Inguinal Subcutaneous Adipose Tissue Research Articles

A long-term high-fat diet induces differential gene expression changes in spatially distinct adipose tissue of male mice.

The accumulation of visceral adipose tissue (VAT) is strongly associated with cardiovascular disease and diabetes. In contrast, individuals with increased subcutaneous adipose tissue (SAT) without corresponding increases in VAT are associated with a metabolic healthy obese phenotype. These observations implicate dysfunctional VAT as a driver of disease processes, warranting investigation into obesity-induced alterations of distinct adipose depots. To determine the effects of obesity on adipose gene expression, male mice (n = 4) were fed a high-fat diet to induce obesity or a normal laboratory diet (lean controls) for 12-14 mo. Mesenteric VAT and inguinal SAT were isolated for bulk RNA sequencing. AT from lean controls served as a reference to obesity-induced changes. The long-term high-fat diet induced the expression of 169 and 814 unique genes in SAT and VAT, respectively. SAT from obese mice exhibited 308 differentially expressed genes (164 upregulated and 144 downregulated). VAT from obese mice exhibited 690 differentially expressed genes (262 genes upregulated and 428 downregulated). KEGG pathway and GO analyses revealed that metabolic pathways were upregulated in SAT versus downregulated in VAT while inflammatory signaling was upregulated in VAT. We next determined common genes that were differentially regulated between SAT and VAT in response to obesity and identified four genes that exhibited this profile: elovl6 and kcnj15 were upregulated in SAT/downregulated in VAT while trdn and hspb7 were downregulated in SAT/upregulated in VAT. We propose that these genes in particular should be further pursued to determine their roles in SAT versus VAT with respect to obesity.NEW & NOTEWORTHY A long-term high-fat diet induced the expression of more than 980 unique genes across subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT). The high-fat diet also induced the differential expression of nearly 1,000 AT genes. We identified four genes that were oppositely expressed in SAT versus VAT in response to the high-fat diet and propose that these genes in particular may serve as promising targets aimed at resolving VAT dysfunction in obesity.

Differential effects of melatonin on adipose tissues under normoestrogenic and estrogen-deficient conditions in rats

In post-menopause, oxidative stress due to the decline of natural antioxidants increases the susceptibility to metabolic syndromes (MetS). Estrogen and melatonin (MEL) share antioxidant properties; however, few studies have reported the impact of estrogen deficiency and MEL treatment on morphology, redox status, and antioxidant defense capacity of diverse adipose tissue (AT) subtypes. To investigate this issue, MEL was administered to ovariectomized (OVX) rats and sham-operated rats for 16 weeks (10 mg/kg). The adipocyte morphology, oxidative stress parameters and antioxidant enzyme activity were evaluated in the visceral retroperitoneal adipose tissue (rVAT), subcutaneous inguinal adipose tissue (iSAT) and brown adipose tissue (BAT). In OVX rats, MEL treatment suppressed rVAT hypertrophy and increased the prevalence of small adipocytes in iSAT, suggesting a better lipid distribution among ATs. MEL treatment increased glutathione reductase and glucose-6-phosphate dehydrogenase activity in iSAT; therefore, restored the glutathione level. In rVAT, MEL increased glutathione peroxidase and glutathione reductase activity. MEL minimized the risks for the development of metabolic abnormalities due to estrogen deficiency. However, under normoestrogenic condition, MEL decreased plasma estradiol levels and uterine mass, raising the concerning of its effect on reproductive functions.

The Effects of Intermittent Fasting on the Size and Number of Subcutaneus Adipocytes in Obese Mouse Models

