Hydroxycitrate delays early mortality in mice and promotes muscle regeneration while inducing a rich hepatic energetic status.

ATP citrate lyase (ACLY) inhibitors have the potential of modulating central processes in protein, carbohydrate, and lipid metabolism, which can have relevant physiological consequences in aging and age-related diseases. Here, we show that hepatic phospho-active ACLY correlates with overweight and Model for End-stage Liver Disease score in humans. Wild-type mice treated chronically with the ACLY inhibitor potassium hydroxycitrate exhibited delayed early mortality. In AML12 hepatocyte cultures, the ACLY inhibitors potassium hydroxycitrate, SB-204990, and bempedoic acid fostered lipid accumulation, which was also observed in the liver of healthy-fed mice treated with potassium hydroxycitrate. Analysis of soleus tissue indicated that potassium hydroxycitrate produced the modulation of wound healing processes. In vivo, potassium hydroxycitrate modulated locomotor function toward increased wire hang performance and reduced rotarod performance in healthy-fed mice, and improved locomotion in mice exposed to cardiotoxin-induced muscle atrophy. Our findings implicate ACLY and ACLY inhibitors in different aspects of aging and muscle regeneration.

Human Omental Mesothelial Cells Impart an Immunomodulatory Landscape Impeding B- and T-Cell Activation.

Mesothelial cells form the mesothelium, a simple epithelium lining the walls of serous cavities and the surface of visceral organs. Although mesothelial cells are phenotypically well characterized, their immunoregulatory properties remain largely unknown, with only two studies reporting their capacity to inhibit T cells through TGF-ß and their consumption of L-arginine by arginase-1. Whether human mesothelial cells can suppress other immune cells and possess additional leukosuppressive mechanisms, remain to be addressed to better delineate their therapeutic potential for cell therapy. Herein, we generated secretomes from omental mesothelial cells (OMC) and assess their capacity to inhibit lymphocytes proliferation, suppress activated T and B cells, as well as to modify macrophage activation markers. The secretome from mesenchymal stromal cells (MSC) served as a control of immuno-suppression. Although OMC and MSC were phenotypically divergent, their cytokine secretion patterns as well as expression of inflammatory and immunomodulary genes were similar. As such, OMC- and MSC-derived secretomes (OMC-S and MSC-S) both polarized RAW 264.7 macrophages towards a M2-like anti-inflammatory phenotype and suppressed mouse and human lymphocytes proliferation. OMC-S displayed a strong ability to suppress mouse- and human-activated CD19+/CD25+ B cells as compared to MSC-S. The lymphosuppressive activity of the OMC-S could be significantly counteracted either by SB-431542, an inhibitor of TGFß and activin signaling pathways, or with a monoclonal antibody against the TGFß1, ß2, and ß3 isoforms. A strong blockade of the OMC-S-mediated lymphosuppressive activity was achieved using L-NMMA, a specific inhibitor of nitric oxide synthase (NOS). Taken together, our results suggest that OMC are potent immunomodulators.

Nuclear Envelope Integrity in Health and Disease: Consequences on Genome Instability and Inflammation.

The dynamic nature of the nuclear envelope (NE) is often underestimated. The NE protects, regulates, and organizes the eukaryote genome and adapts to epigenetic changes and to its environment. The NE morphology is characterized by a wide range of diversity and abnormality such as invagination and blebbing, and it is a diagnostic factor for pathologies such as cancer. Recently, the micronuclei, a small nucleus that contains a full chromosome or a fragment thereof, has gained much attention. The NE of micronuclei is prone to collapse, leading to DNA release into the cytoplasm with consequences ranging from the activation of the cGAS/STING pathway, an innate immune response, to the creation of chromosomal instability. The discovery of those mechanisms has revolutionized the understanding of some inflammation-related diseases and the origin of complex chromosomal rearrangements, as observed during the initiation of tumorigenesis. Herein, we will highlight the complexity of the NE biology and discuss the clinical symptoms observed in NE-related diseases. The interplay between innate immunity, genomic instability, and nuclear envelope leakage could be a major focus in future years to explain a wide range of diseases and could lead to new classes of therapeutics.

Physical Forces and Transient Nuclear Envelope Rupture during Metastasis: The Key for Success?

