Human Adipose Tissue-derived Stem Research Articles

Acquisition of durable insulin-producing cells from human adipose tissue-derived mesenchymal stem cells as a foundation for cell- based therapy of diabetes mellitus

This study aimed to identify the suitable induction protocol to produce highly qualified insulin producing cells (IPCs) from human adipose tissue derived stem cells (ADSCs) and evaluate the efficacy of the most functionally IPCs in management of diabetes mellitus (DM) in rats. The ADSCs were isolated and characterized according to the standard guidelines. ADSCs were further induced to be IPCs in vitro using three different protocols. The success of trans-differentiation was assessed in vitro through analysis of pancreatic endocrine genes expression, and insulin release in response to glucose stimulation. Then, the functionalization of the generated IPCs was evaluated in vivo. The in vitro findings revealed that the laminin-coated plates in combination with insulin-transferrin-selenium, B27, N2, and nicotinamide could efficiently up-regulate the expression of pancreatic endocrine genes. The in vivo study indicated effectual homing of the PKH-26-labelled IPCs in the pancreas of treated animals. Moreover, IPCs infusion in diabetic rats induced significant improvement in the metabolic parameters and prompted considerable up-regulation in the expression of the pancreatic related genes. The regenerative effect of infused IPCs was determined through histological examination of pancreatic tissue. Conclusively, the utilization of laminin–coated plates in concomitant with extrinsic factors promoting proliferation and differentiation of ADSCs could efficiently generate functional IPCs.

Generation and characterization of two iPSC lines from human adipose tissue-derived stem cells of healthy donors.

Here we have generated two induced pluripotent stem cell (iPSC) lines, hASC-iPSC-1A and hASC-iPSC-2A, by reprogramming human adipose tissue derived stem cells of 28 and 23years old healthy donors. Reprogramming was achieved using nonintegrative Sendai viral vector system containing the reprogramming factors Klf4, Oct3/4, Sox2, c-Myc. Though the karyotypes of these cells were normal (46, XX) and (46, XY), their pluripotency potentials were confirmed by the expression of factors of pluripotency markers in vitro and teratoma formation in vivo. These iPSCs differentiated cells can serve as control for disease modeling and drug screening.

Differentiation potential of human adipose tissue derived stem cells into photoreceptors through explants culture and enzyme methods.

To investigate the retinal photoreceptor differentiation potential of human orbital adipose tissue-derived stem cells (ADSCs) generated by enzyme (EN) and explant (EX) culture methods. We investigated potentials of human orbital ADSCs to differentiate into photoreceptors through EN and EX culture methods. EN and EX orbital ADSCs were obtained from the same donor during rehabilitative orbital decompression, and then were subject to a 3-step induction using Noggin, DKK-1, IGF-1 and b-FGF at different time points for 38d. Stem cell, eye-field and photoreceptor-related gene and protein markers were measured by reverse transcription-polymerase chain reaction (RT-PCR) and immunofluorescent (IMF) staining. Both EX and EN orbital ADSCs expressed CD133, a marker of cell differentiation. Moreover, PAX6 and rhodopsin, markers of the retinal progenitor cells, were detected from EX and EN orbital ADSCs. In EX orbital ADSCs, PAX6 mRNA was detected on the 17th day and then the rhodopsin mRNA was detected on the 24th day. In contrast, the EN orbital ADSCs expressed PAX6 and rhodopsin mRNA on the 31st day. EX orbital ADSCs expressed rhodopsin protein on the 24th day, while EN orbital ADSCs expressed rhodopsin protein on the 31st day. Orbital ADSCs isolated by direct explants culture show earlier and stronger expressions of markers towards eye field and retinal photoreceptor differentiation than those generated by conventional EN method.

H2S preconditioning of human adipose tissue-derived stem cells increases their efficacy in an in vitro model of cell therapy for simulated ischemia

AimsA major limitation of cell-based therapies for ischemia–reperfusion injury is the excessive loss of administered cells. We investigated whether H2S can improve the survival and efficacy of therapeutic cells in an in vitro model of cell-based therapy for simulated ischemia. Main methodsH9c2 rat cardiomyoblasts were exposed to oxygen–glucose deprivation and NaHS (3–30μM) pretreated human adipose tissue derived stem cells (hASCs) were added after reoxygenization. Viability of both cell lines was assessed with flow cytometry after 24h. The effects of H2S on antioxidant defense, proliferation, AKT and ERK1/2 phosphorylation and mitochondrial activity were analyzed in hASCs. Proliferation was evaluated using propargylglycine, an inhibitor of endogenous H2S synthesis. Key findingsNaHS pretreatment decreased the ratio of necrotic therapeutic cells by 41.8% in case of 3μM NaHS and by 34.3% with 30μM NaHS. The ratio of necrotic postischemic cardiomyocytes decreased by 35%, but only with the use of 3μM NaHS. Antioxidant defense mechanisms and ERK-phosphorylation were enhanced after 3μM NaHS treatment while AKT-phosphorylation was suppressed. NaHS dose-dependently increased the proliferation of hASCs while pretreatment with propargylglycine decreased it. SignificanceNaHS pretreatment can increase the survival of therapeutically used human adipose tissue-derived stem cells via increased antioxidant defense and improves the postischemic cardiac derived cells' survival as well. Proliferation of human adipose tissue-derived stem cells is enhanced by H2S. The underlying mechanisms involve enhanced ERK-phosphorylation and decreased AKT-phosphorylation. Pretreatment with NaHS may represent a simple pharmacological step that may enhance the efficacy of cell-based therapies.

