title: Proliferation of small cell lung cancer cell line reduced by knocking-down PROX1 via shRNA in lentivirus.
abstract: The present study aimed to find whether PROX1 is expressed in small cell lung cancer (SCLC) cell lines, and whether PROX1 knockdown with shRNA via lentivirus resulted in decreased cell proliferation. SCLC cell lines H69, H82, H187 and H889 were selected for the study. PROX1 mRNA and protein levels were determined with real-time reverse-transcription polymerase chain reaction(RT-PCR) and western blot, respectively. The localization and distribution of PROX1 was mapped by immunocytochemistry with a specific antibody. Three pairs of shRNA were selected from a pool of shRNA pairs, and packaged into lentivirus particles to infect the above cell lines. The non-target sequence (NT) and a house-keeping gene, glyceraldehyde 3 phosphate dehydrogenase (GAPDH), were employed as controls. SCLC cell proliferation rates were measured with bromine deoxyuridine (BrdU) incorporation method. The results indicated levels of that PROX1 mRNA were detected in SCLC cell lines in the following rank order H69>H889>H187>H82. A similar profile for PROX1 protein expression was captured. The majority of PROX1 was concentrated at the cell nucleus. H69 was selected to represent the above SCLC cell lines. The PROX1 level in H69 cells was successfully reduced with shRNA lentivirus, and the cell proliferation rate of infected H69 cells was dramatically reduced by 20-50%. Hence, it is concluded that PROX1 expression in SCLC cell line is high, and can be reduced with shRNA lentivirus, thereby reducing the cell proliferation rate.
authors.affiliation: DABT, Assistant Professor, Department of Pulmonary and Critical Care Medicine, College of Medicine, Mayo Clinic, 200 First Street SW, Rochester, MN 55902, USA. xushzh@yahoo.com
authors.forename: Si-Hai
authors.initials: SH
authors.lastname: Zhu
authors.forename: Chun-Juan
authors.initials: CJ
authors.lastname: Shan
authors.forename: Zhi-Fang
authors.initials: ZF
authors.lastname: Wu
authors.forename: Shang-Zhi
authors.initials: SZ
authors.lastname: Xu
mesh_terms: D015153:Blotting, Western; D001973:Bromodeoxyuridine / Q000378:metabolism; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D016086:Lentivirus / Q000378:metabolism*; D008175:Lung Neoplasms / Q000235:genetics / Q000473:pathology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D055752:Small Cell Lung Carcinoma / Q000235:genetics / Q000473:pathology*; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D000098551:Prospero-Related Homeobox 1 Protein
chemical_list: D018398:Homeodomain Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D000098551:Prospero-Related Homeobox 1 Protein; D001973:Bromodeoxyuridine
keywords: PROX1; cell proliferation; lentivirus; shRNA; small cell lung cancer; viral knockdown