Visceral Adipose-derived Stem Cells Research Articles

Isolation methods, proliferation, and adipogenic differentiation of adipose-derived stem cells from different fat depots in bovines.

The adipose-derived stem cells (ASCs) are a valuable resource for regenerative medicine and essential materials for research in fat deposition. However, the isolation procedure of ASCs has not been standardized and needs to be harmonized; differences in proliferation and adipogenic differentiation of ASCs obtained from different fat depots have not been well characterized. In the present study, we compared the efficiency of ASCs isolation by enzymatic treatment and explant culture methods and the proliferation ability and adipogenic differentiation potential of ASCs isolated from subcutaneous and visceral fat depots. The explant culture method was simple and with no need for expensive enzymes while the enzymatic treatment method was complex, time consuming and costly. By the explant culture method, a larger number of ASCs were isolated from subcutaneous and visceral fat depots. By contrast, fewer ASCs were obtained by the enzymatic treatment method, especially from visceral adipose. ASCs isolated by the explant culture method performed well in cell proliferation and adipogenic differentiation, though they were slightly lower than those by the enzymatic treatment method. ASCs isolated from visceral depot demonstrated higher proliferation ability and adipogenic differentiation potential. In total, the explant culture method is simpler, more efficient, and lower cost than the enzymatic treatment method for ASCs isolation; compared with visceral adipose, subcutaneous adipose is easier to isolate ASCs; however, the visceral ASCs are superior to subcutaneous ASCs in proliferation and adipogenic differentiation.

Visceral mesenchymal stem cells from type 2 diabetes donors activate triglycerides synthesis in healthy adipocytes via metabolites exchange and cytokines secretion.

In recent years, there has been an increase in the prevalence of obesity and type 2 diabetes mellitus (T2DM). Development of visceral instead of subcutaneous adipose tissue is pathogenic and increases the risk of metabolic abnormalities. We hypothesize that visceral adipocytes and stromal cells are able to deteriorate other fat depots metabolism via secretory mechanisms. We study the regulatory role of visceral adipose-derived stem cells (vADSC) from donors with obesity and T2DM or normal glucose tolerance (NGT) on healthy subcutaneous ADSC (sADSC) in the Transwell system. Lipid droplets formation during adipogenesis was assessed by confocal microscopy. Cell metabolism was evaluated by 14C-glucose incorporation analysis and western blotting. vADSC secretome was assessed by Milliplex assay. We showed that both NGT and T2DM vADSC had mesenchymal phenotype, but expression of CD29 was enhanced, whereas expressions of CD90, CD140b and IGF1R were suppressed in both NGT and T2DM vADSC. Co-differentiation with T2DM vADSC increased lipid droplet size and stimulated accumulation of fatty acids in adipocytes from healthy sADSC. In mature adipocytes T2DM vADSC stimulated triglyceride formation, whereas NGT vADSC activated oxidative metabolism. Secretome of NGT vADSC was pro-inflammatory and pro-angiogenic in comparison with T2DM vADSC. The present study has demonstrated the critical role of secretory interactions between visceral and subcutaneous fat depots both in the level of progenitor and mature cells. Mechanisms of these interactions are related to direct exchange of metabolites and cytokines secretion.

Piceatannol Is Superior to Resveratrol at Suppressing Adipogenesis in Human Visceral Adipose-Derived Stem Cells.

Resveratrol (3,4′,5-trans-trihydroxystilbene) and piceatannol (3,3′,4′,5-trans-tetraphydroxystilbene) are major stilbene compounds that are predominantly present in various natural foods, such as berries and fruits. Both phytochemical compounds are consumed as dietary supplements to prevent various metabolic diseases and for their anti-aging properties. Adipose-derived stem cells from human visceral adipose tissue (vASCs) are a useful in vitro model for evaluating their adipogenic effect. Treatment with resveratrol and piceatannol significantly inhibited lipid accumulation in vASCs. Their effective concentrations were 5, 10, and 20 μM for inhibiting adipogenesis of vASCs. Interestingly, despite the similar chemical structures of the two compounds, piceatannol showed a higher anti-adipogenic effect at 20 μM than resveratrol in vASCs. Moreover, the inhibitory capacity of lipid droplet generation was higher for piceatannol at 20 μM than that of resveratrol. Piceatannol significantly attenuated the expression level of adipogenic markers (e.g., CCAAT/enhanced binding protein α (C/EBPα), peroxisome proliferator-activated receptor γ (PPARγ), and adipocyte fatty acid binding protein (aP2)) compared to resveratrol at the mRNA and protein levels. These results suggest that piceatannol is a superior anti-adipogenic compound compared to resveratrol in the vASC model of visceral obesity.

