發(fā)布時(shí)間：2018-05-02 17:55

  本文選題：組織工程 + 種子細(xì)胞�。� 參考：《山西醫(yī)科大學(xué)》2016年碩士論文

【摘要】：背景:關(guān)節(jié)軟骨由于無血管、神經(jīng)及淋巴通過,其一旦損傷則很難自行修復(fù),已知組織工程技術(shù)促進(jìn)軟骨的損傷修復(fù),研究證實(shí):在體外,軟骨細(xì)胞作為種子細(xì)胞在體外培養(yǎng)極易發(fā)生去分化;在體內(nèi),經(jīng)由其發(fā)育而來的修復(fù)組織的表型及力學(xué)特性均較正常關(guān)節(jié)軟骨差,而且隨著體內(nèi)修復(fù)時(shí)間的推移,修復(fù)組織顯現(xiàn)出現(xiàn)快速退變的特征,制約其臨床應(yīng)用的前景。軟骨單位(chondron)被作為關(guān)節(jié)軟骨的基本功能結(jié)構(gòu),其主要由軟骨細(xì)胞與細(xì)胞周基質(zhì)(PCM)組成,其中PCM對軟骨細(xì)胞代謝和力學(xué)傳導(dǎo)起著重要作用。體外實(shí)驗(yàn)證實(shí):軟骨單位與軟骨細(xì)胞以1:1在海藻酸鈉凝膠球中共培養(yǎng),其生物學(xué)特性優(yōu)于其他比例;然而,軟骨單位和1:1共培養(yǎng)組對體內(nèi)軟骨缺損修復(fù)效果如何并不是很清楚。目的:探討使用海藻酸鈉立體培養(yǎng)的軟骨單位、立體共培養(yǎng)的軟骨單位與軟骨細(xì)胞(1:1)移植對兔膝關(guān)節(jié)軟骨缺損修復(fù)的作用,并對比分析兩者修復(fù)效果。方法:1.選取2月齡新西蘭兔5只,通過酶解法將膝關(guān)節(jié)軟骨消化成軟骨細(xì)胞和軟骨單位,與1.2%海藻酸鈉凝膠混合,分別制備成密度為5.8×106/mL的軟骨細(xì)胞、軟骨單位以及軟骨細(xì)胞和軟骨單位(1:1)混懸液,最后使用模具分別制成內(nèi)含單純軟骨細(xì)胞、單純軟骨單位和軟骨細(xì)胞與軟骨單位(1:1)共培養(yǎng)的三種海藻酸鈉凝膠球(大小約25μL/個(gè));2.選取4月齡新西蘭兔45只(4.0±0.2 Kg),通過取雙側(cè)股骨滑車內(nèi)外側(cè)髁連線中點(diǎn)行直徑為4mm,深度為3mm全層軟骨缺損,隨機(jī)分為3組,將軟骨細(xì)胞、軟骨單位與軟骨單位和軟骨細(xì)胞(1:1)混合體植入缺損處;3.分別于植入后6、12、24周,通過mri評估、大體觀察、he染色、番紅o染色、免疫組化染色、rt-pcr檢測以及tunel原位檢測凋亡狀況,以此來綜合判斷缺損修復(fù)情況。結(jié)果:1.mriroberts評分顯示:自6周至24周,三組評分均呈增高趨勢;12周時(shí)共培養(yǎng)組較軟骨細(xì)胞組和軟骨單位組均明顯增高(p0.05);24周時(shí)共培養(yǎng)組較軟骨單位組和軟骨細(xì)胞組增高顯著(p0.05)。2.ii型膠原免疫組化:自6周到24周,軟骨單位組和共培養(yǎng)組其表達(dá)量呈增多趨勢;在6周和12周軟骨單位組其表達(dá)量均高于共培養(yǎng)組,而24周其共培養(yǎng)組高于軟骨單位組。3.wakitani組織修復(fù)評分顯示:隨著時(shí)間遞增,軟骨單位組及共培養(yǎng)組評分均呈逐漸降低趨勢,且12周與6周相比有統(tǒng)計(jì)學(xué)意義(p0.05);各時(shí)間組共培養(yǎng)組均明顯低于軟骨單位組(p0.05)。4.rt-pcr顯示:6周至24周,共培養(yǎng)組中col-ii,sox-9mrna表達(dá)量均逐漸增多,軟骨單位組col-iimrna表達(dá)量逐漸增多;6周至12周,軟骨單位組和共培養(yǎng)組col-x,mmp-13mrna表達(dá)量呈降低趨勢,24周時(shí)兩者均增高,但共培養(yǎng)組較軟骨單位組增高幅度較小;6周和12周軟骨單位組col-ii及sox-9均高于共培養(yǎng)組,而24周兩指標(biāo)共培養(yǎng)組均高于軟骨單位組;三個(gè)時(shí)間點(diǎn)內(nèi),col-x及mmp-13共培養(yǎng)組均低于軟骨單位組。5.tunel細(xì)胞凋亡檢測顯示:自6周至24周,軟骨細(xì)胞組細(xì)胞凋亡率呈增高趨勢,且24周較12周增高有統(tǒng)計(jì)學(xué)意義(p0.05);軟骨單位組細(xì)胞凋亡率有增高趨勢;自6周至12周共培養(yǎng)組細(xì)胞凋亡率明顯降低(p0.05),而24周較12周共培養(yǎng)組細(xì)胞凋亡率增高顯著(p0.05);在三個(gè)時(shí)間點(diǎn)共培養(yǎng)組細(xì)胞凋亡率均低于軟骨單位組和軟骨細(xì)胞組。結(jié)論:1.軟骨單位與軟骨細(xì)胞(1:1)共培養(yǎng)作為種子細(xì)胞,早期對缺損組織有好的修復(fù)作用,晚期可延緩修復(fù)組織退化。2.軟骨單位具有延緩軟骨細(xì)胞凋亡的作用。
[Abstract]:Background: it is difficult to repair the articular cartilage because of no blood vessel, nerve and lymph, and it is difficult to repair itself once the injury is damaged. It is known that tissue engineering technology promotes the repair of cartilage damage. The study confirms that chondrocytes are easily differentiated in vitro as seed cells in vitro; in vivo, the phenotype and force of the repair of tissue through its development in the body. The characteristics of the study were worse than that of the normal articular cartilage, and with the time of repair in the body, the characteristics of rapid degeneration of the repair tissue appeared and the prospect of its clinical application was restricted. The cartilage unit (chondron) was used as the basic functional structure of articular cartilage, which was mainly composed of chondrocytes and cell Zhou Jizhi (PCM), and PCM was used for chondrocytes. Metabolism and mechanical conduction play an important role. In vitro experiments have proved that chondrocytes and chondrocytes are cultured with 1:1 in sodium alginate gel ball, and their biological characteristics are superior to other proportions; however, how the cartilage defect repair effect of cartilage unit and 1:1 co culture group is not very clear. The effects of cartilage unit, co cultured cartilaginous unit and chondrocyte (1:1) transplantation on the repair of articular cartilage defect of the knee in rabbits were studied and compared and analyzed. Methods: 1. 