cbx8 shrna  (OriGene)

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: Ectopic expression of CBX6 or CBX8, but not of other CBX members, represses HKESC1 cells invasion ability, and CBX8 expression is reduced in patients with lymph node metastasis. (A) HKESC1 cells were transfected with Vector or Flag-tagged CBXs for 36 h and then subjected to Western blotting. GAPDH was the loading control. (B) Cell invasion assay was performed in the indicated cells as described in the Materials and methods. Representative images (100 ×) are presented (right panel). The results are expressed as the mean ± SD of three independent experiments (left panel). *** p < 0.001 (Student's t-test). (C) The relative mRNA levels of CBX8 in 27 and 22 patients with (LN+) and without (LN-) lymph node metastasis was detected by qRT-PCR, respectively. p = 0.0132 (Mann-Whitney test).

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques: Expressing, Transfection, Plasmid Preparation, Western Blot, Invasion Assay, Quantitative RT-PCR, MANN-WHITNEY

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: CBX8 plays paradoxical roles in ESCC, suppressing cell migration and invasion while promoting proliferation. (A) The expression of CBX8 in ESCC cell lines was detected at mRNA and protein levels by qRT-PCR and Western blotting (insertion panel), respectively. An immortalized esophageal epithelial cell line NE-1 was used as control. Data from three independent experiments are expressed as mean ± SD. ** p < 0.01 and *** p < 0.001, Student's t-test. (B, C) Both the cell migration and invasion assays were performed in the indicated stable cell lines as described in the Materials and methods. Representative images (100 ×) are shown. The columns, mean of three independent experiments; the bars, SD. ** p < 0.01 and *** p < 0.001 using Student's t-test. (D, E) The cell viability of the indicated stable cell lines was measured in different time points as indicated by MTT assay. Data from three independent experiments are expressed as mean ± SD. * p < 0.05 and ** p < 0.01, Student's t-test.

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques: Migration, Expressing, Quantitative RT-PCR, Western Blot, Stable Transfection, MTT Assay

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: CBX8 suppresses the metastasis of ESCC cells in nude mice. (A, D) The effects of CBX8 on tumor metastasis were evaluated by the tail vein-lung metastasis model. Left panel, representative images of lungs derived from mice injected with the indicated stable cell lines; right panel, H&E staining (Scale bars = 50 μm) of pulmonary sections from the tumor-bearing mice. (B, C, E, F) Numbers (B, E) and wet weights (C, F) of metastatic nodules per lung in the nude mice were summarized (n = 6). The bars indicate the SD. A two-tailed Student's t-test was used for statistical analysis.

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques: Derivative Assay, Injection, Stable Transfection, Staining, Two Tailed Test

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: CBX8 inhibits EMT in ESCC cells. (A) TE-1 cells stably expressing NC or shRNA-CBX8 as indicated were analyzed by immunofluorescence assay for phalloidin (red) with DAPI (blue) counterstaining. (B, C) Relative expressions of the indicated molecules were detected by qRT-PCR (B) and Western blotting (C) in the indicated stable cell lines, respectively. The results are expressed as the mean ± SD of three independent experiments. * p < 0.05, ** p < 0.01 and *** p < 0.001, Student's t-test.

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques: Stable Transfection, Expressing, shRNA, Immunofluorescence, Quantitative RT-PCR, Western Blot

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: CBX8 directly binds to and down-regulates the Snail promoter activity. (A, B) The indicated stable cell lines were transfected with Snail or Slug promoter linked to luciferase for 48 h and then subjected to the luciferase assay as described in Materials and Methods. (C) The upper panel, a schematic illustration of the Snail promoter regions (1-16) with (+) or without (-) binding affinity for CBX8. The arrow indicates the transcriptional start site (TSS). The lower panel, the result of ChIP-quantitative PCR analysis of CBX8 binding to the distinct regions in the Snail promoter, showing enrichment with CBX8 antibody compared with IgG control. The p16 and GAPDH promoters were used as the positive and negative controls, respectively. The results are expressed as the mean ± SD of individual samples from three independent experiments. ** p < 0.01 and *** p < 0.001, Student's t-test. (D) There was a reverse correlation between CBX8 and Snail at mRNA level in ESCC tissue samples. p = 0.0215, linear regression analysis.