Decursin and Decursinol Angelate Suppress Adipogenesis through Activation of β-catenin Signaling Pathway in Human Visceral Adipose-Derived Stem Cells.

Visceral adiposity is closely associated with metabolic disorders and cardiovascular diseases. Angelica gigas Nakai (AGN) has been reported to possess anti-obesity effects and higher amounts of coumarin compounds are present in AGN. However, the active compounds suppressing adipogenesis in AGN and mechanisms of action have not been investigated in adipose-derived stem cells (ASCs) isolated from visceral adipose tissue (VAT). Among four coumarin compounds of AGN, decursin (D) and decursinol angelate (DA) significantly inhibited adipocyte differentiation from ASCs. D and DA downregulated CCAAT/enhancer binding protein α (C/EBPα), peroxisome proliferator-activated receptor γ (PPARγ), adipocyte fatty acid binding protein (aP2), fatty acid synthase (FAS), and acetyl-CoA carboxylase (ACC) at both mRNA and protein levels. Next, treatment with adipogenic differentiation medium (ADM) on ASCs downregulated β-catenin expression at protein level, while addition of D and DA could restore protein expression and nuclear translocation of β-catenin suppressed by ADM. D and DA treatment on ADM treated ASCs increased inhibitory phosphorylation of Glycogen synthase kinase (GSK)-3β, thereby preventing β-catenin from degradation. Additionally, si-β-catenin transfection significantly upregulated protein expression of C/EBPα and PPARγ, alleviating the anti-adipogenic effect of D and DA on ADM treated ASCs. Overall, D and DA, active compounds from AGN, suppressed adipogenesis through activation of β-catenin signaling pathway in ASCs derived from human VAT, possibly using as natural anti-visceral adiposity agents.

CD90 serves as differential modulator of subcutaneous and visceral adipose-derived stem cells by regulating AKT activation that influences adipose tissue and metabolic homeostasis

BackgroundWhite adipose tissue includes subcutaneous and visceral adipose tissue (SAT and VAT) with different metabolic features. SAT protects from metabolic disorders, while VAT promotes them. The proliferative and adipogenic potentials of adipose-derived stem cells (ADSCs) are critical for maintaining adipose tissue homeostasis through driving adipocyte hyperplasia and inhibiting pathological hypertrophy. However, it remains to be elucidated the critical molecules that regulate different potentials of subcutaneous and visceral ADSCs (S-ADSCs, V-ADSCs) and mediate distinct metabolic properties of SAT and VAT. CD90 is a glycosylphosphatidylinositol-anchored protein on various cells, which is also expressed on ADSCs. However, its expression patterns and differential regulation on S-ADSCs and V-ADSCs remain unclear.MethodsS-ADSCs and V-ADSCs were detected for CD90 expression. Proliferation, colony formation, cell cycle, mitotic clonal expansion, and adipogenic differentiation were assayed in S-ADSCs, V-ADSCs, or CD90-silenced S-ADSCs. Glucose tolerance test and adipocyte hypertrophy were examined in mice after silencing of CD90 in SAT. CD90 expression and its association with CyclinD1 and Leptin were analyzed in adipose tissue from mice and humans. Regulation of AKT by CD90 was detected using a co-transfection system.ResultsCompared with V-ADSCs, S-ADSCs expressed high level of CD90 and showed increases in proliferation, mitotic clonal expansion, and adipogenic differentiation, together with AKT activation and G1-S phase transition. CD90 silencing inhibited AKT activation and S phase entry, thereby curbing proliferation and mitotic clonal expansion of S-ADSCs. In vivo CD90 silencing in SAT inhibited S-ADSC proliferation, which caused adipocyte hypertrophy and glucose intolerance in mice. Furthermore, CD90 was highly expressed in SAT rather than in VAT in human and mouse, which had positive correlation with CyclinD1 but negative correlation with Leptin. CD90 promoted AKT activation through recruiting its pleckstrin homology domain to plasma membrane.ConclusionsCD90 is differentially expressed on S-ADSCs and V-ADSCs, and plays critical roles in ADSC proliferation, mitotic clonal expansion, and hemostasis of adipose tissue and metabolism. These findings identify CD90 as a crucial modulator of S-ADSCs and V-ADSCs to mediate distinct metabolic features of SAT and VAT, thus proposing CD90 as a valuable biomarker or target for evaluating ADSC potentials, monitoring or treating obesity-associated metabolic disorders.