2 month old New Zealand rabbits were selected to digest the articular cartilage of the knee into chondrocytes and cartilaginous units by enzymolysis and mixed with 1.2% sodium alginate gel. The chondrocytes, cartilaginous cells and chondrocytes and cartilage units (1:1) were not prepared with a density of 5.8 x 106/mL. Finally, three kinds of sodium alginate gel balls (size 25 mu L/) co cultured with pure cartilage unit and chondrocyte and cartilage unit (1:1) were made by using molds, respectively; 2. selected 4 month old new West. 45 rabbits (4 + 0.2 Kg) were randomly divided into 3 groups by taking the middle point of the internal and external condyle of the bilateral femur trochlear and the depth of 3mm full layer cartilage defect into 3 groups. The cartilage cells, cartilage units and chondrocytes (1:1) were implanted into the defect. 3. points were compared to 6,12,24 weeks after the implantation, and were evaluated by MRI, gross observation, he staining, O staining, immunohistochemical staining, RT-PCR detection and TUNEL in situ detection of apoptosis were used to determine the defect repair. Results: the scores of the three groups were increased from 6 to 24 weeks, and at 12 weeks, the co culture group was significantly higher than that of the cartilage and cartilage units (P0.05), and the co culture group at 24 weeks. The cartilage unit group and cartilage cell group increased significantly (P0.05).2.ii collagen immunohistochemical staining: the expression of cartilage unit group and co culture group increased from 6 to 24 weeks, and the expression of cartilage unit group in 6 and 12 weeks was higher than that of the co culture group, and the co culture group was higher than the.3.wakitani tissue repair score of the cartilage unit group at the 24 week. With the increase of time, the score of cartilage unit group and co culture group decreased gradually, and the 12 weeks compared with 6 weeks was statistically significant (P0.05). All time group co culture groups were significantly lower than the cartilage unit group (P0.05).4.rt-pcr display: 6 weeks to 24 weeks, the co culture group of col-ii, sox-9mrna expression increased gradually, cartilage unit group col-iimrna The expression amount increased gradually. The expression of col-x and mmp-13mrna in the cartilage unit group and co culture group decreased in 6 weeks to 12 weeks, and both increased at 24 weeks, but the increase in the co culture group was smaller than that of the cartilage unit group. The col-ii and Sox-9 in the cartilage unit group were higher than that of the co culture group at 6 and 12 weeks, and the two index co culture group was higher than the cartilage unit group at the 6 and 12 weeks. Three time points, col-x and MMP-13 co culture group were lower than the cartilage unit group.5.tunel cell apoptosis detection showed that from 6 weeks to 24 weeks, the apoptosis rate of chondrocyte group increased, and 24 weeks higher than 12 weeks (P0.05), apoptosis rate of cartilage unit group increased; from 6 to 12 weeks, Co culture group cell apoptosis The rate of apoptosis was significantly lower (P0.05), while the apoptosis rate of cells in the 24 week group was significantly higher than that in the 12 week group (P0.05). The apoptosis rate in the co culture group at three time points was lower than that of the cartilage unit group and the chondrocyte group. Conclusion: 1. cartilage unit and chondrocyte (1:1) co culture as seed cells, early repair of the defect tissue and late delay can be extended. .2. cartilage units with delayed repair of tissue degradation can retard chondrocyte apoptosis.

