Cell Populations In Adipose Tissue Research Articles

Adipose tissue lymphocytes and obesity.

Obesity is associated with chronic inflammation in adipose tissue (AT), mainly evidenced by infiltration and phenotypic changes of various types of immune cells. Macrophages are the major innate immune cells and represent the predominant immune cell population within AT. Lymphocytes, including T cells and B cells, are adaptive immune cells and constitute another important immune cell population in AT. In obesity, CD8+ effector memory T cells, CD4+ Th1 cells, and B2 cells are increased in AT and promote AT inflammation, while regulatory T cells and Th2 cells, which usually function as immune regulatory or type 2 inflammatory cells, are reduced in AT. Immune cells may regulate the metabolism of adipocytes and other cells through various mechanisms, contributing to the development of metabolic diseases, including insulin resistance and type 2 diabetes. Efforts targeting immune cells and inflammation to prevent and treat obesity-linked metabolic disease have been explored, but have not yielded significant success in clinical studies. This review provides a concise overview of the changes in lymphocyte populations within AT and their potential role in AT inflammation and the regulation of metabolic functions in the context of obesity.

Factors Affecting the Population of Mesenchymal Stem Cells in Adipose-Derived Stromal Vascular Fraction

Recently, the adipose tissue-derived stromal vascular fraction has become the most popular source for obtaining mesenchymal stem cells because it is less expensive and is easier to perform than bone marrow concentrate harvesting. However, no study has investigated the factors affecting the mesenchymal stem cell population in adipose tissue derived stromal vascular fraction. Understanding the interaction of patient factors with the mesenchymal stem cell count and cell viability in adipose tissue-derived stromal vascular fraction could provide crucial information for surgeons to improve patient selection and outcomes. To evaluate the factors affecting the mesenchymal stem cell count, total cell count, and cell viability in adipose tissue-derived stromal vascular fraction. Retrospective cross-sectional study. This study retrospectively reviewed the medical records of 30 patients who underwent liposuction to harvest adipose tissue-derived stromal vascular fraction at our stem cell center. Operative variables, such as lipoaspirate amount and donor areas from the stromal vascular fraction harvesting site, included the entire abdomen and lower abdomen. We recorded the mesenchymal stem cell population, cell viability, and cell count of stromal vascular fraction, and we analyzed the results to determine statistical significance. The factors that were found to be significantly related are as follows; between cell number and age (p=0.001) and amount of lipoaspirate (p<0.001); between cell viability and body mass index (p=0.005) and hypertension (p=0.047); and between coronary artery disease and mesenchymal stem cell counts (p = 0.028). The relationship of patient factors (age, body mass index, hypertension, and coronary artery disease) with cell viability and mesenchymal stem cell counts may be important for clinical applications. However, the effect of medications on these relationships should be investigated in larger studies.

Single-cell transcriptomics reveals an angiogenic cell population for therapeutic angiogenesis in adipose tissue

