本文選題：增生性瘢痕 + 成纖維細(xì)胞　； 參考：《太原理工大學(xué)》2015年碩士論文

【摘要】：增生性瘢痕是人體皮膚真皮層受到創(chuàng)傷后引起的一系列真皮層增生。在創(chuàng)傷修復(fù)過(guò)程中,成纖維細(xì)胞起到至關(guān)重要的作用,了解和控制成纖維細(xì)胞的生物學(xué)行為是促進(jìn)傷口愈合和預(yù)防瘢痕形成的基礎(chǔ)和關(guān)鍵。近年來(lái),隨著現(xiàn)代細(xì)胞生物學(xué)和分子生物學(xué)在瘢痕領(lǐng)域的深入研究和迅速發(fā)展,對(duì)成纖維細(xì)胞、細(xì)胞外基質(zhì)和細(xì)胞因子三者之間的相互作用有了進(jìn)一步的認(rèn)識(shí)。成纖維細(xì)胞可以分泌細(xì)胞外基質(zhì),而成纖維細(xì)胞的大量增殖、細(xì)胞外基質(zhì)中膠原合成與降解、部分細(xì)胞因子的大量產(chǎn)生以及三者之間的密切關(guān)系構(gòu)成了增生性瘢痕形成的生物學(xué)基礎(chǔ)。研究發(fā)現(xiàn),在創(chuàng)傷初期,表皮細(xì)胞和真皮細(xì)胞將分泌白介素-1β(Interleukin-1β, IL-1β),且IL-1p具有促進(jìn)成纖維細(xì)胞、血管內(nèi)皮細(xì)胞增殖和細(xì)胞外基質(zhì)沉積作用。本文實(shí)驗(yàn)在細(xì)胞水平,并結(jié)合生物學(xué)的方法和手段,研究了人皮膚成纖維細(xì)胞(human skin fibroblasts, HFB)和瘢痕成纖維細(xì)胞(hypertrophic scar fibroblasts, HSF)在離體狀態(tài)下,不同濃度外源性的IL-1β刺激后,其增殖情況和細(xì)胞外基質(zhì)中膠原蛋白代謝的改變,有助于進(jìn)一步了解瘢痕形成的生物學(xué)機(jī)制以及為增生性瘢痕的治療方法及評(píng)估提供重要參考。本文的主要工作及結(jié)論如下： (1)采用組織塊貼壁法提取并培養(yǎng)HSF,采用膠原酶消化法提取并培養(yǎng)HFB, HSF與HFB在形態(tài)上無(wú)明顯差異,均呈長(zhǎng)梭形；本實(shí)驗(yàn)均使用第3-5代生長(zhǎng)狀態(tài)良好且處于對(duì)數(shù)增殖期的細(xì)胞,以每孔1.5×104個(gè)細(xì)胞的密度接種于6孔培養(yǎng)板或103個(gè)細(xì)胞的密度接種于96孔培養(yǎng)板,24h后,分別用2.5、5、10與20ng/mL濃度的IL-1β培養(yǎng)細(xì)胞24h,以0ng/mL濃度的IL-1β培養(yǎng)細(xì)胞24h為對(duì)照組；檢測(cè)細(xì)胞增殖以及細(xì)胞外基質(zhì)中膠原蛋白代謝的相關(guān)指標(biāo)。 (2)采用MTT比色法測(cè)定各組細(xì)胞的增殖情況,流式細(xì)胞分析法測(cè)定各組細(xì)胞的細(xì)胞周期,并分析各組成纖維細(xì)胞S期所占比例。上述兩種檢測(cè)方法均得出：IL-1β可以促進(jìn)HFB增殖,抑制HSF增殖,從而抑制瘢痕的形成；并且隨著培養(yǎng)液中IL-1β濃度的升高,其促進(jìn)HFB的增殖作用越明顯,抑制HSF的增殖也越明顯。 (3)采用ELISA法檢測(cè)各組成纖維細(xì)胞上清液中Ⅰ型膠原蛋白和Ⅲ型膠原蛋白的濃度。上述檢測(cè)方法得出：IL-1β可以促進(jìn)HFB合成并分泌Ⅰ、Ⅲ型膠原蛋白,抑制HSF合成分泌Ⅰ、Ⅲ型膠原蛋白,從而抑制瘢痕發(fā)生；并且隨著培養(yǎng)液中IL-1β濃度升高,其促進(jìn)HFB膠原蛋白代謝的作用越明顯,抑制HSF膠原蛋白代謝的作用也越明顯。IL-1β刺激能夠影響成纖維細(xì)胞Ⅰ、Ⅲ型膠原蛋白的合成與分泌,且與IL-1β的濃度有關(guān),期望該結(jié)論能為相關(guān)疾病臨床用藥劑量控制提供理論依據(jù)。
[Abstract]:Hypertrophic scar is a series of cortical hyperplasia caused by trauma of human dermis. Fibroblasts play an important role in wound healing. Understanding and controlling the biological behavior of fibroblasts is the basis and key to promote wound healing and prevent scar formation. In recent years, with the deep research and rapid development of modern cell biology and molecular biology in the field of scar, the interaction among fibroblasts, extracellular matrix and cytokines has been further understood. Fibroblasts secrete extracellular matrix (ECM), and fibroblast proliferation, collagen synthesis and degradation in ECM. The production of some cytokines and their close relationship constitute the biological basis of hypertrophic scar formation. It was found that in the early stage of trauma, the epidermal cells and dermal cells secreted interleukin-1 尾 -interleukin-1 尾, IL-1 尾, and IL-1p promoted fibroblast, vascular endothelial cell proliferation and extracellular matrix deposition. In this paper, at the cell level, combined with biological methods and methods, we studied the effects of exogenous IL-1 尾 on human skin fibroblasts skin fibroblasts, HFB) and scar fibroblasts stimulated by hypertrophic scar fibroblasts, HSF) in vitro. Its proliferation and changes of collagen metabolism in extracellular matrix are helpful to further understand the biological mechanism of scar formation and provide important reference for the treatment and evaluation of hypertrophic scar. The main work and conclusions are as follows: (1) HSF and HFB were extracted and cultured by tissue mass adherence and collagenase digestion, respectively. There was no significant difference in morphology between HSF and HFB, and the cells in good growth state and logarithmic proliferative phase were used in this experiment. The density of 1.5 脳 104 cells per well was inoculated in 6-well culture plate or 103 cell density in 96-well culture plate for 24 hours. The cells were cultured with IL-1 尾 at the concentration of 2.5 渭 g / 10 and 20ng/mL for 24 h, respectively, and IL-1 尾 with 0ng/mL concentration for 24 h as control group. Cell proliferation and collagen metabolism in extracellular matrix were measured. (2) MTT colorimetric assay was used to determine cell proliferation, flow cytometry was used to determine cell cycle, and the percentage of fibroblasts in S phase was analyzed. The above two methods showed that IL-1 尾 could promote the proliferation of HFB, inhibit the proliferation of HSF and inhibit the formation of scar, and with the increase of the concentration of IL-1 尾 in the culture medium, the more obvious the effect of promoting the proliferation of HFB, the more obvious the inhibition of the proliferation of HSF. ELISA assay was used to detect the concentration of collagen type 鈪，

本文編號(hào)：1804512