N-3 polyunsaturated fatty acids provoke a specific transcriptional profile in rabbit adipose-derived stem cells in vitro.

Adipose-derived stem cells (ADSCs) possess multipotent properties, and their proper functionality is essential for further development of metabolic disorders. In the current study, we explored the impact of two n-3 LC-PUFAs (long-chain polyunsaturated fatty acids, DHA-docosahexaenoic; C22:6, and EPA-eicosapentaenoic; C20:5) on a specific profile of lipolytic-related gene expressions in the in vitro-differentiated subcutaneous and visceral ADSCs from rabbits. The subcutaneous and visceral ADSCs were obtained from 28-day-old New Zealand rabbits. The primary cells were cultured up to passage 4 and were induced for adipogenic differentiation. Thereafter, the differentiated cells were treated with 100µg EPA or DHA for 48hr. The total mRNA was isolated and target genes expression evaluated by real-time RCR. The results demonstrated that treatment of rabbit ADSCs with n-3 PUFAs significantly enhanced mRNA expression of Perilipin A, while the upregulation of leptin and Rab18 genes was seen mainly in ADSCs from visceral adipose tissue. Moreover, the EPA significantly enhanced PEDF (Pigment Derived Epithelium Factor) mRNA expression only in visceral cells. Collectively, the results suggest activation of an additional lipolysis pathway most evident in visceral cells. The data obtained in our study indicate that in vitro EPA up-regulates the mRNA expression of the studied lipolysis-associated genes stronger than DHA mainly in visceral rabbit ADSCs.

Resveratrol derivative BTM-0512 mitigates obesity by promoting beige remodeling of subcutaneous preadipocytes.

Recent studies revealed that sirtuin 1 (SIRT1) is involved in the regulation of energy metabolism and its agonist resveratrol showed anti-obesity effect. This study aims to determine whether BTM-0512, a novel derivative of resveratrol, acts as an antagonist of obesity and to explore its possible mechanisms. High-fat diet (HFD)-induced obese mice were intragastrically administered with BTM-0512 (5, 10, and 20 mg/kg/day) or resveratrol (10 mg/kg/day). It was found that the body weight, Lee's index, ratio of visceral adipose tissue (VAT) to body weight, and blood glucose were significantly reduced in BTM-0512-treated mice when compared with those in mice treated with resveratrol. BTM-0512 up-regulated the expressions of SIRT1, full length PRDM16 (fPRDM16), total PRDM16 (tPRDM16, including fPPRDM16 and other PRDM16 isoforms), and uncoupling protein 1 (UCP1) in both brown and subcutaneous adipose tissues. Although BTM-0512 and resveratrol also up-regulated SIRT1 and tPRDM16 levels in VAT of HFD-induced obese mice, the expressions of fPRDM16, UCP1, and TMEM26 were down-regulated. In mouse primary subcutaneous preadipocytes cultured with or without adipogenic medium, BTM-0512 up-regulated fPRDM16, tPRDM16, and UCP1 expressions, which was reversed by SIRT1 antagonists. But in cultured brown and visceral adipocytes, the UCP1 protein level showed no significant change after treatment with 1 μM of BTM-0512. Moreover, transfection with human SIRT1 plasmid reduced lipid deposit, as well as the mRNA levels of fPRDM16, UCP1, and TMEM26, in cultured human visceral adipose-derived stem cells. In conclusion, BTM-0512 has stronger anti-obesity effect than resveratrol, which might be associated with activation of beige remodeling in subcutaneous adipose tissue.

Gene expression profiles of human subcutaneous and visceral adipose-derived stem cells.

Subcutaneous and visceral adipose tissues show a different risk effect on metabolic disorders because they have distinct cellular properties. We isolated stem cells from the separate human adipose tissues to investigate that subcutaneous and visceral fat depots have metabolic differences. Adipose-derived stem cells (ASCs) were characterized by immunophenotype and differentiation potentials into adipogenic, osteogenic, and chondrogenic lineages. Although subcutaneous and visceral ASCs (S-ASC and V-ASC) express same surface markers (CD31- , CD34- , CD45- , CD73+ , CD90+ , and CD105+ ) and have differentiation potentials, S-ASCs had higher capacity to proliferate and to differentiate into adipogenic lineage than V-ASCs. Next, we identified that S-ASC and V-ASC were genetically distinct based on microarray analysis. Among a total of 810 genes detected in ASCs of both depots, the differentially expressed genes were involved in energy and lipid metabolism. These data show the existence of the intrinsic difference between S-ASC and V-ASC and suggest the differences of anatomically separated adipose tissue. On the basis of the differentially expressed gene profiles between S-ASC and V-ASC, we suggested significant evidence that adipose tissues originating from different anatomic regions are distinguished at the level of the undifferentiated stem cells such as mature adipocytes. V-ASCs had the upregulated clusters of genes related to lipid biosynthesis and metabolism. By contrast, S-ASCs highly expressed genes involved in DNA-dependent transcription, contributing to proliferation. We provide further insights for ASCs with the different origins to understand fat accumulation and distribution and a possibility of ASCs as a therapeutic target against metabolic disorders or cancer.