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques: Activity Assay, Stable Transfection, Transfection, Luciferase, Binding Assay, Real-time Polymerase Chain Reaction

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: The inhibitory effect of CBX8 on cell migration and invasion depends on the down-regulation of Snail. (A, B) Both the cell migration and invasion assays were performed in the indicated stable cell lines as described in the Materials and methods. Representative images (100 ×) are shown. The columns, mean of three independent experiments; The bars indicate the SD. ** p < 0.01 and *** p < 0.001 using Student's t-test. (C-E) An in vivo lung metastasis model was established in nude mice using TE-1 cells stably expressing NC, shControl, shCBX8, shSnail or both, as indicated, as described in Materials and Methods. (C) Representative images of gross and H&E staining (Scale bars = 50 μm) of metastatic lung nodules. Numbers (D) and wet weights (E) of metastatic nodules per lung in the nude mice were summarized (n = 5). The bars indicate the SD. A two-tailed Student's t-test was used for statistical analysis.

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques: Migration, Stable Transfection, In Vivo, Expressing, Staining, Two Tailed Test

Journal: Theranostics

Article Title: CBX8 Suppresses Tumor Metastasis via Repressing Snail in Esophageal Squamous Cell Carcinoma

doi: 10.7150/thno.20717

Figure Lengend Snippet: The proposed model for the functions of CBX8 in ESCC metastasis and proliferation. CBX8 may down-regulate Snail to increase E-cadherin and to decrease N-cadherin, Fibronectin and Vimentin, which in turn impair EMT and consequently repress metastasis in ESCC. In the meanwhile, CBX8 may positively modulate the ESCC cell cycle progression and proliferation as indicated by Xiao et al . .

Article Snippet: The CBX8 and Snail short hairpin RNA (shRNA) pLKO1 lentiviral constructs were purchased from Sigma Addrich.

Techniques:

Journal: Cancer Cell International

Article Title: Correlations between chromobox homolog 8 and key factors of epithelial–mesenchymal transition in hepatocellular carcinoma

doi: 10.1186/s12935-019-1063-z

Figure Lengend Snippet: CBX8 is correlated with HCC metastasis. a Representative CBX8 protein expression was detected in adjacent non-tumor tissues, not metastatic and metastatic HCC tissues by immunohistochemical analysis at ×50 (upper panel) and ×400 magnification (lower panel). b Comparison of mRNA expression of CBX8 in adjacent non-tumor tissues, not metastatic and metastatic HCCs. ** P < 0.01 is based on the Student t test compared to adjacent non-tumor tissues. c Comparison of protein expression of CBX8 in adjacent non-tumor tissues, not metastatic and metastatic HCCs. ** P < 0.01 is based on the Student t test compared to adjacent non-tumor tissues

Article Snippet: Four lentiviral shRNAs targeting CBX8 (shCBX8) were synthesized on a GV248-puro vector (Genechem, Shanghai, China).

Techniques: Expressing, Immunohistochemical staining

Journal: Cancer Cell International

Article Title: Correlations between chromobox homolog 8 and key factors of epithelial–mesenchymal transition in hepatocellular carcinoma

doi: 10.1186/s12935-019-1063-z

Figure Lengend Snippet: CBX8 promotes the metastasis of HCC cells. a CBX8 protein expression in L02 cell line and HCC cell lines as indicated was detected by Western blot (right panel, gray scan results normalized to β-actin). b CBX8 mRNA expression in SK-Hep-1 cells introduced with four specific shRNAs targeted CBX8 and a control shGFP by semi-quantitative RT-PCR (right panel, gray scan results normalized to GAPDH). c Invasion and migration ability of CBX8 knock-down cells was analyzed by wound healing. ** P < 0.01 is based on the Student t test compared to shGFP cells. d Invasion and migration ability of CBX8 knock-down cells was analyzed by Transwell assay. ** P < 0.01 is based on the Student t test compared to shGFP cells. All results are from three independent experiments

Article Snippet: Four lentiviral shRNAs targeting CBX8 (shCBX8) were synthesized on a GV248-puro vector (Genechem, Shanghai, China).

