Liver Cancer
Copyright ©2005 Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. May 14, 2005; 11(18): 2704-2708
Published online May 14, 2005. doi: 10.3748/wjg.v11.i18.2704
Expression of lysosome-associated protein transmembrane 4B-35 in cancer and its correlation with the differentiation status of hepatocellular carcinoma
Cong Peng, Rou-Li Zhou, Gen-Ze Shao, Jing-An Rui, Shao-Bin Wang, Ming Lin, Sha Zhang, Zi-Feng Gao
Cong Peng, Rou-Li Zhou, Gen-Ze Shao, Ming Lin, Sha Zhang, Department of Cell Biology, School of Basic Medical Sciences, Peking University, Beijing 100083, China
Jing-An Rui, Shao-Bin Wang, Department of General Surgery/Pathology, Peking Union Medical Hospital, Beijing 100032, China
Zi-Feng Gao, Department of Pathology, School of Basic Medical Sciences, Peking University, Beijing 100083, China
Author contributions: All authors contributed equally to the work.
Correspondence to: Rou-Li Zhou, Department of Cell Biology, School of Basic Medical Sciences, Peking University, Beijing 100083, China. rlzhou@bjmu.edu.cn
Telephone: +86-10-82801034 Fax: +86-10-62358270
Received: April 4, 2004
Revised: April 5, 2004
Accepted: May 24, 2004
Published online: May 14, 2005
Abstract

AIM: To produce high-quality polyclonal antibody to lysosome-associated protein transmembrane 4B-35 and to identify LAPTM4B-35 expression in cancer tissues and its correlation with differentiation status of hepatocellular carcinoma (HCC).

METHODS: The 297 bp 5’ end of LAPTM4B cDNA was obtained by PCR and inserted into prokaryotic expression vector pGEX-KG. Then the recombinant pGEX-KG-N1-99 was transformed into E.coli JM109 to express GST-fusion protein. The fusion protein was purified by glutathione sepharoseTM 4B agarose. The purified GST-LAPTM4B-N1-99 was characterized by SDS-PAGE, and used to immunize rabbits. The titer and specificity of antisera were detected by ELISA and Western blot, respectively. The correlation between the expression levels of LAPTM4B-35 and the differentiation status of HCC was analyzed via Western blot. The expression of LAPTM4B-35 in HCC and other six cancer tissues was investigated via tissue chip and immunohistochemical analysis.

RESULTS: About 6.2 mg of pure GST-LAPTM4B-N1-99 was isolated from 1 L of bacteria. The GST-LAPTM4B-N1-99 produced high titer antisera in rabbits and showed good immunity. Western blot showed specific reactions for the antibody to the LAPTM4B-35 in the total proteins from HCC tissues and BEL-7402 cells, also to the fusion protein purified or in the transformed bacteria. LAPTM4B-35 was remarkably expressed in several cancers, such as HCC, breast cancer, gastric carcinoma, lung cancer, and colon carcinoma, but not commonly expressed in esophageal cancer and rectum carcinoma. Notably, the expression levels of LAPTM4B-35 were significantly and inversely correlated to the differentiation of HCCs in a 20 case analysis.

CONCLUSION: Specific polyclonal antibody (LAPTM4B-N1-99-pAb) to LAPTM4B-35 was produced. It identified the expression of LAPTM4B-35 in some cancer tissues originated from single layer cuboidal and columnar epithelial cells and firmly demonstrated that the expression of LAPTM4B-35 in HCC was inversely correlated with the differentiation of HCC.

Keywords: LAPTM4B; LAPTM4B-N1-99-pAb; HCC; Tissue microarray