Xin Ma1, Yang Fan1, Yu Gao1, Yu Zhang1, Qingbo Huang1, Qing Ai1, Dong Ni2, Weihao Chen1, Peng Zhang1, Erlin Song1, Baojun Wang1, Taoping Shi1, Tao Zheng1, Xu Zhang3. 1. State Key Laboratory of Kidney Diseases, Department of Urology, Military Postgraduate Medical College, Chinese People's Liberation Army General Hospital, Beijing, People's Republic of China. 2. Medical School, Nankai University, Tianjin, People's Republic of China. 3. State Key Laboratory of Kidney Diseases, Department of Urology, Military Postgraduate Medical College, Chinese People's Liberation Army General Hospital, Beijing, People's Republic of China. Electronic address: xzhang@foxmail.com.
Abstract
OBJECTIVES: Although emerging evidence has shown that the deregulation of micro-ribonucleic acid (RNA) biogenesis machinery is involved in various human malignancies, this role has not been investigated in clear cell renal cell carcinoma (ccRCC). This study aims to determine whether Dicer, a key enzyme responsible for biogenesis of microRNA, is deregulated in ccRCC. The biological roles of Dicer in vitro are also determined. MATERIALS AND METHODS: The expression of Dicer at messenger RNA and protein levels was detected by real-time quantitative polymerase chain reaction and western blot, respectively, in human kidney tubule epithelial cell line, nonmetastatic 786-O ccRCC cell line, and metastatic ACHN ccRCC cell line, as well as in 42 cases of ccRCC surgical specimens including 14 cases with distant metastasis and their corresponding adjacent normal renal tissues. Dicer expression levels in specimens were also measured by immunohistochemical staining. Knockdown of Dicer expression in 786-O and ACHN ccRCC cell lines was achieved by transfecting short interfering RNA against Dicer. The effects of Dicer on cell proliferation, migration, and invasion were detected by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay, flow cytometric analyses, and Boyden chamber Transwell assay, respectively. RESULTS: Compared with human kidney tubule epithelial cell line, Dicer expression levels were significantly down-regulated in 786-O and ACHN ccRCC cell lines, with the metastatic ACHN ccRCC cell line having even lower levels. Meanwhile, Dicer expression levels were significantly down-regulated in ccRCC surgical specimens compared with adjacent normal renal tissues, with the metastatic ones further reduced, and Dicer messenger RNA levels were significantly correlated with overall tumor-node-metastasis stage of ccRCC. In vitro, the knockdown of Dicer significantly promoted cell proliferation, migration, and invasion. CONCLUSIONS: Reduced expression of Dicer may play a role in the tumorigenesis of ccRCC and further decline may be associated with distant metastasis of ccRCC.
OBJECTIVES: Although emerging evidence has shown that the deregulation of micro-ribonucleic acid (RNA) biogenesis machinery is involved in various humanmalignancies, this role has not been investigated in clear cell renal cell carcinoma (ccRCC). This study aims to determine whether Dicer, a key enzyme responsible for biogenesis of microRNA, is deregulated in ccRCC. The biological roles of Dicer in vitro are also determined. MATERIALS AND METHODS: The expression of Dicer at messenger RNA and protein levels was detected by real-time quantitative polymerase chain reaction and western blot, respectively, in human kidney tubule epithelial cell line, nonmetastatic 786-O ccRCC cell line, and metastatic ACHN ccRCC cell line, as well as in 42 cases of ccRCC surgical specimens including 14 cases with distant metastasis and their corresponding adjacent normal renal tissues. Dicer expression levels in specimens were also measured by immunohistochemical staining. Knockdown of Dicer expression in 786-O and ACHN ccRCC cell lines was achieved by transfecting short interfering RNA against Dicer. The effects of Dicer on cell proliferation, migration, and invasion were detected by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay, flow cytometric analyses, and Boyden chamber Transwell assay, respectively. RESULTS: Compared with human kidney tubule epithelial cell line, Dicer expression levels were significantly down-regulated in 786-O and ACHN ccRCC cell lines, with the metastatic ACHN ccRCC cell line having even lower levels. Meanwhile, Dicer expression levels were significantly down-regulated in ccRCC surgical specimens compared with adjacent normal renal tissues, with the metastatic ones further reduced, and Dicer messenger RNA levels were significantly correlated with overall tumor-node-metastasis stage of ccRCC. In vitro, the knockdown of Dicer significantly promoted cell proliferation, migration, and invasion. CONCLUSIONS: Reduced expression of Dicer may play a role in the tumorigenesis of ccRCC and further decline may be associated with distant metastasis of ccRCC.
Authors: Rileen Sinha; Andrew G Winer; Michael Chevinsky; Christopher Jakubowski; Ying-Bei Chen; Yiyu Dong; Satish K Tickoo; Victor E Reuter; Paul Russo; Jonathan A Coleman; Chris Sander; James J Hsieh; A Ari Hakimi Journal: Nat Commun Date: 2017-05-10 Impact factor: 14.919