Abstract Background Therapeutic angiogenesis mediated by stem/progenitor cells is an attractive therapeutic option against cardiovascular disease (CVD). Adipose tissue (AT) can be safely obtained even in CVD patients with anti-platelet medications, and it is a readily available source of culture-expanded adipose-derived stem cells (ADSCs) for transplantation. Single-cell transcriptome enables us to screen all the surface markers at once, while conventional strategies have been limited for the number of target markers. Furthermore, gene profiling at single-cell resolution can be used for the quantification of each marker by how many favorable cells can be purified without mixing of detrimental cells. Purpose We aimed to identify and characterize a cell population with in vivo angiogenic potential by single-cell RNA sequencing (scRNA-seq) analysis and xenograft experiments. Methods We revisited scRNA-seq datasets of single cell fraction from AT, bone-marrow (BM), and umbilical-cord blood (UCB, n=6/organ) to find cell populations with pro-angiogenic potential. Next, we collected AT from CVD patients (n=23) and used multicolor flow cytometry to quantify and sort the specific populations. PBS, the specific marker-negative and unsorted ADSCs were used as controls. Xenograft models of PKH26 pre-labeled human ADSC transplantation in limb ischemia were used to evaluate the lectin capillary density, PKH+ engrafted ADSCs, and blood flow recovery. Results Clustering divided CD45–CD31–CD34+ progenitor fraction into 3 clusters. We identified pro-/anti-angiogenic clusters based on the expressions of well-known pro-/anti-angiogenic factors. All genes encoding cell-surface proteins were compared in this functional clustering, resulted in 17 markers screened (Fig. 1A, B). Taken together with enrichment analysis, CD271+ cells showed predominant and pro-angiogenic gene profile from the other top candidates including CD36 and CD54 (Fig. 1C, D). Next, we evaluated the number and gene profile of CD271+ cells in well-known stem cell sources including BM and UCB. Surprisingly, the number of CD271 expressing cells were significantly lower and did not show angiogenic gene profile in BM and UCB (Fig. 2A). In analysis of AT from 23 CVD patients, CD271+ cells were significantly decreased by donor insulin resistance (Fig. 2B). Cell therapy using CD271+ ADSCs demonstrated in vivo angiogenic capacity compared to those of CD271– ADSCs and PBS in limb ischemia model. Furthermore, CD271+ ADSC transplantation showed enhanced efficacy compared to unsorted ADSCs from the same donors (Fig. 2C–E). Conclusion In this study, we identified CD271+ cell population in AT as an angiogenic cell population through scRNA-seq analysis and cell therapy experiments. AT obtained from donors without insulin resistance would be the most suitable for CD271+ ADSC isolation. CD271+ ADSC transplantation with a promising angiogenic capacity could contribute better cell-based therapy tackling CVD. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Japan Society for the Promotion of Science (JSPS) KAKENHI (Tokyo, Japan)

Immune Cell Regulation of White Adipose Progenitor Cell Fate.

Adipose tissue is essential for energy storage and endocrine regulation of metabolism. Imbalance in energy intake and expenditure result in obesity causing adipose tissue dysfunction. This alters cellular composition of the stromal cell populations and their function. Moreover, the individual cellular composition of each adipose tissue depot, regulated by environmental factors and genetics, determines the ability of the depots to expand and maintain its endocrine and storage function. Thus, stromal cells modulate adipocyte function and vice versa. In this mini-review we discuss heterogeneity in terms of composition and fate of adipose progenitor subtypes and their interactions with and regulation by different immune cell populations. Immune cells are the most diverse cell populations in adipose tissue and play essential roles in regulating adipose tissue function via interaction with adipocytes but also with adipocyte progenitors. We specifically discuss the role of macrophages, mast cells, innate lymphoid cells and T cells in the regulation of adipocyte progenitor proliferation, differentiation and lineage commitment. Understanding the factors and cellular interactions regulating preadipocyte expansion and fate decision will allow the identification of novel mechanisms and therapeutic strategies to promote healthy adipose tissue expansion without systemic metabolic impairment.

Mechanistic Links Between Obesity, Diabetes, and Blood Pressure: Role of Perivascular Adipose Tissue.

Obesity is increasingly prevalent and is associated with substantial cardiovascular risk. Adipose tissue distribution and morphology play a key role in determining the degree of adverse effects, and a key factor in the disease process appears to be the inflammatory cell population in adipose tissue. Healthy adipose tissue secretes a number of vasoactive adipokines and anti-inflammatory cytokines, and changes to this secretory profile will contribute to pathogenesis in obesity. In this review, we discuss the links between adipokine dysregulation and the development of hypertension and diabetes and explore the potential for manipulating adipose tissue morphology and its immune cell population to improve cardiovascular health in obesity.

Leveraging cell-specific differentially methylated regions to identify leukocyte infiltration in adipose tissue.

Obesity is understood to be an inflammatory condition characterized in part by changes in resident immune cell populations in adipose tissue. However, much of this knowledge has been obtained through experimental animal models. Epigenetic mechanisms, such as DNA methylation may be useful tools for characterizing the changes in immune cell populations in human subjects. In this study, we introduce a simple and intuitive method for assessing cellular infiltration by blood into other heterogeneous, admixed tissues such as adipose tissue, and apply this approach in a large human cohort study. Associations between higher leukocyte infiltration, measured by evaluating a distance measure between the methylation signatures of leukocytes and adipose tissue, and increasing body mass index (BMI) or android fat mass (AFM) were identified and validated in independent replication samples for CD4 (pBMI = 0.009, pAFM = 0.020), monocytes (pBMI = 0.001, pAFM = 4.3 × 10-4 ), and dendritic cells (pBMI = 0.571, pAFM = 0.012). Patterns of depletion with increasing adiposity were observed for plasma B (pBMI = 0.430, pAFM = 0.004) and immature B (pBMI = 0.022, pAFM = 0.042) cells. CD4, dendritic, monocytes, immature B, and plasma B cells may be important agents in the inflammatory process. Finally, the method used to assess leukocyte infiltration in this study is straightforwardly extended to other cell types and tissues in which infiltration might be of interest.

