bluuebunny/crossref_metadata_2025 huggingface dataset:
1540-P: LRP1 in GABAergic Neurons Is a Key Link between Obesity and Memory Function
Authors: KIM YOUNG-BUM , RODRIGUES KELLEN , CHAN KIM SEUNG , CAMPOLIM CLARA , YOUNG JENNIE , YANG WON-MO
Published: June 2024

Low-density lipoprotein receptor-related protein-1 (LRP1) regulates energy homeostasis, and its dysfunction has been linked to cognitive decline, dementia, and Alzheimer’s disease. However, the impact of LRP1 in inhibitory neurons on memory function and cognition in obesity remains unexplored. Mice lacking LRP1 in GABAergic neurons (Vgat-Cre; LRP1loxP/loxP) are subjected to conduct behavioral tests of locomotor activity and motor coordination, short/long-term and spatial memory, and fear learning/memory. We evaluated the relationships between behavior outcomes and metabolic risk factors. Deletion of LRP1 in GABAergic neurons caused a significant impairment in memory function. In the spatial Y-maze test, Vgat-Cre; LRP1loxP/loxP mice exhibited decreased travel distance and duration in the novel arm compared with controls (LRP1loxP/loxP mice). In addition, GABAergic neuron-specific LRP1-deficient mice had a diminished capacity for performing learning and memory tasks during the water T-maze test. Moreover, reduced freezing time was observed in the contextual and cued fear conditioning tests, accompanied by increased neuronal necrosis and neuroinflammation in the hippocampus. Importantly, these memory deficits correlate negatively with metabolic risk parameters, including body weight, serum leptin, insulin, and apolipoprotein J. Our findings demonstrate that LRP1 from GABAergic neurons is important in maintaining normal memory function. Metabolically, obesity caused by LRP1 deletion in GABAergic neurons negatively regulates memory and cognitive function. Thus, LRP1 in GABAergic neurons emerges as a key link between obesity and memory function, emphasizing its potential significance in maintaining neural system integrity associated with metabolism.

Disclosure

Y. Kim: None. K. Rodrigues: None. S. Kim: None. C. Campolim: None. J. Young: None. W. Yang: None.

Funding

National Institutes of Health (R01DK106076 and R01AG080842)
1543-P: Feeding Regulatory Mechanism of Leptin Action in the Nucleus of the Solitary Tract and Area Postrema
Authors: BRUCE KYLA , ZHANG SONG-YANG , KUAH RACHEL , LI ROSA J.W. , LAM TONY K.
Published: June 2024

Obesity is associated with brain leptin resistance. Leptin action in the hypothalamus activates STAT3 to lower feeding and weight, but whether extrahypothalamic regions such as the nucleus of the solitary tract (NTS) and area postrema (AP) mediate leptin action and resistance remain elusive. Here, we conducted refeeding and signaling studies to determine the impact of leptin infusion into the NTS or AP of male rats on chow or 3-day HF diet. Leptin vs vehicle infusion into the NTS (0.5μg; 0.2μL for 5min) increased STAT3 phosphorylation [lep: 1.7±0.1 vs. veh: 1.0±0.1 pSTAT3/total STAT3 intensity; p<0.01; n=6,6] and lowered food intake [lep: 77±5 vs. veh: 100±4 kCal; p<0.01; n=11,10] within 24h of refeeding in chow rats, while leptin infusion into the AP failed to lower feeding [lep: 93±6 vs. veh: 92±5 kCal; n=10,6]. When leptin was co-infused with STAT3 cell-permeable peptide (9.8ng dose that blocks the enzyme's phosphorylation) into the NTS, leptin failed to exert effects on STAT3 [lep + inhibitor: 0.9±0.1 vs. lep: 1.7±0.1 pSTAT3/total STAT3 intensity; p<0.01; n=5,6] and food intake [lep + inhibitor: 96±3 vs. lep: 77±5 kCal; p<0.05; n=13,11]. In 3d HF-induced hyperphagic [HF: 275±2 vs chow: 225±7 kCal; p<0.05; n=16,34] rats, NTS leptin vs vehicle failed to activate STAT3 [lep: 0.8±0.2 vs veh: 1±0.3 pSTAT3/total STAT3 intensity; n=5,5] or inhibit feeding [lep: 110±4 vs. veh: 113±3 kCal; n=10,6] in association with a reduction in leptin receptor expression in the NTS [HF: 0.68±0.02 vs. chow: 1±0.08; p<0.01; n=7,10]. In summary, we discover that 1) leptin administration into the NTS, but not the AP, lowers feeding and the NTS effect is depended on STAT3 activation, 2) 3d of HF feeding induces leptin resistance in the NTS to activate STAT3 and lower feeding in parallel to a reduction in leptin receptor expression. Thus, we unveil NTS as a site for leptin-STAT3 signaling to lower feeding and HF-induced leptin resistance. In contrast, AP appears do not sense leptin to regulate feeding.

Disclosure

K. Bruce: None. S. Zhang: None. R. Kuah: None. R.J.W. Li: None. T.K. Lam: None.

Funding

CIHR PJT-189957
1544-P: Fructooligosacharides Supplementation Prevents Western Diet–Induced Hypothalamic Inflammation
Authors: MUNOZ MARCOS D. , YANG XIAOHUA , LUO PEI , TORRES IRIZARRY VALERIA C. , CARRILLO-SÁENZ LESLIE , PRIYADARSHINI MEDHA , LAYDEN BRIAN T. , XU PINGWEN
Published: June 2024

Obesity is linked to 30-53% of new diabetes cases in the U.S. yearly. This project investigates how supplementation with fructo-oligosaccharides (FOS), a widely used dietary fiber, protects against diet induced obesity (DIO) by decreasing bodyweight (BW) and fat accumulation. The beneficial effects of FOS are largely attributed to the production of Short-Chain Fatty Acids (SCFA) through gut microbiota-mediated fermentation. Our findings indicate that supplementing a Western diet (WD) with 10% FOS for 8 weeks increases SCFA levels in the bloodstream, prevents weight gain in mice, and reduces markers of hypothalamic inflammation and microglial activation. Building on these findings, we further investigated the short-term effects of 7-day FOS supplementation. We found that adding 10% FOS to the WD completely prevented body weight gain and fat mass accumulation, while also lowering fasting blood glucose levels. Notably, mice treated with FOS maintained circadian oscillation of fuel utilization, as observed through the respiratory exchange ratio (RER), in contrast to WD-treated mice whose RER oscillation was disrupted within the first day of the diet. These protective effects were accompanied by reduced expression of hypothalamic interleukin-6 and iNOS after just 1 day of diet treatment. Taken together, our results suggest that early suppression of hypothalamic inflammation may be responsible for the beneficial effects of FOS supplementation. Understanding the mechanisms underlying the protective effects of FOS is a crucial step in developing new dietary strategies and therapeutic targets to address the obesity epidemic and its associated comorbidities.

