Fan et al. BMC Cancer 2011, 11:320 http://www.biomedcentral.com/1471-2407/11/320 Table 3 Prognostic variables for survival in 23 patients with CC Variables Number of cases Median survival in months (range) Univariate analysis (P value) Multivariate analysis (P value) Age < 60 > 60 Gender Male Female Poorly Staging I/II III/IV Differentiation status Well Moderately CD133 expression Positive Negative P < 0.05 is considered significant 14 9 13 10 8 6 9 6 17 14 9 14.00 (11.12-16.88) 11 (3.82-18.21) 11 (7.15-14.85) 17 (10.49-23.51) 15 (12.8-17.1) 8 (2.24-13.7) 8 (0-19.7) 14 (12.5-15.5) 11 (4.9-17.1) 14 (11.9-16.1) 4 (0-8.2) 0.058 0.005 0.025 0.033 0.001 Page 5 of 8 0.086 0.116 0.010 0.321 0.007 Our study and other researches have found that CD133 expression pattern varies in different cancers. In colorectal carcinoma, CD133 positivity is restricted only to the luminal surface [21,28]. In our study, besides the luminal surface pattern in most well-differentiated CC cases, most of moderately or poorly differentiated CC cases showed cytoplasmic or both cytoplasmic and membranous staining (Table 2). It is believed that the different subcellular localization of CD133 (cytoplasm or membrane) may confer different cellular functions [20]. The protein is expressed apically on the membrane of epithelial cells when a lumen has been formed but it is expressed in the cytoplasm of solidly arranged malignant tissues of non-epithelial origin [20]. It is well known that if a formed lumen is observed in an epithelial tumor, it usually indicates for better differentiation of the glandular tumor. In our study, CD133 showed cyto- plasmic expression in many CC cases, suggesting that cytoplasmic CD133 expression should be not just restricted to malignant tissues of non-epithelial origin [20] but also occurs in malignant epithelial cells of CC. The statistical analysis showed that CD133 expression significantly correlated with the differentiation status of CC (P = 0.004). It is possible that the expression of CD133 on the apical membrane may indicate a particu- lar stage of the cell differentiation connected with the formation of lumen and ducts as same as the normal bile ducts, and this deduction is confirmed by Shmelkov, S.V. et al in their study [28]. In addition, we noticed the CD133 positive debris in normal bile ducts and the shedding of CD133 positive tumor cells and apparent non-cellular, CD133-positive materials in the malignant ducts. This observation is in accordance with the pre- vious report that small membrane vesicles containing prominin-1 were found in the human normal saliva and tears as well as malignant lumen of colorectal cancer [29,30]. The concentration of CD133, presented within the apical membrane of glands or in extracellular mem- branous vesicles, suggests that this cholesterol-binding glycoprotein might play a role in regulating the secreting process [29,31]. Similarly, CD133 expression in normal bile ducts and its immunoreactivity associated with the secretion therein indicate that CD133 protein might exist in bile, thus, a quantitive analysis of CD133 in bile might be helpful for distinguishing CC from hepatocel- lular carcinoma.