During metastasis, invading tumor cells and circulating tumor cells (CTC) face multiple mechanical challenges during migration through narrow pores and cell squeezing. However, little is known on the importance and consequences of mechanical stress for tumor progression and success in invading a new organ. Recently, several studies have shown that cell constriction can lead to nuclear envelope rupture (NER) during interphase. This loss of proper nuclear compartmentalization has a profound effect on the genome, being a key driver for the genome evolution needed for tumor progression. More than just being a source of genomic alterations, the transient nuclear envelope collapse can also support metastatic growth by several mechanisms involving the innate immune response cGAS/STING pathway. In this review we will describe the importance of the underestimated role of cellular squeezing in the progression of tumorigenesis. We will describe the complexity and difficulty for tumor cells to reach the metastatic site, detail the genomic aberration diversity due to NER, and highlight the importance of the activation of the innate immune pathway on cell survival. Cellular adaptation and nuclear deformation can be the key to the metastasis success in many unsuspected aspects.

Thyroid hormones in diabetes, cancer, and aging.

Thyroid function is central in the control of physiological and pathophysiological processes. Studies in animal models and human research have determined that thyroid hormones modulate cellular processes relevant for aging and for the majority of age-related diseases. While several studies have associated mild reductions on thyroid hormone function with exceptional longevity in animals and humans, alterations in thyroid hormones are serious medical conditions associated with unhealthy aging and premature death. Moreover, both hyperthyroidism and hypothyroidism have been associated with the development of certain types of diabetes and cancers, indicating a great complexity of the molecular mechanisms controlled by thyroid hormones. In this review, we describe the latest findings in thyroid hormone research in the field of aging, diabetes, and cancer, with a special focus on hepatocellular carcinomas. While aging studies indicate that the direct modulation of thyroid hormones is not a viable strategy to promote healthy aging or longevity and the development of thyromimetics is challenging due to inefficacy and potential toxicity, we argue that interventions based on the use of modulators of thyroid hormone function might provide therapeutic benefit in certain types of diabetes and cancers.

Therapeutic Potential of Mesenchymal Stem Cells for Cancer Therapy.

Mesenchymal stem cells (MSCs) are among the most frequently used cell type for regenerative medicine. A large number of studies have shown the beneficial effects of MSC-based therapies to treat different pathologies, including neurological disorders, cardiac ischemia, diabetes, and bone and cartilage diseases. However, the therapeutic potential of MSCs in cancer is still controversial. While some studies indicate that MSCs may contribute to cancer pathogenesis, emerging data reported the suppressive effects of MSCs on cancer cells. Because of this reality, a sustained effort to understand when MSCs promote or suppress tumor development is needed before planning a MSC-based therapy for cancer. Herein, we provide an overview on the therapeutic application of MSCs for regenerative medicine and the processes that orchestrates tissue repair, with a special emphasis placed on cancer, including central nervous system tumors. Furthermore, we will discuss the current evidence regarding the double-edged sword of MSCs in oncological treatment and the latest advances in MSC-based anti-cancer agent delivery systems.

LRH-1 agonism favours an immune-islet dialogue which protects against diabetes mellitus.

Type 1 diabetes mellitus (T1DM) is due to the selective destruction of islet beta cells by immune cells. Current therapies focused on repressing the immune attack or stimulating beta cell regeneration still have limited clinical efficacy. Therefore, it is timely to identify innovative targets to dampen the immune process, while promoting beta cell survival and function. Liver receptor homologue-1 (LRH-1) is a nuclear receptor that represses inflammation in digestive organs, and protects pancreatic islets against apoptosis. Here, we show that BL001, a small LRH-1 agonist, impedes hyperglycemia progression and the immune-dependent inflammation of pancreas in murine models of T1DM, and beta cell apoptosis in islets of type 2 diabetic patients, while increasing beta cell mass and insulin secretion. Thus, we suggest that LRH-1 agonism favors a dialogue between immune and islet cells, which could be druggable to protect against diabetes mellitus.

The type 2 diabetes-associated HMG20A gene is mandatory for islet beta cell functional maturity.