Abstract 426: Single Cell RNA-seq Analysis of Dynamic LIF Control of Adipose Derived Stem Cells Transition to Cardiomyocytes

Heart failure continues to be a leading cause of human mortality worldwide and is the consequence of various forms of myocardial injuries that compromise the pumping ability of the heart. Cellular fate transition through manipulation of modifying factors holds great promise for large-scale production of cardiomyocytes required for therapy and for revealing principles of gene regulation in the process. In this study, we found leukemia inhibitory factor (LIF) acts directly on adipose derived cells to induce cardiomyogenesis by JAK/STAT3. LIF induces adipose-cardiac transdifferentiation in adipose derived stem cells, which exhibits morphology and function typical of beating cardiomyocytes. However, the cell population in adipose tissue that generates cardiomyocyte is not from adipocytes or stromal cells, as examined by Cre-Loxp system and FACs. Here, using single cell RNA sequencing, we identified a cell population in adipose tissue that undergoes a gradual and simultaneous switch form relatively quiescent fate to proliferate cardiomyocyte like cells. By trajectory alignment, we compared the gene expression kinetics across fresh adipose cells and LIF induced cardiac transition process to identify crucial genes involved in the transition. Our findings may provide a better understanding of the mechanism underlying LIF-induced adipose-cardiac transdifferentiation, therefore reveal new therapeutic strategies for the treatment of myocardial injuries, and provide crucial information for regenerative medicine.

P-61 - Integration of oxidized lipids in pro-inflammatory metabolic networks targeting obesity and related complications

Pathogenesis of obesity and related metabolic diseases is closely associated with redox disbalance and production of oxidized lipids. Oxidized lipids affect all main cell populations in adipose tissue, their adipokine secretion and themselves can act as an active lipokines triggering autocrine and paracrine signaling. Obesity associate adipocyte hypertrophy and M2 to M1 adipose tissue macrophage (ATMs) shift was associated with oxidized lipids signaling. Despite large amount of data on the role of oxidized lipids and especially eicosanoids in pro-inflammatory signaling, no unified metabolic networks required for systems medicine data integration are available till now. Based on the literature meta-study, networks of enzymatic and free-radical-driven oxidation were reconstructed for the most abundant polyunsaturated fatty acids (PUFAs) and included 570 reactions covering 480 oxidized lipid species. Using networks of PUFA oxidation as a scaffold, adipocytes and ATMs specific regulatory metabolic networks covering four eicosanoids, namely prostaglandin E2, lipoxin A4, 6,15-deoxy prostaglandin J3, and epoxyeicosatetraenoic acids were reconstructed and enriched to uncover their roles in obesity related adipose tissue lipolysis, cytokines/adipokine production and secretion.

Eicosapentaenoic acid and 5-HEPE enhance macrophage-mediated Treg induction in mice

Eicosapentaenoic acid (EPA) is an omega-3 fatty acid with immunomodulatory and anti-inflammatory effects. Beyond its direct effects, the metabolic products of EPA also regulate various immune responses. Animal experiments demonstrated that EPA reduces adipose inflammation in high fat diet-induced obese mouse. However, the effects of EPA on infiltrated immune cell populations in adipose tissue and underlying mechanisms remain to be elucidated. We performed flow cytometry of stromal vascular fraction of epididymal adipose tissues from C57BL/6J and ob/ob mice fed normal chow mixed with or without 5% EPA. The numbers of hematopoietic cells, including Tregs, were higher in both C57BL/6J and ob/ob mice fed EPA diet compared with control diet. EPA enhanced the induction of Tregs in co-cultures of adipose tissue macrophages (ATMs) and naïve T cells. Among EPA metabolites, 5-HEPE was the most potent inducer of Tregs. GPR119 and GPR120 are receptors for 5-HEPE and EPA, respectively, and antagonist of GPR119 blocked Treg induction by EPA in the presence of ATMs. Alox5 gene encodes 5-lipoxygenase enzyme catalyzing EPA into 5-HEPE, and inhibitor of 5-lipoxygenase down-regulated EPA-mediated induction of adipose tissue Tregs in ob/ob mice. The study findings demonstrated that both EPA and 5-HEPE enhance ATM-mediated Treg induction.

