Detection of laminin receptor mRNA in human cancer cell lines and colorectal tissues by in situ hybridization.
The 67-kd high-affinity laminin receptor (67 LR) is a gene product whose expression appears to be associated with the invasive and metastatic phenotype of a variety of human cancer cells. Northern blot hybridization has been routinely used to quantify the level of 67 LR mRNA from total cellular RNA extracts of homogenized tissue specimens or in vitro grown cell populations. This technique is useful to assess the average expression of the 67 LR mRNA of a particular sample but does not provide information about expression in specific cell types nor about heterogeneity of expression from cell to cell. In this study, we analyzed the expression of 67 LR mRNA in four human cancer cell lines with varying degrees of expression of 67 LR protein (renal cancer A-704, breast carcinoma MCF-7/4 and MCF-7/7, and pancreatic cancer Panc-1) using in situ hybridization performed with 67 LR riboprobes. Total cellular RNA was simultaneously extracted from the cell lines and hybridized on Northern blots with a 67 LR cDNA probe to assess the validity of the mRNA detection by in situ hybridization. Sixty-seven LR mRNA expression was higher in Panc-1 and MCF-7/4 cells than in MCF-7/7 and renal carcinoma A-704. There was a direct correlation (R2 = 0.88) between the in situ hybridization analysis and the mRNA levels detected by Northern blot analysis. The in situ hybridization method showed a heterogeneous expression of the 67 LR mRNA in the four cell lines with different subpopulations of cells showing a range from negative to high levels of the message. Sixteen freshly frozen human colorectal tissues (seven adenocarcinomas, five matched normal mucosae, and four adenomas) were also analyzed by in situ hybridization. The 67 LR mRNA was localized in normal and neoplastic epithelial cells. Adenocarcinoma cells showed a 1.6- to 5-fold higher expression (P < 0.02 according to the Wilcoxon-Mann-Whitney test) than did epithelial colonic cells from normal mucosae or adenomas. The signal tended to be stronger in poorly differentiated carcinomas and carcinomas with metastases than in moderately differentiated and nonmetastatic tumors. We conclude that the high expression of 67 LR mRNA in colorectal tumors is due to an increased production by tumor cells. Furthermore, in situ hybridization is an effective method to detect the expression of LR mRNA in cultured cell lines as well as in frozen tissue sections.
Campo
E
E
Laboratory of Pathology, National Cancer Institute, Bethesda, Maryland 20892.
Monteagudo
C
C
Castronovo
V
V
Claysmith
A P
AP
Fernandez
P L
PL
Sobel
M E
ME
D000236:Adenoma / Q000737:chemistry* / Q000473:pathology* / Q000648:ultrastructure; D015152:Blotting, Northern; D001943:Breast Neoplasms / Q000737:chemistry* / Q000473:pathology* / Q000648:ultrastructure; D003106:Colon / Q000737:chemistry / Q000166:cytology / Q000648:ultrastructure; D015179:Colorectal Neoplasms / Q000737:chemistry* / Q000473:pathology* / Q000648:ultrastructure; D005347:Fibroblasts / Q000737:chemistry* / Q000473:pathology* / Q000648:ultrastructure; D006801:Humans; D017403:In Situ Hybridization; D010190:Pancreatic Neoplasms / Q000737:chemistry* / Q000473:pathology* / Q000648:ultrastructure; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics; D017474:Receptors, Laminin / Q000235:genetics*; D013462:Sulfur Radioisotopes; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D017474:Receptors, Laminin; D013462:Sulfur Radioisotopes