Disclosure

M.D. Munoz: None. X. Yang: None. P. Luo: None. V.C. Torres Irizarry: None. L. Carrillo-Sáenz: None. M. Priyadarshini: None. B.T. Layden: None. P. Xu: None.

Funding

National Institutes of Health (R01DK123098)
1547-P: SGLT Inhibitor Canagliflozin Modulates Hypothalamic Function with Aging
Authors: SADAGURSKI MARIANNA , JAYARATHNE HASHAN M.
Published: June 2024

The hypothalamus, a critical brain region intricately involved in metabolic regulation and homeostasis, undergoes substantial changes with aging, influencing age-related metabolic alterations. Sodium-glucose co-transporter 2 inhibitors (SGLT2i), such as Canagliflozin, are anti-diabetic agents that enhance glucose elimination, resulting in a net calorie loss. Previously, we demonstrated Canagliflozin's ability to extend the median lifespan of genetically heterogeneous UM-HET3 male mice and improve hypothalamic insulin responsiveness in aging males. Here, we sought to evaluate Canagliflozin's impact on hypothalamic metabolic function during aging. Energy homeostasis was measured in 12- and 26-month-old UM-HET3 mice treated with Canagliflozin from 7 months of age. Canagliflozin-fed mice exhibited increased food consumption and water intake. Notably, 12-month-old Canagliflozin-fed males, but not females, displayed a significant increase in energy expenditure compared to controls. By 26 months, both male and female Canagliflozin-treated mice exhibited elevated metabolic rates, indicating sustained effects. This shift in energy expenditure correlated with alterations in hypothalamic orexigenic and anorexigenic projections in aged mice. Subsequently, bulk RNA-seq analysis of the hypothalamus from Canagliflozin-treated mice at 12 and 26 months revealed sex-specific transcriptional signatures, impacting glycolysis, oxidative phosphorylation, and insulin receptor signaling in aged mice leading to alterations in metabolic pathways. Our findings suggest that Canagliflozin's metabolic and health span effects in genetically diverse mice may be linked to its central effects, presenting it as a promising geroprotective intervention with implications for age-related metabolic changes.

Disclosure

M. Sadagurski: None. H.M. Jayarathne: None.

Funding

National Institutes of Health (RF1AG078170NIH, R01ES033171)
1551-P: Impaired ß-Cell Responsiveness to Glucagon in Type 2 Diabetes Mellitus Assessed by a New C-Peptide Minimal Model
Authors: BOSCOLO FEDERICA , FAGGIONATO EDOARDO , VELLA ADRIAN , DALLA MAN CHIARA
Published: June 2024

Glucagon (Gn) stimulates insulin secretion. However, tools to quantify β-cell responsivity to Gn are lacking. We therefore developed a novel method that provides quantitative indexes of β-cell function in response to a Gn bolus in people without (ND) and with type 2 diabetes (T2D).Eight ND individuals (3M, age = 55±9 yrs, BMI = 32±4 kg/m2) and 6 with T2D (1M, age = 59±5 yrs, BMI = 35±6 kg/m2) underwent a 210-min hyperglycemic clamp (~9 mmol/L). After 180 min, a 1 mg bolus of Gn was administered over 1 min and plasma Gn and C-peptide concentrations were frequently measured over 30 min. After testing a battery of mathematical models describing Gn-stimulated C-peptide secretion (CS), we selected one that assumes that CS is made up of two components, one proportional to the above basal Gn, through parameter Γs (static), and one proportional to Gn rate of change, through parameter Γd (dynamic responsivity to Gn). An index of total β-cell responsivity to Gn (Γtot) can also be calculated from Γs andΓd.Model estimated Γs, Γd, and Γtot were significantly lower in T2D compared to ND (p<0.05), as shown in Fig. 1.

In conclusion, our findings reveal notable differences in both static, dynamic and total insulin secretory response between individuals with and without T2D, demonstrating that decreased β-cell responsiveness in T2D includes impaired responses to Gn as well as glucose.

Disclosure

F. Boscolo: None. E. Faggionato: None. A. Vella: Research Support; Novo Nordisk. Consultant; Hanmi Pharm. Co., Ltd., Crinetics Pharmaceuticals, Inc. Advisory Panel; Rezolute, Inc. C. Dalla Man: None.
1554-P: Insights Into Insulin Synthesis and Turnover Using 13C515N-Valine to Label C-Peptide in Humans
Authors: EGAN AOIFE M. , DYER ROY B. , LAURENTI MARCELLO C. , DALLA MAN CHIARA , VELLA ADRIAN
Published: June 2024

