Role Of Oxidative Damage And Mechanism Of Chromium Picolinate On Chick Embryo Fibroblast

Cr is well known as a kind of essential micronutrients of human and mammals, Cr(Ⅲ)affects lipids, carbohydrates, proteins, and nucleic acid metabolism by synergistic effect ofGTF and improvement of insulin. Nowadays, Cr(Ⅲ) has been widely used in the health careproducts, pharmaceutical and food. From the1990s, the multifunction of Cr(Ⅲ) preparationin animal nutrition is more and more concerned by people. Crpic as a kind of nutritionadditive is widely employed. As cytotoxicity of Crpic reported in1995, Safety problem for ithas become the research hot region. Although some countries approved the adding ofchromium picolinate in animal rations, Minimum requirements and maximum tolerated doseare not reported.The toxicity testing of Crpic are mainly concentrated in human and mouse in vito up tonow, while the damage effects of CrPic to poultry has no research. This reseach was in vitroconducted to investigate the influence of different concentrations of Crpic for growth ofchicken embryo fibroblast (CEF), and to provide a theory basis for rational utilization. In thisstudy, chicken embryo from the9to11hatching day were used in preparation of cell culture,after2following passages, cultured with nutrient solution containing Crpic which finalconcentration was0,8,16,160,400,600μM respectively, cultured for24,48,72hours, cellviability was determined with MTT method. Meanwhile, morphological detection wasdetermined with fluoresence staining, ROS generation, calcium content, the change ofmitochondrial membrane protential and apoptosis rate were determined with FCM (flowcytometry) at48hours. The activity of antioxidase and content of glutathione andmalondialdehyde (MDA) was determined with colorimetric analysis. All these experimentscould evaluate the effect of CrPic on chicken embryo fibroblast activity.By morphology analysis and biochemical analysis on the materials, CrPic is toxic tochicken embryo fibroblasts when which concentration reach400μM and above, while theother concentration had no obvious effect. Cell viability detecting indicated that CrPicinduced cell death in a dose-time relationship, and effects were significantly higher than those in control subjects on400μM and above. The morphology analysis indicated that cellmorphology change, chromatin agglutination and cell apoptosis body at higher concentration(400μM,600μM). And cell apoptosis rate was significantly higher than control subjects. Notsignificantly at8μM，16μM，160μM. We detected a marked increase in ROS (P<0.01), Ca2+(P<0.01) and significant decrease in Mitochondrial membrane potential at160μM、400μMand600μM, not significantly at8μM,16μM. superoxide dismutase, glutathion peroxidase,CAT and glutathione than control subjects slightly raising or not significant difference at8μM,16μM, decrease or significant decrease at160μM,400μM,600μM (P<0.01).Malondialdehyde content was not significant difference at8μM,16μM and160μM, butmarked increase at400μM,600μM than control subjects (P<0.01). Validated by theexperiments, CrPic could change oxidation states of chicken embryo fibroblast, induce celldeath in a dose-time relationship. High concentrations of CrPic might induce cell death byoxidative damage mitochondrial dysfunction.