Apelin-VEGF-C mRNA delivery as therapeutic for the treatment of secondary lymphedema.

Secondary lymphedema (LD) corresponds to a severe lymphatic dysfunction leading to the accumulation of fluid and fibrotic adipose tissue in a limb. Here, we identified apelin (APLN) as a powerful molecule for regenerating lymphatic function in LD. We identified the loss of APLN expression in the lymphedematous arm compared to the normal arm in patients. The role of APLN in LD was confirmed in APLN knockout mice, in which LD is increased and associated with fibrosis and dermal backflow. This was reversed by intradermal injection of APLN-lentivectors. Mechanistically, APLN stimulates lymphatic endothelial cell gene expression and induces the binding of E2F8 transcription factor to the promoter of CCBE1 that controls VEGF-C processing. In addition, APLN induces Akt and eNOS pathways to stimulate lymphatic collector pumping. Our results show that APLN represents a novel partner for VEGF-C to restore lymphatic function in both initial and collecting vessels. As LD appears after cancer treatment, we validated the APLN-VEGF-C combination using a novel class of nonintegrative RNA delivery LentiFlash® vector that will be evaluated for phase I/IIa clinical trial.

Postoperative delirium is a risk factor of institutionalization after hip fracture: an observational cohort study.

Introduction: Hip fracture is a common clinical problem in geriatric patients often associated with poor postoperative outcomes. Postoperative delirium (POD) and postoperative neurocognitive disorders (NCDs) are particularly frequent. The consequences of these disorders on postoperative recovery and autonomy are not fully described. The aim of this study was to determine the role of POD and NCDs on the need for institutionalization at 3 months after hip fracture surgery. Method: A population-based prospective cohort study was conducted on hip fracture patients between March 2016 and March 2018. The baseline interview, which included a Mini-Mental State Examination (MMSE), was conducted in the hospital after admission for hip fracture. NCDs were appreciated by MMSE scoring evolution (difference between preoperative MMSE and MMSE at day 5 >2 points). POD was evaluated using the Confusion Assessment Method. The primary endpoint was the rate of new institutionalization at 3 months. We used a multivariate analysis to assess the risk of new institutionalization. Results: A total of 63 patients were included. Thirteen patients (20.6%) were newly institutionalized at 3 months. Two factors were significantly associated with the risk of postoperative institutionalization at 3 months: POD (OR = 5.23; 95% CI 1.1-27.04; p = 0.04) and IADL evolution (OR = 1.8; 95% CI 1.23-2.74; p = 0.003). Conclusion: Only POD but not NCDs was associated with the risk of dependency and institutionalization after hip fracture surgery. The prevention of POD appears to be essential for improving patient outcomes and optimizing the potential for returning home.

The Tyrosine Phosphatase SHP2: A New Target for Insulin Resistance?

The SH2 containing protein tyrosine phosphatase 2(SHP2) plays essential roles in fundamental signaling pathways, conferring on it versatile physiological functions during development and in homeostasis maintenance, and leading to major pathological outcomes when dysregulated. Many studies have documented that SHP2 modulation disrupted glucose homeostasis, pointing out a relationship between its dysfunction and insulin resistance, and the therapeutic potential of its targeting. While studies from cellular or tissue-specific models concluded on both pros-and-cons effects of SHP2 on insulin resistance, recent data from integrated systems argued for an insulin resistance promoting role for SHP2, and therefore a therapeutic benefit of its inhibition. In this review, we will summarize the general knowledge of SHP2's molecular, cellular, and physiological functions, explaining the pathophysiological impact of its dysfunctions, then discuss its protective or promoting roles in insulin resistance as well as the potency and limitations of its pharmacological modulation.

Upregulated Apelin Signaling in Pancreatic Cancer Activates Oncogenic Signaling Pathways to Promote Tumor Development.

