bluuebunny/crossref_metadata_2025_split huggingface dataset:
1699-P: The Conversion and Potential Mechanism of Conditionally Immortalized Human Classical Brown Preadipocytes from Proliferative to Adipogenic States
Authors: YOU LIANGHUI , HUA SIYU , YU TINGTING , JI CHENBO 
Published: June 2024

Introduction & Objective: Considering to the anti-obesity and diabetic effect of brown adipose tissue (BAT), different immortalized brown preadipocytes cell lines have been established. Notably, murine preadipocyte cell lines expressed Simian virus 40 large tumor antigen (SV40LT) possess adipogenic capacity, whereas human preadipocyte cell lines with the same method failed. This study aimed to establish conditionally immortalized human classical brown preadipocytes (hcBPAs) and elucidate the mechanism underlying SV40LT-mediated regulation of adipogenesis.
Methods: hcBPAs was achieved by controlling SV40LT with doxycycline-inducible Tet-on system. Proliferation ability, adipogenic capacity, and brown adipocytes identity were compared between hcBPAs and primary brown preadipocytes. hcBPAs were transplanted into HFD-induced nude mice. Metabolic index and adipogenic capacity were evaluated after 8 weeks. ATAC-seq and RNA-seq were adopted between doxycycline-removal and -inducible hcBPAs to elucidate changes in chromatin accessibility and adipogenic genes.
Results: A hcBPAs was obtained with similar adipogenic capacity, thermogenic function, and molecular characteristics compared with primary preadipocytes. hcBPAs transplantation models have shown improvements in glucose metabolism and reconstituted functional brown adipose tissue. Differentiated doxycycline-removal hcBPAs showed increased ATAC sensitivity and mRNA expressions in adipogenic related genes and transcription factor (Fosl2 and CTCF) occupancy. RNA-seq and qPCR revealed equal expression levels of these regulatory factors in both groups, raising the possibility that downregulated of SV40TL expression promotes the binding of these factors to chromatin.
Conclusion: The model of hcBPAs was constructed and provided a reliable tool for prevention and treatment of obesity.
Keywords: hcBPAs, SV40TL, ATAC sensitivity, Fosl2, CTCF

Disclosure
L. You: None. S. Hua: None. T. Yu: None. C. Ji: None.

Funding
National Natural Science Foundation of China (82170880)
1700-P: Weight Reduction Is Associated with Improved Quality of Life in Participants with Obesity or Overweight and Type 2 Diabetes—Results from Phase 3 SURMOUNT-2 Trial
Authors: HUNTER GIBBLE THERESA , CAO DACHUANG , ZHANG MICHELLE X. , AGARWAL XAVIER NEENA , LING POON JIAT 
Published: June 2024

Introduction & Objective: Tirzepatide is a glucose-dependent insulinotropic polypeptide and glucagon-like peptide-1 receptor agonist approved in the US for treatment of type 2 diabetes and obesity. In the phase 3, randomized, double-blind, placebo (PBO)-controlled SURMOUNT-2 (NCT04657003) trial, tirzepatide-treated adults showed substantial weight reduction vs PBO. This post-hoc analysis assessed the association between achieving weight reduction targets and improvements in health-related quality of life (HRQoL).
Methods: HRQoL was assessed at baseline and week (W) 72 or early treatment discontinuation using 36-Item Short Form Health Survey, Version2 (SF-36v2), Impact of Weight on Quality of Life-Lite-Clinical Trials (IWQOL-Lite-CT), and EQ-5D-5L with higher scores representing better HRQoL. Mean changes from baseline to W72 were summarized by weight reduction targets (≥5%, ≥10%, ≥15%, ≥20%) in pooled tirzepatide-treated adults with last observed post-baseline value carried forward.
Results: At W72, adults with weight reduction showed improvements from baseline in all domains of SF-36v2 including Physical Functioning (norm-based) (≥5%, 3.78; ≥10%, 3.95; ≥15%, 4.74; ≥20%, 4.63) and Mental Health (norm-based) (≥5%, 1.65; ≥10%, 1.75; ≥15%, 2.34; ≥20%, 2.63). Adults who achieved most higher weight targets showed greater improvements from baseline in IWQOL-Lite-CT total score (≥5%, 14.68; ≥10%, 15.45; ≥15%, 18.53; ≥20%, 20.10), and Physical and Psychosocial composite scores. Greater improvement from baseline in EQ-5D VAS scores was observed in adults meeting greater weight reduction targets (≥5%, 9.75; ≥10%, 10.60; ≥15%, 12.33; ≥20%, 12.67).
Conclusion: Weight reduction in tirzepatide-treated adults was associated with improvement in all HRQoL domains, including physical function and mental health. Adults who lost the most weight generally showed the greatest HRQoL improvements.

