Heterogeneity of osteosarcoma cell lines led to variable responses in reprogramming.

Four osteosarcoma cell lines, Saos-2, MG-63, G-292 and U-2 OS, were reprogrammed to pluripotent state using Yamanaka factors retroviral transduction method. Embryonic stem cell (ESC)-like clusters started to appear between 15 to 20 days post transduction. Morphology of the colonies resembled that of ESC colonies with defined border and tightly-packed cells. The reprogrammed sarcomas expressed alkaline phosphatase and pluripotency markers, OCT4, SSEA4, TRA-1-60 and TRA-1-81, as in ESC up to Passage 15. All reprogrammed sarcomas could form embryoid body-like spheres when cultured in suspension in a low attachment dish for up to 10 days. Further testing on the directed differentiation capacity of the reprogrammed sarcomas showed all four reprogrammed sarcoma lines could differentiate into adipocytes while reprogrammed Saos-2-REP, MG-63-REP and G-292-REP could differentiate into osteocytes. Among the 4 osteosarcoma cell lines, U-2 OS reported the highest transduction efficiency but recorded the lowest reprogramming stability under long term culture. Thus, there may be intrinsic differences governing the variable responses of osteosarcoma cell lines towards reprogramming and long term culture effect of the reprogrammed cells. This is a first report to associate intrinsic factors in different osteosarcoma cell lines with variable reprogramming responses and effects on the reprogrammed cells after prolonged culture.

Reprogramming cancer cells: overview & current progress.

INTRODUCTION: Cancer is a disease with genetic and epigenetic origins, and the possible effects of reprogramming cancer cells using the defined sets of transcription factors remain largely uninvestigated. In the handful of publications available so far, findings have shown that reprogramming cancer cells changed the characteristics of the cells to differ from the parental cancer cells. These findings indicated the possibility of utilizing reprogramming technology to create a disease model in the laboratory to be used in studying the molecular pathogenesis or for drug screening of a particular cancer model. AREAS COVERED: Despite numerous methods employed in generating induced pluripotent stem cells (iPSCs) from cancer cells only a few studies have successfully reprogrammed malignant human cells. In this review we will provide an overview on i) methods to reprogram cancer cells, ii) characterization of the reprogrammed cancer cells, and iii) the differential effects of reprogramming on malignancy, epigenetics and response of the cancer cells to chemotherapeutic agents. EXPERT OPINION: Continued technical progress in cancer cell reprogramming technology will be instrumental for more refined in vitro disease models and ultimately for the development of directed and personalized therapy for cancer patients in the future.

Time-dependent dynamic mobilization of circulating progenitor cells during percutaneous coronary intervention in diabetics.

BACKGROUND: Circulating progenitor cells (CPC) especially endothelial progenitor cell (EPC) levels and functions are attenuated in diabetic patients. This may explain the poorer outcome of diabetics undergoing percutaneous coronary intervention (PCI). We aim to study the dynamic changes of these cells in these patients. METHODS: Blood of 8 diabetics with stable coronary artery disease who underwent elective PCI, were obtained at baseline, 1, 4 and 24 h after PCI. Fluorescence activated cell sorting (FACS) analysis was performed to quantitate CD34+ and CD34+/ KDR+ cells. Patients with recent acute coronary syndrome were excluded. RESULTS: After PCI, decreases in CPC from baseline were detected in 7 out of the 8 patients. In these 7 patients, mean CD34+ and CD34+/KDR+ cells were 182+/-99/1 x 10(5) and 18+/-16/1 x 10(5) cells respectively. Maximal decrease of CD34+ and CD34+/KDR+ cells were 47.8% and 53.3% at 1 h and 4 h respectively. At 24 h, CPC levels returned to baseline but were not elevated. The only patient with raised cardiac enzymes has instead, 2 to 3 fold increase in CPCs at 1 and 4 h. CONCLUSIONS: We found a transient dip in circulating progenitors early during PCI. This suggests incorporation of the cells into the sites of vascular denudation. The absence of subsequent CPC elevation post-PCI in diabetes may be associated with known poorer outcome of these patients. With myocardial injury, more progenitors may be mobilized from the bone marrow into the circulation and abolish the hyperacute reduction in circulating levels.

Inhibition of CD44 expression in hepatocellular carcinoma cells enhances apoptosis, chemosensitivity, and reduces tumorigenesis and invasion.

PURPOSE: CD44 is overexpressed in various tumors including hepatocellular carcinoma (HCC). The purpose of this study was to examine the effects of CD44 antisense oligonucleotide (ASO) alone or combination with doxorubicin on HCC cells in vitro. METHODS: Cytotoxicity was measured by use of a cell viability assay in HCC cell line SNU-449. Tumorigenesis and invasion were accessed by colony formation, growth in soft agar and ECMatrix invasion assay. Apoptosis and necrosis were evaluated by using double staining with Hoechst 33342 and propidium iodide. Protein expression and mRNA level were detected by Western blot and RT-PCR. RESULTS: We have designed novel CD44 ASO, which can effectively down-regulate CD44 expression in SNU-449. Colony formation, growth in soft agar and invasion were significantly impaired after CD44 ASO treatment in SNU-499. In company with CD44 down-regulated by CD44 ASO, MDR-1 and Bcl-2 expression were also greatly reduced. CD44 ASO also increased chemosensitivity to doxorubicin significantly, lowered IC(50 )by one order of magnitude. Apoptosis and necrosis were also induced by CD44 ASO alone or in combination treatment with doxorubicin. CONCLUSIONS: Inhibition of CD44 expression by CD44 ASO significantly induced apoptosis, decreased tumorigenesis and invasion, and increased chemosensitivity. Thus, CD44 ASO is potentially a therapy that is worth investigating in the clinical setting.