Divergent functions of endotrophin on different cell populations in adipose tissue.

Endotrophin is a cleavage product of collagen 6 (Col6) in adipose tissue (AT). Previously, we demonstrated that endotrophin serves as a costimulator to trigger fibrosis and inflammation within the unhealthy AT milieu. However, how endotrophin affects lipid storage and breakdown in AT and how different cell types in AT respond to endotrophin stimulation remain unknown. In the current study, by using a doxycycline-inducible mouse model, we observed significant upregulation of adipogenic genes in the white AT (WAT) of endotrophin transgenic mice. We further showed that the mice exhibited inhibited lipolysis and accelerated hypertrophy and hyperplasia in WAT. To investigate the effects of endotrophin in vitro, we incubated different cell types from AT with conditioned medium from endotrophin-overexpressing 293T cells. We found that endotrophin activated multiple pathological pathways in different cell types. Particularly in 3T3-L1 adipocytes, endotrophin triggered a fibrotic program by upregulating collagen genes and promoted abnormal lipid accumulation by downregulating hormone-sensitive lipolysis gene and decreasing HSL phosphorylation levels. In macrophages isolated from WAT, endotrophin stimulated higher expression of the collagen-linking enzyme lysyl oxidase and M1 proinflammatory marker genes. In the stromal vascular fraction isolated from WAT, endotrophin induced upregulation of both profibrotic and proinflammatory genes. In conclusion, our study provides a new perspective on the effect of endotrophin in abnormal lipid accumulation and a mechanistic insight into the roles played by adipocytes and a variety of other cell types in AT in shaping the unhealthy microenvironment upon endotrophin treatment.

Angiopoietin Like Protein 2 (ANGPTL2) Promotes Adipose Tissue Macrophage and T lymphocyte Accumulation and Leads to Insulin Resistance.

ObjectivesAngiopoietin-like protein 2 (ANGPTL2), a recently identified pro-inflammatory cytokine, is mainly secreted from the adipose tissue. This study aimed to explore the role of ANGPTL2 in adipose tissue inflammation and macrophage activation in a mouse model of diabetes.Methodology/Principal FindingsAdenovirus mediated lacZ (Ad-LacZ) or human ANGPTL2 (Ad-ANGPTL2) was delivered via tail vein in diabetic db/db mice. Ad-ANGPTL2 treatment for 2 weeks impaired both glucose tolerance and insulin sensitivity as compared to Ad-LacZ treatment. Ad-ANGPTL2 treatment significantly induced pro-inflammatory gene expression in white adipose tissue. We also isolated stromal vascular fraction from epididymal fat pad and analyzed adipose tissue macrophage and T lymphocyte populations by flow cytometry. Ad-ANGPTL2 treated mice had more adipose tissue macrophages (F4/80+CD11b+) and a larger M1 macrophage subpopulation (F4/80+CD11b+CD11c+). Moreover, Ad-ANGPTL2 treatment increased a CD8-positive T cell population in adipose tissue, which preceded increased macrophage accumulation. Consistent with our in vivo results, recombinant human ANGPTL2 protein treatment increased mRNA levels of pro-inflammatory gene products and production of TNF-α protein in the human macrophage-like cell line THP-1. Furthermore, Ad-ANGPTL2 treatment induced lipid accumulation and increased fatty acid synthesis, lipid metabolism related gene expression in mouse liver.ConclusionANGPTL2 treatment promotes macrophage accumulation and activation. These results suggest potential mechanisms for insulin resistance.