Highlights:1. This study helps bridge the gap between systemic effects and tissue-level changes, providing a deeper understanding of how histological analysis can be used to explore the effects of intermittent fasting on adipocytes and body weight regulation.2. This study contributes to obesity management through lifestyle modification, specifically intermittent fasting, by focusing on histological changes in adipose tissue. Abstract The accumulation of adipose tissue can have deleterious effects and lead to obesity. Intermittent fasting (IF), an approach that involves time-restricted eating, has gained popularity as an obesity treatment option because it enhances insulin sensitivity and promotes beneficial changes in glucose metabolism. This study used a time-restricted meal intake (TRM) approach to assess the effects of IF on the histological features of obese mouse models' subcutaneous inguinal adipose tissue. The investigation employed an in vivo experimental posttest-only control group design. Twenty male mice were divided into four groups: a normal control group, an obese control group, a TRM group with a high-fat diet (TRM-HF), and a TRM group with a standard diet (TRM-S). The TRM treatment was administered for fourteen days, with a fasting window from 4 p.m. to 8 a.m. The pre- and post-treatment weight analyses were conducted using the paired t-test for normally distributed data and the Wilcoxon test for non-normally distributed data (p<0.05). One-way analysis of variance (ANOVA) was employed for unpaired data on the post-treatment weight. Per field of view, there were an average of 120,500 cells (49,700–136,200) in the normal control group, 68,380±9,194 cells in the obese control group, 70,860±11,029 cells in the TRM-HF group, and 79,360±5,112 cells in the TRM-S group. The average cell sizes (μm3) were 56,730.142±19,273.257 in the normal control group, 138,934.331±27,670.558 in the obese control group, 106,827.767±20,580.501 in the TRM-HF group, and 68,689.114±8,219.727 in the TRM-S group. The number of cells in each group did not differ significantly, but there were significant variations in cell size. The mice receiving TRM treatment did not exhibit substantial body weight changes, whereas the obese control group showed a significant body weight increase. In conclusion, TRM has an effect on cell size but does not affect the quantity of adipocytes in subcutaneous inguinal fat tissue.

Nanofat Accelerates and Improves the Vascularization, Lymphatic Drainage and Healing of Full-Thickness Murine Skin Wounds.

The treatment of wounds using the body's own resources is a promising approach to support the physiological regenerative process. To advance this concept, we evaluated the effect of nanofat (NF) on wound healing. For this purpose, full-thickness skin defects were created in dorsal skinfold chambers of wild-type mice. These defects were filled with NF generated from the inguinal subcutaneous adipose tissue of green fluorescent protein (GFP)+ donor mice, which was stabilized using platelet-rich plasma (PRP). Empty wounds and wounds solely filled with PRP served as controls. Wound closure, vascularization and formation of granulation tissue were repeatedly analyzed using stereomicroscopy, intravital fluorescence microscopy, histology and immunohistochemistry over an observation period of 14 days. PRP + NF-treated wounds exhibited accelerated vascularization and wound closure when compared to controls. This was primarily due to the fact that the grafted NF contained a substantial fraction of viable GFP+ vascular and lymph vessel fragments, which interconnected with the GFP- vessels of the host tissue. Moreover, the switch from inflammatory M1- to regenerative M2-polarized macrophages was promoted in PRP + NF-treated wounds. These findings indicate that NF markedly accelerates and improves the wound healing process and, thus, represents a promising autologous product for future wound management.

TMT Labeling Reveals the Effects of Exercises on the Proteomic Characteristics of the Subcutaneous Adipose Tissue of Growing High-Fat-Diet-Fed Rats.

Aim: Growing period is an important period for fat remodeling. High-fat diet and exercise are reasons for adipose tissue (AT) remodeling, but existing evidence is not enough. Therefore, the effects of moderate-intensity continuous training (MICT) and high-intensity interval training (HIIT) on the proteomic characteristics of the subcutaneous AT of growing rats on normal diet or high-fat diet (HFD) were determined. Methods: Four-week-old male Sprague-Dawley rats (n = 48) were subdivided into six groups: normal diet control group, normal diet-MICT group, normal diet-HIIT group, HFD control group, HFD-MICT group, and HFD-HIIT group. Rats in the training group ran on a treadmill 5 days a week for 8 weeks (MICT: 50 min at 60-70% VO2max intensity; HIIT: 7 min of warm-up and recovery at 70% VO2max intensity, 6 sets of 3 min of 30% VO2max followed by 3 min 90% VO2max). Following physical assessment, inguinal subcutaneous adipose tissue (sWAT) was collected for proteome analysis using tandem mass tag labeling. Results: MICT and HIIT attenuated body fat mass and lean body mass but did not affect weight gain. Proteomics revealed the impact of exercise on ribosome, spliceosome, and the pentose phosphate pathway. However, the effect was reversed on HFD and normal diet. The differentially expressed proteins (DEPs) affected by MICT were related to oxygen transport, ribosome, and spliceosome. In comparison, the DEPs affected by HIIT were related to oxygen transport, mitochondrial electron transport, and mitochondrion protein. In HFD, HIIT was more likely to cause changes in immune proteins than MICT. However, exercise did not seem to reverse the protein effects of HFD. Conclusion: The exercise stress response in the growing period was stronger but increased the energy metabolism and metabolism. MICT and HIIT can reduce fat, increase muscle percentage, and improve maximum oxygen uptake in rats fed with HFD. However, in rats with normal diet, MICT and HIIT triggered more immune responses of sWAT, especially HIIT. In addition, spliceosomes may be the key factors in AT remodeling triggered by exercise and diet.

