【摘要】：背景:基于溫敏培養(yǎng)皿的細(xì)胞膜片技術(shù)在組織工程研究中受到越來(lái)越多的關(guān)注,但是溫敏培養(yǎng)皿價(jià)格高昂,有必要研究替代方法。目的:應(yīng)用普通培養(yǎng)皿進(jìn)行細(xì)胞膜片培養(yǎng),并分析軟骨細(xì)胞膜片的成軟骨性能。方法:取1例鼻中隔偏曲患者手術(shù)切除的鼻中隔軟骨消化提取原代軟骨細(xì)胞,擴(kuò)增培養(yǎng)。取第3代細(xì)胞在體外密集培養(yǎng),使之分泌細(xì)胞外基質(zhì),形成單層細(xì)胞膜片;另取第2代軟骨細(xì)胞,疊加接種于單層細(xì)胞膜片上,使其貼覆生長(zhǎng),形成雙層細(xì)胞膜片。應(yīng)用細(xì)胞刮刀收獲細(xì)胞膜片,研究其性狀,進(jìn)一步在體外培養(yǎng)、植入裸鼠體內(nèi)構(gòu)建出人工軟骨塊。結(jié)果與結(jié)論:(1)單層細(xì)胞膜片和雙層細(xì)胞膜片均形成質(zhì)軟的凝膠樣結(jié)構(gòu),大體觀察兩者無(wú)明顯區(qū)別;(2)新刮起的細(xì)胞膜片具有良好的可流動(dòng)性和成膠性;植入裸鼠體內(nèi)培育8周之后,均形成成熟的軟骨塊;(3)組織學(xué)觀察見(jiàn)細(xì)胞膜片是由成層排列的軟骨細(xì)胞和細(xì)胞外基質(zhì)組成的薄片,在體外培養(yǎng)過(guò)程中即有糖胺聚糖和Ⅱ型膠原的初步表達(dá),植入體內(nèi)8周之后形成的軟骨呈現(xiàn)為成熟的軟骨組織;(4)生化定量檢測(cè)結(jié)果表明,DNA含量高于天然鼻中隔軟骨,而糖胺聚糖和羥基脯氨酸含量與天然軟骨差異無(wú)顯著性意義;(5)結(jié)果提示,應(yīng)用普通培養(yǎng)皿可成功構(gòu)建并獲取細(xì)胞膜片,細(xì)胞膜片呈現(xiàn)良好的成軟骨性能。
[Abstract]:Background: cell membrane technology based on thermo-sensitive petri dish has attracted more and more attention in tissue engineering, but the price of thermo-sensitive dish is high, so it is necessary to study alternative methods. Aim: to study the chondrogenic properties of chondrocytes in vitro. Methods: the primary chondrocytes were extracted from a patient with nasal septum deviation. The third generation of chondrocytes were cultured in vitro to secrete extracellular matrix to form monolayer cell membrane, and the second generation chondrocytes were inoculated on the monolayer cell membrane to form double layer cell membrane. The cell scraper was used to harvest the cell membrane and to study its properties. The artificial chondrocytes were further cultured in vitro and implanted into nude mice to construct artificial chondrocytes. Results and conclusions: (1) soft gelatin-like structures were formed in monolayer and bilayer cell membranes, and there was no significant difference between them, (2) the newly scraped membranes had good fluidity and gelling ability. After 8 weeks of implantation, mature chondrocytes were formed in nude mice. (3) histological observation showed that the membrane slices were composed of chondrocytes and extracellular matrix. The primary expression of glycosaminoglycan and type 鈪，

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