabolic effects in patients with type 2 diabetes and obesity. Although the high efficacy of liraglutide as an anti-diabetic and anti-obesity drug is well known, liraglutide-induced metabolic alterations in diverse tissues remain largely unexplored. Here, we report the changes in metabolic profiles induced by a 2-week subcutaneous injection of liraglutide in diet-induced obese mice fed a high-fat diet for 8 weeks. Our comprehensive metabolomic analyses of the hypothalamus, plasma, liver, and skeletal muscle showed that liraglutide intervention led to various metabolic alterations in comparison with diet-induced obese or nonobese mice. We found that liraglutide remarkably coordinated not only fatty acid metabolism in the hypothalamus and skeletal muscle but also amino acid and carbohydrate metabolism in plasma and liver. Comparative analyses of metabolite dynamics revealed that liraglutide rewired intertissue metabolic correlations. Our study points to a previously unappreciated metabolic alteration by liraglutide in several tissues, which may underlie its therapeutic effects within and across the tissues.

Liver-Specific Deletion of Mouse CTCF Leads to Hepatic Steatosis via Augmented PPARγ Signaling.

BACKGROUND & AIMS: The liver is the major organ for metabolizing lipids, and malfunction of the liver leads to various diseases. Nonalcoholic fatty liver disease is rapidly becoming a major health concern worldwide and is characterized by abnormal retention of excess lipids in the liver. CCCTC-binding factor (CTCF) is a highly conserved zinc finger protein that regulates higher-order chromatin organization and is involved in various gene regulation processes. Here, we sought to determine the physiological role of CTCF in hepatic lipid metabolism. METHODS: We generated liver-specific, CTCF-ablated and/or CD36 whole-body knockout mice. Overexpression or knockdown of peroxisome proliferator-activated receptor (PPAR)γ in the liver was achieved using adenovirus. Mice were examined for development of hepatic steatosis and inflammation. RNA sequencing was performed to identify genes affected by CTCF depletion. Genome-wide occupancy of H3K27 acetylation, PPARγ, and CTCF were analyzed by chromatin immunoprecipitation sequencing. Genome-wide chromatin interactions were analyzed by in situ Hi-C. RESULTS: Liver-specific, CTCF-deficient mice developed hepatic steatosis and inflammation when fed a standard chow diet. Global analysis of the transcriptome and enhancer landscape revealed that CTCF-depleted liver showed enhanced accumulation of PPARγ in the nucleus, which leads to increased expression of its downstream target genes, including fat storage-related gene CD36, which is involved in the lipid metabolic process. Hepatic steatosis developed in liver-specific, CTCF-deficient mice was ameliorated by repression of PPARγ via pharmacologic blockade or adenovirus-mediated knockdown, but hardly rescued by additional knockout of CD36. CONCLUSIONS: Our data indicate that liver-specific deletion of CTCF leads to hepatosteatosis through augmented PPARγ DNA-binding activity, which up-regulates its downstream target genes associated with the lipid metabolic process.

Histone acylation marks respond to metabolic perturbations and enable cellular adaptation.

Acetylation is the most studied histone acyl modification and has been recognized as a fundamental player in metabolic gene regulation, whereas other short-chain acyl modifications have only been recently identified, and little is known about their dynamics or molecular functions at the intersection of metabolism and epigenetic gene regulation. In this study, we aimed to understand the link between nonacetyl histone acyl modification, metabolic transcriptional regulation, and cellular adaptation. Using antibodies specific for butyrylated, propionylated, and crotonylated H3K23, we analyzed dynamic changes of H3K23 acylation upon various metabolic challenges. Here, we show that H3K23 modifications were highly responsive and reversibly regulated by nutrient availability. These modifications were commonly downregulated by the depletion of glucose and recovered based on glucose or fatty acid availability. Depletion of metabolic enzymes, namely, ATP citrate lyase, carnitine acetyltransferase, and acetyl-CoA synthetase, which are involved in Ac-CoA synthesis, resulted in global loss of H3K23 butyrylation, crotonylation, propionylation, and acetylation, with a profound impact on gene expression and cellular metabolic states. Our data indicate that Ac-CoA/CoA and central metabolic inputs are important for the maintenance of histone acylation. Additionally, genome-wide analysis revealed that acyl modifications are associated with gene activation. Our study shows that histone acylation acts as an immediate and reversible metabolic sensor enabling cellular adaptation to metabolic stress by reprogramming gene expression.

Tanycytic TSPO inhibition induces lipophagy to regulate lipid metabolism and improve energy balance.

