Immunophenotyping and transcriptional profiling of in vitro cultured human adipose tissue derived stem cells

Abstract

Adipose-derived stem cells (ASCs) have become an important research model in regenerative medicine. However, there are controversies regarding the impact of prolonged cell culture on the ASCs phenotype and their differentiation potential. Hence, we studied 10 clinical ASCs replicates from plastic and oncological surgery patients, in six-passage FBS supplemented cultures. We quantified basic mesenchymal cell surface marker transcripts and the encoded proteins after each passage. In parallel, we investigated the differentiation potential of ASCs into chondrocytes, osteocytes and adipocytes. We further determined the effects of FBS supplementation and subsequent deprivation on the whole transcriptome by comprehensive mRNA and miRNA sequencing. Our results show that ASCs maintain differentiation potential and consistent profile of key mesenchymal markers, with apparent expression of distinct isoforms, in long-term cultures. No significant differences were observed between plastic and oncological surgery cohorts. ASCs in FBS supplemented primary cultures are almost committed to mesenchymal lineages as they express key epithelial-mesenchymal transition genes including early mesenchymal markers. Furthermore, combined mRNA/miRNA expression profiling strongly supports a modulatory role for the miR-30 family in the commitment process to mesenchymal lineages. Finally, we propose improvements to existing qPCR based assays that address alternative isoform expression of mesenchymal markers.

Adipose Tissue-Derived Stem Cells: The Biologic Basis and Future Directions for Tissue Engineering

“…Moreover, harvesting from adipose tissue is less invasive when compared to the bone marrow, resulting in less risk of severe complications and no ethical limitations [ 6 ]. It was established that the positive expression of surface markers CD13, CD29, CD44, CD73, CD90 and CD105, and negative or low production of HLA-DR characterizes MSCs [ 8 , 9 ]. Adipose tissue (AT)-derived MSCs tend to be more heterogeneous [ 10 ] and exhibit immunomodulatory characteristics, in addition to their differentiation ability similar to bone marrow-derived MSCs (BM-MSCs) [ 11 ].…”

Regenerative and Transplantation Medicine: Cellular Therapy Using Adipose Tissue-Derived Mesenchymal Stromal Cells for Type 1 Diabetes Mellitus

“…Therefore, the optimal numbers of cellular passages for ADMSC induction should be investigated to maintain the multipotentiality of the cells, and prevent malignant activation. Among human ADMSCs, cells up to passage 10 appear to retain almost all the characteristics and properties of cells of earlier passage numbers [153,154]. And, although no DNA fragmentation has been found in vitro after expansion past 10 passages, the safety (i.e., chromosome stability) of ADMSCs cannot be ensured [155].…”

Skin giant neoplastic ulcers – treatment options and clinical challenges

“…by polarization of macrophages (M1/M2) or anti-inflammatory effects. A number of scientific and clinical studies on new biomaterials and therapies are currently being conducted around the world in order to achieve even greater efficiency and safety in the treatment of skin defects [12][13][14].…”