Loss of Short Chain Fatty Acid Receptor GPR43: A Common Molecular Event in Colorectal Carcinoma.
Kristina A Matkowskyj, Jie Liao, Yeon T Chung, Haonan N Li, M Sambasiva Rao, Guang-Yu Yang. Northwestern University, Chicago, IL
Background: GPR43 is a G protein-coupled, short chain fatty acid (SCFA) receptor that activates signal transduction pathways and cellular responses. SCFAs affect colonocyte transport and metabolism, growth and differentiation, hepatic control of lipids and carbohydrates, and provide energy to colonic epithelium, muscle, kidney, heart and brain. The role of SCFAs, particularly butyrate, in colon cancer therapy has been extensively studied, however the role of its receptor GPR43 is unknown. Recently, we found GRP43 frequently loses expression in colonic carcinoma, probably as a tumor suppressor. Here, we analyzed GPR43 expression in cultured human colonic adenocarcinoma and normal cell lines and colorectal resections in order to investigate its functional role in response to SCFAs in regulating tumor cell growth with or without GRP43.
Design: GRP43 expression was determined using 11 cultured cell lines; 10 human colonic adenocarcinomas and 1 normal human colonic epithelial cell line NCM460 using real-time PCR, Western blot and immunohistochemical (IHC) approaches. Colon tissue microarrays (TMA) were created for IHC use and included 15 cases from colonic resection specimens with normal, primary and metastatic colonic adenocarcinoma from each patient, 5 tubular adenomas (TA), and 5 adenocarcinomas arising in TA. The staining intensity was classified as no staining, low intensity staining, or high intensity staining. All tumor cell lines were studied for their response to butyrate using an MTT assay.
Results: High intensity expression of GRP43 in NCM460 cells was determined at both mRNA and protein levels, however only 1 cancer cell line (LS180) of 10 showed GRP43 expression in western blot assays and only the HT29 cell line displayed GRP43 expression using real-time PCR. Decreased response to butyrate-induced inhibition of cell proliferation and differentiation correlated with GRP43 expression in cells. Parallel IHC study on TMAs showed high intensity expression in normal colonic epithelium (n=15) and 65% (13/20) of adenocarcinomas exhibited loss of expression (p<0.01), with moderately/poorly differentiated carcinomas showing no expression (n=13). All TAs (n=5) demonstrated identical levels of expression as in normal mucosa.
Conclusions: This study demonstrates that loss of GPR43 expression is a common molecular event in less differentiated colonic adenocarcinomas and cultured cell lines, indicating it may serve as a useful malignant biomarker. Loss of expression is correlated with response to SCFA treatment, implying regulation of GRP43 can be a potential target for SCFA therapy.
Wednesday, March 2, 2011 9:30 AM
Poster Session V # 35, Wednesday Morning