title: Comparative cytotoxicity of high-osmolar and low-osmolar contrast media on HKCs in vitro.
abstract: BACKGROUND
  Radiocontrast-induced nephropathy is a clinically important complication of intravascularly applied radiocontrast media. A predominant toxic effect of contrast media on renal tubules has been shown in previous clinical trials and animal experiments. Bax and Bcl-2 are members of the Bcl-2 family. Caspases are a family of cell death proteases, caspase-3 is one of the key executioners of apoptosis. In this study, we evaluated the cytotoxicity of high-osmolar contrast media (HOCM; diatrizoate) and low-osmolar contrast media (LOCM; iohexol) on human renal tubular epithelial cells (HKCs), and determined the regulatory roles of Bax/Bcl-2 and caspase-3 on apoptosis induced by contrast media (CM) in HKCs.
  
  
  METHODS
  An HKC line was used. Experiments were divided into 7 groups: the HOCM group with iodine 111 mg/mL, HOCM group with iodine 74 mg/mL, LOCM group with iodine 111 mg/mL, LOCM group with iodine 74 mg/mL, mannitol high-osmolar control group, mannitol low-osmolar control group and a culture media control group . The cytotoxicity of HOCM and LOCM were evaluated by cell proliferation and viability assay (MTT assay) and lactate dehydrogenase (LDH) release. Apoptosis were assessed by Hochest 33258 fluorescence-stained cytospins, TUNEL staining, DNA agarose gel electrophoresis, electron microscope and flow cytometric DNA analysis. The protein ex-pression of Bax/Bcl-2 was determined by Western blot analysis, and caspase-3 activity was also determined by the fluo-rometric method.
  
  
  RESULTS
  Compared with the control group, LDH levels increased significantly (p<0.05) and cell viability decreased in cells treated with HOCM or LOCM (p<0.05) in an osmotic pressure-, iodinated ion- and time-dependent manner; in the HOCM groups, diatrizoate induced cultured HKC apoptosis. In the LOCM groups, iohexol did not induce apoptosis. Compared with equal osmotic pressure mannitol, apoptosis increased in HKCs incubated with diatrizoate (p<0.05). Bax/Bcl-2 production and caspase-3 activity were up-regulated in cultured HKCs treated with HOCM iodine 74 or 111 mg/mL meglumine diatrizoate.
  
  
  CONCLUSIONS
  Both HOCM and LOCM had toxic effects on HKCs, HOCM was more cytotoxic than LOCM; HOCM induced cultured HKC apoptosis while LOCM did not induce cultured HKC apoptosis in the indicated concentrations. The regulation of apoptosis induced by HOCM in HKCs may be regulated by Bax/Bcl-2 and caspase-3.
authors.affiliation: Institute of Nephrology, The Second Xiangya Hospital, Central South University, Changsha, Hunan, China. DuanSB528@hotmail.com
authors.forename: Shaobin
authors.initials: S
authors.lastname: Duan
authors.forename: Xiaorong
authors.initials: X
authors.lastname: Zhou
authors.forename: Fuyou
authors.initials: F
authors.lastname: Liu
authors.forename: Youming
authors.initials: Y
authors.lastname: Peng
authors.forename: Yinyin
authors.initials: Y
authors.lastname: Chen
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Pei
authors.forename: Guanghui
authors.initials: G
authors.lastname: Ling
authors.forename: Letian
authors.initials: L
authors.lastname: Zhou
authors.forename: Ying
authors.initials: Y
authors.lastname: Li
authors.forename: Yihua
authors.initials: Y
authors.lastname: Pi
authors.forename: Ke
authors.initials: K
authors.lastname: Tang
authors.forename: Ruihong
authors.initials: R
authors.lastname: Liu
authors.forename: Guiyan
authors.initials: G
authors.lastname: Li
mesh_terms: D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism*; D002460:Cell Line; D003287:Contrast Media / Q000633:toxicity*; D053938:DNA Fragmentation / Q000187:drug effects; D004847:Epithelial Cells / Q000378:metabolism*; D006801:Humans; D007684:Kidney Tubules / Q000378:metabolism*; D009994:Osmolar Concentration; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
chemical_list: D003287:Contrast Media; D051028:bcl-2-Associated X Protein; D053148:Caspase 3