【學(xué)位授予單位】：山西醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2016
【分類號】：R687.4

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 何清義,李起鴻,許建中,楊柳;轉(zhuǎn)化軟骨細(xì)胞與正常軟骨細(xì)胞的生物特性比較[J];中國臨床康復(fù);2002年10期

2 王小虎;衛(wèi)小春;陳維毅;;軟骨細(xì)胞力學(xué)特性的研究進(jìn)展[J];中華醫(yī)學(xué)雜志;2006年21期

3 沈雁,唐毅,李斯明,鐘燦燦,梁佩紅;堿性成纖維細(xì)胞生長因子與透明質(zhì)酸對培養(yǎng)兔軟骨細(xì)胞的作用[J];中華創(chuàng)傷雜志;2000年06期

4 林建華,吳朝陽,許衛(wèi)紅;不同培養(yǎng)時(shí)間軟骨細(xì)胞的生物學(xué)特性[J];福建醫(yī)科大學(xué)學(xué)報(bào);2000年02期

5 張文濤,盧世璧,黃英,李楠;軟骨細(xì)胞形態(tài)對其縫隙連接的影響[J];中國創(chuàng)傷骨科雜志;2000年04期

6 楊物鵬,許建中;軟骨細(xì)胞培養(yǎng)及其調(diào)控[J];中國矯形外科雜志;2000年08期

7 楊彩榮;軟骨細(xì)胞培養(yǎng)及其調(diào)控因素研究進(jìn)展[J];國外醫(yī)學(xué).耳鼻咽喉科學(xué)分冊;2002年03期

8 張文濤,盧世璧,黃英,李楠;軟骨細(xì)胞形態(tài)、表型與胞間通訊研究[J];中國修復(fù)重建外科雜志;2002年05期

9 楊運(yùn)東 ,陳基長;不同施加因素對軟骨細(xì)胞的影響[J];中醫(yī)正骨;2002年09期

10 張艷,柴崗,崔磊,劉偉,曹誼林;不同類型人軟骨細(xì)胞體外生物學(xué)特性比較[J];中華實(shí)驗(yàn)外科雜志;2003年06期

相關(guān)會議論文 前10條

1 張春雷;孫美樂;姚洪菊;;抗氧化劑和透明質(zhì)酸對軟骨細(xì)胞的保護(hù)作用[A];中國細(xì)胞生物學(xué)學(xué)會第五次會議論文摘要匯編[C];1992年

2 任宏造;;軟骨細(xì)胞的超生結(jié)構(gòu)病理[A];第六次全國電子顯微學(xué)會議論文摘要集[C];1990年

3 張楊;崔麗;郭悅;孫曉雷;李秀蘭;;滑膜細(xì)胞微環(huán)境對軟骨細(xì)胞生物學(xué)活性的影響[A];第十八屆全國中西醫(yī)結(jié)合骨傷科學(xué)術(shù)研討會論文匯編[C];2011年

4 柏濤;;骨髓間充質(zhì)干細(xì)胞誘導(dǎo)表達(dá)軟骨細(xì)胞表型的研究進(jìn)展[A];玉溪市第三屆二次骨科學(xué)術(shù)研討會論文匯編[C];2009年

5 周紅輝;萬福生;;人骨髓間充質(zhì)干細(xì)胞分化為軟骨細(xì)胞的實(shí)驗(yàn)研究[A];華東六省一市生物化學(xué)與分子生物學(xué)學(xué)會2006年學(xué)術(shù)交流會論文集[C];2006年

