Overexpression of hTERT increases stem-like properties and decreases spontaneous differentiation in human mesenchymal stem cell lines

Abstract

To overcome loss of stem-like properties and spontaneous differentiation those hinder the expansion and application of human mesenchymal stem cells (hMSCs), we have clonally isolated permanent and stable human MSC lines by ectopic overexpression of primary cell cultures of hMSCs with HPV 16 E6E7 and human telomerase reverse transcriptase (hTERT) genes. These cells were found to have a differentiation potential far beyond the ordinary hMSCs. They expressed trophoectoderm and germline specific markers upon differentiation with BMP4 and retinoic acid, respectively. Furthermore, they displayed higher osteogenic and neural differentiation efficiency than primary hMSCs or hMSCs expressed HPV16 E6E7 alone with a decrease in methylation level as proven by a global CpG island methylation profile analysis. Notably, the demethylated CpG islands were highly associated with development and differentiation associated genes. Principal component analysis further pointed out the expression profile of the cells converged toward embryonic stem cells. These data demonstrate these cells not only are a useful tool for the studies of cell differentiation both for the mesenchymal and neurogenic lineages, but also provide a valuable source of cells for cell therapy studies in animal models of skeletal and neurological disorders.

Knockdown of p21Cip1/Waf1 enhances proliferation, the expression of stemness markers, and osteogenic potential in human mesenchymal stem cells

“…The current approach successfully improves the properties of expanded MSCs and therefore can be applied in clinical practices. Methods enhancing the proliferation capacity and potentiality of MSCs include retroviral transduction of the human telomerase gene (Shi et al, 2002), HPV16 E6E7 (Hung et al, 2004) or both (Tsai et al, 2010), and the combined use of different growth factors in a serum-free culture (Reyes & Verfaillie, 2001). All these methods increase the possibilities of enriching cells with oncogenic potential (Serakinci et al, 2004).…”

Immortalization of Human Fetal Hepatocyte by Ectopic Expression of Human Telomerase Reverse Transcriptase, Human Papilloma Virus (E7) and Simian Virus 40 Large T (SV40 T) Antigen Towards Bioartificial Liver Support

“…Further, over expression of hTERT Increase stem cell properties. 15 Recently it has been demonstrated that expression of human telomerase alone is sufficient for the immortalization of various cell types. [16][17][18][19] But cell population without hTERT expression in even in SV 40 expressing will enter into crisis, hence immortalization is imcomplete.…”

Midazolam inhibits chondrogenesis via peripheral benzodiazepine receptor in human mesenchymal stem cells

“…Given that KP cells have been transfected using exogenous genes to maintain MSC stemness,28, 29 whether the incorporated genes produce different responses in these cells remain unclear. To test the inhibitory effects of Midazolam in chondrogenesis, we performed primary hMSCs to evaluate the results that were obtained from the KP MSC cell line.…”

“…KP cells, an immortalized human MSC line, were established from primary human MSCs, which were transduced using a retroviral vector LXSN‐16E6E7 28, 29. KP cells display similar types of cell surface marker profiles as parental primary human MSCs do and retain the stem‐like properties.…”