Despite decades of effort in understanding pancreatic ductal adenocarcinoma (PDAC), there is still a lack of innovative targeted therapies for this devastating disease. Herein, we report the expression of apelin and its receptor, APJ, in human pancreatic adenocarcinoma and its protumoral function. Apelin and APJ protein expression in tumor tissues from patients with PDAC and their spatiotemporal pattern of expression in engineered mouse models of PDAC were investigated by immunohistochemistry. Apelin signaling function in tumor cells was characterized in pancreatic tumor cell lines by Western blot as well as proliferation, migration assays and in murine orthotopic xenograft experiments. In premalignant lesions, apelin was expressed in epithelial lesions whereas APJ was found in isolated cells tightly attached to premalignant lesions. However, in the invasive stage, apelin and APJ were co-expressed by tumor cells. In human tumor cells, apelin induced a long-lasting activation of PI3K/Akt, upregulated ß-catenin and the oncogenes c-myc and cyclin D1 and promoted proliferation, migration and glucose uptake. Apelin receptor blockades reduced cancer cell proliferation along with a reduction in pancreatic tumor burden. These findings identify the apelin signaling pathway as a new actor for PDAC development and a novel therapeutic target for this incurable disease.

Apelin-driven endothelial cell migration sustains intestinal progenitor cells and tumor growth.

Stem and progenitor cells residing in the intestinal crypts drive the majority of colorectal cancers (CRCs), yet vascular contribution to this niche remains largely unexplored. VEGFA is a key driver of physiological and tumor angiogenesis. Accordingly, current anti-angiogenic cancer therapies target the VEGFA pathway. Here we report that in CRC expansion of the stem/progenitor pool in intestinal crypts requires VEGFA-independent growth and remodeling of blood vessels. Epithelial transformation induced expression of the endothelial peptide apelin, directs migration of distant venous endothelial cells towards progenitor niche vessels ensuring optimal perfusion. In the absence of apelin, loss of injury-inducible PROX1+ epithelial progenitors inhibited both incipient and advanced intestinal tumor growth. Our results establish fundamental principles for the reciprocal communication between vasculature and the intestinal progenitor niche and provide a mechanism for resistance to VEGFA-targeting drugs in CRCs.

Periprostatic Adipose Tissue Displays a Chronic Hypoxic State that Limits Its Expandability.

White adipose tissue accumulates at various sites throughout the body, some adipose tissue depots exist near organs whose function they influence in a paracrine manner. Prostate gland is surrounded by a poorly characterized adipose depot called periprostatic adipose tissue (PPAT), which plays emerging roles in prostate-related disorders. Unlike all other adipose depots, PPAT secretes proinflammatory cytokines even in lean individuals and does not increase in volume during obesity. These unique features remain unexplained because of the poor structural and functional characterization of this tissue. This study characterized the structural organization of PPAT in patients compared with abdominopelvic adipose tissue (APAT), an extraperitoneal adipose depot, the accumulation of which is correlated to body mass index. Confocal microscopy followed by three-dimensional reconstructions showed a sparse vascular network in PPAT when compared with that in APAT, suggesting that this tissue is hypoxic. Unbiased comparisons of PPAT and APAT transcriptomes found that most differentially expressed genes were related to the hypoxia response. High levels of the hypoxia-inducible factor 2α confirmed the presence of an adaptive response to hypoxia in PPAT. This chronic hypoxic state was associated with inflammation and fibrosis, which were not further up-regulated by obesity. This fibrosis and inflammation explain the failure of PPAT to expand in obesity and open new mechanistic avenues to explain its role in prostate-related disorders, including cancer.

Nuclear HMGB1 protects from nonalcoholic fatty liver disease through negative regulation of liver X receptor.

Dysregulations of lipid metabolism in the liver may trigger steatosis progression, leading to potentially severe clinical consequences such as nonalcoholic fatty liver diseases (NAFLDs). Molecular mechanisms underlying liver lipogenesis are very complex and fine-tuned by chromatin dynamics and multiple key transcription factors. Here, we demonstrate that the nuclear factor HMGB1 acts as a strong repressor of liver lipogenesis. Mice with liver-specific Hmgb1 deficiency display exacerbated liver steatosis, while Hmgb1-overexpressing mice exhibited a protection from fatty liver progression when subjected to nutritional stress. Global transcriptome and functional analysis revealed that the deletion of Hmgb1 gene enhances LXRα and PPARγ activity. HMGB1 repression is not mediated through nucleosome landscape reorganization but rather via a preferential DNA occupation in a region carrying genes regulated by LXRα and PPARγ. Together, these findings suggest that hepatocellular HMGB1 protects from liver steatosis development. HMGB1 may constitute a new attractive option to therapeutically target the LXRα-PPARγ axis during NAFLD.