The immune cells in adipose tissue

Although the pathological role of the immune system in several metabolic disorders, including type 1 diabetes mellitus (T1DM) and Addison's disease, has long been recognized and studied, only in the last decade has it become apparent that the immune system plays a broad and more subtle role in local and systemic metabolism. It is now apparent that the immune system monitors and responds to specific metabolic cues in both pathologic and non-pathologic settings through a set of processes dubbed immunometabolism. Expansion of adipose tissue mass, activation of lipolysis, eating a high fat diet and even non-shivering thermogenesis all lead to the recruitment and activation of immune cells in key metabolic tissues. The responses are complex and not completely defined, and indeed, as is typical of rapidly evolving research areas, there are some conflicting reports, especially related to the metabolic consequences of manipulation of immune function. However, what is clear is the consensus that metabolic processes, especially obesity and obesity-related complications, activate both the innate and adaptive arms of the immune system. Canonical immune processes consist of discrete steps: surveillance, recognition, effector action and resolution. Over the last decade evidence for each part of the immune response has been found at the intersection of the immune system with metabolism. Although evidence for immune surveillance and modulation of metabolism has been found in the liver, muscle, hypothalamus and pancreas, immune cell function has been most intensively studied and best understood in adipose tissue where studies continue to provide insights into the intersection of the metabolic and immune systems. Here we review the modulation of immune cell populations in adipose tissue and discuss regulatory processes implicated in controlling the interface between metabolism and immunologic function.

Female Mice are Protected against High-Fat Diet Induced Metabolic Syndrome and Increase the Regulatory T Cell Population in Adipose Tissue

Sex differences in obesity-induced complications such as type 2 diabetes have been reported. The aim of the study was to pinpoint the mechanisms resulting in different outcome of female and male mice on a high-fat diet (HFD). Mice fed control or HFD were monitored for weight, blood glucose, and insulin for 14 weeks. Circulating chemokines, islet endocrine function and blood flow, as well as adipose tissue populations of macrophages and regulatory T-lymphocytes (Treg) were thereafter assessed. Despite similar weight (43.8±1.0 and 40.2±1.5 g, respectively), male but not female mice developed hyperinsulinemia on HFD as previously described (2.5±0.7 and 0.5±0.1 pmol/l, respectively) consistent with glucose intolerance. Male mice also exhibited hypertrophic islets with intact function in terms of insulin release and blood perfusion. Low-grade, systemic inflammation was absent in obese female but present in obese male mice (IL-6 and mKC, males: 77.4±17 and 1795±563; females: 14.6±4.9 and 240±22 pg/ml), and the population of inflammatory macrophages was increased in intra-abdominal adipose tissues of high-fat-fed male but not female mice. In contrast, the anti-inflammatory Treg cell population increased in the adipose tissue of female mice in response to weight gain, while the number decreased in high-fat-fed male mice. In conclusion, female mice are protected against HFD-induced metabolic changes while maintaining an anti-inflammatory environment in the intra-abdominal adipose tissue with expanded Treg cell population, whereas HFD-fed male mice develop adipose tissue inflammation, glucose intolerance, hyperinsulinemia, and islet hypertrophy.

The potential of adipose‐derived stem cells in craniofacial repair and regeneration

The recent identification of a mesenchymal stem cell population in adipose tissue has led to an abundance of research focused on the regenerative properties of these cells. As such, adipose-derived stem cells (ASCs) and potential therapies in craniofacial regeneration have been widely studied. This review will discuss the identification and potential of ASCs, and specifically, preclinical and clinical studies using ASCs in craniofacial repair. Studies involving ASCs in the repair of defects caused by craniosynostosis and Treacher Collins syndrome will be discussed. A comprehensive review of the literature will be presented, focusing on fat grafting and biomaterials-based approaches that include ASCs for craniofacial regeneration.

Expression of NLRP3 inflammasome and T cell population markers in adipose tissue are associated with insulin resistance and impaired glucose metabolism in humans

