Biological Effects Of Iodine Preparations On Skin Fibroblasts In Vitro

PART ONEBiological effects of iodine preparations on the adhesion and proliferation of human skin fibroblasts in vitroObjectiveIn this study, we use several kind of techniques including cell culture in vitro, immunofluorescence cytochemistry and MTT for detection, to investigate the biological effects of potassium iodide and povidone-iodine (PI).on the adhesion and proliferation of human skin fibroblasts (HSF) in vitro and to explore the role of iodine in wound healing. The research "may enrich experimental research of iodine, providing an effective experimental support for the human body.Methods1. Cell strain:The cells(Human Skin Fibroblast, HSF)were bought from kunming cellbank of Chinese Academy of Sciences. The following expressions proceed when the cells passage steadily.2. groups:Human skin fibroblasts were cultured in vitro, and were divided into 9 groups:control group,3 mg/L KI group,30 mg/L KI group,300 mg/L KI group,3000 mg/L KI group,1 mg/L PI group,2 mg/L PI group,3 mg/L PI group,4 mg/L PI group.3. observation of morphology:The cells were seeded in 6-well culture plates and cultured in the DMEM according to experimental design. Cell were observed after 96 hours.4. MTT for cell adhesion:The cells were cultured in the DMEM which contains different concentration of iodine according to experimental design. After 96 hours inoculated in 96-well plates, each set up five re-holes, 1h later sucking the medium, using D-Hanks lightly washed three times.Human skin fibroblasts adhesion were measured with MTT5. MTT cell proliferation:The cells were cultured in the DMEM which contains different concentration of iodine according to experimental design, cell proliferation were determined by MTT in2d,4d and 6d.6. immunofluorescence:Coverslips were sterilized and put in 6-well plates. Cells were seeded on the coverslips and cultured in the DMEM which contains different concentration iodine for 96 hour. Stained By immunofluorescence cytochemistrying method,then the cells were observed under fluorescence microscope.the photos were analyzed by Image-pro plus 6.0.ResultsIn 3 mg/L KI group,30 mg/L KI group and 300 mg/L KI group, the adhesion and proliferation of skin fibroblasts were remarkably increased, especially in 3 mg/L group the differences were significant compared with the iodine free control (P< 0.05), but the proliferation activity was progressively retarded in 3000 mg/L concentration. In povidone iodine groups, Fibroblast growth was totally inhibited by povidone iodine solutions.ConclusionProper concentration of KI can promote fibroblast adhesion and proliferation, but high concentration of KI lead to apoptosis; Even dilute solutions of povidone-iodine were toxic to human fibroblasts.PART TWOEffects of KI on collagen synthesis of skin fibroblasts in vitroObjectiveTo study the effects of KI on the collagen synthesis of skin fibroblasts in vitro and to explore the role of iodine in wound healing.MethodsHuman skin fibroblasts were cultured in vitro, then our experiment is divided into 5 groups:control group,3mg/L KI group,30mg/L KI group,300mg/L KI group and bFGF, after being stimulated by KI with different concentration for 96 hours, collagen synthesis of human skin fibroblasts were examined by hydroxyproline colorimetric analysis and Real-Time PCR respectively.Resultsâ‘ 3mg/L KI and 30mg/L KI group remarkably increase collagen content in supernatant of cell and in endochylema, especially in 3mg/L group (P< 0.05).â‘¡Compared with the control group, typeâ…¢collagen mRNA expression in 3mg/L KI group was significantly enhanced (P<0.05),and 30mg/L KI group and 300mg/L KI group had no statistical significance, the difference of typeâ… collagen mRNA expression were not statistically significant between each groups.ConclusionIn the gene level of regulation,proper concentration of KI promote cell synthesis and secretion of collagen. Furthermore without affecting the synthesis of typeâ… collagen, Proper concentration of KI effectively increase typeâ…¢collagen content, thereby it may promote wound healing, and reduce scar formation in the wound healing process.PART THREEEffect of iodine on apoptosis in cultured human skin fibroblasts in vitro ObjectiveTo investigate the influence of iodine on apoptosis of human skin fibroblasts in vitro and to explore the mechanism that iodide ions affect cell proliferation and apoptosis. MethodsHuman skin fibroblasts were cultured in vitro, then our experiment is divided into 4 groups:control group,3mg/L KI group,30mg/L KI group,300mg/L KI group, after being stimulated by KI with different concentration for 96 hours. Apoptosis gene Bcl-2 and Bax expression were detected by Western blotting.Results3 mg/L KI concentrations significantly increased bcl-2 expression, but bcl-2 expression was gradually inhibited when using the higher iodine concentrations. 300mg/L KI increased the expression of Bax expression. ConclusionApoptosis is influenced by iodine in cultured human skin fibroblasts through the modulation of Bcl-2 and Bax expression. Low concentration of iodine inhibits apoptosis, whereas high concentrations of iodine increase the rate of apoptosis.