Physical Activity, Body Mass Index, and Blood Progranulin in Older Adults: Cross-Sectional Associations in the MAPT Study.

Physical activity (PA) has been shown to moderate the negative effects of obesity on pro-inflammatory cytokines but its relationship with the adipokine progranulin (PGRN) remains poorly investigated. This study aimed to examine the cross-sectional main and interactive associations of body mass index (BMI) and PA level with circulating PGRN in older adults. Five-hundred and twelve participants aged 70 years and older involved in the Multidomain Alzheimer Preventive Trial (MAPT) study who underwent plasma PGRN measurements (ng/mL) were included. Self-reported PA levels were assessed using questionnaires. People were classified into 3 BMI categories: normal weight, overweight, or obesity. Further categorization using PA tertiles was used to define highly active, moderately active, and low active individuals. Multiple linear regressions were performed in order to test the associations of BMI, PA level, and their interaction with PGRN levels. Multiple linear regressions adjusted by age, sex, diabetes mellitus status, total cholesterol, creatinine level, and MAPT group demonstrated significant interactive associations of BMI status and continuous PA such that in people without obesity, higher PA levels were associated with lower PGRN concentrations, while an opposite pattern was found in individuals with obesity. In addition, continuous BMI was positively associated with circulating PGRN in highly active individuals but not in their less active peers. This cross-sectional study demonstrated reverse patterns in older adults with obesity compared to those without obesity regarding the relationships between PA and PGRN levels. Longitudinal and experimental investigations are required to understand the mechanisms that underlie the present findings. Clinical Trials Registration Number: NCT00672685.

Apelin expression deficiency in mice contributes to vascular stiffening by extracellular matrix remodeling of the aortic wall.

Numerous recent studies have shown that in the continuum of cardiovascular diseases, the measurement of arterial stiffness has powerful predictive value in cardiovascular risk and mortality and that this value is independent of other conventional risk factors, such as age, cholesterol levels, diabetes, smoking, or average blood pressure. Vascular stiffening is often the main cause of arterial hypertension (AHT), which is common in the presence of obesity. However, the mechanisms leading to vascular stiffening, as well as preventive factors, remain unclear. The aim of the present study was to investigate the consequences of apelin deficiency on the vascular stiffening and wall remodeling of aorta in mice. This factor freed by visceral adipose tissue, is known for its homeostasic role in lipid and vascular metabolisms, or again in inflammation. We compared the level of metabolic markers, inflammation of white adipose tissue (WAT), and aortic wall remodeling from functional and structural approaches in apelin-deficient and wild-type (WT) mice. Apelin-deficient mice were generated by knockout of the apelin gene (APL-KO). From 8 mice by groups, aortic stiffness was analyzed by pulse wave velocity measurements and by characterizations of collagen and elastic fibers. Mann-Whitney statistical test determined the significant data (p < 5%) between groups. The APL-KO mice developed inflammation, which was associated with significant remodeling of visceral WAT, such as neutrophil elastase and cathepsin S expressions. In vitro, cathepsin S activity was detected in conditioned medium prepared from adipose tissue of the APL-KO mice, and cathepsin S activity induced high fragmentations of elastic fiber of wild-type aorta, suggesting that the WAT secretome could play a major role in vascular stiffening. In vivo, remodeling of the extracellular matrix (ECM), such as collagen accumulation and elastolysis, was observed in the aortic walls of the APL-KO mice, with the latter associated with high cathepsin S activity. In addition, pulse wave velocity (PWV) and AHT were increased in the APL-KO mice. The latter could explain aortic wall remodeling in the APL-KO mice. The absence of apelin expression, particularly in WAT, modified the adipocyte secretome and facilitated remodeling of the ECM of the aortic wall. Thus, elastolysis of elastic fibers and collagen accumulation contributed to vascular stiffening and AHT. Therefore, apelin expression could be a major element to preserve vascular homeostasis.

Metabolic and cardiovascular adaptations to an 8-wk lifestyle weight loss intervention in younger and older obese men.

