Herbal formula Astragali Radix and Rehmanniae Radix exerted wound healing effect on human skin fibroblast cell line Hs27 via the activation of transformation growth factor (TGF-β) pathway and promoting extracellular matrix (ECM) deposition.
Astragali Radix (AR) and Rehmanniae Radix (RR) have long been used in traditional Chinese Medicine and as the principal herbs in treating diabetic foot ulcer. In this study, we investigated the effect of NF3, which comprises of AR and RR in the ratio of 2:1(w/w), on skin fibroblast cell migration and the activation of selected genes and proteins related to wound healing. Human skin fibroblast cell line Hs27 was treated with NF3 at 4 mg/ml for 24h, and in vitro scratch wound healing and quantitative cell migration assays were performed, respectively. The expression of transformation growth factor (TGF-β1) and bone morphogenetic protein 6 (BMP6) in Hs27 cells with or without NF3 treatment was analyzed by western blot analysis. In addition, the expression of a panel of genes involved in human TGF-β signaling pathway was analyzed in Hs27 cells upon NF3 treatment (4 mg/ml, 24 h) by quantitative real-time PCR (qRT-PCR). Furthermore, the expression of several genes and proteins associated with ECM synthesis was investigated by qRT-PCR analysis or/and ELISA techniques. The results suggested that NF3 promoted the migration of human skin fibroblast cells. Western blot analysis demonstrated that NF3 up-regulated TGF-β1 and BMP-6 synthesis. qRT-PCR analysis revealed that the expression of 26 genes in Hs27 cells was changed upon NF3 induction, including TGF-β superfamily ligands and down stream effectors genes, and genes involved in TGF/Smad pathway, and Ras/MAPK (non-Smad) pathway. Among the extracellular matrix (ECM)-related molecules, it was found that NF3 up-regulated the expression of type I and III collagens, fibronectin as well as TIMP-1, and down-regulated the MMP-9 expression in skin fibroblast cells. This study demonstrated that herb formula NF3 could enhance skin fibroblast cell migration and activated genes involved in TGF-β1 pathway. NF3 could regulate gene transcription for extracellular matrix synthesis via the Smad pathway, and gene transcription for cell motility via the Ras/MAPK (non-Smad) pathway.
Department of Biology and Chemistry, City University of Hong Kong, Kowloon, Hong Kong.
Qi
Q
Zhang
Chi Chun
CC
Fong
Wai Kin
WK
Yu
Yao
Y
Chen
Fan
F
Wei
Chi Man
CM
Koon
Kit Man
KM
Lau
Ping Chung
PC
Leung
Clara Bik San
CB
Lau
Kwok Pui
KP
Fung
Mengsu
M
Yang
D027883:Astragalus Plant; D055418:Bone Morphogenetic Protein 6 / Q000096:biosynthesis; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D003094:Collagen / Q000187:drug effects; D048909:Diabetes Complications / Q000188:drug therapy; D004359:Drug Therapy, Combination; D004365:Drugs, Chinese Herbal / Q000494:pharmacology* / Q000627:therapeutic use; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008517:Phytotherapy; D018517:Plant Roots; D032267:Rehmannia; D015398:Signal Transduction; D012867:Skin / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015533:Transcriptional Activation; D053773:Transforming Growth Factor beta1 / Q000096:biosynthesis*; D014945:Wound Healing / Q000187:drug effects* / Q000235:genetics; D014947:Wounds and Injuries / Q000235:genetics / Q000628:therapy
C525816:BMP6 protein, human; D055418:Bone Morphogenetic Protein 6; D004365:Drugs, Chinese Herbal; D005353:Fibronectins; D053773:Transforming Growth Factor beta1; D003094:Collagen