cells culture human osteosarcoma hos cells  (ATCC)

Journal: International Journal of Clinical and Experimental Pathology

Article Title: TREX1 suppression imparts cancer-stem-cell-like characteristics to CD133 - osteosarcoma cells through the activation of E2F4 signaling

doi:

Figure Lengend Snippet: Expression of TREX1 in human osteosarcoma samples and cell lines. (A) Quantitative real-time PCR analysis of TREX1 mRNA expression was carried out in human osteosarcoma samples classified by differentiation status and histologic grade. Non-metastatic: low-grade (IA [n=5] and IB [n=4]), high-grade (IIA [n=21] and IIB [n=13]); Metastatic: IIIA [n=33] and IIIB [n=18]. For every sample the PCR was carried out in triplicate. Results are expressed as means ± SEM and normalized to IA. *P<0.05 and **P<0.01 versus IA. (B) Immunohistochemical analysis of paraffin-embedded human osteosarcoma samples with polyclonal antibody against TREX1. Representative images are shown. Scale bar indicates 100 μm. The numbers of osteosarcoma samples with varied degrees of TREX1-positive staining are shown in a proportional graph. (C, D) Quantitative real-time PCR (C) and western blotting (D) of TREX1 expression in CD133+ cancer stem cells (CSCs) and CD133- non-CSCs derived from different osteosarcoma cell lines. For (C), results are expressed as means ± SEM and normalized to CD133+ CSCs. n=3, **P<0.01. For (D), representative images are shown. (E) Immunofluorescent staining for TREX1 (red) in CD133+ CSCs and CD133- non-CSCs from the osteosarcoma cell line HOS; Nuclei were stained with DAPI (blue). Representative images are shown. The scale bar indicates 200 μm.

Article Snippet: Cell culture Human osteosarcoma cell lines HOS, MG63, SW1353 and U2OS were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in Dulbecco’s modified Eagle’s medium (DMEM; Sigma-Aldrich, St Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS; Atlanta Biologicals, Lawrenceville, GA, USA), 100 U/ml penicillin and 100 μg/ml streptomycin (Invitrogen, Carlsbad, CA, USA).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Immunohistochemical staining, Staining, Western Blot, Derivative Assay

Journal: International Journal of Clinical and Experimental Pathology

Article Title: TREX1 suppression imparts cancer-stem-cell-like characteristics to CD133 - osteosarcoma cells through the activation of E2F4 signaling

doi:

Figure Lengend Snippet: FACS for CD133- non-CSCs and CD133+ CSCs from human osteosarcoma cell line HOS. Representative images of FACS for CD133- HOS cells and CD133+ HOS cells. P2 represents CD133+ cells and P3 represents CD133- cells.

Article Snippet: Cell culture Human osteosarcoma cell lines HOS, MG63, SW1353 and U2OS were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in Dulbecco’s modified Eagle’s medium (DMEM; Sigma-Aldrich, St Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS; Atlanta Biologicals, Lawrenceville, GA, USA), 100 U/ml penicillin and 100 μg/ml streptomycin (Invitrogen, Carlsbad, CA, USA).

Techniques:

Journal: International Journal of Clinical and Experimental Pathology

Article Title: TREX1 suppression imparts cancer-stem-cell-like characteristics to CD133 - osteosarcoma cells through the activation of E2F4 signaling

doi:

Figure Lengend Snippet: TREX1 knockdown (KD) promotes CSC-like phenotypes of CD133- HOS cells in the dependence of E2F4. (A) The frequencies of CD133+ cells and CD117+Stro-1+ cells were analyzed by flow cytometry (FCM) in CD133- HOS cells transfected with shTREX1 or shScramble, respectively in the presence of siE2F4 or control siRNA (siCon). Representative images are shown. Results are expressed as means ± SEM. n=3, *P<0.05 and **P<0.01. (B) Cell cycle distribution of CD133- HOS cells transfected with or shTREX1 or shScramble were analyzed using propidium iodide staining and FCM. Results are expressed as means ± SEM. n=3. (C-E) Quantitative real-time PCR (C), western blotting (D) and representative immunofluorescent staining (E) of E2F4 expression in CD133- HOS cells transfected with shTREX1 or shScramble. For (C), results are expressed as means ± SEM and normalized to the cells transfected with shScramble. n=3, **P<0.01. For (D) & (E), representative images are shown; the scale bar in (E) indicates 200 μm. (F) Immunohistochemical analysis of paraffin-embedded human osteosarcoma tissue samples with anti-E2F4 antibody. Representative images are shown. The scale bar indicates 100 μm. The numbers of osteosarcoma samples with varied degrees of E2F4-positive staining are shown in a proportional graph.

Article Snippet: Cell culture Human osteosarcoma cell lines HOS, MG63, SW1353 and U2OS were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA), and maintained in Dulbecco’s modified Eagle’s medium (DMEM; Sigma-Aldrich, St Louis, MO, USA) supplemented with 10% fetal bovine serum (FBS; Atlanta Biologicals, Lawrenceville, GA, USA), 100 U/ml penicillin and 100 μg/ml streptomycin (Invitrogen, Carlsbad, CA, USA).

Techniques: Knockdown, Flow Cytometry, Transfection, Control, Staining, Real-time Polymerase Chain Reaction, Western Blot, Expressing, Immunohistochemical staining