Int J Stem Cells.  2020 Jul;13(2):268-278. 10.15283/ijsc20028.

Adipose Tissue-Derived Stem Cells from Type 2 Diabetics Reveal Conservative Alterations in Multidimensional Characteristics

Affiliations
  • 1Organ Transplant Center, Tianjin First Central Hospital, Nankai University, Tianjin, China
  • 2NHC Key Laboratory for Critical Care Medicine, Tianjin, China
  • 3Tianjin Clinical Research Center for Organ Transplantation, Tianjin, China
  • 4The Postdoctoral Research Station, School of Medicine, Nankai University, Tianjin, China

Abstract

Background and Objectives
Adipose tissue-derived mesenchymal stem cells (ASCs) are recognized as an advantaged source for the prevention and treatment of diverse diseases including type 2 diabetes mellitus (T2DM). However, alterations in characteristics of ASCs from the aforementioned T2DM patients are still obscure, which also hinder the rigorous and systematic illumination of progression and pathogenesis.
Methods and Results
In this study, we originally isolated peripancreatic adipose tissue-derived mesenchymal stem cells from both human type 2 diabetic and non-diabetic donors (T2DM-ASCs, ND-ASCs) with the parental consent, respectively. We noticed that T2DM-ASCs exhibited indistinguishable immunophenotype, cell vitality, chondrogenic differentiation and stemness as ND-ASCs. Simultaneously, there’s merely alterations in migration and immunoregulatory capacities in T2DM-ASCs. However, differing from ND-ASCs, T2DM-ASCs exhibited deficiency in adipogenic and osteogenic differentiation, and in particular, the delayed cell cycle and different cytokine expression spectrum.
Conclusions
The conservative alterations of T2DM-ASCs in multifaceted characteristics indicated the possibility of autologous application of ASCs for cell-based T2DM treatment in the future.

Keyword

Adipose tissue-derived mesenchymal stem cells; Type 2 diabetes mellitus; Cell vitality; Immunoregulation

Figure

  • Fig. 1 T2DM-ASCs show similar cytomorphology and immunophenotype. (A) Morphology of ND-ASCs and T2DM-ASCs under same conditions (left, 40× magnification; right, 100× magnification,). (B, C) Immunophenotype of ND-ASCs (n=5) and T2DM-ASCs (n=3) at P6, analysed by flow cytometry. The markers including CD44, CD73, CD90, CD105, CD34 and CD45. The data were shown as the mean±SEM, each experiment was repeated at least twice, one representative example shown. NS: not significant.

  • Fig. 2 T2DM-ASCs exhibit indiscriminate signatures in cell viability. (A) Cell count of ND-ASCs (n=4) and T2DM-ASCs (n=3) at P5. (B) CCK-8 assay of ND-ASCs (n=3) and T2DM-ASCs for 4, 24, 48 and 72 h. (C) PI staining profiles of ND-ASCs (n=4) and T2DM-ASCs (n=3) by flow cytometry. (D) Histograms statistics of cell cycle status populations. (E) 7-AAD and Annexin V staining profiles of ND-ASCs (n=4) and T2DM-ASCs (n=3) by flow cytometry. (F) Histograms statistics of cell apoptosis, including early apoptosis (Annexin Ⅴ+7-AAD−) and total apoptosis (Annexin Ⅴ+) populations. The data were shown as the mean± SEM, each experiment was repeated at least twice, one representative example shown. NS: not significant. ***p<0.001.

  • Fig. 3 T2DM-ASCs possess impaired adipogenic and osteogenic differentiation potential. (A) The morphology of adipogenic differentiation of ND-ASCs (n=3) and T2DM-ASCs (n=3) by Oil Red O staining (200× magnification). (B) qRT-PCR assessment of adipogenic marker PPAR-γ. (C) The morphology of osteogenic differentiation of ND-ASCs (n=3) and T2DM-ASCs (n=3) by von Kossa staining (200× magnification). (D) qRT-PCR assessment of osteogenic marker BGLAP. (E) The morphology of chondrogenic differentiation of ND-ASCs (n=3) and T2DM-ASCs (n=3) by Alcian Blue staining (200× magnification). (F) qRT-PCR assessment of chondrogenic marker ACAN. All values are normalized to the negative control (NC) group. The data were shown as the mean±SEM, each experiment was repeated at least twice, one representative example shown. *p<0.05, **p<0.01.

  • Fig. 4 T2DM-ASCs show comparable migration and immunosuppressive capacity. (A) Cell migration ability of ND-ASCs (n=3) and T2DM-ASCs (n=3) at 0, 12 and 24 hours (40×magnification). (B) Statistical analysis of cell migration area by the ImageJ. (C) ND-ASCs (n=3) and T2DM-ASCs (n=3) cocultured with PBMCs for 3 days, flow cytometry analysis of PBMCs with CD3 staining. (D) Percentages of CD3+ lymphocyte proliferation. The data were shown as the mean±SEM, each experiment was repeated at least twice, one representative example shown. NS: not significant. **p<0.01, ***p<0.001.

  • Fig. 5 The characteristics of cytokine expression and stemness of T2DM-ASCs and ND-ASCs. (A, B) The comparation of immunomodulation-associated factors (A) and islet function-related factors (B) between T2DM-ASCs (n=3) and ND-ASCs (n=3). (C) The comparation of stemness-associated gene expression (NANOG, POU5F1) between T2DM-ASCs (n=3) and ND-ASCs (n=3). The data were shown as the mean± SEM, each experiment was repeated at least twice. NS: not significant. **p<0.01, ***p<0.001.


Reference

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