Retinoic Acid Mediates Visceral-Specific Adipogenic Defects of Human Adipose-Derived Stem Cells

Increased visceral fat, rather than subcutaneous fat, during the onset of obesity is associated with a higher risk of developing metabolic diseases. The inherent adipogenic properties of human adipose-derived stem cells (ASCs) from visceral depots are compromised compared with those of ASCs from subcutaneous depots, but little is known about the underlying mechanisms. Using ontological analysis of global gene expression studies, we demonstrate that many genes involved in retinoic acid (RA) synthesis or regulated by RA are differentially expressed in human tissues and ASCs from subcutaneous and visceral fat. The endogenous level of RA is higher in visceral ASCs; this is associated with upregulation of the RA synthesis gene through the visceral-specific developmental factor WT1. Excessive RA-mediated activity impedes the adipogenic capability of ASCs at early but not late stages of adipogenesis, which can be reversed by antagonism of RA receptors or knockdown of WT1. Our results reveal the developmental origin of adipocytic properties and the pathophysiological contributions of visceral fat depots.

Adipose-derived stem cells from both visceral and subcutaneous fat deposits significantly improve contractile function of infarcted rat hearts.

Adipose-derived stem cells (ASCs) from subcutaneous and visceral adipose tissues have been studied individually. No studies have compared their abilities in treatment of heart failure. This study was designed to evaluate whether ASCs from the two sources could provide a long-term improvement of cardiac function in infarcted hearts. Rat subcutaneous and visceral adipose tissues were excised for isolation of ASCs. Morphology, yield, proliferation, surface markers, differentiation, and cytokine secretion of the subcutaneous ASCs (S-ASCs) and visceral ASCs (V-ASCs) were analyzed. Then a rat model of myocardial infarction (MI) was established by a coronary occlusion. Seven days after occlusion, S-ASCs (n = 22), V-ASCs (n = 22), and Dulbecco's modified Eagle medium (DMEM, n = 20) were injected into the infarct rim, respectively. Cardiac function was then monitored with MRI for up to 6 months. The hearts were then removed for histological assessments. The yield of V-ASCs per gram of the visceral adipose depot was significantly greater than that of S-ASCs in 1 g of the subcutaneous adipose depot. On the other hand, the S-ASCs showed a greater proliferation rate and colony-forming unit relative to the V-ASCs. In addition, the infarcted hearts treated with either S-ASCs or V-ASCs showed a significantly greater left ventricular ejection fraction (LVEF) than those treated with DMEM at 4 weeks and 6 months following the cell/DMEM transplantation. Moreover, the infarct sizes of both S-ASC- and V-ASC-treated hearts were significantly smaller than that in the DMEM-treated hearts. MRI showed the implanted ASCs at the end of 6 months of recovery. Despite the differences in cell yield, proliferation, and colony formation capacity, both S-ASCs and V-ASCs provide a long-lasting improvement of cardiac contractile function in infarcted hearts. We conclude that the subcutaneous and visceral adipose tissues are equally effective cell sources for cell therapy of heart failure.

Characterization of adipose-derived stem cells from subcutaneous and visceral adipose tissues and their function in breast cancer cells.

Adipose-derived stem cells are capable of differentiating into multiple cell types and thus considered useful for regenerative medicine. However, this differentiation feature seems to be associated with tumor initiation and metastasis raising safety concerns, which requires further investigation. In this study, we isolated adipose-derived stem cells from subcutaneous as well as from visceral adipose tissues of the same donor and systematically compared their features. Although being characteristic of mesenchymal stem cells, subcutaneous adipose-derived stem cells tend to be spindle form-like and are more able to home to cancer cells, whereas visceral adipose-derived stem cells incline to be "epithelial"-like and more competent to differentiate. Moreover, compared to subcutaneous adipose-derived stem cells, visceral adipose-derived stem cells are more capable of promoting proliferation, inducing the epithelial-to-mesenchymal transition, enhancing migration and invasion of breast cancer cells by cell-cell contact and by secreting interleukins such as IL-6 and IL-8. Importantly, ASCs affect the low malignant breast cancer cells MCF-7 more than the highly metastatic MDA-MB-231 cells. Induction of the epithelial-to-mesenchymal transition is mediated by the activation of multiple pathways especially the PI3K/AKT signaling in breast cancer cells. BCL6, an important player in B-cell lymphoma and breast cancer progression, is crucial for this transition. Finally, this transition fuels malignant properties of breast cancer cells and render them resistant to ATP competitive Polo-like kinase 1 inhibitors BI 2535 and BI 6727.