The insulin secretory response to an intravenous, as opposed to an oral, glucose challenge is characterized by 2 phases the first of which is thought to represent preformed insulin whereas the second represents provisioning from newly synthesized insulin or newly mobilized insulin granules. The first phase is absent in people with type 2 diabetes (T2DM). Whether this reflects defects in insulin synthesis, insulin storage or both remains to be ascertained. To address these questions, we studied 10 subjects with and without T2DM. Subjects were admitted to the Clinical Research Unit at 1700 and consumed a standard meal and fasted overnight. At 2100 5g of arginine was given. An infusion of 13C515N-Valine (Prime 1.5mg/Kg body mass and then 1.5mg/Kg/hr) commenced and continued till 0600 the following morning. At 0800 subjects underwent a 75g OGTT followed by 5g I.V. arginine at 1200. Blood was sampled at frequent intervals to measure hormone and substrate concentrations as well as C-peptide labelled with 13C515N-Valine. This was purified via repetitive cycling through an affinity resin conjugated with C-peptide antibody in a custom manufactured mass spectrometric immunoassay pipette tip. Peptides were detected in positive mode with a high resolution, Q Exactive Plus Mass Spectrometer optimized by infusion of synthetic peptides. During the overnight fast, average C-peptide enrichment in the peripheral circulation was higher in T2DM (0.3 ± 0.3 vs. 2.3 ± 0.2 %, nondiabetic vs T2DM respectively, p<0.01). After oral glucose, C-peptide enrichment was lower in T2DM (6.7 ± 0.5 vs. 5.0 ± 0.4 %, p=0.05). In contrast after I.V. arginine challenge, enrichment did not differ (5.6 ± 1.2 vs. 3.5 ± 0.4 %, p=0.08). These data imply that insulin turnover is higher in T2DM: a greater proportion of newly synthesized insulin is secreted during fasting. This may explain the depletion of 1st phase insulin and other easily releasable granule pools in T2DM.

Disclosure

A.M. Egan: None. R.B. Dyer: None. M.C. Laurenti: Employee; Angelini Pharma. C. Dalla Man: None. A. Vella: Research Support; Novo Nordisk. Consultant; Hanmi Pharm. Co., Ltd., Crinetics Pharmaceuticals, Inc. Advisory Panel; Rezolute, Inc.

Funding

National Institutes of Health (DK078646)
156-OR: Genome-Wide CRISPR Screen Identified That Alternative Splicing Regulates in Insulin Secretion
Authors: WEI SIYING , APARECIDA DA SILVA PEREIRA JESSICA , LEE-PAPASTAVROS ALEXANDER , YI PENG
Published: June 2024

Pancreatic β cell functionality, especially glucose-stimulated insulin secretion (GSIS), is pivotal in diabetes pathophysiology. The impairment of GSIS is not only evident in Type 2 diabetes (T2D) but also during the early stages of Type 1 diabetes (T1D). In addition, enhancing GSIS also holds significant promise for improving the functionality of stem cell-derived β cells (SC- β cells). Addressing this limitation offers new opportunities in cell-based diabetes therapy. To uncover the intricate regulatory mechanisms of insulin secretion, we developed a unique reporter cell line carrying NPY-pHluorin transgene to visualize insulin exocytosis at single-cell level and conducted a genome-wide CRISPR screen using flow cytometry system in NIT-1 cells. Notably, our study identifies alternative splicing (AS) as a pivotal factor in insulin secretion regulation. Employing genetic Sf3b1 knockout - a key component in AS machinery in NIT-1 cells, results in a surprising outcome: increased insulin secretion and improved GSIS. Consistently, using chemical inhibitors such as FR901464 to suppress AS activities, we observed the same phenotype in β cell lines and pancreatic islets. To unravel the underlying mechanisms, we performed comprehensive RNA-seq and alternative splicing-seq analyses in Sf3b1 knockout cell lines and cells treated with AS inhibitor, revealing alterations in genes related to the insulin production machinery and synaptic/exocytic vesicle traffic pathways. Our findings emphasize the significant role of AS in modulating β cell function. This research bridges critical gaps in our current understanding and sets the stage for innovative approaches to address the intricate challenges posed by diabetes.

Disclosure

S. Wei: None. J. da Silva Pereira: None. A. Lee-Papastavros: None. P. Yi: None.
1567-P: Integrating Bioinformatics and Machine Learning to Identify Lactylation-Related Diagnostic Biomarkers and Characterize Immune Cell Infiltration in Nonalcoholic Fatty Liver Disease
Authors: ZHANG JUNYI , WU XUEYAN
Published: June 2024

Background: This study sought to explore potential lactylation-related targets for non-alcoholic fatty liver disease (NAFLD) and examine the role of immune cell infiltration in the disease's progression.

Methods: Gene expression datasets were sourced from the Gene Expression Omnibus collection, and genomic enrichment analysis was conducted utilizing differentially expressed genes (DEGs). Overlapping lactylation and NAFLD DEGs were determined using a Venn diagram, and a protein-protein interaction (PPI) network was developed. To ascertain immune patterns, the ConsensusClusterPlus package in R was employed. Key genes were identified using three machine-learning algorithms: random forest, support vector machine-recursive feature elimination, and least absolute shrinkage and selection operator. The infiltration of immune cells was assessed by the GSVA package. The upstream transcriptional factors (TFs) and microRNAs (miRNAs) were predicted using the Regnetwork database and NetworkAnalyst.

Results: The intersection of key genes and DEGs yielded 34 genes associated with lactylation and NAFLD, comprising 22 upregulated and 12 downregulated genes. Eight genes, namely FABP5, SIRT1, TERF2, ADNP, BTF3, CDC5L, DDX17, and MNDA, were identified as diagnostic indicators with relatively high diagnostic specificity for NAFLD. The interaction among key genes were shown by PPI network. A spectrum of immune cells was found to potentially contribute to NAFLD development. The association between key genes and immune infiltration were revealed by the correlation analysis. Also, we predicted the upstream TFs and miRNAs of hub genes.

Conclusions: Our research identified 8 key genes related to lactylation and analyzed the immune cell infiltration characteristics in NAFLD patients, thereby offering additional insights into the potential mechanisms influencing NAFLD prognosis.

