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. 2015 Jun;185(6):1724–1739. doi: 10.1016/j.ajpath.2015.02.010

Figure 3.

Figure 3

Human cholangiocarcinoma (CCA) primary cultures. A: Immunohistochemical analyses of CCA primary cultures after 2 to 3 passages (P2/P3) and 20 to 30 passages (P > 20). The expression of mesenchymal and epithelial-mesenchymal transition markers (vimentin, α-SMA, SNAIL, Twist, S100A4, P-cadherin) largely predominate over that of epithelial markers (CD133, EpCAM, LGR5, E-cadherin) in all passages examined. During progression of primary cultures (P > 20 versus P2/P3) SNAIL- and Twist-positive cells tend to increase, whereas EpCAM-, LGR5-, and CD133-positive cells tend to decrease. Similar diffuse positivity for vimentin, α-SMA, and S100A4 is observed in all passages. B–D: Flow cytometric (FC) analyses of primary cultures (20 to 30 passages) obtained from mixed-IHCCA (IH-mixed), mucin-IHCCA (IH-mucin), and mucin-pCCA (p-mucin) subtypes; representative FC plots. B: Cells positive for CD13, CD44, and CD90 largely predominate with respect to CD133, EpCAM, and LGR5. CD13+ and CD44+ cells predominate in mixed-IHCCA with respect to mucin-IHCCA or mucin-pCCA, whereas the opposite is found for CD90+ cells. C: Cells double-positive for CD13 and CD44 (CD13+/CD44+) predominate in mixed-IHCCA with respect to mucin-IHCCA or mucin-pCCA. D: CD90+/CD44−/CD13− cells predominate in mucin-IHCCAs and mucin-pCCA with respect to mixed-IHCCAs. In these experiments, CD90+ cells are gated and further analyzed for the expression of CD44 and CD13 markers. ∗P < 0.05, ∗∗P < 0.01 versus mucin-CCAs. Original magnification: ×40 (A).