轉(zhuǎn)染CDMP-2基因的成肌細(xì)胞與PLGA材料的相容性
發(fā)布時(shí)間：2018-10-10 14:35
【摘要】：目的 探討轉(zhuǎn)染CDMP-2(軟骨源性形態(tài)發(fā)生蛋白2)基因后的成肌細(xì)胞與PLGA（聚乳酸/乙醇酸共聚物）材料的相容性。 方法 取新生5d的SD大鼠骨骼肌，采用混合酶一步消化分離獲取成肌細(xì)胞，將差速貼壁法和胰酶消化法相結(jié)合進(jìn)行純化，觀察形態(tài)學(xué)特點(diǎn)，生長(zhǎng)狀況,并檢測(cè)desmin表達(dá)。采用脂質(zhì)體轉(zhuǎn)染方法將CDMP2基因轉(zhuǎn)染P3代成肌細(xì)胞，，將成肌細(xì)胞和轉(zhuǎn)染CDMP-2基因的成肌細(xì)胞分別接種于PLGA支架上。分4組：A成肌細(xì)胞;B轉(zhuǎn)染CDMP-2基因的成肌細(xì)胞;C成肌細(xì)胞+PLGA支架;D轉(zhuǎn)染CDMP-2基因的成肌細(xì)胞+PLGA支架。采用MTT法檢測(cè)各組成肌細(xì)胞的生物活性，繪制生長(zhǎng)曲線，在掃描電鏡下觀察細(xì)胞形態(tài)。 結(jié)果 細(xì)胞接種于PLGA支架后早期，倒置相差顯微鏡視野內(nèi)見不到細(xì)胞在支架表面生長(zhǎng)、附著，只能看到縱橫交錯(cuò)的纖維網(wǎng)狀結(jié)構(gòu)。72小時(shí)后可見細(xì)胞增殖，細(xì)胞數(shù)量顯著增多。MTT示各組成肌細(xì)胞在492nm波長(zhǎng)時(shí)分別為0.046414,0.057665,0.038945,0.049911；在629nm波長(zhǎng)時(shí)分別為0.132158，0.178811，0.108284，0.173810，ABCD各組在統(tǒng)計(jì)學(xué)上都無(wú)統(tǒng)計(jì)學(xué)意義（P0.05）。 結(jié)論 成肌細(xì)胞能夠在PLGA支架上生長(zhǎng)良好，說(shuō)明PLGA材料對(duì)成肌細(xì)胞無(wú)細(xì)胞毒性，具有優(yōu)異的生物相容性，可用于組織修復(fù)，為進(jìn)一步研究軟骨損傷的修復(fù)提供了新的思路和方法。
[Abstract]:Objective to investigate the compatibility of CDMP-2 (chondrogenic morphogenetic protein-2) gene transfected myoblasts with PLGA (poly (lactic acid / glycolic acid) material. Methods the skeletal muscle of 5 days old SD rats was isolated and isolated by one step digestion of mixed enzymes. The differential adherent method and trypsin digestion method were combined to purify the myoblasts. The morphological characteristics, growth status and expression of desmin were observed. The CDMP2 gene was transfected into P3 generation myoblast by liposome transfection. The myoblast and the CDMP-2 gene transfected myoblast were inoculated on the PLGA scaffold respectively. Four groups were divided into four groups: myoblast A, myoblast transfected with CDMP-2 gene B, PLGA scaffold with C myoblast and PLGA scaffold transfected with CDMP-2 gene. The biological activity of muscle cells was measured by MTT method, and the growth curve was drawn. The morphology of the cells was observed by scanning electron microscope (SEM). Results at the early stage after inoculation with PLGA scaffold, the cells could not grow and attach on the scaffold surface in inverted phase contrast microscope, only the crisscross fibrous reticular structure could be seen. After 72 hours, cell proliferation could be seen. MTT showed that the number of muscle cells was 0.0464140.0576650.038945and 0.1321580.1788110.1082840.1738101010 at the wavelength of 492nm, respectively, and there was no statistical significance in each group (P0.05). Conclusion myoblast can grow well on PLGA scaffold, which indicates that PLGA has no cytotoxicity to myoblasts and has excellent biocompatibility, and can be used for tissue repair. It provides a new idea and method for the further study of cartilage repair.
【學(xué)位授予單位】：遼寧醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R318.08;R687
[Abstract]:Objective to investigate the compatibility of CDMP-2 (chondrogenic morphogenetic protein-2) gene transfected myoblasts with PLGA (poly (lactic acid / glycolic acid) material. Methods the skeletal muscle of 5 days old SD rats was isolated and isolated by one step digestion of mixed enzymes. The differential adherent method and trypsin digestion method were combined to purify the myoblasts. The morphological characteristics, growth status and expression of desmin were observed. The CDMP2 gene was transfected into P3 generation myoblast by liposome transfection. The myoblast and the CDMP-2 gene transfected myoblast were inoculated on the PLGA scaffold respectively. Four groups were divided into four groups: myoblast A, myoblast transfected with CDMP-2 gene B, PLGA scaffold with C myoblast and PLGA scaffold transfected with CDMP-2 gene. The biological activity of muscle cells was measured by MTT method, and the growth curve was drawn. The morphology of the cells was observed by scanning electron microscope (SEM). Results at the early stage after inoculation with PLGA scaffold, the cells could not grow and attach on the scaffold surface in inverted phase contrast microscope, only the crisscross fibrous reticular structure could be seen. After 72 hours, cell proliferation could be seen. MTT showed that the number of muscle cells was 0.0464140.0576650.038945and 0.1321580.1788110.1082840.1738101010 at the wavelength of 492nm, respectively, and there was no statistical significance in each group (P0.05). Conclusion myoblast can grow well on PLGA scaffold, which indicates that PLGA has no cytotoxicity to myoblasts and has excellent biocompatibility, and can be used for tissue repair. It provides a new idea and method for the further study of cartilage repair.
【學(xué)位授予單位】：遼寧醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R318.08;R687
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