Rat embryonic fibroblasts improve reprogramming of human keratinocytes into induced pluripotent stem cells.

Patient-specific human induced pluripotent stem (hiPS) cells not only provide a promising tool for cellular disease models in general, but also open up the opportunity to establish cell-type-specific systems for personalized medicine. One of the crucial prerequisites for these strategies, however, is a fast and efficient reprogramming strategy from easy accessible somatic cell populations. Keratinocytes from plucked human hair had been introduced as a superior cell source for reprogramming purposes compared with the widely used skin fibroblasts. The starting cell population is, however, limited and thereby further optimization in terms of time, efficiency, and quality is inevitable. Here we show that rat embryonic fibroblasts (REFs) should replace mouse embryonic fibroblasts as feeder cells in the reprogramming process. REFs enable a significantly more efficient reprogramming procedure as shown by colony number and total amount of SSEA4-positive cells. We successfully produced keratinocyte-derived hiPS (k-hiPS) cells from various donors. The arising k-hiPS cells display the hallmarks of pluripotency such as expression of stem cell markers and differentiation into all 3 germ layers. The increased reprogramming efficiency using REFs as a feeder layer occurred independent of the proliferation rate in the parental keratinocytes and acts, at least in part, in a non-cell autonomous way by secreting factors known to facilitate pluripotency such as Tgfb1, Inhba and Grem1. Hence, we provide an easy to use and highly efficient reprogramming system that could be very useful for a broad application to generate human iPS cells.

Telomere length, telomerase activity and osteogenic differentiation are maintained in adipose-derived stromal cells from senile osteoporotic SAMP6 mice.

Adipose tissue provides for a rich and easily accessible source of multipotent stromal cells and thus offers the potential for autologous cell-based therapy for a number of degenerative diseases. Senile osteoporosis is characterized by a reduction in bone quality, which is associated with inadequacies in bone marrow stromal cell (BMSC) differentiation. In the present study, we have characterized adipose-derived stromal cells (ASCs) isolated from aged osteoporotic mice and evaluated their suitability as a source of osteogenic precursor cells. Significant reductions in both tibia bone quality and telomere length in liver tissue were observed in the senescence-accelerated mouse prone 6 strain (SAMP6), as compared to the control age-matched senescence-accelerated mouse resistant 1 strain (SAMR1), thus confirming osteoporosis and accelerated ageing traits in this model. ASCs isolated from inguinal fat expressed mesenchymal surface markers and were capable of differentiating along the osteoblast, adipocyte and chondrocyte lineages. Telomere length was not compromised in ASCs from SAMP6 mice but was actually found to be significantly increased as compared to control SAMR1 mice. Furthermore, ASCs from both strains were comparable in terms of telomerase activity, p21 mRNA expression, SA-ß-gal activity and proliferative capacity. The overall osteogenic and adipogenic potential of ASCs was comparable between SAMP6 and SAMR1 strains, as determined by quantitative molecular, biochemical and histological analyses. In conclusion, adipose tissue may represent a promising autologous cell source for the development of novel bone regenerative therapeutic strategies in the treatment of age-related osteoporosis.

Telomerase gene mutations are associated with cirrhosis formation.

UNLABELLED: Telomere shortening impairs liver regeneration in mice and is associated with cirrhosis formation in humans with chronic liver disease. In humans, telomerase mutations have been associated with familial diseases leading to bone marrow failure or lung fibrosis. It is currently unknown whether telomerase mutations associate with cirrhosis induced by chronic liver disease. The telomerase RNA component (TERC) and the telomerase reverse transcriptase (TERT) were sequenced in 1,121 individuals (521 patients with cirrhosis induced by chronic liver disease and 600 noncirrhosis controls). Telomere length was analyzed in patients carrying telomerase gene mutations. Functional defects of telomerase gene mutations were investigated in primary human fibroblasts and patient-derived lymphocytes. An increased incidence of telomerase mutations was detected in cirrhosis patients (allele frequency 0.017) compared to noncirrhosis controls (0.003, P value 0.0007; relative risk [RR] 1.859; 95% confidence interval [CI] 1.552-2.227). Cirrhosis patients with TERT mutations showed shortened telomeres in white blood cells compared to control patients. Cirrhosis-associated telomerase mutations led to reduced telomerase activity and defects in maintaining telomere length and the replicative potential of primary cells in culture. CONCLUSION: This study provides the first experimental evidence that telomerase gene mutations are present in patients developing cirrhosis as a consequence of chronic liver disease. These data support the concept that telomere shortening can represent a causal factor impairing liver regeneration and accelerating cirrhosis formation in response to chronic liver disease.