Time Course of Fat Accumulation in Various Organs of Young OLETF Rats

PurposeExcessive fat accumulation induces chronic inflammation in the white adipose tissue (WAT), which leads and accelerates to obesity‐related health problems such as glucose intolerance, insulin resistance, and dyslipidemia. Determination of the time course of fat accumulation in obesity might allow identification of target tissues to treat the health problems in each stage. The purpose of the present study was to investigate the relationship between inflammatory changes and time course of fat accumulation in various organs.MethodsMale Otsuka Long‐Evans Tokushima fatty (OLETF) rats were used as spontaneous obesity animals. All rats were fed a standard diet containing 55.3% carbohydrate and water ad libitum. From 4‐ to 20‐week‐old, blood samples from vena cava and samples of the epididymal adipose tissue, retroperitoneal adipose tissue, mesenteric adipose tissue, and inguinal subcutaneous adipose tissue as WAT, brown adipose tissue (BAT) from interscapular region, liver, pancreas, and plantaris muscles were analyzed histologically every 2 weeks. Collected serum was used to detect triglyceride (TG) and free fatty acid (FFA) concentrations. Tissue samples were sliced and stained with hematoxylin and eosin or oil red O for histological observation.ResultsBody weight in OLETF rats increased linearly from 4‐ to 20‐week‐old. Although the wet weight and adipocyte diameter of the epididymal adipose tissue also increased throughout the experimental period, the values of inguinal subcutaneous adipose tissue increased by 10‐week‐old and were almost the same after 10‐week‐old. The liver TG levels increased from 4‐ to 8‐week‐old. However, hypertriglyceridemia, BAT whitening, and hepatic steatosis were not observed at this period. The serum TG levels increased after 10‐week‐old. Additionally, the number of whitened unilocular adipocytes with a diameter >10 μm in the BAT increased after 10‐week‐old and oil red O positive lipid droplets were usually observed in the liver at this period. Not only the serum TG levels but also the serum FFA levels increased after 18‐week‐old. Additionally, enlarged adipocytes with a diameter >130 μm and crown like structure indicating inflammatory change were often observed in the epididymal adipose tissue and retroperitoneal adipose tissue, not in mesenteric adipose tissue and inguinal subcutaneous adipose tissue, at this period. Fatty pancreas and intramuscular fat deposition were not observed throughout the experimental period.ConclusionAdipocyte hypertrophy in the visceral WAT and fatty liver could precede dyslipidemia in young OLETF rats and continuous obesity subsequently proceed to ectopic fat accumulation including BAT whitening and chronic inflammation in the epididymal adipose tissue and retroperitoneal adipose tissue with higher FFA levels. Hyperphagia mainly on carbohydrate could contribute to ectopic fat accumulation in the liver and BAT whitening, not to fatty pancreas and intramuscular fat deposition in young stage.

Distinct Properties of Adipose Stem Cell Subpopulations Determine Fat Depot‐Specific Characteristics

In mammals, white adipose tissues are largely divided into visceral epididymal adipose tissue (EAT) and subcutaneous inguinal adipose tissue (IAT) with distinct metabolic properties. Although emerging evidence suggests that subpopulations of adipose stem cells (ASCs) would be important to explain fat depot differences, ASCs of two fat depots have not been comparatively investigated. Here, we characterized heterogeneous ASCs and examined the effects of intrinsic and tissue micro‐environmental factors on distinct ASC features. We demonstrated that ASC subpopulations in EAT and IAT exhibited different molecular features with three adipogenic stages. ASC transplantation experiments revealed that intrinsic ASC features primarily determined their adipogenic potential. Upon obesogenic stimuli, EAT specific SDC1+ ASCs promoted fibrotic remodeling, whereas IAT‐specific CXCL14+ ASCs suppressed macrophage infiltration. Moreover, IAT‐specific BST2high ASCs exhibited a high potential to become beige adipocytes. Collectively, our data broaden the understanding of ASCs with new insights into the origin of white fat depot differences.

Maternal Exercise Programs Glucose and Lipid Metabolism and Modulates Hepatic miRNAs in Adult Male Offspring.

