ObjectiveTo detect the expressions of CD133 in four gastric cancer cell lines (KATO-Ⅲ, SGC7901, AGS, and MKN-45) and investigate the different biological characteristics of CD133 positive (CD133+) cells and CD133 negative (CD133-) cells in the KATO-Ⅲ cell lines. MethodsThe CD133 gene and protein expressions of four gastric cancer cell lines were evaluated by semiquantitative RT-PCR and Western blot, respectively. CD133+ cells in KATO-Ⅲ cell lines were isolated by magnetic activated cell sorting (MACS) and examined in morphology, growth characteristics, proliferation, and differentiation in vitro. The sensitivity of different concentrations (0.05, 0.10, 0.20, 0.50, and 1.00 mg/ml) 5-fluoropyrimidinedione (5-FU) were contrasted by drug sensitivity testing in vitro (Cell Counting Kit 8-assay) between CD133+ cells and CD133- cells. ResultsThe expressions of CD133 gene and protein in the KATO-Ⅲ cell lines were significantly higher than those in the other three cell lines (Plt;0.05). CD133+ cells produced spheroid colonies in serum-free medium culture and were ber abilities of proliferation and differentiation than those of CD133- cells in vitro. The inhibitor rate of the CD133+ cells at concentration of 0.50 mg/ml was lower than that of CD133- cells (Plt;0.05). ConclusionsCell population with CD133+ in the KATO-Ⅲ cell lines have b ability of cloning, better capability of proliferation and differentiation, as well as anticancer drug resistance to 5-FU. CD133 can be applied as one of surface markers for the detection to gastric cancer initiator cells.
Objective To investigate the prognostic value of epithelial-mesenchymal transition （EMT） related proteins （Snail， E-cadherin， and N-cadherin） in gastric cancer and its relationship with tumor initiating cells （TICs） marker （CD133）. Methods The expressions of EMT-related proteins and CD133 protein in the gastric cancer tissues and normal gastric mucosa tissues adjacent to gastric cancer were detected by Western blot method. The relations between the expressions of EMT-related factors proteins and CD133 protein and the clinicopathologic characters were analyzed. The correlations between EMT-related factors and CD133 were analyzed by Spearman. The correlations between EMT-related factors expressions and CD133 expression and survival were analyzed by Kaplan-Meier method and Log-rank test. Results ① The protein expression levels of Snail， N-cadherin， and CD133 in the gastric cancer tissues were significantly higher than those in the normal gastric mucosa tissues adjacent to gastric cancer （Snail：0.599±0.114 versus 0.259±0.108， P=0.020；N-cadherin：0.754±0.154 versus 0.329±0.134， P=0.001；CD133：0.635±0.119 versus 0.485±0.116， P=0.029）， while the protein expression level of E-cadherin was lower than that in the normal gastric mucosa tissues adjacent to gastric cancer （0.378±0.123 versus 0.752±0.156， P=0.003）.② The expression levels of Snail and N-cadherin in the gastric cancer patients with vascular invasion， lymphatic vessel invasion，N3 lymph node metastasis， diameter more than 5 cm， and Ⅲ＋Ⅳ staging were significantly higher than those in the patients without vascular invasion， lymphatic vessel invasion， N0-N2 lymph node metastasis， diameter less than 5 cm， andⅠ＋Ⅱ staging（P＜0.05）， while E-cadherin protein expression was lower than that in the patients without vascular invasion， lymphatic vessel invasion， N0-N2 lymph nodes metastasis， andⅠ＋Ⅱstaging （P＜0.05）. The expression levels of CD133 in the gastric cancer patients with lymphatic vessel invasion， diameter more than 5 cm， and Ⅲ＋Ⅳ staging were significantly higher than those in the patients without lymphatic vessel invasion， diameter less than 5 cm， andⅠ＋Ⅱ staging （P＜0.05）. ③The Snail and N-cadherin protein expressions were significantly positive correlated with CD133 protein expression， respectively （rs=0.278， P=0.048；rs=0.406， P=0.003）， while E-cadherin protein expression was significantly negative correlated with CD133 protein expression （rs=-0.504， P=0.000）.④ The survival time in the patients with lower expressions of Snail， N-cadherin， and CD133 were significantly longer than those in the patients with higher expressions of Snail， N-cadherin， and CD133 （P＜0.05）. The combination of Snail， N-cadherin， E-cadherin， and CD133 could effectively predict survival. Conclusions There is a significant correlation between EMT and gastric cancer TICs， and which are correlated with aggressive clinicopathologic features of gastric cancer. The combination of Snail， E-cadherin， N-cadherin， and CD133 may be effectively predict the prognosis of gastric cancer patients.