Recent studies in rodents indicate that the nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3 (NLRP3) inflammasome and a proinflammatory shift in the T cell population in adipose tissue (AT) contribute to AT inflammation and insulin resistance. We investigated: (1) the interplay between the NLRP3 inflammasome and T cell populations in abdominal subcutaneous AT in obese and lean humans in relation to AT inflammatory processes, and (2) involvement of the NLRP3 inflammasome and T cell populations in insulin resistance. Abdominal subcutaneous AT biopsies were collected in 10 obese men with impaired glucose tolerance and 9 lean normal glucose tolerant age-matched controls. AT gene expression of NLRP3 inflammasome-related genes and markers of T cell populations, chemoattraction, macrophage infiltration and other aspects of inflammation were examined. Furthermore, we examined systemic adaptive immune activation and insulin sensitivity (hyperinsulinemic-euglycemic clamp). CASPASE-1 mRNA and the proportion of Th1 transcripts (TBX21/CD3ɛ) were significantly higher in AT from obese compared with lean subjects. CASPASE-1 expression and a relative increase in Th1 transcripts in AT were strongly associated with insulin resistance and impairments in glucose homeostasis. Gene expression of NLRP3, CASPASE-1, CD3ɛ (pan T cells), TBX21 (Th1 cells) and RORC (Th17 cells) was positively, whereas GATA3 (Th2 cells) was inversely correlated with AT inflammation. Our data suggest that NLRP3 inflammasome activation and a Th1 shift in the T cell population in AT of obese subjects is related to insulin resistance and impaired glucose metabolism, which may be explained by AT inflammatory processes.

NLRP3 inflammasome regulates obesity induced systemic inflammation and insulin signaling (117.4)

Abstract Activation of adipose tissue macrophages and T cells within fat depots is coupled with the development of obesity-induced proinflammatory state and insulin resistance. The release of IL-1β and IL-18 from macrophages is dependent on autocatalytic activation of procaspase-1 into active 20-kDa caspase-1. However, an upstream initiating event for obesity-induced immune cell activation in adipose tissue is not established. We found that Nlrp3 inflammasome, member of the Nod-like receptor (NLR) family of innate immune cell sensors is activated by obesity-associated lipotoxic factors. The obesity-induced macrophage activation and IL-1b production was lowered in absence of Nlrp3 signaling. Furthermore, elimination of Nlrp3 in obese mice reduced IL-18 and adipose tissue Interferon-g expression and improved insulin signaling. However, blocking the obesity-induced Nlrp3 inflamma¬some function in mice did not affect Treg cell homeostasis in adipose tissue but specifically reduced the numbers of effector memory cells and increased the naive CD4+ and CD8+ T cell populations in adipose tissue. Notably, weight loss associated improvement in type 2 diabetes in obese patients was associated with reduced NLRP3 expression and inflammation in adipose tissue. Our data establishes that Nlrp3 inflamamsome participates in origin of inflammation in obesity and development of insulin-resistance.

Obesity Increases the Production of Proinflammatory Mediators from Adipose Tissue T Cells and Compromises TCR Repertoire Diversity: Implications for Systemic Inflammation and Insulin Resistance

Emerging evidence suggests that increases in activated T cell populations in adipose tissue may contribute toward obesity-associated metabolic syndrome. The present study investigates three unanswered questions: 1) Do adipose-resident T cells (ARTs) from lean and obese mice have altered cytokine production in response to TCR ligation?; 2) Do the extralymphoid ARTs possess a unique TCR repertoire compared with lymphoid-resident T cells and whether obesity alters the TCR diversity in specific adipose depots?; and 3) Does short-term elimination of T cells in epididymal fat pad without disturbing the systemic T cell homeostasis regulate inflammation and insulin-action during obesity? We found that obesity reduced the frequency of naive ART cells in s.c. fat and increased the effector-memory populations in visceral fat. The ARTs from diet-induced obese (DIO) mice had a higher frequency of IFN-gamma(+), granzyme B(+) cells, and upon TCR ligation, the ARTs from DIO mice produced increased levels of proinflammatory mediators. Importantly, compared with splenic T cells, ARTs exhibited markedly restricted TCR diversity, which was further compromised by obesity. Acute depletion of T cells from epididymal fat pads improved insulin action in young DIO mice but did not reverse obesity-associated feed forward cascade of chronic systemic inflammation and insulin resistance in middle-aged DIO mice. Collectively, these data establish that ARTs have a restricted TCR-Vbeta repertoire, and T cells contribute toward the complex proinflammatory microenvironment of adipose tissue in obesity. Development of future long-term T cell depletion protocols specific to visceral fat may represent an additional strategy to manage obesity-associated comorbidities.