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Migration, Transwell Assay

Journal: Cancer Cell International

Article Title: Correlations between chromobox homolog 8 and key factors of epithelial–mesenchymal transition in hepatocellular carcinoma

doi: 10.1186/s12935-019-1063-z

Figure Lengend Snippet: CBX8 induces EMT. a Expression of EMT markers as indicated were detected in SK-Hep-1-shCBX8 cells by Western blot. b Expression of EMT markers as indicated were detected in SK-Hep-1-shCBX8 cells by immunofluorescence

Article Snippet: Four lentiviral shRNAs targeting CBX8 (shCBX8) were synthesized on a GV248-puro vector (Genechem, Shanghai, China).

Techniques: Expressing, Western Blot, Immunofluorescence

Journal: Cancer Cell International

Article Title: Correlations between chromobox homolog 8 and key factors of epithelial–mesenchymal transition in hepatocellular carcinoma

doi: 10.1186/s12935-019-1063-z

Figure Lengend Snippet: The correlation between CBX8 and EMT factors in HCC tissues

Article Snippet: Four lentiviral shRNAs targeting CBX8 (shCBX8) were synthesized on a GV248-puro vector (Genechem, Shanghai, China).

Techniques:

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a The expression of CBX8 in HCC and adjacent normal liver tissues of 83 paired samples was assessed by qRT-PCR, representative results are shown, and the comparison of the expression levels of CBX8 mRNA between non-metastatic ( n = 42) and metastatic ( n = 41) HCCs. b CBX8 protein expression for the 83 paired samples was analyzed by western blotting, representative results are shown, and the comparison of the relative expression levels of CBX8 protein between non-metastatic and metastatic HCCs. c CBX8 protein expression was analyzed by IHC analysis of 153 paired HCC and normal liver tissues, epresentative results are shown. Semiquantification of CBX8 expression was assessed by IHC of 153 adjacent tissues and primary HCC tissues with or without distant metastasis. d The association between CBX8 expression in HCC as determined by IHC and survival time of 153 patients was analyzed by Kaplan–Meier survival analysis. ** p < 0.01 compared with adjacent tissues; ## p < 0.01 compared with the tissues without metastasis. CBX8 Chromobox homolog 8, HCC hepatocellular carcinoma, qRT-PCR quantitative reverse transcription-PCR, mRNA messenger RNA, IHC immunohistochemistry

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Expressing, Quantitative RT-PCR, Comparison, Western Blot, Reverse Transcription, Immunohistochemistry

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: Correlation between CBX8 expression and clinical pathological characteristics of HCC patients

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Expressing, Staining

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a Proliferation of HepG2-CBX8, Huh7-CBX8, and their control cells was examined by CCK-8 assay. b Proliferation of SMMC-7721-shCBX8, Sk-Hep-1-shCBX8, and their control cells was examined by CCK-8 assay. c Images (left), growth (middle), and weight (right) of tumors following subcutaneous injection of HepG2-CBX8 or control cells. d Images (left), growth (middle), and weight (right) of tumors following subcutaneous injection of SMMC-7721-shCBX8 or control cells. e Representative images of CBX8 and Ki-67 immunostaining in HepG2-CBX8 and the control xenografts. f Ki-67 immunostaining in SMMC-7721-shCBX8 and the control xenografts. ** p < 0.01. CBX8 Chromobox homolog 8, HCC hepatocellular carcinoma, CCK-8 Cell Counting Kit-8

