Death Signal"Transduction And Modulation Mediated By Fas On Fibroblasts Derived From Pathological Scars

Our Previous studies have shown that fibroblasts derived from keloids showed highly expressed Fas antigen and resistance to apoptosis induced by FasMcab in contrast to hypertrophic scars and normal skins. At the same time, low expression of Bcl-2 protein were testified in all samples. Then we indicated that Fas signal transduction must be blocked in firoblasts derived from keloids.. Purpose: To explore the concrete occlusive location, we investigate the the gap junctional communication and responses of intercellular Ca2~ LPO and ceramide to different concentration of FasMcab in fibroblasts derived from hypertrophic Scar and keloid. Our following purpose is to identify if the mutation of Fas exist which be responsible for lack of Fas- mediated apoptosis. Method: 1 .Each of six samples were collected from hypertrophic scars.. keloids. The gap junctional communication among ftbroblasts derived from these samples was investigated by use of the adherent cell analysis and sorting interactive laser coytometer(ACAS 570) after being cell-cultured. 2. Fibroblasts derived from these samples were exposed to different concentration of FasMcab for 24 hours to investigate the Fas Induced apoptosis .With the help of the adherent cell analysis and sorting interactive laser coytometer(ACAS 570), the responses of intercellular Ca2~.. H~ LPO were detected at the same time.3. Fibroblasts derived from these samples were exposed to different concentration of FasMcab for 20 minutes and then intercellular ceramide was quantified by the diacyigiycerol kinase assay. 4. keloids and hypertrophic scar tissue and the peripheral blood of each Patient were collected. After DNA isolated from the samples, we used the single-stranded conformation polymorphism (SSCP) analysis and DNA sequencing, to examined the structure of the Fas gene Result: 1 .The result indicated that fibroblasts derived from the normal 8 skins showed normal gap junction; gap junctional communication was restrained in fibroblasts derived from the hypertropic scars and the intracellular communication was completely occLuded in fibroblasts derived from the keloids. 2. Exposed to different concentration of FasMcab, the intercellular Ca2~ ~. LPO and ceramide rise prominently in hypertophic scar derived fibroblasts but no changes were seen in keloids derived ftbroblasts. 3. Approximate fall of H~ was seen in both kinds of fibroblasts. 4. Mutation in the Fas gene were identified and confirmed in the keloid tissue of 20%(2110) of the keloid patientwhich may cause loss of the function and contribute to the pathogenesis of keloid. A deletion of the nucleotide 揂?at the same position was found, which caused a frameshift mutations in exon 9. Conclusion: 1 .The occlusion of intracellular communication may play an important role in the pathogenesis of excessive and invasive growth of fibroblasts derived from the keloids. 2. As important messengers, the deficiency of the production of intracellular Ca2~ and LPO may be responsible for the abnormal apoptosis of keloid. 3. as the second messenger of the 揇eath?signal transduction mediated by Fas , the production of ceramide may be deficiect in keloid derived fibroblasts exposed to FasMcab. The occlusio...