Disclosure

J. Zhang: None. X. Wu: None.
1568-P: Liver-specific Expression of Activated G Protein Gsα Leads to Hyperglycemia with Reduced Pancreatic α-Cell Mass and Function
Authors: GUPTA MEENAKSHI , SUN HUI , LAYTON DANIEL , ZHANG JIANHUA , LEE JI-EUN , CHEN MIN , WEINSTEIN LEE
Published: June 2024

Gsα couples hormones to stimulate intracellular cAMP generation. We previously showed that mice with liver-specific Gsα deficiency (LGsKO) have improved glucose tolerance and enlarged pancreatic islets. Here we generated mice with liver-specific expression of an activated Gsα by breeding mice containing a Lox-STOP-Lox-GsαR201C transgene with albumin-Cre mice (LGsR201C). Compared to controls, male LGsR201C mice had normal survival but increased hepatic cAMP levels and gluconeogenetic gene expression, confirming the activation of liver Gsα/cAMP signaling in these mice. Consequently, LGsR201C mice showed elevated blood glucose, enhanced glucose response to pyruvate administration and hyperglycemic response to glucagon. LGsR201C mice had glucose intolerance and lacked insulin response to i.p. glucose injection, although having normal pancreatic β-cell mass and intact glucose-stimulated insulin secretion from isolated islets ex vivo . On the other hand, LGsR201C mice exhibited normal insulin sensitivity but lacked counterregulatory action of glucagon responding to lower blood glucose levels in vivo and had less pancreatic α-cell mass, reduced islet glucagon content and impaired arginine-stimulated glucagon secretion from isolated islets. Notably, LGsR201C mice showed significantly lower serum levels in all tested 17 amino acids. RNAseq analysis revealed that an array of genes was oppositely regulated in LGsR201C vs. LGsKO mice, including the genes of several amino acid transporters upregulated in the liver and islets of LGsR201C mice and downregulated in the liver of LGsKO mice. Thus, our data indicate that activation of Gsα/cAMP signaling in the liver triggers a liver-islet circuit that plays divergent roles in the control of pancreatic islet α- and β-cell biogenesis and functions.

Disclosure

M. Gupta: None. H. Sun: None. D. Layton: None. J. Zhang: None. J. Lee: None. M. Chen: None. L. Weinstein: None.

Funding

NIDDK Intramural Research Program
1576-P: Evaluation of Hepatic Steatosis and Adiposity in People with Type 1 Diabetes and Obesity
Authors: TRIKUDANATHAN SUBBULAXMI , AMIN KATHAN , BAMBHA KIRAN , MELHORN SUSAN J. , SEKHON MANDEEP , NAVARRO SANDI , SCHUR ELLEN A. , HIRSCH IRL B.
Published: June 2024

Introduction: Prevalence of obesity in T1D is rising, contributing to the development of hepatic steatosis (HS) with the associated risk for metabolic dysfunction associated steatotic liver disease (MASLD). While T2 D and MASLD are reliably linked, prevalence of MASLD in T1D varies by the imaging modality employed. We therefore estimated the prevalence of HS and examined the key determinants of HS in people with T1D with BMI≥ 30 kg/m2.

Methods: In a cross-sectional study, 42 individuals with T1D on a continuous glucose monitor (CGM) with duration of diabetes >10 years and BMI ≥ 30 kg/m2 were studied. MRI-Proton Density Fat Fraction (PDFF) was used to estimate HS. Visceral (VAT) and subcutaneous adiposity (SAT) measures were obtained. Linear regression models were used to evaluate the association between HS, VAT, and SAT, with outcomes of glycemic control and insulin resistance.

Results: Participants had a mean (SD) age of 47.8 (13.3) y, 59.5% were men with BMI -32.9 (3.1) kg/m2 and 30.6% had a family history of T2D. Mean A1c was 7.1 (0.77), TIR of 65% (12.5), and TAR of 32.8(12.2) and GMI of 7.2(0.45). Prevalence of HS was 9.5% in the study sample. No statistically significant associations were found between any of the parameters evaluated and HS although a trend was observed for total daily dose of insulin in units (TDDI) (p=0.06) with higher insulin doses positively associated HS. Higher A1c (p =0.004), lower TIR (p=0.02), higher TAR (p =0.009) and higher TDDI (p<0.0001) were associated with higher VAT in the unadjusted models and remained significant for TDDI and VAT (p=0.01) when adjusted for age, sex, and WC.

Conclusion: Our study, which incorporates CGM metrics in a distinctive subpopulation of T1D with obesity, suggests a lower prevalence rate of HS when a sensitive imaging modality such as MRI-PDFF was used. Lack of portal hyperinsulinemia may contribute to lower HS in T1D. The results suggest that markers of insulin resistance and glycemic exposure correlate with VAT but not HS in adults with T1D and obesity.

Disclosure

S. Trikudanathan: Research Support; Insulet Corporation. K. Amin: None. K. Bambha: None. S.J. Melhorn: None. M. Sekhon: None. S. Navarro: None. E.A. Schur: Consultant; Amgen Inc. I.B. Hirsch: Advisory Panel; Abbott. Research Support; Dexcom, Inc. Advisory Panel; Roche Diabetes Care. Research Support; MannKind Corporation, Tandem Diabetes Care, Inc. Advisory Panel; embecta. Research Support; Tandem Diabetes Care, Inc. Advisory Panel; Vertex Pharmaceuticals Incorporated.

Funding

UWMDI Pilot & Feasibility Award
1579-P: Altered Serum Ceramides as Part of Dyslipidemia in Adolescents with Metabolic Dysfunction–Associated Steatotic Liver Disease
Authors: SHORT KEVIN R. , FRIEDMAN JACOB E. , WANG LIPING , SUMMERS SCOTT A.
Published: June 2024

Introduction & Objective: Metabolic dysfunction-associated steatotic liver disease (MASLD) in adults is associated with altered plasma ceramides, and may serve as liver metabolic biomarkers. However, less is known about whether similar changes are present in adolescents. We performed a lipidomic analysis to test whether adolescents with MASLD have altered plasma ceramides.

Methods: Plasma was acquired from boys and girls, 11-20y, classified as, 1) obese with biopsy-confirmed MASLD (n=44); 2) obese without MASLD (Ob, n=28); and 3) normal weight without MASLD (NW, n=29). MASLD patients had moderate to severe liver steatosis and 91% had ≥ stage 1 liver fibrosis.