HMG20A (also known as iBRAF) is a chromatin factor involved in neuronal differentiation and maturation. Recently small nucleotide polymorphisms (SNPs) in the HMG20A gene have been linked to type 2 diabetes mellitus (T2DM) yet neither expression nor function of this T2DM candidate gene in islets is known. Herein we demonstrate that HMG20A is expressed in both human and mouse islets and that levels are decreased in islets of T2DM donors as compared to islets from non-diabetic donors. In vitro studies in mouse and human islets demonstrated that glucose transiently increased HMG20A transcript levels, a result also observed in islets of gestating mice. In contrast, HMG20A expression was not altered in islets from diet-induced obese and pre-diabetic mice. The T2DM-associated rs7119 SNP, located in the 3' UTR of the HMG20A transcript reduced the luciferase activity of a reporter construct in the human beta 1.1E7 cell line. Depletion of Hmg20a in the rat INS-1E cell line resulted in decreased expression levels of its neuronal target gene NeuroD whereas Rest and Pax4 were increased. Chromatin immunoprecipitation confirmed the interaction of HMG20A with the Pax4 gene promoter. Expression levels of Mafa, Glucokinase, and Insulin were also inhibited. Furthermore, glucose-induced insulin secretion was blunted in HMG20A-depleted islets. In summary, our data demonstrate that HMG20A expression in islet is essential for metabolism-insulin secretion coupling via the coordinated regulation of key islet-enriched genes such as NeuroD and Mafa and that depletion induces expression of genes such as Pax4 and Rest implicated in beta cell de-differentiation. More importantly we assign to the T2DM-linked rs7119 SNP the functional consequence of reducing HMG20A expression likely translating to impaired beta cell mature function.

Targeting pancreatic expressed PAX genes for the treatment of diabetes mellitus and pancreatic neuroendocrine tumors.

INTRODUCTION: Four members of the PAX family, PAX2, PAX4, PAX6 and PAX8 are known to be expressed in the pancreas. Accumulated evidences indicate that several pancreatic expressed PAX genes play a significant role in pancreatic development/functionality and alterations in these genes are involved in the pathogenesis of pancreatic diseases. Areas covered: In this review, we summarize the ongoing research related to pancreatic PAX genes in diabetes mellitus and pancreatic neuroendocrine tumors. We dissect the current knowledge at different levels; from mechanistic studies in cell lines performed to understand the molecular processes controlled by pancreatic PAX genes, to in vivo studies using rodent models that over-express or lack specific PAX genes. Finally, we describe human studies associating variants on pancreatic-expressed PAX genes with pancreatic diseases. Expert opinion: Based on the current literature, we propose that future interventions to treat pancreatic neuroendocrine tumors and diabetes mellitus could be developed via the modulation of PAX4 and/or PAX6 regulated pathways.

Endoplasmic Reticulum Stress Links Oxidative Stress to Impaired Pancreatic Beta-Cell Function Caused by Human Oxidized LDL.

Elevated plasma concentration of the pro-atherogenic oxidized low density lipoprotein cholesterol (LDL) triggers adverse effects in pancreatic beta-cells and is associated with type 2 diabetes. Here, we investigated whether the endoplasmic reticulum (ER) stress is a key player coupling oxidative stress to beta-cell dysfunction and death elicited by human oxidized LDL. We found that human oxidized LDL activates ER stress as evidenced by the activation of the inositol requiring 1α, and the elevated expression of both DDIT3 (also called CHOP) and DNAJC3 (also called P58IPK) ER stress markers in isolated human islets and the mouse insulin secreting MIN6 cells. Silencing of Chop and inhibition of ER stress markers by the chemical chaperone phenyl butyric acid (PBA) prevented cell death caused by oxidized LDL. Finally, we found that oxidative stress accounts for activation of ER stress markers induced by oxidized LDL. Induction of Chop/CHOP and p58IPK/P58IPK by oxidized LDL was mimicked by hydrogen peroxide and was blocked by co-treatment with the N-acetylcystein antioxidant. As a conclusion, the harmful effects of oxidized LDL in beta-cells requires ER stress activation in a manner that involves oxidative stress. This mechanism may account for impaired beta-cell function in diabetes and can be reversed by antioxidant treatment.

Adipose mesenchymal stromal cells isolated from type 2 diabetic patients display reduced fibrinolytic activity.

Stem cells have been successfully used for the treatment of critical limb ischemia (CLI). We conducted a clinical trial to determine the feasibility of using autologous adipose-derived mesenchymal stromal cells (AdMSCs) for the treatment of CLI. Unexpectedly, two diabetic patients developed peripheral microthrombosis. This adverse effect, which contrasts with the reported antithrombotic properties of MSCs, may stem from the diabetic environment that alters the fibrinolytic activity of AdMSCs, thereby increasing the probability of developing thrombosis. Here, we confirm this premise by demonstrating that diabetic AdMSCs cultured in the presence of blood sera expressed and released higher levels of plasminogen activator inhibitor type 1, reduced levels of tissue plasminogen activator, and lower d-dimer formation compared with nondiabetic AdMSCs. Thus, to establish an appropriate cell therapy for diabetic patients, we recommend including new preclinical safety tests, such as the d-dimer and/or the tissue plasminogen activator-to-plasminogen activator inhibitor type 1 ratio tests, to assess fibrinolytic activity of cells before implantation.

