核心蛋白多糖在瘢痕组织及成纤维细胞中的表达及意义
摘要
一、论文核心蛋白多糖在瘢痕组织及成纤维细胞中的表达及意义
【目的】对病理性瘢痕组织及成纤维细胞中Decorin的含量及表达进行检测,深入探讨并揭示Decorin对瘢痕的作用及机制,为临床预防和治疗瘢痕提供分子及病理学水平的理论依据。
【方法】(1)对病理性瘢痕成纤维细胞进行体外培养,采用光镜、扫描电镜观察成纤维细胞形态、活性及凋亡等;提取细胞RNA,应用RT-PCR对Decorin、TGF-β1的mRNA表达进行检测分析。(2)将病理性瘢痕组织进行固定、包埋、切片,采用免疫组织化学染色对Decorin、TGF-β1进行检测。通过以上实验观察及统计学分析,以研究Decorin对病理性瘢痕成纤维细胞生物学行为的影响及病理改变,Decorin在病理性瘢痕发生发展中的作用。
【结果】(1)光镜下:成纤维细胞为贴壁生长,长梭形或多边形,呈栅栏状排列;与正常皮肤成纤维细胞相比,病理性瘢痕成纤维细胞形态不规则、体积较大,细胞排列较乱;(2)电镜下:与正常皮肤对照组相比,瘢痕成纤维细胞线粒体增多,粗面内质网增多并肿胀、扩张呈囊状,胞质内微丝微管增多,细胞核内常染色质丰富,表明其合成蛋白的功能活跃;(3)RT-PCR结果显示:瘢痕疙瘩成纤维细胞中Decorin mRNA含量较正常瘢痕或正常皮肤成纤维细胞降低;瘢痕疙瘩成纤维细胞中TGF-β1 mRNA表达较正常皮肤及瘢痕组织成纤维细胞升高;(4)免疫组化结果显示:Decorin主要表达于正常皮肤真皮层细胞外基质中及胶原纤维表面,与正常皮肤相比,Decorin在瘢痕组织中表达量明显降低;而TGF-β1在瘢痕组织中表达量明显高于正常皮肤中,具有统计学意义(P＜0.05)。
【结论】Decorin在病理性瘢痕组织及成纤维细胞内含量较正常皮肤明显减少,提示在创面愈合早期由于Decorin含量降低,使TGF-β1活性上调,其对成纤维细胞的刺激作用也随之增加,引起成纤维细胞的大量增生、迁移,并合成过量胶原,这可能是病理性瘢痕形成的一个重要因素。
【Objective】Measure the expression of Decorin on scar tissues and fibroblast to depth explore and reveal the role of Decorin and mechanism for the prevention and treatment of clinical and pathological on scar therapy as the theoretical basis at the molecular level.
【Methods】(1) Hypertrophic scar fibroblasts were cultured in vitro,using light microscopy and electron microscopy to observe the fibroblasts' morphology, activity,apoptosis.Using the real-time quantitative PCR to detecte and analyze the expression of mRNA of Decorin and TGF-β1 in fibroblasts.(2) Scar tissue were fixed,embedded,sliced,using immunohistochemical detect the expression of the Decorin and TGF-β1 in scar tissue.By analyzing the experimental observation of above and we would study scar fibroblasts behavior and pathological,and further more to discuss the pathological role of decorin in the develoPment of scar formation.
【Results】(1)Light microscopy showed:Scar fibroblasts adherent growth,spindle or polygonal,showing palisade arrangement.Compared with normal skin fibroblasts,scar fibroblasts are irregular,larger cells and arranged in disorder.(2)Electron microscopy shows:Compared with normal skin control group, with a large number of mitochondria,rough endoplasmic reticulum was reduced and see more rough endoplasmic reticulum swelling,expansion.Microtubule and microfilament in cytoplasm increased and euchromatin-rich in nucleus.Above show that the function of its protein synthesis is very active.(3)The result of real-time quantitative PCR showed that:Compared with normal skin,the expression of Decorin levels were significantly reduced in scar fibroblasts.On the other hand,the expression of TGF-β1 levels were significantly increased in scar fibroblasts and was statistically significant.(4)Immunohistochemistry on analysis showed that:Decorin was mainly expressed in normal skin dermis and collagen extracellular matrix in the fiber surface.Compared with normal skin control group,Decorin expression in scar tissue decreased;TGF-β1 expression in the scar tissue was significantly higher than in normal skin and statistically significant(P＜0.05).
【Conclusions】Compared with normal skin,Decorin levels in scar tissue and fibroblast decreased significantly.Decreasing of Decorin in early wound healing prompted activity of TGF-β,which caused a large number of fibroblast proliferation,migration and excessive collagen synthesis.This may be an important factor in scar formation.