Objective To study the expression of CD133 mRNA in peripheral blood mononuclear cells （PBMCs） of patients with gastric adenocarcinoma （GC） before operation or after operation and its clinical significance. To learn the relationship of CD133 expressions in PBMCs to primary lesion of GC. Methods Fifty patients with GC，10 patients with gastric ulcer （GU）， and 10 healthy volunteers were registered in this research. Expressions of CD133 mRNA in PBMCs and in primary lesion of GC by semi-quantitative RT-PCR and expression of CD133 protein in primary lesion of GC by immunohistochemical staining were detected. Correlations of CD133 mRNA expression with clinicopathologic parameters and postoperative survival rate were analyzed. Relation between CD133 mRNA level and CD133 protein expression or lymphatic metastasis were assessed too. Results The brightness scale value （ABSV） of CD133 mRNA in PBMCs of the patients with GC before operation （0.270±0.163） was higher than that in the healthy volunteers （0.029±0.060） or in the patients with GU （0.059±0.099） （P=0.000）. The ABSV of CD133 mRNA in the PBMCs of patients with GC before operation was related to poor cell differentiation （P=0.002）， lymph vessel invasion （P=0.028），deeper tumor invasion （P=0.041），later lymph node metastasis stage （P=0.010），later TNM stage （P=0.006），and positive expression of CD133 protein in the primary lesion （P=0.011）. Relative analysis revealed that the ABSV of CD133 mRNA in the PBMCs of patients with GC before operation was related positively to metastatic lymphatic nodes ratio （rs=0.422，P=0.002），metastatic lymph node number （rs=0.398，P=0.004），and ABSV of CD133 mRNA in the primary lesion of GC （rs=0.337，P=0.017）. The ABSV of CD133 mRNA in the PBMCs of patients with GC after operation obtained from blood sample at 1 week after curative resection was higher than that in the patients before operation （P=0.021）. Patients suffered from deeper invasion inclined to have a higher ABSV of CD133 mRNA after surgery （P=0.039）. Higher expression of CD133 mRNA in patients after operation demonstrated a much poorer survival rate （P=0.013）. Conclusions Higher expressive level of CD133 mRNA in GC before operation is associated to poor cell differentiation，lymph vessel invasion，deeper invasion，higher lymph node metastasis，later TNM stage，and positive expression of CD133 protein. It is also related positively to metastatic lymphatic nodes ratio，metastatic lymph node number，and ABSV of CD133 mRNA in primary lesion of GC. The level of CD133 mRNA in PBMCs of patients with GC is higher after operation as compared with before operation，which demonstrates deeper invasion and poorer survival.
ObjectiveTo investigate the expression of CD133 protein in primary lesions of gastric cancer and its clinical significance. MethodsThe expressions of CD133 protein in the primary lesion of tumor and normal gastric mucosa tissues confirmed by using histopathologic examination of 99 patients were detected by immunohistochemical staining. The correlation of CD133 protein expression with the clinicopathologic parameters and features after operation were analyzed. ResultsPositive cells of CD133 protein were localized in the gland parietal and cell membrane surface. The expression of CD133 protein in the cancer and normal gastric mucosa tissues were 29.29% (29/99) and zero, respectively (P=0.000). Expression of CD133 protein in tumor with diameter gt;5 cm was significantly higher than that in the tumor with diameter ≤5 cm (P=0.041). The expression of CD133 protein was correlated with TNM stage (P=0.044), lymph node metastasis (P=0.017), lymphatic vessel invasion (P=0.000), and vascular invasion (P=0.000). Logistic regression analysis revealed that invasion depth of tumor (P=0.011), lymph node metastasis (P=0.043), and TNM stage (P＝0.049) were independent risk factors for CD133 protein expression. Survival time of patients with positive expression of CD133 protein was significantly shorter than that negative expression of CD133 protein (P＝0.046). Cox proportial hazard regression model analysis demonstrated that lymph node metastasis (P=0.042), TNM stage (P=0.046), and positive expression of CD133 protein (P=0.046) were independent risk factors for patients survival. ConclusionThe CD133 protein expression in primary lesions is closely related with development, metastasis, and prognosis of gastric cancer.