Hypothalamic glial cells named tanycytes, which line the 3rd ventricle (3V), are components of the hypothalamic network that regulates a diverse array of metabolic functions for energy homeostasis. Herein, we report that TSPO (translocator protein), an outer mitochondrial protein, is highly enriched in tanycytes and regulates homeostatic responses to nutrient excess as a potential target for an effective intervention in obesity. Administration of a TSPO ligand, PK11195, into the 3V, and tanycyte-specific deletion of Tspo reduced food intake and elevated energy expenditure, leading to negative energy balance in a high-fat diet challenge. Ablation of tanycytic Tspo elicited AMPK-dependent lipophagy, breaking down lipid droplets into free fatty acids, thereby elevating ATP in a lipid stimulus. Our findings suggest that tanycytic TSPO affects systemic energy balance through macroautophagy/autophagy-regulated lipid metabolism, and highlight the physiological significance of TSPO in hypothalamic lipid sensing and bioenergetics in response to overnutrition. ABBREVIATIONS: 3V: 3rd ventricle; ACAC: acetyl-Coenzyme A carboxylase; AGRP: agouti related neuropeptide; AIF1/IBA1: allograft inflammatory factor 1; AMPK: AMP-activated protein kinase; ARC: arcuate nucleus; Atg: autophagy related; Bafilo: bafilomycin A1; CAMKK2: calcium/calmodulin-dependent protein kinase kinase 2, beta; CCCP: carbonyl cyanide m-chlorophenylhydrazone; CNS: central nervous system; COX4I1: cytochrome c oxidase subunit 4I1; FFA: free fatty acid; GFAP: glial fibrillary acidic protein; HFD: high-fat diet; ICV: intracerebroventricular; LAMP2: lysosomal-associated membrane protein 2; LD: lipid droplet; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MBH: mediobasal hypothalamus; ME: median eminence; MEF: mouse embryonic fibroblast; NCD: normal chow diet; NEFM/NFM: neurofilament medium; NPY: neuropeptide Y; OL: oleic acid; POMC: pro-opiomelanocortin-alpha; PRKN/Parkin: parkin RBR E3 ubiquitin protein ligase; Rax: retina and anterior neural fold homeobox; RBFOX3/NeuN: RNA binding protein, fox-1 homolog (C. elegans) 3; RER: respiratory exchange ratio; siRNA: small interfering RNA; SQSTM1: sequestosome 1; TG: triglyceride; TSPO: translocator protein; ULK1: unc-51 like kinase 1; VCO2: carbon dioxide production; VMH: ventromedial hypothalamus; VO2: oxygen consumption.

Palmitate-induced autophagy liberates monounsaturated fatty acids and increases Agrp expression in hypothalamic cells.

Fatty acids regulate food intake, although the exact mechanism remains unknown. Emerging evidence suggests that intracellular free fatty acids generated by starvation-induced autophagy regulate food intake. Starvation for 6 h elevated fatty acids such as palmitate, oleate, arachidonate, eicosatrienoate, and docosahexaenoate in the mouse serum. Among them, palmitate induced lipophagy, an autophagic degradation of cellular lipid droplets, in agouti-related peptide (Agrp)-expressing hypothalamic cells. Palmitate-induced lipophagy increased both Agrp expression and the contents of monounsaturated fatty acids such as palmitoleate, oleate, and (E)-9-octadecanoate, whereas these effects were blunted by autophagy deficiency. These findings support the role of free fatty acids in hypothalamic autophagy that regulates the appetite by changing the expression of orexigenic neuropeptides.

Alternative splicing induces cytoplasmic localization of RBFOX2 protein in calcific tendinopathy.

BACKGROUND: Calcific tendinopathy (CT) is characterized by deposits of calcium, most commonly found in the shoulder tendons. The exact cause and pathogenesis of CT are not fully understood. This study analyzed the expression pattern of RNA-binding protein fox-1 homolog 2 (RBFOX2), a crucial splicing regulator in tissue differentiation. METHODS: Normal and calcific tendons were compared for RBFOX2 mRNA level using quantitative reverse-transcription polymerase chain reaction. Intracellular localization of RBFOX2 protein was investigated using immunofluorescence microscopy. Normal and calcific tendon cDNAs were used to clone RBFOX2. Sequencing analysis identified coding sequences of the RBFOX2 isoform. RESULTS: The intracellular localization of RBFOX2 protein differed with disease status, with RBFOX2 localized in the cytoplasm in calcific tendons and the nucleus in normal tendons. Analysis of the RBFOX2 protein-coding sequence showed that exon 10, responsible for nuclear localization, was absent in calcific tendons. Splicing of RBFOX2 target genes CHD2 and MBNL1 was significantly affected by cytoplasmic localization of RBFOX2 in calcific tendons. DISCUSSION: Given the function of RBFOX2 as a splicing regulator in the nucleus, cytoplasmic localization of RBFOX2 protein in calcific tendons may have affected overall splicing events and altered gene expression. These results provide insights for comprehension of CT pathogenesis.