Islet ß-Cell Mass Preservation and Regeneration in Diabetes Mellitus: Four Factors with Potential Therapeutic Interest.

Islet ß-cell replacement and regeneration are two promising approaches for the treatment of Type 1 Diabetes Mellitus. Indeed, the success of islet transplantation in normalizing blood glucose in diabetic patients has provided the proof of principle that cell replacement can be employed as a safe and efficacious treatment. Nonetheless, shortage of organ donors has hampered expansion of this approach. Alternative sources of insulin-producing cells are mandatory to fill this gap. Although great advances have been achieved in generating surrogate ß-cells from stem cells, current protocols have yet to produce functionally mature insulin-secreting cells. Recently, the concept of islet regeneration in which new ß-cells are formed from either residual ß-cell proliferation or transdifferentiation of other endocrine islet cells has gained much interest as an attractive therapeutic alternative to restore ß-cell mass. Complementary approaches to cell replacement and regeneration could aim at enhancing ß-cell survival and function. Herein, we discuss the value of Hepatocyte Growth Factor (HGF), Glucose-Dependent Insulinotropic Peptide (GIP), Paired box gene 4 (Pax4) and Liver Receptor Homolog-1 (LRH-1) as key players for ß-cell replacement and regeneration therapies. These factors convey ß-cell protection and enhanced function as well as facilitating proliferation and transdifferentiation of other pancreatic cell types to ß-cells, under stressful conditions.

Immunohistochemical assessment of Pax8 expression during pancreatic islet development and in human neuroendocrine tumors.

The paired box transcription factor Pax8 is critical for development of the eye, thyroid gland as well as the urinary and reproductive organs. In adult, Pax8 overexpression is associated with kidney, ovarian and thyroid tumors and has emerged as a specific marker for these cancers. Recently, Pax8 expression was also reported in human pancreatic islets and in neuroendocrine tumors, identifying Pax8 as a novel member of the Pax family expressed in the pancreas. Herein, we sought to provide a comprehensive analysis of Pax8 expression during pancreogenesis and in adult islets. Immunohistochemical analysis using the most employed Pax8 polyclonal antibody revealed strong nuclear staining in the developing mouse pancreas and in mature human and mouse islets. Astonishingly, Pax8 mRNA in mouse islets was undetectable while human islets exhibited low levels. These discrepancies raised the possibility of antibody cross-reactivity. This premise was confirmed by demonstrating that the polyclonal Pax8 antibody also recognized the islet-enriched Pax6 protein both by Western blotting and immunohistochemistry. Thus, in islets polyclonal Pax8 staining corresponds mainly to Pax6. In order to circumvent this caveat, a novel Pax8 monoclonal antibody was used to re-evaluate whether Pax8 was indeed expressed in islets. Surprisingly, Pax8 was not detected in neither the developing pancreas or in mature islets. Reappraisal of pancreatic neuroendocrine tumors using this Pax8 monoclonal antibody exhibited no immunostaining as compared to the Pax8 polyclonal antibody. In conclusion, Pax8 is not expressed in the pancreas and cast doubts on the value of Pax8 as a pancreatic neuroendocrine tumor marker.

The liver receptor homolog-1 (LRH-1) is expressed in human islets and protects {beta}-cells against stress-induced apoptosis.

Liver receptor homolog (LRH-1) is an orphan nuclear receptor (NR5A2) that regulates cholesterol homeostasis and cell plasticity in endodermal-derived tissues. Estrogen increases LRH-1 expression conveying cell protection and proliferation. Independently, estrogen also protects isolated human islets against cytokine-induced apoptosis. Herein, we demonstrate that LRH-1 is expressed in islets, including ß-cells, and that transcript levels are modulated by 17ß-estradiol through the estrogen receptor (ER)α but not ERß signaling pathway. Repression of LRH-1 by siRNA abrogated the protective effect conveyed by estrogen on rat islets against cytokines. Adenoviral-mediated overexpression of LRH-1 in human islets did not alter proliferation but conferred protection against cytokines and streptozotocin-induced apoptosis. Expression levels of the cell cycle genes cyclin D1 and cyclin E1 as well as the antiapoptotic gene bcl-xl were unaltered in LRH-1 expressing islets. In contrast, the steroidogenic enzymes CYP11A1 and CYP11B1 involved in glucocorticoid biosynthesis were both stimulated in transduced islets. In parallel, graded overexpression of LRH-1 dose-dependently impaired glucose-induced insulin secretion. Our results demonstrate the crucial role of the estrogen target gene nr5a2 in protecting human islets against-stressed-induced apoptosis. We postulate that this effect is mediated through increased glucocorticoid production that blunts the pro-inflammatory response of islets.