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Control, CCK-8 Assay, Injection, Immunostaining, Cell Counting

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a Representative phase-contrast images of HepG2 and Huh7 cells showing that CBX8 overexpression modulates the expression of E-cadherin and N-cadherin as analyzed by immunofluorescence staining. b Expression of epithelial (E-cadherin) and mesenchymal (N-cadherin, vimentin, Slug, and Snail) markers were analyzed by western blotting in HepG2 and Huh7 cells. c Expression of epithelial (E-cadherin) and mesenchymal (N-cadherin, vimentin, Slug, and Snail) markers were analyzed by western blotting in SMMC-7721 and Sk-Hep-1 cells. d Representative phase-contrast images of SMMC-7721 and Sk-Hep-1 cells showing that CBX8 shRNA modulates the expression of E-cadherin and N-cadherin as analyzed by immunofluorescence staining. CBX8 Chromobox homolog 8, HCC hepatocellular carcinoma, shRNA short hairpin RNA

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Over Expression, Expressing, Immunofluorescence, Staining, Western Blot, shRNA

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a HepG2-CBX8 and the control cells were subjected to transwell migration and Matrigel invasion assays. Quantification of migrated cells through the membrane and invaded cells through either the membrane or Matrigel for each cell line are shown as a proportion of the vector control. b SMMC-7721-shCBX8 and the control cells were subjected to transwell migration and Matrigel invasion assays. Quantification of migrated cells through either the membrane or Matrigel for each cell lines is shown as proportions of their controls. c Number of metastatic foci per section in lungs from individual mouse with injection of HepG2-CBX8 or its control cells were determined. A representative stain and quantification of the results are shown. d Number of metastatic foci per section in lungs from individual mice injected with SMMC-7721-shCBX8 or its control cells were determined. A representative stain and quantification of the results are shown. ** p < 0.01. CBX8 Chromobox homolog 8, HCC hepatocellular carcinoma

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Control, Migration, Membrane, Plasmid Preparation, Injection, Staining

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a Holoclone assays in HepG2 cells transfected with pBabe or pBabe-CBX8, and SMMC-7721 cells transfected with pSuper or pSuper-shCBX8 were used in three experiments (Exp. I, 100 cells/well scored on d 9; Exp. II, 100 cells/well scored on d 13; Exp. III, 500 cells/well scored on d 7). b Clonogenic assay (Exp. I, 1250 cells/well scored on d 5; Exp. II, 25,000 cells/well scored on d 5) in HepG2 cells transfected with pBabe or pBabe-CBX8 and SMMC-7721 cells transfected with pSuper or pSuper-shCBX8. Two experiments were performed (Exp. I, 1250 cells/well scored on d 5; Exp. II, 25,000 cells/well scored on d 5). c SP population in CBX8 overexpression and silenced HCC cells was determined by Hoechst 33342 efflux assay. d The expression of CBX8 mRNA in sorted primary EPCAM− and EPCAM+ liver cancer cells was determined by qRT-PCR (left). The expression of CBX8 mRNA of sorted primary CD133− and CD133+ liver cancer cells was determined by qRT-PCR (right). e Levels of CBX8 mRNA in magnetically sorted EPCAM+ and EPCAM− were measured by qRT-PCR in HepG2 and SMMC-7721 cells (left). Levels of CBX8 mRNA in magnetically sorted CD133+ and CD133− were measured by qRT-PCR in HepG2 and SMMC-7721 cells (right). ** p < 0.01. CBX8 Chromobox homolog 8, HCC hepatocellular carcinoma, qRT-PCR quantitative reverse transcription-PCR, mRNA messenger RNA

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Transfection, Clonogenic Assay, Over Expression, Expressing, Quantitative RT-PCR, Reverse Transcription