The number of older obese adults is increasing worldwide. Whether obese adults show similar health benefits in response to lifestyle interventions at different ages is unknown. The study enrolled 25 obese men (body mass index: 31-39 kg/m2) in two arms according to age (30-40 and 60-70 yr old). Participants underwent an 8-wk intervention with moderate calorie restriction (∼20% below individual energy requirements) and supervised endurance training resulting in ∼5% weight loss. Body composition was measured using dual energy X-ray absorptiometry. Insulin sensitivity was assessed during a hypersinsulinemic-euglycemic clamp. Cardiometabolic profile was derived from blood parameters. Subcutaneous fat and vastus lateralis muscle biopsies were used for ex vivo analyses. Two-way repeated-measure ANOVA and linear mixed models were used to evaluate the response to lifestyle intervention and comparison between the two groups. Fat mass was decreased and bone mass was preserved in the two groups after intervention. Muscle mass decreased significantly in older obese men. Cardiovascular risk (Framingham risk score, plasma triglyceride, and cholesterol) and insulin sensitivity were greatly improved to a similar extent in the two age groups after intervention. Changes in adipose tissue and skeletal muscle transcriptomes were marginal. Analysis of the differential response to the lifestyle intervention showed tenuous differences between age groups. These data suggest that lifestyle intervention combining calorie restriction and exercise shows similar beneficial effects on cardiometabolic risk and insulin sensitivity in younger and older obese men. However, attention must be paid to potential loss of muscle mass in response to weight loss in older obese men.NEW & NOTEWORTHY Rise in obesity and aging worldwide are major trends of critical importance in public health. This study addresses a current challenge in obesity management. Do older obese adults respond differently to a lifestyle intervention composed of moderate calorie restriction and supervised physical activity than younger ones? The main conclusion of the study is that older and younger obese men similarly benefit from the intervention in terms of cardiometabolic risk.

SHP2 drives inflammation-triggered insulin resistance by reshaping tissue macrophage populations.

Insulin resistance is a key event in type 2 diabetes onset and a major comorbidity of obesity. It results from a combination of fat excess-triggered defects, including lipotoxicity and metaflammation, but the causal mechanisms remain difficult to identify. Here, we report that hyperactivation of the tyrosine phosphatase SHP2 found in Noonan syndrome (NS) led to an unsuspected insulin resistance profile uncoupled from altered lipid management (for example, obesity or ectopic lipid deposits) in both patients and mice. Functional exploration of an NS mouse model revealed this insulin resistance phenotype correlated with constitutive inflammation of tissues involved in the regulation of glucose metabolism. Bone marrow transplantation and macrophage depletion improved glucose homeostasis and decreased metaflammation in the mice, highlighting a key role of macrophages. In-depth analysis of bone marrow-derived macrophages in vitro and liver macrophages showed that hyperactive SHP2 promoted a proinflammatory phenotype, modified resident macrophage homeostasis, and triggered monocyte infiltration. Consistent with a role of SHP2 in promoting inflammation-driven insulin resistance, pharmaceutical SHP2 inhibition in obese diabetic mice improved insulin sensitivity even better than conventional antidiabetic molecules by specifically reducing metaflammation and alleviating macrophage activation. Together, these results reveal that SHP2 hyperactivation leads to inflammation-triggered metabolic impairments and highlight the therapeutical potential of SHP2 inhibition to ameliorate insulin resistance.

The Chemokine Receptor CCR3 Is Potentially Involved in the Homing of Prostate Cancer Cells to Bone: Implication of Bone-Marrow Adipocytes.

Bone metastasis remains the most frequent and the deadliest complication of prostate cancer (PCa). Mechanisms leading to the homing of tumor cells to bone remain poorly characterized. Role of chemokines in providing navigational cues to migrating cancer cells bearing specific receptors is well established. Bone is an adipocyte-rich organ since 50 to 70% of the adult bone marrow (BM) volume comprise bone marrow adipocytes (BM-Ads), which are likely to produce chemokines within the bone microenvironment. Using in vitro migration assays, we demonstrated that soluble factors released by human primary BM-Ads are able to support the directed migration of PCa cells in a CCR3-dependent manner. In addition, we showed that CCL7, a chemokine previously involved in the CCR3-dependent migration of PCa cells outside of the prostate gland, is released by human BM-Ads. These effects are amplified by obesity and ageing, two clinical conditions known to promote aggressive and metastatic PCa. In human tumors, we found an enrichment of CCR3 in bone metastasis vs. primary tumors at mRNA levels using Oncomine microarray database. In addition, immunohistochemistry experiments demonstrated overexpression of CCR3 in bone versus visceral metastases. These results underline the potential importance of BM-Ads in the bone metastatic process and imply a CCR3/CCL7 axis whose pharmacological interest needs to be evaluated.