【目的】对病理性瘢痕组织及成纤维细胞中Decorin的含量及表达进行检测,深入探讨并揭示Decorin对瘢痕的作用及机制,为临床预防和治疗瘢痕提供分子及病理学水平的理论依据。
【方法】(1)对病理性瘢痕成纤维细胞进行体外培养,采用光镜、扫描电镜观察成纤维细胞形态、活性及凋亡等;提取细胞RNA,应用RT-PCR对Decorin、TGF-β1的mRNA表达进行检测分析。(2)将病理性瘢痕组织进行固定、包埋、切片,采用免疫组织化学染色对Decorin、TGF-β1进行检测。通过以上实验观察及统计学分析,以研究Decorin对病理性瘢痕成纤维细胞生物学行为的影响及病理改变,Decorin在病理性瘢痕发生发展中的作用。
【结果】(1)光镜下:成纤维细胞为贴壁生长,长梭形或多边形,呈栅栏状排列;与正常皮肤成纤维细胞相比,病理性瘢痕成纤维细胞形态不规则、体积较大,细胞排列较乱;(2)电镜下:与正常皮肤对照组相比,瘢痕成纤维细胞线粒体增多,粗面内质网增多并肿胀、扩张呈囊状,胞质内微丝微管增多,细胞核内常染色质丰富,表明其合成蛋白的功能活跃;(3)RT-PCR结果显示:瘢痕疙瘩成纤维细胞中Decorin mRNA含量较正常瘢痕或正常皮肤成纤维细胞降低;瘢痕疙瘩成纤维细胞中TGF-β1 mRNA表达较正常皮肤及瘢痕组织成纤维细胞升高;(4)免疫组化结果显示:Decorin主要表达于正常皮肤真皮层细胞外基质中及胶原纤维表面,与正常皮肤相比,Decorin在瘢痕组织中表达量明显降低;而TGF-β1在瘢痕组织中表达量明显高于正常皮肤中,具有统计学意义(P＜0.05)。
【结论】Decorin在病理性瘢痕组织及成纤维细胞内含量较正常皮肤明显减少,提示在创面愈合早期由于Decorin含量降低,使TGF-β1活性上调,其对成纤维细胞的刺激作用也随之增加,引起成纤维细胞的大量增生、迁移,并合成过量胶原,这可能是病理性瘢痕形成的一个重要因素。
【Objective】Measure the expression of Decorin on scar tissues and fibroblast to depth explore and reveal the role of Decorin and mechanism for the prevention and treatment of clinical and pathological on scar therapy as the theoretical basis at the molecular level.
【Methods】(1) Hypertrophic scar fibroblasts were cultured in vitro,using light microscopy and electron microscopy to observe the fibroblasts' morphology, activity,apoptosis.Using the real-time quantitative PCR to detecte and analyze the expression of mRNA of Decorin and TGF-β1 in fibroblasts.(2) Scar tissue were fixed,embedded,sliced,using immunohistochemical detect the expression of the Decorin and TGF-β1 in scar tissue.By analyzing the experimental observation of above and we would study scar fibroblasts behavior and pathological,and further more to discuss the pathological role of decorin in the develoPment of scar formation.
【Results】(1)Light microscopy showed:Scar fibroblasts adherent growth,spindle or polygonal,showing palisade arrangement.Compared with normal skin fibroblasts,scar fibroblasts are irregular,larger cells and arranged in disorder.(2)Electron microscopy shows:Compared with normal skin control group, with a large number of mitochondria,rough endoplasmic reticulum was reduced and see more rough endoplasmic reticulum swelling,expansion.Microtubule and microfilament in cytoplasm increased and euchromatin-rich in nucleus.Above show that the function of its protein synthesis is very active.(3)The result of real-time quantitative PCR showed that:Compared with normal skin,the expression of Decorin levels were significantly reduced in scar fibroblasts.On the other hand,the expression of TGF-β1 levels were significantly increased in scar fibroblasts and was statistically significant.(4)Immunohistochemistry on analysis showed that:Decorin was mainly expressed in normal skin dermis and collagen extracellular matrix in the fiber surface.Compared with normal skin control group,Decorin expression in scar tissue decreased;TGF-β1 expression in the scar tissue was significantly higher than in normal skin and statistically significant(P＜0.05).
【Conclusions】Compared with normal skin,Decorin levels in scar tissue and fibroblast decreased significantly.Decreasing of Decorin in early wound healing prompted activity of TGF-β,which caused a large number of fibroblast proliferation,migration and excessive collagen synthesis.This may be an important factor in scar formation.
引文
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