In vivo conditional Pax4 overexpression in mature islet ß-cells prevents stress-induced hyperglycemia in mice.

OBJECTIVE: To establish the role of the transcription factor Pax4 in pancreatic islet expansion and survival in response to physiological stress and its impact on glucose metabolism, we generated transgenic mice conditionally and selectively overexpressing Pax4 or a diabetes-linked mutant variant (Pax4R129W) in ß-cells. RESEARCH DESIGN AND METHODS: Glucose homeostasis and ß-cell death and proliferation were assessed in Pax4- or Pax4R129W-overexpressing transgenic animals challenged with or without streptozotocin. Isolated transgenic islets were also exposed to cytokines, and apoptosis was evaluated by DNA fragmentation or cytochrome C release. The expression profiles of proliferation and apoptotic genes and ß-cell markers were studied by immunohistochemistry and quantitative RT-PCR. RESULTS: Pax4 but not Pax4R129W protected animals against streptozotocin-induced hyperglycemia and isolated islets from cytokine-mediated ß-cell apoptosis. Cytochrome C release was abrogated in Pax4 islets treated with cytokines. Interleukin-1ß transcript levels were suppressed in Pax4 islets, whereas they were increased along with NOS2 in Pax4R129W islets. Bcl-2, Cdk4, and c-myc expression levels were increased in Pax4 islets while MafA, insulin, and GLUT2 transcript levels were suppressed in both animal models. Long-term Pax4 expression promoted proliferation of a Pdx1-positive cell subpopulation while impeding insulin secretion. Suppression of Pax4 rescued this defect with a concomitant increase in pancreatic insulin content. CONCLUSIONS: Pax4 protects adult islets from stress-induced apoptosis by suppressing selective nuclear factor-κB target genes while increasing Bcl-2 levels. Furthermore, it promotes dedifferentiation and proliferation of ß-cells through MafA repression, with a concomitant increase in Cdk4 and c-myc expression.

PDX1 deficiency causes mitochondrial dysfunction and defective insulin secretion through TFAM suppression.

Mutations in the transcription factor Pdx1 cause maturity-onset diabetes of the young 4 (MODY4). Islet transduction with dominant-negative Pdx1 (RIPDN79PDX1) impairs mitochondrial metabolism and glucose-stimulated insulin secretion (GSIS). Transcript profiling revealed suppression of nuclear-encoded mitochondrial factor A (TFAM). Herein, we show that Pdx1 suppression in adult mice reduces islet TFAM expression coinciding with hyperglycemia. We define TFAM as a direct target of Pdx1 both in rat INS1 cells and human islets. Adenoviral overexpression of TFAM along with RIPDN79PDX1 in isolated rat islets rescued mitochondrial DNA (mtDNA) copy number and restored respiratory chain activity as well as glucose-induced ATP synthesis and insulin secretion. CGP37157, which blocks the mitochondrial Na(+)/Ca(2+) exchanger, restored ATP generation and GSIS in RIPDN79PDX1 islets, thereby bypassing the transcriptional defect. Thus, the genetic control by the beta cell-specific factor Pdx1 of the ubiquitous gene TFAM maintains beta cell mtDNA vital for ATP production and normal GSIS.

Pancreatic insulin content regulation by the estrogen receptor ER alpha.

The function of pancreatic beta-cells is the synthesis and release of insulin, the main hormone involved in blood glucose homeostasis. Estrogen receptors, ER alpha and ER beta, are important molecules involved in glucose metabolism, yet their role in pancreatic beta-cell physiology is still greatly unknown. In this report we show that both ER alpha and ER beta are present in pancreatic beta-cells. Long term exposure to physiological concentrations of 17beta-estradiol (E2) increased beta-cell insulin content, insulin gene expression and insulin release, yet pancreatic beta-cell mass was unaltered. The up-regulation of pancreatic beta-cell insulin content was imitated by environmentally relevant doses of the widespread endocrine disruptor Bisphenol-A (BPA). The use of ER alpha and ER beta agonists as well as ER alphaKO and ER betaKO mice suggests that the estrogen receptor involved is ER alpha. The up-regulation of pancreatic insulin content by ER alpha activation involves ERK1/2. These data may be important to explain the actions of E2 and environmental estrogens in endocrine pancreatic function and blood glucose homeostasis.