Antibody-based delivery of interleukin-9 to neovascular structures: Therapeutic evaluation in cancer and arthritis.

Interleukin-9 is a cytokine with multiple functions, including the ability to activate group 2 innate lymphoid cells, which has been postulated to be therapeutically active in mouse models of arthritis. Similarly, interleukin-9 has been suggested to play an important role in tumor immunity. Here, we describe the cloning, expression, and characterization of three fusion proteins based on murine interleukin-9 and the F8 antibody, specific to the alternatively spliced EDA domain of fibronectin. EDA is strongly expressed in cancer and in various arthritic conditions, while being undetectable in the majority of healthy organs. Interleukin-9-based fusion proteins with an irrelevant antibody specific to hen egg lysozyme served as negative control in our study. The fusion proteins were characterized by quantitative biodistribution analysis in tumor-bearing mice using radioiodinated protein preparations. The highest tumor uptake and best tumor:organ ratios were observed for a format, in which the interleukin-9 moiety was flanked by two units of the F8 antibody in single-chain Fv format. Biological activity of interleukin-9 was retained when the payload was fused to antibodies. However, the targeted delivery of interleukin-9 to the disease site resulted in a modest anti-tumor activity in three different murine models of cancer (K1735M2, CT26, and F9), while no therapeutic benefit was observed in a collagen induced model of arthritis. Collectively, these results confirm the possibility to deliver interleukin-9 to the site of disease but cast doubts about the alleged therapeutic activity of this cytokine in cancer and arthritis, which has been postulated in previous publications.

Obesity of mice lacking VAP-1/SSAO by Aoc3 gene deletion is reproduced in mice expressing a mutated vascular adhesion protein-1 (VAP-1) devoid of amine oxidase activity.

The product of Aoc3 gene is known as vascular adhesion protein-1 (VAP-1), a glycoprotein contributing to leukocyte extravasation and exhibiting semicarbazide-sensitive amine oxidase activity (SSAO). Regarding the immune functions of VAP-1/SSAO, it is known that mice bearing Aoc3 gene knock-out (AOC3KO) exhibit defects in leukocyte migration similar to those of mice expressing a mutated VAP-1 lacking functional SSAO activity (knock-in, AOC3KI). However, it has not been reported whether these models differ regarding other disturbances. Thus, we further compared endocrine-metabolic phenotypes of AOC3KO and AOC3KI mice to their respective control. Special attention was paid on adiposity, glucose and lipid handling, since VAP-1/SSAO is highly expressed in adipose tissue (AT). In both mouse lines, no tissue SSAO activity was found, while Aoc3 mRNA was absent in AOC3KO only. Although food consumption was unchanged, both AOC3KO and AOC3KI mice were heavier and fatter than their respective controls. Other alterations commonly found in adipocytes from both lines were loss of benzylamine insulin-like action with unchanged insulin lipogenic responsiveness and adiponectin expression. A similar downregulation of inflammatory markers (CD45, IL6) was found in AT. Glucose handling and liver mass remained unchanged, while circulating lipid profile was distinctly altered, with increased cholesterol in AOC3KO only. These results suggest that the lack of oxidase activity found in AOC3KI is sufficient to reproduce the metabolic disturbances observed in AOC3KO mice, save those related with cholesterol transport. Modulation of SSAO activity therefore constitutes a potential target for the treatment of cardiometabolic diseases, especially obesity when complicated by low-grade inflammation.

Postoperative Delirium is a Risk Factor of Poor Evolution Three Years After Cardiac Surgery: An Observational Cohort Study.

