title: [Characterization of a Clonorchis sinensis antigen, calmodulin, and its relationship with liver fibrosis].
abstract: OBJECTIVE
  To characterize the biological function of calmodulin (CaM) from Clonorchis sinensis (C. sinensis, Cs) and investigate its role in clonorchiasis-associated hepatic fibrosis.
  
  
  METHODS
  The full-length sequence of CsCaM gene was isolated from Cs cDNA library and its homologues were searched using BLASTx for comparison. Bioinformatics analysis was performed to compare the homologues and predict the physiochemical characteristics and functional domains. The gene was cloned in a prokaryotic plasmid and expressed in E. coli, and the recombinant protein was purified by affinity chromatography for immunizing rats to produce polyclonal antibodies, whose titer was determined using ELISA analysis. Immunoblotting analysis was carried out to determine of the purity and antibody recognition of CsCaM. Immunofluorescence assay was employed to analyze the tissue location of the protein. A rat model of liver fibrosis was established by introperitoneal injection of the recombinant protein.
  
  
  RESULTS
  The recombinant CsCaM protein obtained contained 150 amino acids with a theoretical molecular mass of 23.4 kD. CsCaM homologue had EF hand motifs. The recombinant pET-30a-CsCaM plasmid expressed in BL21 E. coli was about 23.4 kD. The total IgG antibody titer in the immunized mice reached the peak level (over 1: 51200) 2 to 4 weeks after the first injection. Immunohistochemistry showed that CsCaM located in the testis of adult C. sinensis. The rats receiving intraperitoneal injection of CsCaM showed severe liver inflammation with mild to moderate liver fibrosis.
  
  
  CONCLUSIONS
  The pro-inflammation and pro-fibrosis effects of CsCaM in rat liver suggest its involvement in clonorchiasis- associated hepatic fibrosis.
authors.affiliation: Department of Clinical Laboratory, Sun Yat-sen Memorial Hospital of Sun Yat-sen University, Guangzhou 510120, China.E-mail: 7874439@163.com.
authors.forename: Minghui
authors.initials: M
authors.lastname: Zheng
authors.forename: Kunhua
authors.initials: K
authors.lastname: Hu
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.forename: Xinbing
authors.initials: X
authors.lastname: Yu
mesh_terms: D000818:Animals; D000909:Antibodies, Helminth / Q000097:blood; D000947:Antigens, Helminth / Q000276:immunology*; D002147:Calmodulin / Q000276:immunology*; D003003:Clonorchiasis / Q000276:immunology*; D003004:Clonorchis sinensis / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D015723:Gene Library; D007074:Immunoglobulin G / Q000097:blood; D007249:Inflammation; D008103:Liver Cirrhosis / Q000469:parasitology*; D008297:Male; D051379:Mice; D051381:Rats; D011994:Recombinant Proteins / Q000276:immunology
chemical_list: D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D002147:Calmodulin; D007074:Immunoglobulin G; D011994:Recombinant Proteins