Dracohodin Perochlorate Stimulates Fibroblast Proliferation via EGFR Activation and Downstream ERK/CREB and PI3K/Akt/mTOR Pathways In Vitro.

Lin Liu,Xiaowen Jiang,Wenhui Yu

doi:10.1155/2019/6027186

Abstract

In recent years, an increasing number of natural plant extracts have been determined to be potential drugs for various illnesses. In this study, we investigated the effects of dracorhodin perchlorate (DP) on fibroblast proliferation, which is crucial for wound healing. Cell proliferation assays were performed by different concentrations of DP, and the cell viability was detected by CCK-8 kits. After DP treatment for 24 h, the cell cycle was checked by flow cytometer. EGFR and downstream signaling pathways ERK1/2 and PI3K were examined with DP treatment by western blot. We further determined the effects of the related inhibitors on DP-induced relative protein phosphorylation and cell proliferation. The results showed that 3 μg/mL of DP promoted cell proliferation most significantly at treatment lengths of 24 h, and the percentage of cells in the S + G2 phase increased compared to those of the control group. In western blot detection, we found that DP significantly upregulated EGFR phosphorylation and activated the downstream ERK/CREB and PI3K/Akt/mTOR signaling pathway. Moreover, the results also showed that AG1478 abolished DP-induced relative protein activation and cell proliferation. When U0126 or LY294002 pretreated cells alone, DP-induced p-ERK or p-PI3K downstream proteins and cell proliferation were suppressed compared to those of the control group, but EGFR was not affected. In addition, ICG001 and BEZ235 collectively eliminated DP-induced fibroblast proliferation. Our findings suggest that DP-promoted fibroblast proliferation is stimulated by p-EGFR-induced activation of the ERK1/2-CREB and PI3K/Akt/mTOR pathways. Our present study explored the mechanism of DP-promoted fibroblast proliferation and provided a new basis for wound healing.

Highlights

Skin damage is common in clinical surgery
We found that inhibitors did not affect cell proliferation when cells were not treated by dracorhodin perchlorate (DP) (P > 0.05, Figure 6(a)). e results of subsequent detections are shown in Figure 6(b), which demonstrates significantly low cell viability due to the inhibition of EGFR, ERK1/2, PI3K-AKT, CREB, and mTOR (92%, 49%, 47%, 48%, and 50%, respectively) compared to the DP-treated group without inhibitor pretreatment
In the detections of EGFR and FGFR after DP treatment, we found that EGFR phosphorylation level was significantly increased, but FGFR has no obvious change. us, we speculated that EGFR may be the molecular target of DP acting on fibroblasts

Summary

Introduction

Skin damage is common in clinical surgery. Further, wound healing is a complex biological process. Fibroblasts are the most vital cells in the wound healing process because they synthesize and secrete a large number of collagen fibers and matrix components, which combine with the capillaries to form granulation tissue. It is beneficial for the epidermal cells to cover the wound [4]. E EGFR signaling pathway plays an important role in cell processes, such as cell growth, proliferation, and differentiation [21]. Cell proliferation caused by EGFR activation mediates two main transmission methods to the nucleus: Ras/ Raf/MAPK and the PI3K/PKC/IKK signaling pathways. The relative signaling pathway was detected with or without the inhibitor pretreatment. e purpose of this study was to determine the mechanism of DP-induced fibroblast proliferation and provide a basis for further clinical application

Materials and Methods

Results

Discussion

Conclusion