Cell density-dependent transcriptional activation of endocrine-related genes in human adipose tissue-derived stem cells

Adipose tissue is recognized as an endocrine organ that plays an important role in human diseases such as type II diabetes and cancer. Human adipose tissue-derived stem cells (ASCs), a distinct cell population in adipose tissue, are capable of differentiating into multiple lineages including adipogenesis. When cultured in vitro under a confluent condition, ASCs reach a commitment stage for adipogenesis, which can be further induced into terminally differentiated adipocytes by a cocktail of adipogenic factors. Here we report that the confluent state of ASCs triggers transcriptional activation cascades for genes that are responsible for the endocrine function of adipose tissue. These include insulin-like growth factor 1 (IGF-1) and aromatase (Cyp19), a key enzyme in estrogen biosynthesis. Despite similar adipogenic potentials, ASCs from different individuals display huge variations in activation of these endocrine-related genes. Bioinformatics and experimental data suggest that transcription factor Foxo1 controls a large number of “early” confluency-response genes, which subsequently induce “late” response genes. Furthermore, siRNA-mediated knockdown of Foxo1 substantially compromises the ability of committed ASCs to stimulate tumor cell migration in vitro. Thus, our work suggests that cell density is an important determinant of the endocrine potential of ASCs.

Obesity increases the production of pro-inflammatory mediators from adipose tissue T cells and compromises TCR repertoire diversity: Implications for systemic inflammation and insulin-resistance (87.32)

Abstract Yun-Hee Youm, Hyunwon Yang, Bolormaa Vandanmagsar, Jeffrey M. Gimble, Frank Greenway, Randall Mynatt and Vishwa Deep Dixit Emerging evidence suggests that increase in activated T cell populations in adipose tissue may contribute towards obesity-associated inflammation and insulin-resistance. Here we asked whether, 1)the adipose-resident T cells(ARTs) produce cytokines in response to T cell receptor(TCR) ligation. 2)If the extralymphoid ARTs possess unique TCR repertoire compared to lymphoid-resident T cells and if obesity alters the TCR diversity in specific adipose depots. 3)Can specific elimination of T cells regulate inflammation and insulin-action in obesity? We found that obesity reduced the frequency of naïve ART cells in subcutaneous fat and increased the effector-memory cells in visceral fat. The purified ARTs from obese mice had higher frequency of IFNγ+, Granzyme B+ cells and upon TCR ligation, produced pro-inflammatory cytokines and chemokines. Importantly extralymphoid ARTs exhibited markedly restricted TCR diversity and obesity further compromised the TCR repertoire in adipose depots. Collectively, these data establish that ARTs have restricted TCR-Vβ repertoire and T lymphocytes contribute towards the complex proinflammatory microenvironment of adipose tissue in obesity. However,specific depletion of extralymphoid T cells in visceral fat alone is insufficient to reverse obesity associated feed-forward cascade of inflammation and insulin-resistance.

Characterization of Osteogenically Induced Adipose Tissue-Derived Precursor Cells in 2-Dimensional and 3-Dimensional Environments

Earlier reports on a putative precursor cell population in adipose tissue showed differentiation along several mesodermal lineages, leading some to think that adipose tissue can be a source of cells applicable in regenerative medicine. However, characterizations of these adipose-derived precursor cells (ADPC) in the 3-dimensional (3-D) environment, especially within the area of bone-specific composite scaffolds, have been lacking. In this study, ADPC plated on culture flasks or seeded on medical grade polycaprolactone-tricalcium phosphate scaffolds (mPCL-CaP) were able to differentiate along the osteogenic lineages in both 2-D and 3-D environments as assessed with immunohistochemistry of osteo-related proteins, reverse transcriptase-polymerase chain reactions and alkaline phosphatase assay. The mPCL-CaP scaffolds provided adipose-derived cells (ADC) with a suitable environment as determined by DNA and metabolic assays, light, confocal and scanning electron microscopy. Flow cytometry revealed ADC to be CD29+, CD44+, CD73+, CD90+ and CD14–, CD31–, CD34–, CD45–, CD71–, and therefore showed the absence of hematopoietic stem cells but possibly the presence of pericytes and mescenchymal stem cells with osteogenic potential. The results of this study demonstrated the potential of using ADPC in combination with mPCL-CaP scaffolds for bone regenerative medicine.