Immediate arthroscopy following ORIF for tibial plateau fractures provide early diagnosis and treatment of the combined intra-articular pathologies.

PURPOSE: To evaluate the effectiveness of immediate arthroscopy and clinical outcomes following open reduction and internal fixation (ORIF) of tibial plateau fractures. METHODS: Sixty patients (36 men and 24 women, median age 56 (20-78) years) were divided into Group I (ORIF only: 26 patients, median age 58 (25-78) years) or Group II (ORIF with immediate arthroscopy: 34 patients, median age 55 (20-75) years) in tibial plateau fractures (Schatzker Type II-VI fractures). In the first part of this study, ORIF only was performed without arthroscopic treatment. In the second part, ORIF with immediate arthroscopic examination and treatment was performed. Clinical outcomes, utilizing range of motion (ROM), International Knee Documentation Committee (IKDC) score and hospital for special knee score (HSS) were assessed. RESULTS: At the final follow-up, HSS score was 81 ± 11 points in Group I and 83 ± 9 points in Group II. The IKDC score was 85 ± 8 points in Group I and 86 ± 6 points in Group II. In Group II, concomitant intra-articular lesions in 10 patients (29%) were found and treated simultaneously. However, there were no significant differences in clinical scores or ROM between the two groups. CONCLUSION: Immediate arthroscopy following ORIF for tibial plateau fracture is an effective procedure that provides accurate information for fracture reduction, leading to immediate treatment of concomitant intra-articular lesions without complications. LEVEL OF EVIDENCE: III.

Delayed Lateral Row Anchor Failure in Suture Bridge Rotator Cuff Repair: A Report of 3 Cases.

Compared to single row repair, use of lateral row anchors in suture bridge rotator cuff repair enhances repair strength and increases footprint contact area. If a lateral knotless anchor (push-in design) is inserted into osteoporotic bone, pull-out of the lateral row anchor can developed. However, failures of lateral row anchors have been reported at several months after surgery. In our cases, even though complete cuff healing occurred, delayed pull-out of the lateral row anchor in the suture bridge repair occurred. In comparison to a conventional medial anchor, further biomechanical evaluation of the pull-out force, design, and insertion angle of the lateral anchor is needed in future studies. We report three cases with delayed pull-out of lateral row anchor in suture bridge rotator cuff repair with a literature review.

A Metabolomic Approach to Understanding the Metabolic Link between Obesity and Diabetes.

Obesity and diabetes arise from an intricate interplay between both genetic and environmental factors. It is well recognized that obesity plays an important role in the development of insulin resistance and diabetes. Yet, the exact mechanism of the connection between obesity and diabetes is still not completely understood. Metabolomics is an analytical approach that aims to detect and quantify small metabolites. Recently, there has been an increased interest in the application of metabolomics to the identification of disease biomarkers, with a number of well-known biomarkers identified. Metabolomics is a potent approach to unravel the intricate relationships between metabolism, obesity and progression to diabetes and, at the same time, has potential as a clinical tool for risk evaluation and monitoring of disease. Moreover, metabolomics applications have revealed alterations in the levels of metabolites related to obesity-associated diabetes. This review focuses on the part that metabolomics has played in elucidating the roles of metabolites in the regulation of systemic metabolism relevant to obesity and diabetes. It also explains the possible metabolic relation and association between the two diseases. The metabolites with altered profiles in individual disorders and those that are specifically and similarly altered in both disorders are classified, categorized and summarized.

Use of scopoletin to inhibit the production of inflammatory cytokines through inhibition of the IkappaB/NF-kappaB signal cascade in the human mast cell line HMC-1.

Scopoletin (6-methoxy-7-hydroxycoumarin) is a coumarin compound and a pharmacologically active agent that has been isolated from several plant species. However, as yet there is no clear explanation of how scopoletin affects the production of inflammatory cytokine. We therefore used cells from the human mast cell line (HMC-1) to investigate this effect. Scopoletin significantly and dose-dependently inhibits the way in which phorbol 12-myristate 13-acetate (PMA) plus A23187 induces the production of inflammatory cytokines such as tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-8 (P<0.05). The maximal rates at which scopoletin (0.2 mM) inhibited the production of TNF-alpha, IL-6, and IL-8 were 41.6%+/-4.2%, 71.9%+/-2.5%, and 43.0%+/-5.7%, respectively. In activated HMC-1 cells, the expression level of nuclear factor (NF)-kappaB/Rel A protein was increased in the nucleus whereas the level of NF-kappaB/Rel A in nucleus was decreased by treatment with scopoletin. Scopoletin decreased PMA plus A23187-induced luciferase activity. Scopoletin also inhibits IkappaBalpha phosphorylation and degradation in cytoplasm. These results indicate that scopoletin has a potential regulatory effect on inflammatory reactions that are mediated by mast cells.