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a Supervised hierarchical clustering of the differentially expressed genes after CBX8 silencing in SMMC-7721 cells. b BMP4 mRNA expression was measured by qRT-PCR and BMP4 protein expression was measured by western blotting in HCC cells with CBX8 overexpression or knockdown. c H3K4me3, H3K9me3, and H3k27me3 was measured by western blotting. d Presentation of five regions relative to the BMP4 transcriptional start site used as primers to test histone occupied abundance. e BMP4 is one of the genes that bind on CBX8 protein by ChIP-seq. f The relative methylation of the BMP4 promoter in HCC tissues. Representative results are shown between adjacent liver tissues and HCC tissues, and the comparison of the relative methylation of the BMP4 promoter between HCC tissues with or without metastasis. ** p < 0.01. CBX8 Chromobox homolog 8, BMP4 bone morphogenetic protein 4, mRNA messenger RNA, qRT-PCR quantitative reverse transcription-PCR, ChIP-seq chromatin immunoprecipitation-sequencing

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Over Expression, Knockdown, ChIP-sequencing, Methylation, Comparison, Reverse Transcription

Journal: Cell Death & Disease

Article Title: CBX8 exhibits oncogenic properties and serves as a prognostic factor in hepatocellular carcinoma

doi: 10.1038/s41419-018-1288-0

Figure Lengend Snippet: a Expression of (p-) Smad1/5/8 were measured by western blotting. b Expression of (p-) ERK and (p-) JNK were measured by western blotting. c – e Expression of BMP4, (p-) ERK, (p-) JNK, (p-) Smad1/5/8, E-cadherin, N-cadherin, Vimentin, Slug, and Snail were measured by western blotting. f BMP4 expression was positively correlated with CBX8 expression in HCC tissues. g Schematic illustration of the proposed function of CBX8 in HCC metastasis and stemness. CBX8 Chromobox homolog 8, BMP4 bone morphogenetic protein 4, EMT epithelial–mesenchymal transition, MAPK mitogen-activated protein kinase, p- phosphorylated, ERK extracellular signal-regulated kinase, JNK c-JUN N-terminal kinase, HCC hepatocellular carcinoma

Article Snippet: pBabe.puro retroviral constructs containing human CBX8 and BMP4 cDNAs and pSuper.retro.puro retroviral constructs containing short hairpin RNA (shRNA) against human CBX8 were prepared by GenePharma, Shanghai, China (shCBX8#1, 5′-GCAUGGAAUACCUCGUGAATT-3′; shCBX8#2, 5′-CGAGUUUCGAAGUGACUCATT-3′; shCBX8#3, 5′-CUUCGAAACAUGGGUUUGUTT-3′).

Techniques: Expressing, Western Blot

Journal: International Journal of Clinical and Experimental Pathology

Article Title: CBX8, a novel DNA repair protein, promotes tumorigenesis in human esophageal carcinoma

doi:

Figure Lengend Snippet: Expression of CBX8 in esophageal tissues and cell lines. A. CBX8 mRNA levels were measured in 59 pairs of esophageal carcinoma and corresponding adjacent normal tissues by real-time RT-PCR, where 18S was used as an internal reference. The line in each box is the median -ΔCt value, while the upper and lower edges of each box show the 75th and 25th percentile, respectively. P value was calculated by paired t-test (P<0.01). B. The difference in CBX8 expression was tested by western blot analysis. β-Actin was the loading control. N represents adjacent normal tissues, T represents carcinoma tissues. C. Representative immunohistochemical detection of the CBX8 protein expression in paired esophageal tissues, N represents adjacent normal tissues, T represents carcinoma tissues. Left: hematoxylin-eosin staining (H&E); Right: immunohistochemistry. D. CBX8 expression patterns in normal cell lines and ESCC cell lines.

Article Snippet: For stable CBX8 depletion, the cells were infected with lentivirus (LV) that expressed CBX8-shRNA (Genchem, China) or its negative control at 30-50% confluence.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Immunohistochemical staining, Staining, Immunohistochemistry

Journal: International Journal of Clinical and Experimental Pathology

Article Title: CBX8, a novel DNA repair protein, promotes tumorigenesis in human esophageal carcinoma

doi:

Figure Lengend Snippet: Association between CBX8 expression and clinicopathologic information of ESCC patients

Article Snippet: For stable CBX8 depletion, the cells were infected with lentivirus (LV) that expressed CBX8-shRNA (Genchem, China) or its negative control at 30-50% confluence.

