Impact of type 2 diabetes mellitus on the immunoregulatory characteristics of adipose tissue-derived mesenchymal stem cells.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is a chronic metabolic disorder associated with several complications. Adipose tissue-derived mesenchymal stem cells (AT-MSCs) represent an emerging type of MSCs with high plasticity and immunoregulatory capabilities and are useful for treating inflammation-related disorders such as T2DM. However, the pathogenic microenvironment of T2DM may affect their therapeutic potential. We aimed to examine the impact of the diabetic milieu on the immunomodulatory/anti-inflammatory potential of AT-MSCs. METHODS: We assessed the proliferation potential, cell surface expression of MSC-characteristic markers and immunomodulatory markers, along with the gene expression and protein secretion of pro-inflammatory and anti-inflammatory cytokines and adipokines in AT-MSCs derived from T2DM patients (dAT-MSCs) vs. those derived from non-diabetic volunteers (ndAT-MSCs). Furthermore, we evaluated the IFN-γ priming effect on both groups. RESULTS: Our data revealed comparable proliferative activities in both groups. Flow cytometric analysis results showed a lower expression of CD200 and CD276 on dAT-MSCs vs. ndAT-MSCs. qPCR demonstrated upregulation of IL-1ß associated with a downregulation of IL-1RN in dAT-MSCs vs. ndAT-MSCs. IFN-γ priming induced an elevation in CD274 expression associated with IDO1 and ILRN overexpression and IL-1ß downregulation in both groups. ELISA analysis uncovered elevated levels of secreted IL-1ß, TNF, and visfatin/NAMPT in dAT-MSCs, whereas IL-1RA and IDO levels were reduced. ELISA results were also evident in the secretome of dAT-MSCs upon IFN-γ priming. CONCLUSIONS: This study suggests that the T2DM milieu alters the immunomodulatory characteristics of AT-MSCs with a shift towards a proinflammatory phenotype which may restrain their autologous therapeutic use. Furthermore, our findings indicate that IFN-γ priming could be a useful strategy for enhancing dAT-MSC anti-inflammatory potential.

Adipose tissue-derived mesenchymal stem cells and chitosan/poly (vinyl alcohol) nanofibrous scaffolds for cartilage tissue engineering.

Osteoarthritis (OA) has been defined as a chronic inflammatory joint disease characterized by progressive articular cartilage degeneration. Recently growing interest in regenerative medicine, using cell therapy and tissue engineering, where cellular components in combination with engineered scaffolds and bioactive materials were used to induce functional tissue regeneration. In the present study, nanofibrous scaffold based on chitosan (CS)/poly (vinyl alcohol) (PVA) were used to develop biologically functionalized biomaterial to mimic the extracellular matrix, allowing the human adipose tissue derived mesenchymal stem cells (ADSCs) to proliferate and differentiate to chondrogenic cells. The morphology of the nanofibrous mat was examined using field emission scanning electron microscope (FE/SEM). The characteristic functional groups and the nature of the chemical bonds between atoms were evaluated using Fourier transform infrared spectroscopy (FTIR) spectrum. Characterization of the seeded cells was morphologically evaluated by scanning electron microscopy and by flow cytometry for the expression of the stem cell surface markers. The differentiation potential was verified after chondrogenic induction by analyzing the expression of chondrogenic marker genes using real-time (RT PCR). Current study suggest significant potential for the use of ADSCs with the nanofibrous scaffolds in improving the osteoarthritis pathology.

Non-target Genes Regulate miRNAs-Mediated Migration Steering of Colorectal Carcinoma.

MicroRNAs (miRNAs) trigger a two-layer regulatory network directly or through transcription factors and their co-regulators. Unlike miR-375, the role of miR-145 and miR-224 in inhibiting or driving cancer cell migration is controversial. This study is a step towards addressing the potential of miR-375, miR-145 and miR-224 expression modulation to inhibit colorectal carcinoma (CRC) cells migration in vitro through regulation of non-target genes VEGFA, TGFß1, IGF1, CD105 and CD44. Transwell migration assay results revealed a significant subdue of migration ability of cells transfected with miR-375 and miR-145 mimics and miR-224 inhibitor. Real time PCR data showed that expression of VEGFA, TGFß1, IGF1, CD105 and CD44 was downregulated as a consequence of exogenous re-expression of miR-375 and inhibition of miR-224. On the other hand, ectopic expression of miR-145 did not affect VEGFA, TGFß1 and CD44 expression, while it elevated CD105 and suppressed IGF1 expression. MAP4K4, a predicted target of miR-145, was validated as a target that could play a role in miR-145-mediated regulation of migration. At mRNA level, no change was observed in expression of MAP4K4 in cells with restored expression of miR-145, while western blotting analysis revealed a 25% reduction of protein level. By applying luciferase reporter assay, a significant decrease in luciferase activity was observed, supporting that miR-145 directly target 3' UTR of MAP4K4. The study highlighted the involvement of non-target genes VEGFA, TGFß1, IGF1, CD105 and CD44 in mediating anti- and pro-migratory effect of miR-375 and miR-224, respectively, and validated MAP4K4 as a direct target of anti-migratory miR-145.

MicroRNA expression analysis in endometriotic serum treated mesenchymal stem cells.

Endometriosis is defined by presence of endometrial-like-tissue outside the uterus. Recently, ectopic endometriotic lesions have been suggested to originate by abnormal differentiation of endometrial mesenchymal stem cells (eMSCs). MicroRNAs (miRNAs) play an important role in the pathophysiology of endometriosis. Through a PCR array approach, we aimed to assess the differential expression of microRNAs in human eMSC treated in culture with sera derived from women with severe endometriosis. Sera were collected from five patients with severe endometriosis and three control women and added individually in the culture medium to conduct experimental and control eMSC sets, respectively. Regular microscopic follow-up for cell morphology was performed. SYBR Green based real-time PCR array was used to assess the expression of 84 miRNAs. Bioinformatics analysis was done to predict the target genes of the significantly dysregulated miRNAs and their enriched biological processes and pathways. Thirty-two miRNAs were found significantly dysregulated in experimental cultures. Functional enrichment analysis revealed several endometriosis associated biological processes and pathways were enriched by target genes of these miRNAs. In conclusion, treatment of human eMSCs with sera of severe endometriosis cases affects the expression of certain miRNAs and their target genes. This may result in altering cell functions and consequently, endometriosis development.