6 王正輝;吳寶俊;許珉;;殼聚糖/明膠復(fù)合不同軟骨細(xì)胞體外構(gòu)建組織工程軟骨的實(shí)驗(yàn)研究[A];全國耳鼻咽喉頭頸外科中青年學(xué)術(shù)會議論文匯編[C];2012年

7 馬劍雄;馬信龍;張華鋒;張園;王志鋼;楊陽;;生物力學(xué)因素在激素性股骨頭壞死中對軟骨細(xì)胞的作用[A];2009第十七屆全國中西醫(yī)結(jié)合骨傷科學(xué)術(shù)研討會論文匯編[C];2009年

8 李建華;黃建榮;康奕飛;許繼德;涂永生;;體外誘導(dǎo)人骨髓間充質(zhì)干細(xì)胞分化為軟骨細(xì)胞的研究[A];中南地區(qū)第六屆生理學(xué)學(xué)術(shù)會議論文摘要匯編[C];2004年

9 石印玉;曹月龍;馮偉;鄭昱新;石瑛;王翔;;補(bǔ)腎、柔肝中藥對軟骨細(xì)胞生物功能的影響[A];2004'中華中醫(yī)藥科技成果專輯[C];2004年

10 許道榮;金丹;肖庭輝;余斌;;長時(shí)間拉伸應(yīng)變對軟骨細(xì)胞生化環(huán)境的影響[A];第20屆中國康協(xié)肢殘康復(fù)學(xué)術(shù)年會論文選集[C];2011年

相關(guān)重要報(bào)紙文章 前10條

1 聶翠蓉;英科學(xué)家從成人骨骼中找到軟骨干細(xì)胞[N];科技日報(bào);2008年

2 吳一福;西安交大：用人胎兒軟骨細(xì)胞培養(yǎng)成功軟骨組織工程種子細(xì)胞[N];中國醫(yī)藥報(bào);2006年

3 中文;人體軟骨舉足輕重[N];廣東科技報(bào);2000年

4 劉霞;英用患者組織細(xì)胞成功培育出再生軟骨[N];科技日報(bào);2010年

5 保健時(shí)報(bào)特約記者 方序;體外“養(yǎng)”一塊軟骨補(bǔ)膝蓋[N];保健時(shí)報(bào);2011年

6 記者 張可喜;培養(yǎng)軟骨細(xì)胞[N];新華每日電訊;2002年

7 健康時(shí)報(bào)記者 李海清;軟骨破了打“補(bǔ)丁”[N];健康時(shí)報(bào);2009年

8 楊春;藥物增高不可信[N];大眾衛(wèi)生報(bào);2005年

9 黃楓 謝國平;中醫(yī)藥治療膝骨關(guān)節(jié)炎實(shí)驗(yàn)研究進(jìn)展[N];中國中醫(yī)藥報(bào);2006年

10 記者 何屹;英國用干細(xì)胞成功培育出軟骨組織[N];科技日報(bào);2005年

相關(guān)博士學(xué)位論文 前10條

1 王曉鳳;自噬在軟骨發(fā)育不全與軟骨生成中的作用與機(jī)制研究[D];第三軍醫(yī)大學(xué);2015年

2 王勝楠;杜仲苷對IL-1β誘導(dǎo)的軟骨細(xì)胞分解代謝和凋亡的影響及其作用機(jī)制[D];南方醫(yī)科大學(xué);2015年

3 張?jiān)?MiR-145在原發(fā)性膝關(guān)節(jié)骨性關(guān)節(jié)炎軟骨中的表達(dá)及其意義[D];天津醫(yī)科大學(xué);2015年

4 張龍強(qiáng);整合素β1促進(jìn)GIT1表達(dá)而影響軟骨細(xì)胞增殖凋亡的研究[D];山東大學(xué);2015年

5 馬中希;周期性張應(yīng)力對大鼠生長板軟骨細(xì)胞的增殖和凋亡作用[D];華中科技大學(xué);2015年

6 劉艷;BMP-2在骨性關(guān)節(jié)炎中表達(dá)的意義及其誘導(dǎo)凋亡及增殖的研究[D];吉林大學(xué);2016年

7 梁錦前;瘦素對于人OA軟骨細(xì)胞RhoA/LIMK1/Cofilin細(xì)胞骨架調(diào)控通路的作用[D];中國協(xié)和醫(yī)科大學(xué);2010年

8 楊運(yùn)東;骨炎定對體外培養(yǎng)軟骨細(xì)胞的影響[D];廣州中醫(yī)藥大學(xué);2001年

9 張e，

本文編號：1834907