Techniques: Expressing

Journal: International Journal of Clinical and Experimental Pathology

Article Title: CBX8, a novel DNA repair protein, promotes tumorigenesis in human esophageal carcinoma

doi:

Figure Lengend Snippet: CBX8 is required for the proliferation of ESCC cells and cell cycle progression. (A) The interference efficiency of two CBX8-siRNAs was tested by both RT-PCR and western blot. (B, C) The relative cell proliferation was measured by MTT (B) and colony formation assays (C) in EC109 and EC9706 cells. (D) Xenograft mouse model. Tumor volume and weight were measured for 30 days after subcutaneous injection. n=5 per group. (E) FACS analysis showed that CBX8-depleted cells accumulated in the G2M phase in EC109. (F) G2M checkpoint-related proteins were detected in EC109 cells. (NC: Negative Control).

Article Snippet: For stable CBX8 depletion, the cells were infected with lentivirus (LV) that expressed CBX8-shRNA (Genchem, China) or its negative control at 30-50% confluence.

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Injection, Negative Control

Journal: International Journal of Clinical and Experimental Pathology

Article Title: CBX8, a novel DNA repair protein, promotes tumorigenesis in human esophageal carcinoma

doi:

Figure Lengend Snippet: CBX8 is related to DNA damage. (A) CBX8-depletion spontaneously induces DNA damage. EC109 cells were transfected with the indicated siRNA for 48 h, exposed to 10 Gy of IR or 0.1 mM H2O2 for 20 min, and subjected to neutral comet analysis. (B) Quantification of tail moments using cells from (A). Tail moments for each condition were calculated on a minimum of 300 cells for each data point. Three independent experiments have been done for each result. *P<0.05, without IR or H2O2 treatment; **P<0.01, IR treatment or H2O2 treatment. (C) A marker for DNA DSBs, γ-H2AX, was tested by western blot both in EC109 and EC9706 cells. (D) EC109 cells transfected with NC or siCBX8 were fixed and immunostained with anti-γ-H2AX antibody.

Article Snippet: For stable CBX8 depletion, the cells were infected with lentivirus (LV) that expressed CBX8-shRNA (Genchem, China) or its negative control at 30-50% confluence.

Techniques: Transfection, Marker, Western Blot

Journal: International Journal of Clinical and Experimental Pathology

Article Title: CBX8, a novel DNA repair protein, promotes tumorigenesis in human esophageal carcinoma

doi:

Figure Lengend Snippet: Depletion of CBX8 decreases resistance to H2O2 or IR. A, B. Both EC109 and HeLa cells were treated with H2O2 or IR. For H2O2 treatment, cells were exposed to 0.1 mM H2O2 for 20 min and then maintained in culture for indicated time points at 37°C, 5% CO2. For IR exposure, cells were exposed to 10 Gy IR and maintained in culture for indicated time points at 37°C, 5% CO2. CBX8 and γ-H2AX were determined by Western blot. C. EC109 cells were transfected with siCBX8 for 48 h, then exposed to 0.1 mM H2O2 for 20 min, and subjected to neutral comet assay at the indicated time points. D. EC109 cells were transfected with siCBX8; exposed to 0.1 mM H2O2 for 20 min; and maintained in culture for indicated time points. CBX8 and γ-H2AX levels were monitored by western blot analysis; β-actin was used as an internal reference. Results indicate that CBX8 depletion increases sensitivity of EC109 cells to H2O2.

Article Snippet: For stable CBX8 depletion, the cells were infected with lentivirus (LV) that expressed CBX8-shRNA (Genchem, China) or its negative control at 30-50% confluence.

Techniques: Western Blot, Transfection, Neutral Comet Assay