Disclosure
T. Hunter Gibble: Employee; Eli Lilly and Company. Stock/Shareholder; Eli Lilly and Company. D. Cao: Employee; Eli Lilly and Company. Stock/Shareholder; Eli Lilly and Company. M.X. Zhang: Employee; Eli Lilly and Company. Stock/Shareholder; Eli Lilly and Company. N. Xavier: Employee; Eli Lilly and Company. J. Poon: Employee; Eli Lilly and Company. Stock/Shareholder; Eli Lilly and Company.

Funding
Eli Lilly and Company
1702-P: High Circulating MIF Levels Indicate the Association with Atypical Antipsychotic-Induced Metabolic Adverse Effects
Authors: CHEN XI , GAO PINGYI , QI YADAN , CUI DONGHONG , QI DAKE 
Published: June 2024

Atypical antipsychotics (AAPs) are first-line medications for schizophrenia (SZ). However, their use is frequently associated with the development of metabolic adverse effects, and the mechanisms behind these negative effects remain inadequately elucidated. Macrophage migration inhibitory factor (MIF) is a procytokine and involved in the development of metabolic dysfunction. To investigate the role of MIF in regulating antipsychotic-induced metabolic abnormalities, we recruited 142 healthy individuals and 388 SZ patients who had been receiving either typical antipsychotic (TAP) or AAP treatments. Subsequently, we conducted assessments of metabolic indices and measured plasma MIF levels, followed by a comprehensive statistical analysis to investigate the connection between MIF levels and metabolic dysfunction. A significant increase in plasma MIF levels was observed in groups receiving monotherapies with five major AAPs in comparison to healthy controls (all p &lt; 0.0001). There was no such increase shown in the group receiving TAP treatment (p &gt; 0.05). Elevated plasma MIF levels displayed a notable correlation with insulin resistance (β = 0.024, p = 0.020), as well as with the levels of triglycerides (β = 0.019, p = 0.001) and total cholesterol (β = 0.012, p = 0.038) in the groups receiving AAPs. However, while the TAP group also displayed some degree of metabolic dysfunction compared to healthy controls, no significant association was evident with plasma MIF levels (all p &gt; 0.05).
In conclusion, Plasma MIF levels exhibit a distinctive correlation with metabolic abnormalities triggered by AAPs. Thus, MIF could be further developed as a unique marker to monitor AAP-induced metabolic adverse effects in clinical settings.

Disclosure
X. Chen: None. P. Gao: None. Y. Qi: None. D. Cui: None. D. Qi: None.

Funding
This study was supported by National Sciences and Engineering Research Council of Canada (NSERC: RGPIN-2017-04542) and Canadian Institutes of Health Research (CIHR Project Grant: PJT-156116) for Dr. Qi.
1714-P: SerpinB13 Regulates Proliferation and Cell Survival in Beta Cells
Authors: KRYVALAP YURY , CZYZYK JAN 
Published: June 2024

Our studies with a monoclonal antibody (mAb) to serpinB13, a protease inhibitor of cathepsin L, and serpinB13-deficient mice, demonstrated that during development, this serpin molecule downregulates the number of Ngn3-positive pancreatic endocrine progenitor cells and their conversion to insulin-producing cells. Establishing the precise role of serpinB13 in the pancreas after birth is important, as its inhibition with a mAb may provide a novel approach to augment beta cell neogenesis, proliferation, and survival in diabetes. To address this, we injected 10 micrograms of low-endotoxin, recombinant serpinB13 (or chicken ovalbumin as a control [OVA]) once a week for 4 weeks into healthy 3-week old immunodeficient Balb/c SCID mice. In addition, during the same period, these mice received 5-EdU in two i.p. injections per week (8 injections in total). At 8 weeks of age the animals were sacrificed for microscopic examination of the percentage of EdU+/insulin+ cells. In an alternative approach, to measure the impact of serpinB13 on cell survival in vitro, immortalized MIN6 beta cells were cultured with a 4 micromolar solution of staurosporine in the presence or absence of serpinB13 at 1 microgram/mL for four hours. Early and late apoptosis was examined with antibodies to Annexin V and caspase 3 antibody, respectively. We found that female mice injected with serpinB13 had significantly reduced proliferation of beta cells with a similar trend for alpha cells. This difference was observed in females rather than males and was particularly prominent for pancreatic islets less than 180 microns in diameter. Moreover, our in vitro studies revealed a significant augmentation of apoptosis and death events following exposure of MIN6 cells to serpinB13. Together our data suggest that serpinB13 has detrimental effects on beta-cell biology, both during embryogenesis and after birth, and further suggests that impeding serpinB13 function is potentially desirable to achieve improved clinical outcomes in the treatment of diabetes.