Results: MASLD and Ob had similar body fat (45±7%), and were higher (p<0.01) than NW (31±6%). MASLD had higher triglycerides (p<0.01) than Ob and NW (199±110, 107±48, & 86±40 mg/dl, respectively) and had lower HDL-C than Ob or NW (40±10, 47±12, & 59±12 mg/dl respectively). Of the 6 major ceramide species measured, d18:1/24:0 was ~18% lower (p<0.001) in MASLD and Ob versus NW, while the others did not differ among groups. Of the 6 major dihydroceramide (dhCer) species measured, 4 were 32-185% higher (p<0.001) in the MASLD group versus the Ob and NW groups. The dhCer, but not the ceramide species, were positively correlated with trunk fat (r=0.34 to 0.55), but not with insulin resistance. dhCer 18:0/18:0, the most altered species, was positively correlated with liver steatosis (r=0.36), and along with 3 other dhCer species was positively correlated with MASLD Activity Score (r=0.35 to 0.41). There were no group differences in hexosylceramides, sphingosine, sphinganine, or the major sphingomyelins or dihydrosphingomyelins.

Conclusions: Similar to adults, adolescents with MASLD have altered plasma ceramides and dyslipidemia compared to NW and Ob peers without MASLD. The patterns observed suggest that ceramide flux may be elevated and ceramide-producing cells may be damaged in pediatric MASLD patients.

Disclosure

K.R. Short: None. J.E. Friedman: None. L. Wang: Stock/Shareholder; Centaurus Therapeutics. S.A. Summers: Board Member; Centaurus Therapeutics.

Funding

NIH R01DK129656 and U54GM104938, Presbyterian Health Foundation, Harold Hamm Diabetes Center, and Children's Health Foundation
1590-P: Lian-He-Xiao-Zhi Ointment Improves Lipid Metabolism and Inflammation in MAFLD by Regulating PPARα
Authors: NIE LIJUAN , YANG XINYI , ZHOU XIQIAO
Published: June 2024

Background and Aims: Lian-He-Xiao-Zhi ointment (LHXZO) is an empirical formula, which has a significant therapeutic effect on metabolic associated fatty liver disease (MAFLD) in clinical practice. However, the precise underlying mechanisms remain unclear. The objective of this study is to elucidate the mechanisms through which the LHXZO ameliorates MAFLD, utilizing an animal model.

Methods: C57BL/6J mice (6-8 weeks old, male) were subjected to a high-fat diet (HFD) to induce a MAFLD model. Mice received either LHXZO decoction or control solution intragastrically daily for twelve weeks. Histological and serum analyses were used to assess the liver injury and disorders of glucolipid metabolism. Total RNAseq and pathway analyses were used to identify the potential signaling pathways modulated by LHXZO in the liver. The expression levels of key genes involved in regulating lipid metabolism and inflammation were validated by qRT-PCR or Western blot analysis. The impact on gut microbiota was assessed through 16S rRNA gene sequencing.

Results: LHXZO treatment significantly improved glucolipid metabolic disorders, liver injury and liver inflammation, evidenced by decreased serum levels of fasting blood glucose, free fatty acid, TG, AST, ALT, LPS and IL-1β. HE and Oil Red O staining also showed LHXZO treatment reduced lipid deposition in the liver. Meanwhile, LHXZO significantly reduced the body weight of MAFLD mice. LHXZO mitigated inflammation by downregulating TLR4, NF-KB, NLRP3, IL-1β and IL-18. In addition, LHXZO significantly upregulated the level of PPARa. Furthermore, LHXZO significantly modulated gut microbiota.

Conclusions: LHXZO effectively improve glucolipid metabolism, liver injury and inflammation in MAFLD, indicating its potential as a therapeutic agent for MAFLD. LHXZO's improvement of liver inflammation in MAFLD mice may be achieved by regulating PPARα or gut microbiota.

Keywords: MAFLD; PPARα; inflammation; gut microbiota

Disclosure

L. Nie: None. X. Yang: None. X. Zhou: None.

Funding

the Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD, 035062005002-14), the special research project of Jiangsu Administration of traditional Chinese medicine (zt202105), and the Project Funded by Jiangsu Province Hospital of Chinese Medicine (kkzx01).
1593-P: Symptoms and Correlations in Newly Diagnosed Type 2 Diabetes Mellitus (T2DM) with Nonalcoholic Fatty Liver Disease (NAFLD)
Authors: MUKHERJEE SUPRIYO
Published: June 2024

Introduction & Objective: The study conducted in North Bihar, India, aimed to explore the prevalence of NAFLD among individuals newly diagnosed with T2DM. It investigated the potential correlations between patient symptoms and various factors such as educational background, socioeconomic class, family history of diabetes mellitus (DM), and gender.

Methods: A cross-sectional study, from December 2022 to May 2023, involved 148 newly diagnosed T2DM patients aged 18+. Exclusion criteria comprised alcohol use, pregnancy, traditional medicine, liver-related issues, and malignancy. Sociodemographic, symptom, and clinical data were collected via detailed tools, analysed using Stata 13.0 and IBM SPSS 28 for correlations and differences.

Results: Out of the 148 individuals, 73.7% presented with liver disease (NAFLD), emphasizing the high prevalence of liver involvement among newly diagnosed T2DM cases. Common symptoms included excessive thirst, polyuria, weight loss, and nocturia. Intriguingly, correlations emerged between symptom presentation and education level, socioeconomic class, family history of DM, and gender. Individuals with atypical symptoms tended to have higher education and a family history of DM. Lower-middle-class individuals exhibited a higher percentage of atypical symptoms, suggesting potential disparities in symptom recognition and healthcare-seeking behavior.

Conclusion: This study highlighted high NAFLD rates in newly diagnosed T2DM cases in North Bihar. Correlations with education, socioeconomic factors, and gender emphasize tailored interventions for early detection and management.

Disclosure

S. Mukherjee: None.
160-OR: PI3K Inhibition with Single-Dose Alpelisib as a Model of Insulin Resistance in Healthy Adults
Authors: COOK JOSHUA R. , BEDEIR NUR , SONE ZACHARY D. , WATTACHERIL JULIA , GINSBERG HENRY N. , LAFERRÈRE BLANDINE
Published: June 2024

Introduction & Objective: Understanding the pathophysiology of insulin resistance (IR) is essential to treating type 2 diabetes, but mechanistic studies of IR in human subjects are often complicated by ancestry and phenotypic heterogeneity, variable chronicity, and difficulty in causal inference. We therefore seek to establish a safe, reliable, acute method for modeling IR in a translational setting. Our objective is to test a single dose of the phosphoinositide-3-kinase (PI3K) inhibitor alpelisib, versus placebo, for tolerability and effect on glucose and insulin metabolism.