ObjectiveTo investigate the expressions of CD133 and CD44 protein in primary lesions of gastric cancer and its clinical significance. MethodsThe expressions of CD44 and CD133 protein in gastric cancer tissues of 100 patients with gastric cancer were detected by immunohistocheimcal stainings.The relation between the expressions of CD44 and CD133 protein and the clinicopathologic characters were analyzed. ResultsBoth CD44 and CD133 protein were expressed on the cell membranes.No correlation were found between CD44/CD133 and the clinicopathologic parameters include gender and age (P > 0.05), but the positive expression rate of CD44/CD133 with diameter>5 cm was significantly higher than that tumor with diameter≤5 cm (CD44 P=0.150;CD133 P=0.056), and correlated with the tissue differentiation (CD44 P=0.008;CD133 P=0.007), vascular invasion (CD44 P=0.043;CD133 P=0.023), lymphatic vessel invasion (CD44 P=0.020;CD133 P=0.044), lymph nodes metastasis (CD44 P=0.002;CD133 P=0.004), inva-sion depth of tumor (CD44 P=0.006;CD133 P=0.021), and pTNM stage (CD44 P=0.034;CD133 P=0.001).No correlation were found between the co-expression of CD44 and CD133 protein and the clinicopathologic parameters include gender, age, tissue differentiation, and vascular invasion (P > 0.05), but the positive co-expression rate of CD44 and CD133 with diameter>5 cm was significantly higher than that tumor with diameter≤5 cm (P=0.010), and correlated with lymphatic vessel invasion (P=0.003), lymph nodes metastasis (P=0.045), invasion depth of tumor (P=0.041), and pTNM stage (P=0.049).The Spearman rank correlation analysis showed that there was positive correlation between the expressions of CD44 and CD133 protein (r=0.207, P=0.039).Univariate analysis showed that lymph nodes metastasis (P < 0.001), pTNM stages (P=0.013), CD44 protein expression (P=0.005), CD133 protein expression (P=0.002), and co-expression of CD44 and CD133 protein (P < 0.001) were significantly correlated with 3-year survival rate of pati-ents with gastric cancer respectively.Logistic regression analysis revealed that lymph node metastasis (P=0.038) was independent risk factor for co-expression of CD44 and CD133 protein.Multivariate analysis with the Cox regression models showed that co-expression of CD44 and CD133 protein (P=0.003) and lymph node metastasis (P=0.006) were significantly associated with poor prognosis. ConclusionsCD44 and CD133 protein may be considered as robust cancer stem cell markers in gastric cancer.The co-expression of CD44 and CD133 protein is the independent prognosis factor for gastric cancer and strongly associated with poor prognosis when they are expressed more high.
ObjectiveTo study the mechanism of invasion of CD133 positive population in gallbladder cancer. MethodsThe invasive abilities of the CD133 positive cells and the CD133 negative cells were detected by Transwell.The CXCR4 mRNA and protein in the CD133 positive cells and the CD133 negative cells were detected by the semi-quanti-tative RT-PCR, Western blot method, and immunofluorescence, respectively.SDF-1αand AMD3100 were respectively used to stimulate/inhibit the GBC-SD cells.The invasive ability and the migration force were detected in the CD133 posi-tive cells and the CD133 negative cells.The expressions CD133 mRNA and protein of the GBC-SD cells were detected by semi-quantitative RT-PCR and Western blot method, respectively. Results①The number of invasion cells in the CD133 positive cells was significantly more than that in the CD133 negative cells (23.78±8.74 versus 6.56±3.09, P=0.000 7).②The fluorescent protein of CXCR4 in the CD133 positive cells was stronger than that in the CD133 negative cells.The expression of CXCR4 mRNA in the CD133 positive cells was significantly higher than that in the CD133 negative cells (0.642 4±0.020 4 versus 0.335 9±0.043 2, P=0.004).The expression of CXCR4 protein in the CD133 positive cells was significantly higher than that in the CD133 negative cells (0.765 0±0.106 6 versus 0.409 4±0.019 5, P=0.013).③In the CD133 positive cells, compared with the control group, the number of invasion cells was significantly increased in the SDF-1αgroup (62.89±15.27 versus 23.78±8.74, P=0.000 6) and decreased in the AMD3100 group (10.33±2.00 versus 23.78±8.74, P=0.000 2).In the CD133 negative cells, compared with the control group, the number of invasion cells was not significant change in the SDF-1αgroup (6.89±4.23 versus 6.59±3.09, P=0.41) and in the AMD3100 group (6.11±2.67 versus 6.59±3.09, P=0.38), respectively.④In the CD133 positive cells, compared with the control group, the number of migration cells was significantly increased in the SDF-1αgroup (74.56±15.80 versus 35.56±10.97, P=0.000 3) and decreased in the AMD3100 group (12.67±2.40 versus 35.56±10.97, P=0.000 2).In the CD133 negative cells, compared with the control group, the number of migration cells was not significant change in the SDF-1αgroup (9.78±2.04 versus 9.56±1.74, P=0.43) and in the AMD3100 group (9.54±1.74 versus 9.56±1.74, P=0.42).⑤In the GBC-SD cells, compared with the control group, the CD133 mRNA was significantly increased in the SDF-1αgroup (0.626 5±0.048 7 versus 0.450 0±0.024 3, P=0.004) and decreased in the AMD3100 group (0.359 3±0.047 3 versus 0.450 0±0.024 3, P=0.011);the CD133 protein was significantly increased in the SDF-1αgroup (0.508 9±0.020 7 versus 0.440 9±0.013 0, P=0.016) and decreased in the AMD3100 group (0.317 7±0.013 7 versus 0.440 9±0.013 0, P=0.004). ConclusionThe high invasion ability of CD133 positive population in gallbladder cancer might be due to the high expression of CXCR4.
