Effect Of Selenium-rich Hot Spring Water On The Proliferate Of Hypertrophic Scar Fibroblasts

Background:Hypertrophic scar is result of tissue post-traumatic excessive repair. Any woundhealing is associated with the severity of different degrees of scarring. Fibroblast ismainly influencing factors cells of hypertrophic sacr formation and proliferate, afterburns occurred, the fibroblasts as the main component of the granulation tissue inburn wound were proliferate to fill wound, so as to promote wound healing.Therefore, the functional state of fibroblasts is main factor to affect the hypertrophicscar, development and the outcome, and it is the key link of hypertrophic scar tissueformation. At present, non-surgical treatment of hypertrophic scars contains pressure,laser, silicone application, radiation and drug injections, but their effects are poor, itis necessary to find a safe, effective, economic and convenient method of treatmentfor hypertrophic scar.In clinical practice, we found that selenium-rich hot spring water can promote thehealing of burn wounds and improve the patient’s immune system. The scars in burnpatients were soft, color lighe and rarely affect limb function leading to disability.But its exact mechainism is not clear. To further study the mechanism ofselenium-rich hot spring water bath to inhibit scar formation, we used selenium-richhot spring water intervene hypertrophic scar fibroblasts that cultured in vitro, andused direct counting method and MTT assay observed the proliferate of fibroblasts,initially explore the mechanism of selenium-rich hot spring water to inhibit scarformation.Purpose:The most obvious feature of the hypertrophic scar is fibroblasts proliferattion,clinical trials also confirmed that selenium-rich hot spring water can effectivelyinhibit the growth of scar. To further clarify the mechanism of action ofselenium-rich hot spring water to treatment of hypertrophic scar, we designed fromhypertrophic scar patients to remove scar tissue and culture in vitro, and was intervened by selenium-rich hot spring water, then we observed the proliferate offibroblasts, in order to provide the experimental basis for selenium-rich hot springwater to inhibit scar formation, initially explore the mechanism of selenium-rich hotspring water bath for the treatment of burn wounds.Methods:1. Orthopedic patients hospitalized burn unit in the First Affiliated Hospital ofNanchang University, we taken their hypertrophic scar improved tissue block understerile conditions to performed fibroblast cell culture. These cells was detected theactivity greater than90%, then continue to pass the culture to3-6generations andused them in the experiment.2. Direct cells counting method: good growth3rd generation cells were made to2×10~6/ml cell suspension and divided into four groups, include selenium-rich hotspring water group (group A), tap water group (group B), distilled water group（group C）, without any solution to normal culture medium group (group D, normalcontrol group), respectively. we used the24-well plate to cultured cells, the sameconcentration and the same solute medium in every three holes, the finalconcentration of group A, B, C were10%,20%,30%, respectively. Cells werecultured to a predetermined time under the37℃and the volume fraction of5%CO2were digested and collected, we used the counting board to count directly.3. MTT assay: after counting board to adjust the cell density to5×10~7/L, cellsseeded in96-well plates,100μL of cell suspension and100μL of DMEM mediumwas added in each hole, continued to cultured for24h, discard liquid when the cellswere completely adherent, added with a final concentration of10%,20%,30%ofselenium-rich spring water, tap water and distilled water DMEM medium (volumefraction of10%fetal bovine serum), respectively. Cells in normal DMEM mediumas the blank control group, and that in selenium-rich spring water group and blankcontrol group were observed by inverted microscope. The cultured cells in eachgroup into96-well plates into the incubator and cultured for48h, used the pipette toremove the original media carefully, MTT solution adding into each hole to continueculture for4h and discard the culture medium, then we adding dimethyl sulfoxideinto each hole150μL, taken it into the porous oscillator low-speed oscillation for10 min, measured490nm avsorbance with microplate reader, record data and calculatethe inhibition of cell growth rate.Results:1. cells observed by inverted phase contrast microscope, number of cells willreduce by cultured by selenium-rich hot spring water, cell processes to varyingdegrees will be shorter or deficiencies, cytoplasm and nucleus were condensed. Thecell survival conditions deteriorate with increase of the concentration of hot springwater.2. count results: cultured fibroblast count in each experimental group were lessthan the normal medium group, compare with the normal medium control group,cells in10%concentration of water medium group and10%concentration of thedistilled water medium group were essentially similar trend. Selenium-rich hotspring water group and20%,30%tap water and distilled water group for fibroblastgrowth had significant effect. Compare with selenium-rich hot spring water group,the number of cells in normal medium group, the difference was significant (P﹤0.05).3. MTT result: different concentrations of selenium-rich spring water, tap water,distilled water inhibit the proliferate of fibroblasts.The most obvious inhibitoryeffect was selenium-rich hot spring water medium. and the inhibition rate increaseswith medium concentration increased.Conclusion:Selenium-rich hot spring water with a certain concentration range can dose-dependently inhibited the proliferation of human hypertrophic scar fibroblasts.