BACKGROUND: After cardiac surgery, postoperative delirium (POD) is common and is associated with long-term changes in cognitive function. Impact on health-related quality of life (QOL) and long-term dependence are not well known. This aim of this study is to evaluate the role of POD in poor evolution at three years after surgery including poor QOL and dependence and mortality. PATIENTS AND METHODS: We enrolled and followed 173 patients 60 years of age or older who were planning to undergo cardiac surgery with cardiopulmonary bypass. The primary composite outcome was death of any causes, or patients with either a loss of QOL (evaluated with of EuroQuol verbal 5D EQ5D less than 50), or a loss of two points on the instrumental activities of daily living occurring three years after surgery. POD was diagnosed with the use of Confusion Assessment Method. Multivariate logistic regression was performed. RESULTS: At three years, 74 patients (42.8%) had a poor evolution. Independent risk factors in poor patient evolution were sex (female gender; OR: 3.6; 95%CI: 1.45-8.7; p=0.006), metabolic status (diabetic patients; OR: 4; 95%CI: 1.6-10.2; p=0.002), Euroscore 2 (Euroscore 2 >1.5; OR: 5.2; 95%CI: 1.7-15.4; p=0.003) and POD (OR: 3.3; 95%CI 1.4-7.8; p=0.006). Coronary disease was protective (OR: 0.3; 95%CI: 0.14-0.71; p=0.006). CONCLUSION: After cardiac surgery, POD significantly altered patient evolution and increased risk of dependence and loss of QOL.

Adipocyte Fatty Acid Transfer Supports Megakaryocyte Maturation.

Megakaryocytes (MKs) come from a complex process of hematopoietic progenitor maturation within the bone marrow that gives rise to de novo circulating platelets. Bone marrow microenvironment contains a large number of adipocytes with a still ill-defined role. This study aims to analyze the influence of adipocytes and increased medullar adiposity in megakaryopoiesis. An in vivo increased medullar adiposity in mice caused by high-fat-diet-induced obesity is associated to an enhanced MK maturation and proplatelet formation. In vitro co-culture of adipocytes with bone marrow hematopoietic progenitors shows that delipidation of adipocytes directly supports MK maturation by enhancing polyploidization, amplifying the demarcation membrane system, and accelerating proplatelet formation. This direct crosstalk between adipocytes and MKs occurs through adipocyte fatty acid transfer to MKs involving CD36 to reinforce megakaryocytic maturation. Thus, these findings unveil an influence of adiposity on MK homeostasis based on a dialogue between adipocytes and MKs.

Adipocyte extracellular vesicles carry enzymes and fatty acids that stimulate mitochondrial metabolism and remodeling in tumor cells.

Extracellular vesicles are emerging key actors in adipocyte communication. Notably, small extracellular vesicles shed by adipocytes stimulate fatty acid oxidation and migration in melanoma cells and these effects are enhanced in obesity. However, the vesicular actors and cellular processes involved remain largely unknown. Here, we elucidate the mechanisms linking adipocyte extracellular vesicles to metabolic remodeling and cell migration. We show that adipocyte vesicles stimulate melanoma fatty acid oxidation by providing both enzymes and substrates. In obesity, the heightened effect of extracellular vesicles depends on increased transport of fatty acids, not fatty acid oxidation-related enzymes. These fatty acids, stored within lipid droplets in cancer cells, drive fatty acid oxidation upon being released by lipophagy. This increase in mitochondrial activity redistributes mitochondria to membrane protrusions of migrating cells, which is necessary to increase cell migration in the presence of adipocyte vesicles. Our results provide key insights into the role of extracellular vesicles in the metabolic cooperation that takes place between adipocytes and tumors with particular relevance to obesity.

Human Bone Marrow Is Comprised of Adipocytes with Specific Lipid Metabolism.

Under caloric restriction, bone marrow adipocytes (BM-Ads) do not decrease in size compared to white adipocytes, suggesting they harbor unique metabolic properties. We compare human primary BM-Ads with paired subcutaneous adipocytes (SC-Ads) using proteomic and lipidomic approaches. We find that, although SC-Ads and BM-Ads share similar morphological features, they possess distinct lipid metabolism. Although BM-Ad shows enrichment in proteins involved in cholesterol metabolism, correlating with increased free cholesterol content, proteins involved in lipolysis were downregulated. In particular, monoacylglycerol lipase expression is strongly reduced in BM-Ads, leading to monoacylglycerol accumulation. Consequently, basal and induced lipolytic responses are absent in BM-Ads, affirming their differences in metabolic fitness upon caloric restriction. These specific metabolic features are not recapitulated in vitro using common protocols to differentiate bone marrow mesenchymal stem cells. Thus, contrary to classical SC-Ads, BM-Ads display a specific lipid metabolism, as they are devoid of lipolytic activity and exhibit a cholesterol-orientated metabolism.