Detrimental exposures in mothers are recognized as risk factors for the development of metabolic dysfunction in offspring. In contrast, maternal exercise has been reported to be an effective strategy to maintain offspring health. However, the mechanisms underlying the protective effects of maternal exercise on adult offspring metabolic homeostasis are largely unclear. This study aims to investigate whether maternal exercise before and during pregnancy could combat the adverse effects of maternal high-fat diet (HFD) on metabolism in 24-week-old male offspring and to explore the role of miRNAs in mediating the effects. Female C57BL/6 mice were fed with either control diet or HFD 3-week prior to breeding and throughout pregnancy and lactation, among whom half of the HFD-fed mice were submitted to voluntary wheel running training 3-week before and during pregnancy. Male offspring were sedentary and fed with a control diet from weaning to 24 weeks. Body weight, the content of inguinal subcutaneous adipose tissue and perirenal visceral adipose tissue, glucose tolerance, and serum insulin and lipids in offspring were analyzed. Hepatic tissues were collected for transcriptome and miRNA sequencing and reverse transcription-quantitative polymerase chain reaction validation. The results showed that maternal HFD resulted in significant glucose intolerance, insulin resistance, and dyslipidemia in adult offspring, which were negated by maternal exercise. Transcriptome sequencing showed that maternal exercise reversed perinatal HFD-regulated genes in adult offspring, which were enriched in glucose and lipid metabolic-related signaling pathways. At the same time, maternal exercise significantly rescued the changes in the expression levels of 3 hepatic miRNAs in adult offspring, and their target genes were involved in the regulation of cholesterol biosynthesis and epigenetic modification, which may play an important role in mediating the intergenerational metabolic regulation of exercise. Overall, our research pioneered the role of miRNAs in mediating the programming effects of maternal exercise on adult offspring metabolism, which might provide novel insight into the prevention and treatment of metabolic disorders in early life.

Distinct properties of adipose stem cell subpopulations determine fat depot-specific characteristics

In mammals, white adipose tissues are largely divided into visceral epididymal adipose tissue (EAT) and subcutaneous inguinal adipose tissue (IAT) with distinct metabolic properties. Although emerging evidence suggests that subpopulations of adipose stem cells (ASCs) would be important to explain fat depot differences, ASCs of two fat depots have not been comparatively investigated. Here, we characterized heterogeneous ASCs and examined the effects of intrinsic and tissue micro-environmental factors on distinct ASC features. We demonstrated that ASC subpopulations in EAT and IAT exhibited different molecular features with three adipogenic stages. ASC transplantation experiments revealed that intrinsic ASC features primarily determined their adipogenic potential. Upon obesogenic stimuli, EAT-specific SDC1+ ASCs promoted fibrotic remodeling, whereas IAT-specific CXCL14+ ASCs suppressed macrophage infiltration. Moreover, IAT-specific BST2high ASCs exhibited a high potential to become beige adipocytes. Collectively, our data broaden the understanding of ASCs with new insights into the origin of white fat depot differences.

Adipose expression of CREB3L3 modulates body weight during obesity

We found the hepatic transcription factor Cyclic-AMP Responsive Element Binding Protein 3-like-3 (CREB3L3) to be expressed in adipose tissue, and selectively downregulated in the more metabolically protective subcutaneous adipose tissue in obese mice and humans. We sought to elucidate the specific role of this factor in adipose biology. CREB3L3 fat-specific knockout mice were fed a high-fat diet to induce obesity and metabolic dysfunction. Additionally, we injected a flip-excision adeno-associated virus directly into the subcutaneous inguinal adipose tissue of Adiponectin-Cre mice to create a depot-specific overexpression model for further assessment. Fat-specific ablation of CREB3L3 enhanced weight gain and insulin resistance following high-fat feeding, as fat-specific knockout mice expended less energy and possessed more inflammatory adipose tissue. Conversely, inguinal fat CREB3L3 overexpression deterred diet-induced obesity and ameliorated metabolic dysfunction. Together, this study highlights the relevance of CREB3L3 in obese adipose tissue and demonstrates its role as a powerful body weight modulator.

Transcriptional Landscaping Identifies a Beige Adipocyte Depot in the Newborn Mouse.

The present study sought to identify gene networks that are hallmarks of the developing inguinal subcutaneous adipose tissue (iWAT) and the interscapular brown adipose tissue (BAT) in the mouse. RNA profiling revealed that the iWAT of postnatal (P) day 6 mice expressed thermogenic and lipid catabolism transcripts, along with the abundance of transcripts associated with the beige adipogenesis program. This was an unexpected finding, as thermogenic BAT was believed to be the only site of nonshivering thermogenesis in the young mouse. However, the transcriptional landscape of BAT in P6 mice suggests that it is still undergoing differentiation and maturation, and that the iWAT temporally adopts thermogenic and lipolytic potential. Moreover, P6 iWAT and adult (P56) BAT were similar in their expression of immune gene networks, but P6 iWAT was unique in the abundant expression of antimicrobial proteins and virus entry factors, including a possible receptor for SARS-CoV-2. In summary, postnatal iWAT development is associated with a metabolic shift from thermogenesis and lipolysis towards fat storage. However, transcripts of beige-inducing signal pathways including β-adrenergic receptors and interleukin-4 signaling were underrepresented in young iWAT, suggesting that the signals for thermogenic fat differentiation may be different in early postnatal life and in adulthood.

