Fig 1.
Tumor classification and selection of tumor areas with high density of CK7+ cells.
(A-C) Tumors were classified as well-, moderately- and poorly-differentiated. (D-F) Immunohistochemical detection of CK7 in representative histologically-graded ICCs. Areas with at least 50% of CK7 staining were selected for further RNA extraction and immunofluorescence stainings. White scale bars = 200 μm. Black scale bars = 500 μm
Table 1.
Characteristics of tumors samples.
Fig 2.
Molecular biliary differentiation markers are not differentially expressed among histologically-graded iCC's.
(A) All biliary differentiation markers are expressed (qRT-PCR) at lower levels in ICC as compared to distant non-tumoral tissue. Low levels of HNF4α indicate lack of significant contamination with normal hepatocytes. (Mean ± SEM; *p<0.05, **p<0.01, ***p<0.001). (B-C) Lack of correlation between molecular differentiation marker expression and tumor grade when comparing (B) ICC samples or (C) the ratio of expression in tumor versus non-tumor tissue. Dots represent individual ICC samples and horizontal bars represent the mean of the samples.
Fig 3.
Biliary-specific proteins are not differentially expressed among histologically-graded ICC's.
(A) All markers are expressed (qRT-PCR) at lower levels in ICC as compared to distant non-tumoral tissue. (Mean ± SEM; *p<0.05, **p<0.01, ***p<0.001). (B-C) Lack of correlation between molecular differentiation marker expression and tumor grade when comparing (B) ICC samples or (C) the ratio of expression in tumor versus non-tumor tissue. Dots represent individual ICC samples and horizontal bars represent mean of the samples.
Table 2.
Selection of microRNAs associated with ICC differentiation grade.
Fig 4.
ICC-associated microRNAs are not differentially expressed in histologically-graded ICC.
MicroRNAs known to be dysregulated in ICC and differentially expressed among tumor grades were selected for qRT-PCR analysis. (A) Three microRNAs are up-regulated in ICC when compared to distant non-tumoral tissues. (Mean ± SEM; *p<0.05). (B-C) Lack of correlation between microRNA expression and tumor differentiation grade when (B) comparing ICC samples, or (C) when analysing the ratio of expression in tumor versus non-tumor tissue. Dots represent individual ICC samples.
Fig 5.
Expression of molecular biliary markers in histologically-graded ICC's.
(A-F) SOX9 expression is detected in the cytoplasm of well-differentiated ICC cells (A-B) and both in the cytoplasm and nucleus of moderately and poorly differentiated tumor cells (C-F). (G-L) CK19 is expressed in all types of histologically-graded tumors. (M-R) OPN is detected in all types of histologically graded-ICC's. Its expression is mostly apical in well-differentiated tumor cells (M-N, white arrowheads), but cytoplasmic OPN is detected in moderately and poorly differentiated ICCs and in a subset of well-differentiated tumor cells (M-R, yellow arrowheads). (M’-R’) HNF1β expression inversely correlates with ICC differentiation grade. White scale bar = 100 μm, black scale bar = 500 μm.