ObjectiveTo study the expression and role of homeobox transcription antisense intergenic ribose nucleic acid (HOTAIR) in CD133-positive gastric cancer cells, which was classified to long non-coding RNA (LncRNA). MethodsImmune magnetic cell sorting (MACS) was performed to sort CD133-positive and CD133-negative cells of KATO-Ⅲgastric cancer cells, then reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the expressions of HOTAIR mRNA and CD133 mRNA. After the intervention of small interfering RNA (siRNA) for CD133-positive KATO-Ⅲcells, RT-PCR method was performed to detect the expression of HOTAIR mRNA to select siRNA who had the best silent effect. The selected-siHOTAIR was used to silent the expression of HOTAIR, then the expressions of CD133 mRNA, E-cadherin mRNA, and N-cadherin mRNA were detected by RT-PCR. At last, Transwell experiments were performed to detect the migration ability and invasion ability. Results①?RT-PCR test results showed that, the expression levels of CD133 mRNA and HOTAIR mRNA in CD133-positive group were significantly higher than those of CD133-negative group and no separation group (P < 0.05).②?After interference of siHOTAIR, the expression levels of HOTAIR mRNA in siHOTAIR1 group, siHOTAIR2 group, and siHOTAIR3 group were all significantly lower than those of blank control group and negative control group (P < 0.05), and the expression levels of HOTAIR mRNA in siHOTAIR2 group was lower than those of siHOTAIR1 group and siHOTAIR3 group (P < 0.05). The results indicated that siHOTAIR2 had the best interference efficiency.③?The expression levels of CD133 mRNA and N-cadherin mRNA in siHOTAIR2 group were lower than those corresponding indicators of blank control group and negative control group (P < 0.05), but the expression level of E-cadherin mRNA was higher than those of blank control group and negative control group (P < 0.05). Transwell experiment results showed that, number of cells which through the cell membrane in siHOTAIR2 group was lower than those of blank control group and negative control group (P < 0.05). ConclusionThe expression of HOTAIR mRNA in CD133-positive KATO-Ⅲgastric cancer cells was higher than that of CD133-negative cells, interfering the expression of HOTAIR mRNA can reduce the expression of CD133 mRNA in CD133-positive KATO-Ⅲgastric cancer cells, and can inhibit cell migration and invasion.
ObjectiveTo systematically review the expression and clinical significance of CD133 in gastric cancer. MethodsSearches in the databases such as PubMed, EMbase, Web of Knowledge, The Cochrane Library (Issue 10, 2013), CBM, VIP, CNKI and WanFang Data were performed to collect case-control studies about the association between the CD133 expression and gastric cancer up to October 2013. Two reviewers independently screened the literature according to the inclusion and exclusion criteria, extracted the data, and assessed the methodological quality of the included studies, and then RevMan 5.2 software was used for meta-analysis. ResultsNine case-control studies involving 623 patients were included. The results of the meta-analyses showed that, there were significant differences of CD133 expression in the following comparisons:gastric cancer tissues vs. normal esophageal tissues (OR=3.89, 95%CI 1.87 to 8.11, P=0.000 3), lymph node metastasis vs. non-lymph node metastasis (OR=2.75, 95%CI 1.99 to 3.81, P < 0.000 01), clinical stages Ⅲ-Ⅳ vs. clinical stages Ⅰ-Ⅱ (OR=2.83, 95%CI 2.13 to 3.76, P < 0.000 01), as well as distant metastasis vs. non-distant metastasis (OR=2.38, 95%CI 1.47 to 3.85, P=0.000 4). While no significant difference was found between the cell differentiation G1-G2 vs. cell differentiation G3 (OR=1.70, 95%CI 0.90 to 3.21, P=0.10). ConclusionOver-expression of CD133 is associated with lymph node metastasis, distant metastasis and poor TNM stage, which suggests that CD133 may participate in the carcinogenesis of gastric cancer. However, due to the limitations of the included studies, more large-sample high-quality case-control studies are still needed to verify these results.