Characterization of Adipose Stem Cells through Single Cell RNA‐sequencing Analysis

White adipose tissue is largely divided into visceral epididymal adipose tissue (EAT) and subcutaneous inguinal adipose tissue (IAT). EAT and IAT appear to have their unique roles and different characteristics in wholebody energy homeostasis. Accumulating evidence suggests that adipose stem cells (ASCs) would be a crucial regulator to mediate fat depot-specific characteristics. Although ASC subpopulations have been recently reported, key distinct features of ASCs from two fat depots, EAT and IAT, are largely elusive. In this study, we performed single-cell RNA sequencing analysis to investigate distinct properties of each ASC. UMAP plot of ASCs revealed common and depot-specific gene expression. Trajectory analysis revealed that ASCs from two fat depots were largely divided into 3 stages. Uncommitted stage (Stage I) ASCs were highly proliferative and multipotent cells. Committed stage (Stage ll) and preadipocytes stage (Stage lll) ASCs exhibited high adipogenic potentials. Proportions of Stage II and III ASCs were enriched in EAT, while Stage I ASCs were the major subtype in IAT. Collectively, these results suggest that ASC heterogeneity would explain hierarchy of ASC that mediate different features in EAT and IAT for adipogenesis.

Alleviation of salt-induced exacerbation of cardiac, renal, and visceral fat pathology in rats with metabolic syndrome by surgical removal of subcutaneous fat

ObjectivesEvidence suggests that visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) should be considered as distinct types of white fat. Although VAT plays a key role in metabolic syndrome (MetS), the role of subcutaneous adipose tissue (SAT) has been unclear. DahlS.Z-Leprfa/Leprfa (DS/obese) rats, an animal model of MetS, develop adipocyte hypertrophy and inflammation to similar extents in SAT and VAT. We have now investigated the effects of salt loading and SAT removal on cardiac, renal, and VAT pathology in DS/obese rats.MethodsDS/obese rats were subjected to surgical removal of inguinal SAT or sham surgery at 8 weeks of age. They were provided with a 0.3% NaCl solution as drinking water or water alone for 4 weeks from 9 weeks of age.ResultsSalt loading exacerbated hypertension, insulin resistance, as well as left ventricular (LV) hypertrophy, inflammation, fibrosis, and diastolic dysfunction in DS/obese rats. It also reduced both SAT and VAT mass but aggravated inflammation only in VAT. Although SAT removal did not affect LV hypertrophy in salt-loaded DS/obese rats, it attenuated hypertension, insulin resistance, and LV injury as well as restored fat mass and alleviated inflammation and the downregulation of adiponectin gene expression in VAT. In addition, whereas salt loading worsened renal injury as well as upregulated the expression of renin–angiotensin-aldosterone system-related genes in the kidney, these effects were suppressed by removal of SAT.ConclusionsSAT removal attenuated salt-induced exacerbation of MetS and LV and renal pathology in DS/obese rats. These beneficial effects of SAT removal are likely attributable, at least in part, to inhibition of both VAT and systemic inflammation.

1946-P: Dapagliflozin Suppresses Adipose Fatty Acid Accumulation in Rats Fed on High-Fat Diet but Not on Normal Chow