Abstract 242: Subcutaneous and Visceral Adipose-Derived Stem Cells Have Similar Biological Properties and Both Improve Cardiac Function of Infarcted Rat Hearts

Background: Adipose stem cells (ASC) from subcutaneous and visceral adipose tissues have been studied individually. However, it is unclear whether ASC from the two sources have different biological properties and, more importantly, whether one sub-type of ASC is more effective in treatment of CHF. This study was designed to address these concerns. Methods: Morphology, yield, proliferation, surface markers, and cytokine secretion of rat subcutaneous ASC (S-ASC) and visceral ASC (V-ASC) were analyzed. A rat model of myocardial infarction (MI) was established by occlusion of the LAD. 7 days after MI, S-ASC (n = 11), V-ASC (n = 11), and cell culture medium (Control, n = 7) were injected into the infarct rim, respectively. Cardiac function of the infarcted hearts was monitored with MRI for 6 months. Results: Both S-ASC and V-ASC exhibited a fibroblast-like morphology and expressed stromal cell markers (CD29, CD90 and CD105). No significant expression of hematopoietic markers (CD11b, CD34 and CD45) was found. Under appropriate conditions, both cells could differentiate to adipocyte- and osteocyte-like cells. Both of them expressed a significant level of HGF, IGF-1 and VEGF. As to their differences, V-ASC had approximately 3-times greater cell yield and a lower colony-formation rate (9.8±1.0% vs.13.5±2.6%) relative to S-ASC. In contrast, S-ASC showed a significantly greater growth rate (Doubling Time: 17.9 h vs. 26.0 h) relative to V-ASC. Both S-ASC and V-ASC-treated hearts showed a significantly greater left ventricular ejection fraction (LVEF, 58.3% and 56.7%) than the control group (LVEF, 47.2%) at end of 6 months. LVEF between the two ASC-treated groups was not significantly different. Finally, the implanted stem cells were readily detected in vivo with MRI for at least 6 months. Myocardial tissue sections showed existence of ASC and their locations matched with MRI signals. Conclusions: S-ASC and V-ASC share several biological characteristics. Both provide comparable significant improvement on cardiac function. Moreover, these implanted cells can be reliably tracked for at least 6 months using MRI. We conclude that the S-ASC and V-ASC are equally effective for treatment of heart failure.

Stemness and Osteogenic and Adipogenic Potential are Differently Impaired in Subcutaneous and Visceral Adipose Derived Stem Cells (ASCs) Isolated from Obese Donors

Today adipose tissue is not just considered as the primary energy storage organ, but it is also recognized as an important endocrine tissue and an abundant source of mesenchymal stem cells (adipose-derived stem cells, ASCs). During the last decade, several studies have provided preclinical data on the safety and efficacy of ASCs, supporting their use in cell-based therapy for regenerative medicine purposes. Little is known about the effect of obesity on ASCs properties. Since ASCs differentiation and proliferation are determined by their niche, the differences in body fat distribution and the obesity-related co-morbidities may have several consequences. In this study we compared ASCs of subcutaneous adipose tissue from obese (obS-ASCs) and non-obese (nS-ASCs) donors in order to compare their immunophenotype and osteogenic and adipogenic potential. Moreover, in order to evaluate the possible difference between subcutaneous and visceral fat, obS-ASCs were also compared to ASCs derived from visceral adipose tissue of the same obese donors (obV-ASCs). Our results show that subcutaneous and visceral ASCs derived from obese donors have an impaired cell proliferation, clonogenic ability and immunophenotype. Nevertheless, obS-ASCs are able to differentiate toward osteogenic and adipogenic lineages, although to a small extent with respect to non-obese donors, whereas obV-ASCs lose most of their stem cell characteristics, including multi-differentiation potential. Taken together our findings confirm that not all ASCs present the same behavior, most likely due to their biological microenvironment in vivo. The specific stimuli which can play a key role in ASCs impairment, including the effects of the obesity-related inflammation, should be further investigated to have a complete picture of the phenomenon.

What's in a Name?

What's in a Name?