title: XPD Functions as a Tumor Suppressor and Dysregulates Autophagy in Cultured HepG2 Cells.
abstract: BACKGROUND
  Recent clinical studies have linked polymorphisms in the xeroderma pigmentosum group D (XPD) gene, a key repair gene involved in nucleotide excision repair, to increased risk of hepatocellular carcinoma (HCC). However, the cellular effects of XPD expression in cultured HCC cells remain largely uncharacterized. Therefore, the aim of this study was to characterize the in vitro cellular effects of XPD expression on the HCC cell line HepG2.
  
  
  METHODS
  HepG2 cells were transfected as follows to create four experimental groups: pEGFP-N2/XPD plasmid (XPD) group, EGFP-N2 plasmid (N2) control group, lipofectamine™ 2000 (lipid) control group, and non-transfected (CON) control group. An MTT cell proliferation assay, Annexin V-APC apoptosis assay, colony formation assay, scratch wound migration assay, Transwell migration assay, and Western blotting of the autophagic proteins LC3 and p62 were conducted.
  
  
  RESULTS
  XPD expression significantly inhibited HepG2 cell proliferation (p<0.05), significantly promoted HepG2 cell apoptosis (p<0.05), significantly inhibited HepG2 colony formation (p<0.05), significantly decreased HepG2 cells' migratory ability (p<0.05), and significantly lowered HepG2 cells' invasive capacity (p<0.05). Western blotting showed that XPD expression significantly increased LC3 expression (p<0.05) and significantly reduced p62 expression (p<0.05).
  
  
  CONCLUSIONS
  XPD expression serves as a tumor suppressor and dysregulates autophagic protein degradation in HepG2 cells in vitro. Further in vivo pre-clinical studies and clinical trials are needed to validate XPD's potential as a tumor-suppressive gene therapy.
authors.affiliation: The Second Affiliated Hospital of Nanchang University, The Second Affiliated Hospital of Nanchang University, Nanchang, China (mainland).
authors.forename: Jian-Feng
authors.initials: JF
authors.lastname: Zheng
authors.affiliation: The Second Affiliated Hospital of Nanchang University, The Second Affiliated Hospital of Nanchang University, Nanchang, China (mainland).
authors.forename: Lin-Lin
authors.initials: LL
authors.lastname: Li
authors.affiliation: Jiangxi Children's Hospital, Jiangxi Children's Hospital, Nanchang, China (mainland).
authors.forename: Juan
authors.initials: J
authors.lastname: Lu
authors.affiliation: Department of Medicine, Nanchang University, Nanchang, China (mainland).
authors.forename: Kun
authors.initials: K
authors.lastname: Yan
authors.affiliation: The Second Affiliated Hospital of Nanchang University, The Second Affiliated Hospital of Nanchang University, Nanchang, China (mainland).
authors.forename: Wu-Hua
authors.initials: WH
authors.lastname: Guo
authors.affiliation: Key Laboratory of Molecular Medicine, The Second Affiliated Hospital of Nanchang University, Nanchang, China (mainland).
authors.forename: Ji-Xiang
authors.initials: JX
authors.lastname: Zhang
mesh_terms: D017304:Annexin A5; D017209:Apoptosis / Q000502:physiology; D001343:Autophagy / Q000502:physiology*; D015153:Blotting, Western; D049109:Cell Proliferation / Q000502:physiology; D003114:Colony-Forming Units Assay; D056945:Hep G2 Cells; D006801:Humans; D066298:In Vitro Techniques; D013778:Tetrazolium Salts; D013844:Thiazoles; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000502:physiology*; D051759:Xeroderma Pigmentosum Group D Protein / Q000235:genetics / Q000502:physiology*
chemical_list: D017304:Annexin A5; D013778:Tetrazolium Salts; D013844:Thiazoles; D025521:Tumor Suppressor Proteins; D051759:Xeroderma Pigmentosum Group D Protein; C022616:thiazolyl blue