SGLT2 inhibitor (SGLT2i) exerts not only on enhancement of glucose excretion but also on improvement of serum lipid profile via increase in lipid utilization as an energy source instead of glucose. That is, SGLT2i might affect the lipid storage. In the present study, we evaluated the effects of dapagliflozin (DAPA) on weight and fatty acid contents of adipose tissues. At 8 weeks of age, male rats were divided into four groups: (a) NC (n=6) fed on a standard laboratory chow (LC); (b) NC+DAPA (n=6) on the LC with 1 mg/kg/day DAPA administration; (c) HFD (n=6) on a high-fat diet (HF); (d) HFD+DAPA (n=6) on the HF with 1 mg/kg/day DAPA administration. After 7 weeks, we weighed epididymal (EPI), mesenteric (MES), retroperitoneal (RET), and inguinal subcutaneous (SUB) adipose tissues, and measured 21 adipose fatty-acid contents (μmol/g BW). As a result, adipose tissue weight in the examined four sites in HFD rats were significantly higher (P<0.01) than those, respectively, in NC rats. No significant differences were observed in weights of any adipose tissues or in their contents of fatty acids between the NC and NC+DAPA rats. In contrast, MES weight in HFD+DAPA rats was significantly lower (P<0.05) than that in the HFD rats. Contents of 1, 3, 2 and 1 saturated fatty acids (SFAs) in EPI, MES, RET and SUB, respectively, in the HFD+DAPA rats were significantly lower (P<0.05) than those, respectively, in the HFD rats. Contents of 3 and 2 monounsaturated fatty acids in MES and SUB, respectively, and contents of 2 and 1 polyunsaturated fatty acids in MES and RET, respectively, in the HFD+DAPA rats were significantly lower (P<0.05) than those, respectively, in the HFD rats. These results suggest that DAPA could reduce some SFAs in any adipose tissues in HF-fed rats, but not in LC-fed rats. The reduction might sometimes arise without concomitant decrease in tissue weight. In addition, DAPA might suppress accumulation of a wide range of fatty acids in MES of HF-fed rats in particular, probably due to their consumption. Disclosure D. Sato: None. J. Amarume: None. M. Yano: None. Y. Umehara: None. M. Kusunoki: Other Relationship; Self; Kaken Pharmaceutical Co., Ltd., Ono Pharmaceutical Co., Ltd., Taisho Pharmaceutical Co., Ltd. Z. Feng: None. T. Nakamura: Research Support; Self; AstraZeneca K.K., Ono Pharmaceutical Co., Ltd. Funding Ono Pharmaceutical Co., Ltd.; AstraZeneca K.K.

Adipocyte-Derived Exosomes Increase the Growth of Breast Tumors in Obese Mice

Abstract Objectives 1) We assessed whether the secretion and cargos of exosomes are distinct in obese and lean mice. 2) We assessed whether adipocyte-derived exosomes increase the size of tumors of the mammary gland in mice. Methods Female tamoxifen-inducible adipose-specific TSG101 conditional knockout mice (TSG KO) were initiated on an AIN-93G-based 50% high fat (HFD) or 15.8% low fat (LFD) at age 3 weeks and continued on these diets for 10 weeks. At age 4 weeks, mice were injected with tamoxifen or vehicle (TSG+), followed by the injection of 1.8*106 murine breast adenocarcinoma E0771 cells into the mammary gland fat pad 10 days after tamoxifen injection. Loss and gain of TSG101 and Cre, respectively, in inguinal subcutaneous adipose tissue were confirmed by RT-qPCR. Tumors were excised and their size was assessed using a caliper and by weight. H&E staining was used to assess the tumor type. Exosomes were isolated from plasma by an ExoQuick Plasma Prep kit, authenticated by western blot analysis and counted using a Nanosight tracker. The statistical analysis was performed by one-way ANOVA and Least Significant Differences posthoc test using a significance level of P < 0.05. Results HFD led to an increase in breast tumor size, which was diminished in adipose-specific TSG101 KO fed HFD: The size of mammary tumors in TSG+ fed HFD increased by 198% compared to TSG+ fed LFD (P < 0.05), whereas tumor size decreased by 58% TSG101 KO fed HFD compared to TSG+ fed HFD (P < 0.05). HFD led to an increase in adipose-derived exosomes that was reduced by TSG101 KO: The expression of ASC-1, a marker for adipose-derived exosomes in plasma exosomes increased by 58% in TSG+ fed HFD compared to TSG+ fed LFD, and decreased by 29% in TSG KO fed HFD compared to TSG+ fed HFD (P < 0.05). Plasma exosomes tested positive for exosome markers, CD63, CD81 and HSP70 and negative for a non-exosome marker, calnexin. H&E staining suggested that the tumors observed in all mice were adenocarcinomas. Tumor numbers were not affected by TSG101 and diet. Conclusions HFD increases the growth of mammary tumors in mice and the diet effect depends on exosome biogenesis in adipose tissue. We speculate that adipose-derived exosomes promote tumor growth but not metastasis. Funding Sources NIFA, NIH, and USDA. J.Z. is a consultant for PureTech Health.

Histology of skeletal muscle reconstructed by means of the implantation of autologous adipose tissue: an experimental study.