Disclosure
Y. Kryvalap: None. J. Czyzyk: None.
1715-P: The c-Jun Transcription Factor Regulates Beta-Cell Survival following Prolactin Treatment
Authors: RUIZ OTERO NELMARI , CHUNG JIN-YONG , BANERJEE RONADIP R. 
Published: June 2024

Pregnancy and postpartum states drive dynamic expansion and regression of maternal beta-cells in response to changing metabolic demands. We previously identified a critical role for prolactin receptor (PRLR) signaling in orchestrating the gestational expansion of beta-cells. Using single-cell RNA sequencing of mouse islets, we recently identified increased expression of immediate early genes (IEGs) in late gestation and early postpartum beta-cells. Furthermore, expression of a subset of IEGs, including c-Jun, appeared to be mutually exclusive with beta-cells undergoing apoptosis in the postpartum as assessed by TUNEL staining. The c-Jun transcription factor is a well-characterized IEG implicated in regulating proliferation, apoptosis, and survival in various tissues. Therefore, we hypothesized that c-Jun plays a critical role in beta-cell survival within the postpartum islet via PRLR signaling. Using immunostaining we found that c-Jun is normally expressed in islets from late pregnancy and early postpartum but is not present in mice lacking PRLR within beta-cells. This indicated PRLR signaling is required for induction of c-Jun. Prolactin treatment induces c-Jun expression within beta-cells in cultured murine islets as well as islets from a human female donor. Induction of c-Jun by prolactin is mediated by MAPK/ERK but not Stat5 or PI3K signaling as determined by selective inhibition of these pathways. Prolactin has been shown to promote survival of beta-cells. Inhibition of Jun phosphorylation blocks prolactin-mediated protection from dexamethasone-induced apoptosis of beta-cells. Together, these studies suggest c-Jun acts downstream of PRLR / MAPK signaling contributing to the pro-survival effects of prolactin signaling.

Disclosure
N. Ruiz Otero: None. J. Chung: None. R.R. Banerjee: None.

Funding
National Institutes of Health (RO1 (DK120761))
1717-P: Protein Kinase C δ Activation by Nitric Oxide Mediates Pancreatic ß-Cell Apoptosis
Authors: COLLINS JILLIAN , FARNSWORTH NIKKI L. 
Published: June 2024

In type 1 diabetes (T1D), autoreactive immune cells secrete cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interferon-γ (IFN-γ), that initiate β-cell apoptosis in pancreatic islets and contribute to disease progression. Protein kinase C delta (PKCδ) activation plays a pivotal role in mediating cytokine-induced β-cell apoptosis via nuclear translocation and activation of pro-apoptotic signaling. In other cell types, nitric oxide (NO) produced in response to cytokines can also activate PKCδ via nitrosylation of tyrosine; however, the exact modification sites and effects on apoptotic signaling have not been investigated in β-cells. We hypothesize cytokine-induced production of NO activates PKCδ leading to nuclear translocation and upregulation of apoptotic signaling.
Islets from WT mice, mice with a β-cell specific knockout of PKCδ (PKCδβ-KO), or human islets were cultured for 24hr with a NO donor (SNAP, 1mM), cytokines (10ng/ml TNFα, 5ng/ml IL-1β, 100ng/ml IFN-γ), cytokines with an inducible nitric oxide synthase inhibitor (iNOSinh, 5mM), or an adenovirus to produce GFP-tagged PKCδ. Translocation was determined by GFP colocalization with a nuclear stain. Apoptosis was determined by fluorescent live/dead staining. PKCδ activity was determined with a FRET-based sensor in MIN6 cells. Nitrosylation of PKCδ was determined via a biotin switch assay and western blot.
Increases in PKCδ activity were observed in NO treated islets (p=0.05). Inhibiting PKCδ activation protected against β-cell apoptosis in cytokine and NO treated islets (p=0.004). Overall, our results support a role for cytokine induced NO in activating PKCδ and mediating downstream apoptotic β-cell death. Uncovering mechanisms to inhibit PKCδ activation that are associated with T1D can improve targeted protection of β-cells and reduce off target affects. Identifying novel mechanisms of cytokine-mediated apoptosis will provide insight into novel targets to prevent or delay β-cell death in T1D.

