Effect Of Connective Tissue Growth Factor On The Pathogenesis Of The Human Hypertrophic Scar

Hypertrophic scar (HS), characterized by excessive fibroblast (Fb) proliferation and local collagen deposition, is a very common pathologic phenomenon of scar hyperplasia after human skin injury and wound healing, and it's pathogenesis is still unclear. Connective tissue growth factor (CTGF) is a kind of important cytokine in the process of tissue fibrosis. During the pathologic process, the over expression of the CTGF is closely related with some proliferative and fibrotic diseases, such as scleroderma,renal fibrosis,cirrhosis of liver,pulmonary fibrosis and atherosclerosis, etc. The objective of this study is to explore the effect of CTGF on the pathogenesis of HS through detecting the expression of CTGF in the HS tissue and observing the influence of blocking the expression of CTGF on the proliferation,collagen synthesis,the expression of CTGF protein and messengerribonucleicacid (mRNA) of the cultured hypertrophic scar fibroblast (HSF). The study included: 1. To clarify whether there is high level expression of CTGF in HS tissues, the sirius red stain and polarization microscopy methods were used to detect the degrees of fibrosis in normal skin (NS),flat scar (FS) and HS tissues, and the immunohistochemistry strept avidin-biotin complex (SABC) staining method and image analysis technique were used to detect the expression of connective tissue growth factor protein in them. 2. To clarify the direct effect of CTGF on the cultured HSF, CTGF antisense oligonucleotide (ASODN) was specificly synthesized and phosphorothioated, and part of it was labelled by fluorescein isothiocyanate (FITC) in the 5'end of the chain. With the encapsulation of liposome, the CTGF ASODN was transfected into the cultured HSF. The intracellular distribution of CTGF ASODN after transfection was observed by fluorescence microscopy in the fixed HSF. The proliferation of HSF after transfection was measured by MTT test. The collagen synthesis of HSF after transfection was measured by H3-proline incorporation method. The expression of CTGF protein of HSF after transfection was detected by immunocytochemistry SABC staining method. The expression of CTGF mRNA of HSF after transfection was detected by reverse transcription polymerase chain reaction (RT-PCR). The results were as the following: 1. The area value of CTGF positiveparticles of HS tissue was higher than that of NS and FS tissues, and there were very significant differences among them by image analysis and statistics. 2. After the gene transfection, the CTGF ASODN was fisrt localized in the cytoplasm, then accumulated into the nucleus, and dispeared at last. When compared with the control groups, the proliferation,collagen synthesis and the expression of CTGF protein and mRNA of the cultured HSF were down regulated, and there were very significant differences among them by statistics.We conclude that 1. CTGF protein is over expressed in the HS tissue. 2. CTGF ASODN can markedly inhibit the proliferation,collagen synthesis and the expression of CTGF protein and mRNA of the cultured HSF. 3. CTGF can exert important influences on promoting the fibrotic process of human HS, and it is possible to stop the development of HS by inhibiting the expression of CTGF.