The purpose of this study was to determine the histological characteristics of a skeletal muscle reconstructed by means of the implantation of autologous adipose tissue following an experimentally-induced volumetric muscle loss. A cylindrical piece in the belly of the rat anterior tibial muscle was removed. In the hole, inguinal subcutaneous adipose tissue of the same rat was grafted. Animals were sacrificed 7, 14, 21, 28 and 60 days posttransplantation. Histological, histochemical, immunohistochemical and morphometric techniques were used. At all times analyzed, the regenerative muscle fibers formed from the edges of the muscle tissue showed histological, histochemical and immunohistochemical differences in comparison with the control group. These differences are related to delays in the maturation process and are related to problems in reinnervation and disorientation of muscle fibers. The stains for MyoD and desmin showed that some myoblasts and myotubes seem to derive from the transplanted adipose tissue. After 60 days, the transplant area was 20% occupied by fibrosis and by 80% skeletal muscle. However, the neo-muscle was chaotically organized showing muscle fiber disorientation and centronucleated fibers with irregular shape and size. Our results support the hypothesis that, at least from a morphological point of view, autologous adipose tissue transplantation favors reconstruction following a volumetric loss of skeletal muscle by combining the inherent regenerative response of the organ itself and the myogenic differentiation of the stem cells present in the adipose tissue. However, in our study, the formed neo-muscle exhibited histological differences in comparison with the normal skeletal muscle.

Interleukin-11 regulates the fate of adipose-derived mesenchymal stem cells via STAT3 signalling pathways.

ObjectiveAdipose‐derived mesenchymal stem cells (ADSCs) offer great promise as cell therapy for ischaemic diseases. Due to their poor survival in the ischaemic environment, the therapeutic efficacy of ADSCs is still relatively low. Interleukin‐11 (IL‐11) has been shown to play a key role in promoting cell proliferation and protecting cells from oxidative stress injury. The aim of this study was to determine whether IL‐11 could improve therapeutic efficacy of ADSCs in ischaemic diseases.Methods and ResultsADSCs were prepared from inguinal subcutaneous adipose tissue and exposed to hypoxic environment. The protein expression of IL‐11 was decreased after hypoxic treatment. In addition, ADSCs viability was increased after IL‐11 treatment under hypoxia. Moreover, IL‐11 enhanced ADSCs viability in a dose‐dependent manner under normoxia. Importantly, IL‐11 promoted ADSCs proliferation and migration and protected ADSCs against hydrogen peroxide‐induced cellular death. Notably, IL‐11 enhanced ADSCs proliferation and migration, also promoted cell survival and apoptosis resistance by STAT3 signalling. In vivo, mice were subjected to limb ischaemia and treated with IL‐11 overexpression ADSCs and control ADSCs. IL‐11 overexpression ADSCs improved perfusion recovery in the ischaemic muscles.ConclusionsWe provide the evidence that IL‐11 promoted ADSCs proliferation, stimulated ADSCs migration and attenuated ADSCs apoptosis by activation of STAT3 signalling. These results suggest that IL‐11 facilitated ADSCs engraftment in ischaemic tissue, thereby enhanced ADSCs therapeutic efficacy.

A comparison of the use of adipose-derived and bone marrow-derived stem cells for peripheral nerve regeneration in vitro and in vivo

BackgroundTo date, it has repeatedly been demonstrated that infusing bone marrow-derived stem cells (BMSCs) into acellular nerve scaffolds can promote and support axon regeneration through a peripheral nerve defect. However, harvesting BMSCs is an invasive and painful process fraught with a low cellular yield.MethodsIn pursuit of alternative stem cell sources, we isolated stem cells from the inguinal subcutaneous adipose tissue of adult Sprague–Dawley rats (adipose-derived stem cells, ADSCs). We used a co-culture system that allows isolated adult mesenchymal stem cells (MSCs) and Schwann cells (SCs) to grow in the same culture medium but without direct cellular contact. We verified SC phenotype in vitro by cell marker analysis and used red fluorescent protein-tagged ADSCs to detect their fate after being injected into a chemically extracted acellular nerve allograft (CEANA). To compare the regenerative effects of CEANA containing either BMSCs or ADSCs with an autograft and CEANA only on the sciatic nerve defect in vivo, we performed histological and functional assessments up to 16 weeks after grafting.ResultsIn vitro, we observed reciprocal beneficial effects of ADSCs and SCs in the ADSC–SC co-culture system. Moreover, ADSCs were able to survive in CEANA for 5 days after in vitro implantation. Sixteen weeks after grafting, all results consistently showed that CEANA infused with BMSCs or ADSCs enhanced injured sciatic nerve repair compared to the acellular CEANA-only treatment. Furthermore, their beneficial effects on sciatic injury regeneration were comparable as histological and functional parameters evaluated showed no statistically significant differences. However, the autograft group was roundly superior to both the BMSC- or ADSC-loaded CEANA groups.ConclusionThe results of the present study show that ADSCs are a viable alternative stem cell source for treating sciatic nerve injury in lieu of BMSCs.