Disclosure
J. Collins: None. N.L. Farnsworth: None.

Funding
American Diabetes Association (7-21-JDF-020); Juvenile Diabetes Research Foundation (1-FAC-2020-891-A-N)
172-OR: Lineage Tracing of Vasculogenic Fibroblasts In Vivo and Their Significance in the Rescue of Diabetic Ischemic Tissue
Authors: SINGH KANHAIYA , VERMA SUMIT S. , ROY SASHWATI , SEN CHANDAN K. , KUMAR MANISHEKHAR 
Published: June 2024

Our recent work reported on the identification of the vasculogenic fibroblast (VF) that is capable of generating new blood vessels during tissue repair (Pal et al. Nat Com, 2023). While these cells are physiologic and injury-inducible, the generation of VF is blunted under conditions of diabetes. Such barrier may be overcome by the inhibition of fibroblast miR-200b using tissue nanotransfection technology (PMIDs: 28785092, 34837085, 35819852, 36380587). To understand the mechanisms underlying VF function and significance, in this work we developed miR-200b-429fl/fl COL1A2creER GT(ROSA)mT/mG mice. In these mice, miR-200b regions in fibroblasts (COL1A2+ cells) are knocked out in response to tamoxifen. To gain insight into the significance of VF in vivo, ischemic hind limb studies were performed. Tamoxifen treatment not only led to Cre excision of miR-200b-429 in these mice, but also induced the constitutive expression of ROSA driven tdTomato in fibroblasts with onset of permanent GFP expression. Significant lowering of miR-200b was noted in laser capture dissected dermal fibroblast tissue elements from the wound-edge of tamoxifen treated mice (n=4). Tamoxifen treatment led to significantly enhanced perfusion in animals subjected to ischemic injury as determined by laser speckle perfusion imaging (Perimed Inc.) (n=8). This rescue was associated with increased abundance of GFP+CD31+ VF (n=5) which accounted for 26% of all CD31+ vascular cells in the laser captured blood vessels (n=9). Thus, targeted lowering of miR-200b abundance within fibroblasts resulted in substantial augmentation of VF in vivo which was associated with improved blood flow.

Disclosure
K. Singh: None. S.S. Verma: None. S. Roy: None. C.K. Sen: Consultant; SouthWest Technologies. Board Member; VisopalExo. Consultant; Vomaris Inc. M. Kumar: None.

Funding
National Institute of Diabetes and Digestive and Kidney Diseases (DK136814, DK128845, DK135447, DK125835); Department of Defense (W81XWH-21-1-0033, W81XWH-22-1-0146)
1724-P: DNA Damage Vulnerability of Immature Beta Cells
Authors: VARGHESE SNEHA S. , HERNANDEZ-DE LA PENA ALESSANDRO G. , BHATTACHARYA SUPRIYO , DHAWAN SANGEETA 
Published: June 2024

Introduction & Objective: A healthy reserve of functional β-cells is crucial for maintaining glucose homeostasis and is disrupted in diabetes. Most of this reserve is established in early neonatal life via replication, following which β-cells exit cell-cycle and undergo functional maturation. Although the process of replication is extremely vulnerable to DNA damage and consequent genomic instability, the extent and impact of such vulnerabilities on β-cell maturation and subsequent viability is not clear. Here, we sought to determine the DNA damage vulnerability of replicating β-cells in the neonatal growth phase.
Methods: We quantified β-cell replication (Ki67, pHH3) and DNA damage (ɣH2AX) in early (p7) and late (p21) neonatal B6 mice (N=8 animals/ group). We acquired super-resolution 3D images of DNA damage foci and cohesins (modulators of genomic stability) in replicating β-cells using the Zeiss Elyra 7 Lattice SIM^2 and LSM880 with Airyscan, and analyzed using the Imaris software.
Results: Our data revealed that while the overall number of β-cells with DNA damage decreased as replication declined from p7 to p21, ~20% of replicating β-cells at each stage harbored DNA damage. Notably, the G2 phase had the highest density of DNA damage foci, which were resolved by the M phase. Single-cell RNA-seq of murine neonatal islets showed the enrichment of DNA damage repair pathways in replicating β-cells.
Conclusion: Collectively, our data establish the high DNA damage vulnerability of neonatal β-cells and suggest the existence of robust mechanisms that assist the resolution of DNA damage with cell cycle progression to maintain genomic stability.

Disclosure
S.S. Varghese: None. A.G. Hernandez-De La Pena: None. S. Bhattacharya: None. S. Dhawan: None.