Methods: This single-center, double-blind phase I clinical trial randomized healthy adults, ages 18-65 y with body mass index 18.0-26.9 kg/m2 and HbA1C ≤ 5.6%, to take a single oral dose of alpelisib 300 mg (n = 5) or placebo (n = 6) at bedtime. The next morning, participants had blood drawn for fasting insulin and glucose levels­ (co-primary endpoints) followed by a 3-hour, 75-g oral glucose tolerance test (OGTT). Data are analyzed based on intention to treat using an independent two-sample t test.

Results: Fasting plasma glucose trended higher with alpelisib (mean ± SEM = 93 ± 6 mg/dL) versus placebo (84 ± 2 mg/dL) (p = 0.07). Mean fasting serum insulin increased nearly fivefold: 23 ± 6 μIU/mL for alpelisib versus 5 ± 1 μIU/mL for placebo (p < 0.01), resulting in Homeostasis Model Assessment of Insulin Resistance scores of 5.4 ± 1.4 vs. 1.1 ± 0.2 and Matsuda Index scores of 8.6 ± 2.2 vs. 3.0 ± 1.6, respectively. During OGTT, area under the curve (AUC) for insulin was greater with alpelisib (26.0 ± 7.5 mIU/mL × min) than placebo (5.8 ± 1.0 mIU/mL × min) (p < 0.02); glucose AUC trended higher with alpelisib. Single-dose alpelisib was well tolerated without serious adverse events.

Conclusion: A single dose of alpelisib produced metabolic derangements consistent with acute induction of IR, validating its use as a safe and effective tool for studying insulin action in humans.

Disclosure

J.R. Cook: None. N. Bedeir: None. Z.D. Sone: None. J. Wattacheril: Research Support; Intercept Pharmaceuticals, Inc. Advisory Panel; GlaxoSmithKline plc. Consultant; Alphasights. Research Support; AMRA, Galectin Therapeutics. H.N. Ginsberg: None. B. Laferrère: Consultant; Up-To-Date.
1600-P: Sunflower Oil HFD Feeding in Mice Leads to Gut Dysbiosis, Mild Steatohepatitis, and Impaired Glycemic Control
Authors: SINDHU SARDAR T.A.K. , BAHMAN FATEMAH , ZUBBAIR MALIK MOHAMMAD , AREFANIAN HOSSEIN , ALRASHED FATEMA , NIZAM RASHEEBA , KOCHUMON SHIHAB P. , THOMAS REEBY S. , ALBELOUSHI SHAIMA , HASAN AMAL , ABU-FARHA MOHAMED , AL MADHOUN ASHRAF , THANGAVEL ALPHONSE T. , ALMULLA FAHD , AHMAD RASHEED
Published: June 2024

Introduction & Objective: High-fat diets (HFDs) influence gut microbiota and promote obesity & metabolic dysregulation. Dietary sucrose is a co-player in modifying gut microbial ecosystem towards dysbiosis. Fish & sunflower dietary fats may be consumed with/without sucrose; however, gut bacterial changes from excessive intake of these fats, without the added effect of sucrose, remain unclear.

Methods: To address this objective, we compared effects of sucrose-free, sunflower high-fat diet (S-HFD) & fish (F)-HFD feeding (24 wks) on the gut microbiota, obesity, liver inflammation, and glycemic control in C57BL/6 mice.

Results: The 16S rRNA sequencing of fecal microbiome showed that gut bacterial communities were differentially expressed between two dietary groups. Firmicutes were highly-abundant in S-HFD (40%) compared to F-HFD (3%) mice, while Verrucomicrobia were abundant in F-HFD (26%) compared to S-HFD (˂1%) mice. Firmicutes/Bacteroidetes ratios in S-HFD & F-HFD were 1.32 & 2.45, respectively. MicrobiomeAnalyst & MIAOME analyses identified highly-abundant taxa in S-HFD mice, including Oscillospira, guilliermondii, Allobaculum, Yaniella, Ruminococcus, gnavus, Staphylococcus, Clostridium, Adlercreutzia, Aerococcaceae, Anaeroplasma, Mogibacteriaceae, Christensenellaceae, & RF39, having predicted associations with host genes of metabolic disorders/pathways linked to inflammation & glucose metabolism. Compared to F-HFD mice, the S-HFD mice showed slightly increased body weights (P = 0.06) but significant differences regarding fasting hyperglycemia, hepatic macrophage infiltration (F4/80 expression), macrovascular steatosis, lobular inflammation, & expression of genes associated with de novo lipogenesis (Acaca, Fasn, and Scd1) & monocyte chemotaxis (Ccl2).

Conclusion: Taken together, S-HFD feeding in mice leads to gut dysbiosis and the clinical manifestations of moderate liver inflammation, steatosis and impaired glycemic control.

Disclosure

S.T. Sindhu: None. F. Bahman: None. M. Malik: None. H. Arefanian: None. F. Alrashed: None. R. Nizam: None. S.P. Kochumon: None. R.S. Thomas: None. S. Albeloushi: None. A. Hasan: None. M. Abu-farha: None. A. Al Madhoun: None. A.T. Thangavel: None. F. Almulla: None. R. Ahmad: None.