Programming of a Pro‐Adipogenic Phenotype in Offspring Born to Metabolically Compromised Pregnancies

ObjectivesDetermine the role of adipose tissue dysfunction in programming of cardiovascular disease risk in offspring born to dams with metabolic dysfunction.MethodsThe female mouse heterozygous for leptin receptor deficiency (Hetdb) was used as a model of maternal metabolic dysfunction. These females exhibit higher adiposity, dyslipidemia and hyperinsulinemia during pregnancy. Our data show that this model reproduces programming of early‐onset cardiovascular disease risk reported in human offspring born to obese or diabetic mothers. We sought to test the hypothesis that programming of cardiovascular disease risk is attributable to perturbed adipose tissue development leading to later‐life adipose tissue dysfunction. Wild type (Wt) offspring from both Wt and Hetdb pregnancies were studied as neonates and adults. At 7 weeks of age, offspring were placed on either a control or high fat/sugar (HF/HS) diet. Adipogenic potential was examined in preadipocytes isolated from the stromal vascular fraction (SVF) of inguinal subcutaneous adipose tissue (iSAT) in both neonatal and adult Wt offspring. After induction of differentiation, Oil Red O staining was used to measure lipid droplet accumulation and qPCR was used to measure adipogenic markers. Flow cytometry was utilized to quantify adipogenic progenitors in SVF. Adipokine secretion was measured with ELISA. Several indices of adipose tissue function were measured in adult offspring, including adipose tissue insulin resistance (Adipo‐IR).ResultsNMR whole‐body fat mass was 1.3 fold higher (p < 0.01) and plasma resistin levels were 1.7 fold higher (p < 0.01) in Wt neonates born to Hetdb pregnancies. The iSAT of Wt neonates and adults born to Hetdb vs. Wt dams exhibited a shift in the cell size distribution from smaller adipocytes to larger adipocytes (p < 0.05), indicating that adipogenesis was accelerated in the iSAT of Hetdb neonates. The percentage of CD31‐CD45‐CD29+CD34+ viable SVF cells was higher in neonates from the Hetdb vs. Wt pregnancy. Preadipocytes isolated from Hetdb offspring exhibited a pro‐adipogenic phenotype in early life that persisted into adulthood. After HF/HS feeding, preadipocytes isolated from Wt female (p < 0.05) and male (p < 0.05) adult offspring born to Hetdb pregnancies accumulated more lipid during differentiation. Heightened adipogenesis in adult offspring was also accompanied by impaired insulin‐stimulated inhibition of lipolysis.ConclusionsAdipose tissue dysfunction in offspring born to metabolically adverse pregnancies stems from in utero programming of a pro‐adipogenic phenotype in preadipocytes.Support or Funding InformationCanadian Institutes of Health Research, Alberta Children’s Hospital Research Institute, Heart & Stroke Foundation of Canada and Libin Cardiovascular Institute of Alberta

Cold Induced Depot-Specific Browning in Ferret Aortic Perivascular Adipose Tissue.

Brown adipose tissue is responsible for facultative thermogenesis to produce heat and increase energy expenditure in response to proper stimuli, e.g., cold. Acquisition of brown-like features (browning) in perivascular white adipose tissue (PVAT) may protect against obesity/cardiovascular disease. Most browning studies are performed in rodents, but translation to humans would benefit from a closer animal model. Therefore, we studied the browning response of ferret thoracic aortic PVAT (tPVAT) to cold. We performed global transcriptome analysis of tPVAT of 3-month-old ferrets acclimatized 1 week to 22 or 4°C, and compared the results with those of inguinal subcutaneous adipose tissue. Immunohistochemistry was used to visualize browning. Transcriptome data revealed a stronger cold exposure response of tPVAT, including increased expression of key brown/brite markers, compared to subcutaneous fat. This translated into a clear white-to-brown remodeling of tPVAT, with the appearance of multilocular highly UCP1-stained adipocytes. The pathway most affected by cold exposure in tPVAT was immune response, characterized by down-regulation of immune-related genes, with cardio protective implications. On the other hand, subcutaneous fat responded to cold by increasing energy metabolism based on increased expression of fatty acid oxidation and tricarboxylic acid cycle genes, concordant with lower inguinal adipose tissue weight in cold-exposed animals. Thus, ferret tPVAT responds to cold acclimation with a strong induction of browning and immunosuppression compared to subcutaneous fat. Our results present ferrets as an accessible translational animal model displaying functional responses relevant for obesity and cardiovascular disease prevention.