本文選題：骨結(jié)核 + 脂肪干細(xì)胞��； 參考：《新疆醫(yī)科大學(xué)》2014年碩士論文

【摘要】：目的：基于兔脂肪干細(xì)胞(rabbit adipose-derived stem cells, rADSCs)、利福噴丁聚乳酸緩釋微球、同種異體部分脫鈣骨支架,初步于體外構(gòu)建抗結(jié)核性骨組織工程復(fù)合體。方法：分離、培養(yǎng)rADSCs,經(jīng)檢測(cè)細(xì)胞周期、表面抗原CD44及分化誘導(dǎo)鑒定干細(xì)胞特性。用利福噴丁生長(zhǎng)液干預(yù)rADSCs,按濃度分為0μg/ml、20μg/ml、40μg/ml及60μg/ml四組,分別繪制細(xì)胞生長(zhǎng)曲線、檢測(cè)細(xì)胞凋亡及堿性磷酸酶(alkaline phosphatase, ALP)含量。制備利福噴丁微球,計(jì)算平均直徑、載藥率及包封率。構(gòu)建rADSCs-利福噴丁微球二維復(fù)合物并成骨誘導(dǎo)。制備骨支架并構(gòu)建rADSCs-利福噴丁微球-骨支架三維復(fù)合物,計(jì)算細(xì)胞粘附率；掃描電子顯微鏡及蘇木精-伊紅(HE)染色觀察復(fù)合情況；每3d更換生長(zhǎng)液,檢測(cè)體外釋藥特性。結(jié)果：rADSCs呈長(zhǎng)梭形生長(zhǎng),G0/G1期占(80.3+3.2)%,表面抗原CD44陽(yáng)性表達(dá)。成骨誘導(dǎo)后ALP、茜素紅染色,成脂誘導(dǎo)后油紅O染色及成軟骨誘導(dǎo)后阿利新藍(lán)染色均為陽(yáng)性。生長(zhǎng)曲線示60μg/ml組第4-12d增值能力較其他三組弱(P0.01)；凋亡檢測(cè)示60μg/ml組凋亡率高于其他三組(P0.01)；ALP檢測(cè)示第7、14d60μg/ml組ALP含量較其他三組少(P0.05)；臨界濃度為40μg/ml。微球平均直徑為(26.4+3.6)μm,載藥率為(40.89±0.63)%,包封率為(75.24±0.18)%。二維復(fù)合物成骨誘導(dǎo)后茜素紅染色陽(yáng)性。三維復(fù)合物細(xì)胞粘附率為(96.94±1.24)%；掃描電鏡及HE染色觀察復(fù)合體相容性良好；體外培養(yǎng)第3、6、39d累積釋藥量分別達(dá)總量的8.54%、20.12%及93.66%,每個(gè)時(shí)間點(diǎn)檢測(cè)的釋藥濃度均低于臨界濃度,但高于最低抑菌濃度。結(jié)論：體外構(gòu)建的抗結(jié)核性骨組織工程復(fù)合體具有持續(xù)緩慢釋放藥物的性質(zhì),且該釋藥濃度不影響rADSCs的增值及成骨活性。
[Abstract]:Aim: to construct anti-tuberculous bone tissue engineering complex in vitro based on rabbit adipose stem cells adipose-derived stem cells, rifampicin polylactic acid sustained-release microspheres and allogeneic partial decalcification bone scaffold. Methods: rADSCs were isolated and cultured. Stem cells were identified by cell cycle detection, surface antigen CD44 and differentiation induction. RADSCswere treated with rifapentine growth liquid. The cells were divided into four groups: 0 渭 g / ml, 20 渭 g / ml, 40 渭 g/ml and 60 渭 g/ml. The cell growth curves were plotted, and the cell apoptosis and alkaline phosphatase (ALPaline) content were measured. Rifapentine microspheres were prepared and the mean diameter, drug loading rate and encapsulation efficiency were calculated. Two-dimensional complex of rADSCs- rifapentine microspheres was constructed and induced by osteogenesis. Bone scaffolds were prepared and three dimensional complexes of rADSCsrifopentine microspheres and bone scaffolds were constructed to calculate the cell adhesion rate, scanning electron microscopy and hematoxylin eosin HEhe staining were used to observe the composite, and the growth fluid was changed every 3 days to detect the drug release characteristics in vitro. Results the percentage of CD44 in G _ 0 / G _ 1 phase of G _ 0 / G _ 1 was 80.33.2%, and the expression of surface antigen CD44 was positive. ALP, alizarin red staining, oil red O staining and cartilage induction were all positive after osteogenic induction. The growth curve showed that the increment ability of 60 渭 g/ml group was weaker than that of the other three groups on 4-12 d, and the apoptotic rate of 60 渭 g/ml group was higher than that of the other three groups. The ALP content of 60 渭 g/ml group was lower than that of the other three groups on the 7th day, 14 d, 60 渭 g/ml group, and the critical concentration was 40 渭 g / ml. The average diameter of the microspheres was 26.4.3.6) 渭 m, the drug loading rate was 40.89 鹵0.63 and the encapsulation efficiency was 75.24 鹵0.18. Alizarin red staining was positive after osteogenic induction of two-dimensional complex. The cell adhesion rate of the three dimensional complex was 96.94 鹵1.24%, the compatibility of the complex was well observed by SEM and HE staining, the cumulative release of the complex reached 8.54% and 93.66% of the total drug release at the 639d of culture in vitro, and the drug release concentration detected at each time point was lower than the critical concentration. But above the minimum inhibitory concentration. Conclusion: the anti-tuberculous bone tissue engineering complex constructed in vitro has the properties of sustained slow release of the drug, and the release concentration of the drug does not affect the proliferation and osteogenic activity of rADSCs.
【學(xué)位授予單位】：新疆醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R529.2

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本文編號(hào)：1966781