Funding

Kuwait Foundation for the Advancement of Sciences (KFAS) (Grants #: RA 2010-003)
1604-P: Lrrc2 Preserves Mitochondrial ATP Production in Diabetic Muscle and Promotes Adaptations to Exercise
Authors: PENNIMAN CHRISTIE M. , BHARDWAJ GOURAV , ALLEN RYAN , LIRA VITOR A. , O'NEILL BRIAN T.
Published: June 2024

Uncontrolled diabetes impairs muscle strength and mitochondrial metabolism, leading to disabilities. FoxOs are key mediators of insulin signaling and we demonstrated that FoxOs mediate mitochondrial dysfunction in mice with streptozotocin (STZ) diabetes. Unfortunately, knockout of FoxOs in proliferative tissues causes cancer. We aimed to investigate FoxO targets restricted to skeletal muscle in hopes of improving strength in diabetic conditions without carcinogenic effects. RNA-seq from STZ mice with/without FoxO1/3/4 triple knockout showed LRRC2 (Leucine rich repeating containing 2) is a striated-muscle specific FoxO target that is transiently increased in diabetic muscle. The function of LRRC2 is unknown, but gene expression databases show LRRC2 is restricted to muscle and heart. Adenoviral overexpression of LRRC2 in C2C12 cells localized to mitochondria and showed increase mRNA levels of biogenesis marker PGC1α, and antioxidant genes SOD2 and Gpx3. To determine LRRC2’s effect on exercise tolerance and mitochondrial bioenergetics in vivo, we created muscle specific LRRC2 knockout (LKO) mice and rendered them diabetic with STZ. Diabetic LKO mice had the poorest performance on treadmill exercise to exhaustion, and female LKO mice showed impaired exercise training with free wheel running when compared to controls. Interestingly, soleus muscles from LKO mice were 17% larger than their controls, whereas extensor digitorum longus (EDL) were 25% smaller in the diabetic LKO mice, indicating divergent effects on oxidative vs. glycolytic muscle. Soleus permeabilized fibers showed 20% decreased ATP production in diabetic LKO compared to controls that was uncoupled from respiration with glutamate/malate. Mitochondria from quad/gastroc of LKO mice showed increased H2O2 production compared to controls.

In conclusion, Lrrc2 is a novel FoxO target that helps preserve mitochondrial function and exercise tolerance in a type 1 diabetes mouse model.

Disclosure

C.M. Penniman: None. G. Bhardwaj: None. R. Allen: None. V.A. Lira: None. B.T. O'Neill: None.

Funding

VA Merit Review Award Number (lO1 BXOO4468); Fraternal Order of Eagles Diabetes Research
1605-P: Skeletal Muscle Ubiquitin Protease Function Governs Mitochondrial Quality Control and Insulin Sensitivity in Type 2 Diabetes
Authors: AXELROD CHRISTOPHER L. , DANTAS WAGNER S. , HEINTZ ELIZABETH C. , ZUNICA ELIZABETH R.M. , DAVULURI GANGARAO , KIRWAN JOHN P.
Published: June 2024

Introduction: Skeletal muscle mitochondrial content declines in patients with type 2 diabetes (T2D), which is driven by prolonged hyperactivation of DRP1-mediated organelle fission. However, the mechanism underlying activation of fission and its relation to the regulation of glucose uptake in T2D remains largely unknown. The purpose of this study was to examine the influence of ubiquitin function on mitochondrial quality control and DRP1 hyperactivation in patients with T2D.

Methods: 58 sedentary adults (37±12 years) were enrolled into one of three groups based upon the following criteria: (1) healthy weight without T2D (HW); (2) obesity without T2D (Ob); or (3) obesity with T2D (T2D). Participants underwent a 3-day inpatient stay for measurements of body composition (DXA), aerobic capacity (VO2MAX), and insulin sensitivity (hyperinsulinemic euglycemic clamp). A skeletal muscle biopsy was obtained for determination of mitochondrial dynamics (Western blot) and isolation of primary human skeletal muscle (pHSkM) cells. pHSkM cells were transfected with FLAG-USP15/USP30 to induce overexpression. Glucose uptake was measured by incorporation of [3H]-2-deoxy-d-glucose. Comparisons were made by one-way ANOVA with contrasts.

Results: USP13, USP15, and USP30 expression were increased in T2D relative to Ov/Ob and HW (all P<0.05). USP15 and USP30 interaction with Parkin decreased in T2D relative to Ov/Ob and HW (all P<0.05). USP15 (R2=0.55, P<0.0001) and USP30 (R2=0.40, P<0.0001) expression negatively correlated with peripheral insulin sensitivity. USP15 and USP30 overexpression activated DRP1 while blocking ubiquitin-mediated Parkin activation.

Conclusions: Ubiquitin protease function is heightened in patients with T2D which impairs mitochondrial quality control, promotes excessive mitochondrial fission, depletes mitochondrial volume, and restricts glucose uptake.

Disclosure

C.L. Axelrod: None. W.S. Dantas: None. E.C. Heintz: None. E.R.M. Zunica: None. G. Davuluri: None. J.P. Kirwan: None.

Funding

National Institutes of Health (DK108089 and GM104940)
1606-P: Modelling Muscle Function Loss in Engineered Human Tissues
Authors: HØJFELDT MARTINA , BIRK JESPER , SEGERITZ-WALKO CHARIS , WOJTASZEWSKI JØRGEN F.P. , PEHMØLLER CHRISTIAN , KNUDSEN JONAS R.
Published: June 2024

Introduction and Objective: Loss of muscle function is detrimental to the human body and understanding its molecular underpinnings and identifying therapeutic avenues to modify these is of major importance. To enable this, we aimed to develop a translatable drug discovery model that circumvents limitations by species-driven biological differences and demonstrates high sensitivity to readouts related to muscle function.

Methods: A human primary myoblast-based engineered muscle tissue model was established, differentiated, and cultured for weeks. We characterized tissue maturity through contractility and gene and protein expression longitudinally and under various electrical stimulation regimens. Key physiological features such as force-frequency relationship, post-tetanic potentiation, and fatigue during exercise was assessed. Finally, functionality of the tissues following two different atrophy-inducing interventions was studied.

Results: Differentiated tissues maintained stable maximal force production for over 30 days. Maturity markers (muscle AMPK isoforms and mitochondrial, contractile and Ca2+ handling proteins) increased during a 23-day evaluation. Electrical stimulation also increased maturity, including improved maximal contractile force. The tissues exhibited expected physiological responses (force-frequency relationship, post tetanic potentiation and fatigue following exercise mimicking electrical stimulation). Activation of glucocorticoid (by dexamethasone) and activin (by Activin A) signaling pathways dose dependently caused a decline in maximal force development which was mitigated by antagonism. Intriguingly, the model demonstrated sensitivity to detect significant force changes with effect sizes below 15%.

