Regulation Of Bioelectric Field On Skin Fibroblast Transformation And Its Mechanism

Background: Wound contraction and later scar repair is the focus of clinical wound treatment,and it is also a hot and difficult point of research.Myofibroblasts play an important role in it.Myofibroblasts are transformed from fibroblasts,so regulating the transformation of fibroblasts into myofibroblasts is of great significance to wound healing.The wound bioelectric field is generated immediately after the wound is formed,with the positive pole at the wound edge and the negative pole at the center of the wound.The intensity of the electric field reaches its peak within 3 days and then gradually decreases until the wound is healed.The electric field plays an important role in the healing of the wound,and the healing speed of the wound is slowed down when the electric field is removed.Earlier studies have shown that electric fields can affect microtubules and microfilaments in fibroblast cells to induce their directional migration,but the mechanism and whether the electric field regulates fibroblast transformation are still unclear.In the early stage of the wound,fibroblasts become active,migrate slowly to the wound edge,transform into myofibroblasts and secrete actin and other related proteins,and participate in wound healing by remodeling the extracellular matrix and shrinking the wound.As the wound heals gradually,myofibroblasts undergo apoptosis.Combining the corresponding relationship between the bioelectric field of the wound and the dynamic process of fibroblasts,we speculate that the electric field may be involved in the transformation of wound fibroblasts.RhoA GTPase(RhoA)is a widely expressed membrane-associated guanine nucleotide binding protein,which is essential for regulating various cell functions.Members of the Rho kinase(ROCK)family,including ROCK1 and ROCK2,are key effectors of RhoA.RhoA plays a key role in actin cytoskeleton reorganization,cell shape regulation,adhesion and cell transdifferentiation.We suspect that RhoA/ROCK may be involved in the regulation of fibroblast transformation by electric field.The study puts forward the scientific hypothesis that the bioelectric field regulates the conversion of fibroblasts to myofibroblasts through RhoA/ROCK.By establishing an in vitro cell electrification model that simulates the bioelectric field of the wound,the effect of electric field on the regulation of fibroblasts to myofibroblast conversion is detected.To explore the regulation of the electric field on the RhoA/ROCK of fibroblasts,clarify the regulation mechanism of the electric field on the transformation of fibroblasts,and enrich the theoretical system of the molecular mechanism of electric field promoting wound healing.Objective: To Investigate the role of electric field in the transformation of fibroblasts into myofibroblasts and explore the mechanism of electric field regulating fibroblast transformation..Methods: The HSF cell line of human dermal fibroblasts was selected as the experimental object.First,the changes in the arrangement and migration of fibroblasts under an electric field intensity of 200 m V/mm were observed,and their movement trajectories statistically were analyzed.The expression of α-SMA,COL-1,COL-3,PCNA,RhoA and ROCK1 in the cells after electric field treatment were detected.After using RhoA inhibitor Y27632,the effect of inhibiting the RhoA/ROCK1 signal pathway on the expression of α-SMA,COL-1 and COL-3 under the action of an electric field were detected.Cell movement speed,trajectory and direction were detected by live cell workstation.The protein expression ofα-SMA,COL-1,COL-3,PCNA,RhoA and ROCK1 in fibroblasts were detected by western blotting.The expression and distribution of α-SMA in cells were detected by immunofluorescence staining.Results: The bioelectric field regulated the migration of fibroblasts from the negative electrode to the positive electrode and accelerated the movement speed.At the same time,the secretion of matrix proteins such asα-SMA,COL-1,COL-3 increased.And the expression of α-SMA,COL-1,COL-3 had a certain dependence on the prolongation of treatment time and the increase of electric field intensity.There was no significant change in the number of cells treated by electric field in a short period of time.PCNA proliferation protein decreased with the increase of electric field treatment time and electric field intensity.Electric field treatment can activate RhoA/ROCK activity.The ability of fibroblasts to transform into myofibroblasts under the action of electric field is weakened after inhibiting RhoA/ROCK.Conclusion: Physiological electric field promotes the transformation of fibroblasts into myofibroblasts by regulating the activity of RhoA/ROCK1.The proliferation potential of myofibroblasts is lower than that of fibroblasts.These results provide new ideas for the molecular mechanism of EF-induced skin fibroblast differentiation during wound healing and new ideas for the treatment of wound repair.