The Role of Scaffold Architecture and Composition on the Bone Formation by Adipose-Derived Stem Cells

Scaffold architecture and composition are crucial parameters determining the initial cell spatial distribution and consequently bone tissue formation. Three-dimensional poly-ε-caprolactone (PCL) scaffolds with a 0/90° lay-down pattern were plotted and subjected to (1) an oxygen plasma (PCL O) or (2) a postargon plasma modification with gelatin and fibronectin (PCL Fn). These scaffolds with an open pore structure were compared with more compact scaffolds fabricated by conventional processing techniques: oxidized polylactic acid (LA O) and collagen (COL) scaffolds. Human adipose tissue-derived stem cell/scaffold interactions were studied. The study revealed that the biomimetic surface modification of plotted scaffolds did not increase the seeding efficiency. The proliferation and colonization was superior for PCL Fn in comparison with PCL O. The plotted PCL Fn was completely colonized throughout the scaffold, whereas conventional scaffolds only at the edge. Protein-based scaffolds (PCL Fn and COL) enhanced the differentiation, although plotted scaffolds showed a delay in their differentiation compared with compact scaffolds. In conclusion, protein modification of plotted PCL scaffolds enhances uniform tissue formation, but shows a delayed differentiation in comparison with compact scaffolds. The present study demonstrates that biomimetic PCL scaffolds could serve as a guiding template to obtain a uniform bone tissue formation in vivo.

Sphingosine-1-phosphate mediates proliferation maintaining the multipotency of human adult bone marrow and adipose tissue-derived stem cells

Sphingosine-1-phosphate mediates proliferation maintaining the multipotency of human adult bone marrow and adipose tissue-derived stem cells

Hemangioblastic Characteristics of Human Adipose Tissue-derived Adult Stem Cells In Vivo

Recent studies have demonstrated the existence of a population of adipose tissue-derived adult stem (ADAS) cells that can undergo multilineage differentiation in vitro. However, it remains unclear whether these cells maintain their multilineage potential in vivo. The aim of this study was to investigate whether Flk1(+)CD31(-)CD34(-) ADAS cells have characteristics of hemangioblasts. These human ADAS (hADAS) cells were able to differentiate into endothelial and hematopoietic cells at the single-cell level in vivo. These postnatal Flk1(+)CD31(-)CD34(-) hADAS cells bear characteristics of hemangioblast and may have potential application for the hematopoietic and vascular diseases.

288 CHARACTERIZATION OF PORCINE ADIPOSE TISSUE-DERIVED ADULT STEM CELL SURFACE PROTEINS

Human adipose tissue-derived adult stem (ADAS) cells are a self-renewing population of cells with a multilineage plasticity similar to bone marrow-derived mesenchymal stem cells. Human ADAS have promise for use in combination with various biomaterials for reconstructive tissue engineering. The phenotypic profile of human ADAS cell surface proteins has been partially characterized for stem cell-associated cluster differentiation molecules including CD29, CD44, and CD90. Porcine ADAS cells, an animal model for tissue engineering, also have the ability to self-renew and differentiate into multiple tissue lineages. However, the surface protein phenotype has not been described. Because porcine ADAS are isolated from fat depots likely different from human ADAS liposuction aspirates, it is important to characterize these cells. In this study, we have partially characterized the surface protein phenotype of undifferentiated porcine ADAS cells in comparison with the immunophenotype of undifferentiated human ADAS cells as reported in the literature. Flow cytometry and enhanced chemiluminescence Western blot analysis of early passage (passages 0–4) porcine ADAS cell populations demonstrated that the profiles are not similar to the human ADAS cell surface. Immunoblot detection paired with an enhanced chemiluminescence kit revealed a positive expression for CD44 and CD90 in human ADAS cells as indicated by bands present at the expected sizes and a negative expression for CD44 and CD90 in porcine ADAS cells. Flow cytometric analysis also indicated differences between human and early passage porcine ADAS cell surfaces with a relatively low expression of CD29 (5 cell lines with a mean percent positive of 4.5 ± 1.7 and a range of 2.5–7.2%) and CD44 (5 cell lines with a mean percent positive of 0.66 ± 0.67 and a range of 0.0–1.8%) compared with human ADAS values of 98 ± 1 and 60 ± 15, respectively (Gronthos et al. 2001). Other cell surface proteins analyzed at early passages include CD3 (3 cell lines; 0.07 ± 0.06% positive and 0.0–0.1 range), CD8 (3 cell lines; 0.10 ± 0.10% positive and 0–0.2 range), and CD90 [5 cell lines; 12.7 ± 11.9% positive and 2.4–33 range; human ADAS geometric mean 25.96% (Zuk et al. 2002)]. Analysis of late passage (passages 5–11) porcine ADAS cell populations revealed an increased expression of CD29 (3 cell lines; 26.4 ± 7.2% positive and 21.2–34.6 range). The expression level of CD90 at late passages were 21.3 and 26.9% positive for 2 cell lines and CD44 remained low (3 cell lines; 4.1 ± 3.5% positive and 0.2–7.0 range). Later passages were also analyzed for c-Kit (CD117), which was expressed at low levels (2 cell lines; 0.3 and 0.4% positive). The characterization of adipose tissue-derived adult stem cell surface proteins present at different stages of in vitro culture from a model animal, such as the pig, could have valuable impacts on tissue engineering research. These results suggest that care should be taken when interpreting results from animal models of somatic stem cells.