Conclusion: Our human-based engineered muscle tissue model offers a robust platform with high sensitivity for elucidating mechanisms related to muscle function decline and therapeutic exploration.

Disclosure

M. Højfeldt: Employee; Novo Nordisk A/S. J. Birk: None. C. Segeritz-Walko: None. J.F.P. Wojtaszewski: Consultant; Pfizer Inc. Stock/Shareholder; Pfizer Inc., Novo Nordisk A/S. C. Pehmøller: Employee; Novo Nordisk A/S. J.R. Knudsen: Employee; Novo Nordisk A/S.
1608-P: MyoScreen—A High-Content–Based Discovery Platform to Model Key Metabolic Processes in Human Skeletal Muscle
Authors: MOROZZI GIULIO , ZABLOCKI ANIELA , MARTIN-TISSIER ANTOINE , TRAVARD LAURIANE , MASSERA CÉLINE , FLAENDER MÉLANIE , FARGIER GUILLAUME , LORINTIU OANA , VENTRE ERWANN , YOUNG JOANNE , AUTIER VALERIE , SELIG LUC
Published: June 2024

Introduction & Objective: Loss of muscle mass and function are now recognized as early and common abnormalities in a wide range of chronic diseases especially obesity, diabetes and cardiomyopathies. However, relevant in-vitro screening models mimicking human physiological processes in muscles are still lacking.

Method: Using MyoScreen, a high-throughput and robust skeletal muscle drug discovery platform derived from human primary myoblasts, we have developed a panel of predictive in-vitro image-based functional assays combined with newly developed image analysis scripts, that recapitulate key metabolic disease hallmarks such as autophagy, lipid accumulation, muscle atrophy and mitochondrial health. 

Results: Dose dependent accumulation of LC3b spots (puncta) (EC50= 20.77 μM for CQ, Chloroquine, EC50= 3.9 nM for BafA1, Bafilomycin) and area of vesicles positive for LC3b (EC50= 25.97 μM for CQ, EC50= 6.1 nM for BafA1) can be monitored in fixed starved (8h) cells following inhibition of autophagy flux. Co-labeling of LAMP1-LC3b (EC50= 23.17 μM for CQ, EC50= 2.7 nM for BafA1) follows the autophagic flux and autophagolysosome formation. Oversupply of FAs (fatty acids) increases mitochondrial fragmentation and the relative area of LDs (lipid droplets) in myotubes by 13-23%: this accumulation can be modulated by siRNAs treatment. MyoScreen demonstrated the ability to distinguish phenotypic properties related to stimulatory or inhibitory effects of hypertrophic (IGF-1 EC50=95.5 pM for fusion index) or atrophic (TNF-α IC50=15.9 pM for fusion index) signaling compounds. 

Conclusion: The MyoScreen platform brings new prospects for drug discovery in metabolic disorders facilitating a predictive research strategy for identifying and validating novel targets, understanding mechanism of action, and supporting preclinical development to select innovative drug candidates targeting key aspects of muscle metabolic function.

Disclosure

G. Morozzi: None. A. Zablocki: None. A. Martin-Tissier: None. L. Travard: None. C. Massera: None. M. Flaender: None. G. Fargier: None. O. Lorintiu: None. E. Ventre: Employee; CYTOO. J. Young: None. V. Autier: None. L. Selig: None.
1609-P: Whole-Body Metabolic Flexibility Is Discordant with Skeletal Muscle Metabolism
Authors: KRASSOVSKAIA POLINA , JEVTOVIC FILIP , CHORZEPA MARTA , JONES TERRY E. , TANNER CHUCK J. , ZHENG DONGHAI , PORIES WALTER J. , DOHM LYNIS , HOUMARD JOSEPH A. , BROSKEY NICHOLAS T.
Published: June 2024

Introduction & Objective: Metabolic inflexibility is the inability to appropriately switch substrate use in response to a metabolic challenge and is seen in individuals with metabolic disease. Metabolic inflexibility can be observed in seemingly healthy individuals, potentially predisposing them to metabolic disease. The goal of this study was to examine if whole-body metabolic inflexibility is reflected by differential substrate use at the skeletal muscle (SkM) level.

Methods: This study recruited young (27 ± 8 years) healthy individuals with overweight. Anthropometric measures, VO2max test, hyperinsulinemic-euglycemic clamp, and a vastus lateralis biopsy was performed. Based on difference in the respiratory quotient (ΔRQ) during the clamp, participants were median stratified to a “metabolically flexible” (FLEX) (ΔRQ>0.06; n=10) or “metabolically inflexible” (INFLEX) (ΔRQ<0.06; n=9) group. SkM biopsy samples were assessed for substrate oxidation with 14C-pyruvate and 14C-palmitate, mitochondrial respiration, and muscle fiber type. Data was analyzed with descriptive statistics, Mann-Whitney U-test, and Student’s t-test.

Results: FLEX had a lower BMI than INFLEX (26.4 ± 1.3 vs 28.7 ± 1.5 kg/m2; p=0.002) but had a similar percentage of body fat (p=0.3) or lean mass (p=0.2). Groups did not differ in age (p=0.06), VO2max (p=0.3), M value (p=0.1), or basal RQ (p=0.5). FLEX had lower palmitate oxidation than INFLEX (p=0.03) and trended towards a higher ratio of pyruvate to palmitate oxidation (p=0.06). Incomplete palmitate oxidation was also higher in FLEX (p=0.003). Groups did not differ in mitochondrial respiratory capacity (p=0.3) or fiber type (p=0.9). ΔRQ was significantly associated with the M value (p=0.02, r=0.55) and did not associate with fiber type (p=0.6, r=0.16).

Conclusions: FLEX and INFLEX individuals differ in substrate oxidation in SkM tissue with INFLEX showing greater and more efficient fat oxidation. Differences in ΔRQ are not influenced by muscle fiber type.

Disclosure

P. Krassovskaia: None. F. Jevtovic: None. M. Chorzepa: None. T.E. Jones: None. C.J. Tanner: None. D. Zheng: None. W.J. Pories: None. L. Dohm: None. J.A. Houmard: None. N.T. Broskey: None.

Funding

National Institutes of Health R01DK120296