Funding
Postdoctoral Fellowship from California Institute of Regenerative Medicine (EDU4-12772); NIH-National Institute of Diabetes and Digestive and Kidney Diseases (R01DK120523)
1748-P: NMDA Receptor Is Involved in Amplification Effects of Glycine on Insulin Secretion
Authors: WANG ZIWEI , MENG SHI , HAN DONGNA , LI CHANGHONG , ZHANG JUNQING 
Published: June 2024

The NMDA receptor plays a crucial role in the central nervous system. Glutamate acts as its agonist, while glycine and D-serine play a co-agonist role. Recent studies have indicated that β-cells express the NMDA receptor, and it regulates insulin secretion, but the detailed mechanism is still unclear. Mouse islets were perifused in response to 10 mM glucose stimulation, a ramp of glutamate and glycine (0 to 10 mM) was administered either alone or on top of glucose. The ramp of glutamate and glycine alone have no effects on insulin secretion in perifused mouse islets. However, glycine ramp, not glutamate and D-serine ramp, can potentiate the 10 mM glucose-stimulated insulin secretion (GSIS). Glycine transporter 1, 2 and glycine receptor inhibitors, such as strychnine (5 µM), org25543 (10 µM), and the combination of org25543 (10 µM) and org24598 (10 µM), cannot block the potentiation effects of glycine to GSIS. The NMDA receptor inhibitor, MK801 (10 µM), enhanced the ability of glycine to augment GSIS, resulting in a leftward shift of the glycine amplification curve. Remarkably, D-serine and MK801 significantly enhanced the insulin secretion driven by 10 mM glucose, although glycine no longer exhibited its enhancing effects on GSIS. The synergistic effects of D-serine and MK801 on glucose ramp (0 to 25 mM) stimulated insulin secretion were examined. The combination resulted in an increase in the maximum insulin secretion induced by the glucose ramp, while the threshold remained unchanged. In vivo tests demonstrated that glycine improved glucose tolerance by increasing baseline insulin secretion. Our study has shown that glycine enhances GSIS through the involvement of the NMDA receptor pathway. In contrast to the central nervous system, the NMDA receptor in islets is no longer responsive to stimulation by glutamate. However, it still exhibits a robust response to glycine. While the NMDA receptor is present in islets, it may be regulated and sensed differently compared to the central nervous system.

Disclosure
Z. Wang: None. S. Meng: Employee; Nanjing Ascendrare Pharmaceutical Technology. D. Han: Employee; Nanjing Ascendrare Pharmaceutical Technology. C. Li: Employee; Hua Medicine (Shanghai) LTD., Nanjing AscendRare Pharmaceutical Technology. J. Zhang: None.

Funding
National Natural Science Foundation of China (81970701)
1749-P: Deficient Mitochondrial Activation in Type 2 Diabetic Human ß-Cells during Insulin Secretion
Authors: GERENCSER AKOS A. 
Published: June 2024

Glucose and cellular energy metabolism are central components of glucose sensing in pancreatic β-cells. Conceptually, β-cells operate by converting the rate of glucose metabolism into the rate of insulin secretion. We have identified a novel activation mechanism in healthy human β-cell mitochondria that plays a central role in setting this rate of metabolism during glucose-stimulated insulin secretion (GSIS). Using live islets and dispersed islet cell cultures originating from type 2 diabetic (T2D) and nondiabetic human cadavers, here we show the failure of this activation mechanism and a loss of bioenergetic control in β-cells from T2D individuals. Modular kinetic analysis of oxidative phosphorylation was used to measure pathway activities based on monitoring cell respiration, mitochondrial membrane potential (ΔψM) and NADH/NAD+. We observed an energization-dependent activation of glucose oxidation during GSIS in nondiabetic β-cells, and localized this to intramitochondrial NADH-producing pathways, excluding Ca2+-dependent activation. In T2D β-cells the activation of this activation was impaired. Chronic culturing of nondiabetic β-cells at elevated glucose concentration or in the presence of methylglyoxal recapitulated the suppression of activation observed in T2D β-cells. In high glucose-cultured nondiabetic and in standard-cultured T2D β-cells the magnitude of GSIS was highly variable between individuals, and often increased. GSIS correlated well and was predicted by ΔψM in nondiabetic β-cells, but this bioenergetic fine control was lost in T2D suggesting a rearrangement of stimulus-secretion coupling in T2D, where the role energetics reduces to permissive, and other coupling pathways become dominant with kinetic properties different from nondiabetic.

Disclosure
A.A. Gerencser: None.

Funding
National Institutes of Health (R01DK135807)