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A Comparative Study of the Effect of Anatomical Site on Multiple Differentiation of Adipose-Derived Stem Cells in Rats.

ABSTRACT: Mesenchymal stem cells (MSCs) derived from adipose tissue are evolved into various cell-based regenerative approaches. Adipose-derived stem cells (ASCs) isolated from rats are commonly used in tissue engineering studies. Still, there is a gap in knowledge about how the harvest locations influence and guide cell differentiation. This study aims to investigate how the harvesting site affects stem-cell-specific surface markers expression, pluripotency, and differentiation potential of ASCs in female Sprague Dawley rats. ASCs were extracted from the adipose tissue of the peri-ovarian, peri-renal, and mesenteric depots and were compared in terms of cell morphology. MSCs phenotype was validated by cell surfaces markers using flow cytometry. Moreover, pluripotent gene expression of Oct4, Nanog, Sox2, Rex-1, and Tert was evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR). ASCs multipotency was evaluated by specific histological stains, and the results were confirmed by quantitative polymerase chain reaction (RT-qPCR) expression analysis of specific genes. There was a non-significant difference detected in the cell morphology and immunophenotype between different harvesting sites. ASCs from multiple locations were significantly varied in their capacity to differentiate into adipocytes, osteoblastic cells, and chondrocytes. To conclude, depot selection is a critical element that should be considered when using ASCs in tissue-specific cell-based regenerative therapies research.

SUBMITTER: Hendawy H

PROVIDER: S-EPMC8465004 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Human adipose-derived stem cells (ASCs) may differentiate into cardiomyocytes and this provides a source of donor cells for tissue engineering. In this study, we evaluated cardiomyogenic differentiation protocols using a DNA demethylating agent 5-azacytidine (5-aza), a modified cardiomyogenic medium (MCM), a histone deacetylase inhibitor trichostatin A (TSA) and co-culture with neonatal rat cardiomyocytes. 5-aza treatment reduced both cardiac actin and TropT mRNA expression. Incubation in MCM only slightly increased gene expression (1.5- to 1.9-fold) and the number of cells co-expressing nkx2.5/sarcomeric alpha-actin (27.2% versus 0.2% in control). TSA treatment increased cardiac actin mRNA expression 11-fold after 1 week, which could be sustained for 2 weeks by culturing cells in cardiomyocyte culture medium. TSA-treated cells also stained positively for cardiac myosin heavy chain, alpha-actin, TropI and connexin43; however, none of these treatments produced beating cells. ASCs in non-contact co-culture showed no cardiac differentiation; however, ASCs co-cultured in direct contact co-culture exhibited a time-dependent increase in cardiac actin mRNA expression (up to 33-fold) between days 3 and 14. Immunocytochemistry revealed co-expression of GATA4 and Nkx2.5, alpha-actin, TropI and cardiac myosin heavy chain in CM-DiI labelled ASCs. Most importantly, many of these cells showed spontaneous contractions accompanied by calcium transients in culture. Human ASC (hASC) showed synchronous Ca(2+) transient and contraction synchronous with surrounding rat cardiomyocytes (106 beats/min.). Gap junctions also formed between them as observed by dye transfer. In conclusion, cell-to-cell interaction was identified as a key inducer for cardiomyogenic differentiation of hASCs. This method was optimized by co-culture with contracting cardiomyocytes and provides a potential cardiac differentiation system to progress applications for cardiac cell therapy or tissue engineering.

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A Comparative Study of the Effect of Anatomical Site on Multiple Differentiation of Adipose-Derived Stem Cells in Rats.

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