Abstract
Next-generation sequencing (RNAseq) analysis of gene expression changes during the long-term in vitro culture and osteogenic differentiation of ASCs remains to be important, as the analysis provides important clues toward employing stem cells as a therapeutic intervention. In this study, the cells were isolated from adipose tissue obtained during routine surgical procedures and subjected to 14-day in vitro culture and differentiation. The mRNA transcript levels were evaluated using the Illumina platform, resulting in the detection of 19,856 gene transcripts. The most differentially expressed genes (fold change >|2|, adjusted p value < 0.05), between day 1, day 14 and differentiated cell cultures were extracted and subjected to bioinformatical analysis based on the R programming language. The results of this study provide molecular insight into the processes that occur during long-term in vitro culture and osteogenic differentiation of ASCs, allowing the re-evaluation of the roles of some genes in MSC progression towards a range of lineages. The results improve the knowledge of the molecular mechanisms associated with long-term in vitro culture and differentiation of ASCs, as well as providing a point of reference for potential in vivo and clinical studies regarding these cells’ application in regenerative medicine.
Highlights
It is well known that the advancement of modern medicine will be highly dependent on the development of knowledge regarding stem cells [1,2,3,4,5,6]
RNAseq analysis was used to determine the transcriptomic changes associated with long-term in vitro cultures of adipose-derived stem cells (ASCs), as well as their induced osteogenic differentiation
Next-generation sequencing (NGS) methods, such as RNAseq, allowed deeply investigating the molecular basis of various cell- and tissue-associated processes, especially when it comes to in vitro cultured cells [22]
Summary
It is well known that the advancement of modern medicine will be highly dependent on the development of knowledge regarding stem cells [1,2,3,4,5,6]. While some studies focus on embryonic or induced pluripotent stem cells, the limited knowledge of stem cell regulatory mechanisms, as well as the significant risks associated with their administration, effectively delays any therapeutic approaches based on these cell types [7,8]. Adult progenitor cells, such as those derived from the bone marrow, have been effectively used in treatment of hematopoiesis-associated diseases (e.g., leukemia) since the 1970s [9,10]. The canine (Canis familiaris) source of stem cells is a relatively widely available model for human studies while allowing for understanding canine stem cells as a potentially effective treatment in veterinary medicine [4,20,21]
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