【摘要】：目的：研究移植外源性骨髓間充質(zhì)干細(xì)胞(BMSCs)對博萊霉素致小鼠晚期肺纖維化即肺纖維化形成期的干預(yù)作用。 方法：全骨髓貼壁細(xì)胞分離培養(yǎng)法分離、培養(yǎng)雄性6-8周齡ICR小鼠BMSCs。本實(shí)驗(yàn)中設(shè)計4個分組，共160只實(shí)驗(yàn)小鼠，將這些小鼠隨機(jī)分為：陰性對照組（NC組），陽性對照組（PC組），BMSCs干預(yù)組A（A組），BMSCs干預(yù)組B（B組）。NC組于經(jīng)鼻腔滴入50ulPBS作為陰性對照，PC組、A組、B組則按6ug/g的劑量經(jīng)鼻腔滴入50ul博萊霉素制成小鼠的肺纖維化模型，A組在造模后第14天經(jīng)尾靜脈注射外源性的BMSCs1×107/ml，200ul；B組在造模后第14天起隔日經(jīng)尾靜脈注射外源性的BMSCs1×107/ml，200ul。以上各組小鼠造模后28天統(tǒng)一稱重后處死，取小鼠全肺組織稱重并留取部分肺組織行HE、MASSON染色及羥脯氨酸含量測定，用熒光定量PCR及免疫組化檢測肺組織中SP-C和AQP5表達(dá)情況；提取肺組織的DNA檢測雄鼠的性別決定基因(Sry基因)。 結(jié)果：1、肺系數(shù)： PC組及A、B組肺系數(shù)均較NC組高，PC組、A組、B組3組間無明顯差異。2、肺組織病理：PC組及A組、B組肺泡炎、肺纖維化程度均較NC組明顯加重；A組肺泡炎及纖維化較PC、B組稍減輕，但差異無統(tǒng)計學(xué)意義。3、HYP含量：PC組及A組、B組3組HYP含量均較NC組明顯升高，A組較PC組、B組有所減低，與B組間的差異較為明顯，有統(tǒng)計學(xué)意義，PC組、B組間無明顯差異。4、Sry基因檢測： A組、B組均有雄性小鼠的sry基因存在。5、肺組織SP-C蛋白的表達(dá)：PC組、A組、B組中SP-C蛋白的表達(dá)均低于NC組，A組的表達(dá)高于PC組、B組，而PC組與B組間無明顯差異。6、肺組織AQP-5蛋白的表達(dá)：PC組、A組、B組的SP-C mRNA表達(dá)均低于NC組，PC組、A組、B組3組間無明顯差異。7、肺組織SP-C mRNA表達(dá)情況：PC組、A組、B組3組中SP-C mRNA表達(dá)均低于NC組，，A組的表達(dá)高于PC組、B組，差異有統(tǒng)計學(xué)意義，而PC組與B組間無明顯差異。8、肺組織AQP5mRNA表達(dá)情況：PC組、A組、B組的AQP5mRNA表達(dá)均低于NC組，3組間無明顯差異。 結(jié)論：1、外源性BMSCs經(jīng)尾靜脈注射后可以到達(dá)博來霉素誘導(dǎo)的肺纖維化形成期小鼠的肺組織并可成功實(shí)現(xiàn)在局部定植；2、外源性BMSCs在肺纖維化形成期單次給藥較單純造模組的肺纖維化程度有減輕的趨勢，但差異無明顯統(tǒng)計學(xué)意義，因此目前尚不能說明在肺纖維化形成期給予外源性BMSCs干預(yù)能治療肺纖維化；3、在肺纖維化形成期單次和多次給予外源性BMSCs的干預(yù)效果無統(tǒng)計學(xué)差異，甚至多次給藥有加重肺纖維化的趨勢。
[Abstract]:Aim: to investigate the effects of transplantation of exogenous bone marrow mesenchymal stem cells (BMSCs) on bleomycin-induced late pulmonary fibrosis in mice. Methods: the whole bone marrow adherent cells were isolated and cultured in male 6-8 week old ICR mice BMSCs. In this experiment, four groups were designed for 160 experimental mice. These mice were randomly divided into two groups: negative control group (NC group), positive control group (PC group) BMSCs intervention group (A (A group). The pulmonary fibrosis model of mice was made by bleomycin. Group A was injected with exogenous BMSCs 1 脳 107 / ml / ml 200ul1 脳 10 ~ (7) / ml on the 14th day after the establishment of the model. In group B, exogenous BMSCs 1 脳 107 / ml / ml was injected via caudal vein on the 14th day after modeling. All the above groups were killed after 28 days of unified weighing. The whole lung tissue of the mice was weighed and some of the lung tissues were taken for determination of Hedman son staining and hydroxyproline content. The expression of SP-C and AQP5 in lung tissue was detected by fluorescence quantitative PCR and immunohistochemistry. The male sex determinant gene (Sry gene) was detected by extracting DNA from lung tissue. Results compared with NC group, there was no significant difference in lung coefficient between PC group and Agna B group. There was no significant difference between group A and group B. The lung pathological changes of PC group and group A group were significantly worse than those of NC group. The pulmonary fibrosis degree in group A and group A was significantly higher than that in group C. The pulmonary alveolitis and fibrosis in group A were slightly less than those in group PCB, but there was no significant difference in HYP content between group A and group B, but the content of HYP in group A was significantly higher than that in group C, and the difference between group A and group B was more obvious than that in group B, but the content of HYP in group A was significantly higher than that in group C. There was no significant difference in the expression of SP-C gene between group B and group A: the expression of SP-C protein in lung tissue of group A was lower than that of group A (group A). The expression of SP-C protein in group B was lower than that in group A (group A), and the expression of SP-C protein in group B was lower than that in group A (group A). The expression of SP-C protein in lung tissue in group A was lower than that in group B. The expression of the protein in PC group was higher than that in PC group B group. However, there was no significant difference between PC group and B group. The expression of AQP-5 protein in lung tissue of PC group was significantly lower than that of group A and B group (P < 0.05). The expression of SP-C mRNA in lung tissue of PC group was significantly lower than that of group A and B group (P < 0.05). The expression of SP-C mRNA in lung tissue of PC group was significantly lower than that of group A and B group (P < 0.05). The expression of SP-C mRNA in three groups was lower than that in group A of NC group, and the expression of SP-C mRNA in group A was higher than that in group B of PC group. There was no significant difference in AQP5 mRNA expression between PC group and B group. The expression of AQP5 mRNA in lung tissue of group A was significantly lower than that of group B (P < 0.05). There was no significant difference in AQP5 mRNA expression between group B and group C (P < 0.05). ConclusionThe exogenous BMSCs can reach the lung tissue of bleomycin induced pulmonary fibrosis mice after injection through tail vein and can be successfully colonized locally. 2. The degree of pulmonary fibrosis of exogenous BMSCs at the stage of pulmonary fibrosis formation was alleviated, but the difference was not statistically significant. Therefore, at present, it is not clear that exogenous BMSCs can be used to treat pulmonary fibrosis in the stage of pulmonary fibrosis formation, but there is no statistical difference between single and multiple intervention effects of exogenous BMSCs at the stage of pulmonary fibrosis formation. Even multiple doses of drugs have a tendency to exacerbate pulmonary fibrosis.
【學(xué)位授予單位】：南昌大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R563.9

【參考文獻(xiàn)】

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1 陳寅;馬南;梅舉;肖海波;陸善偉;徐懷陽;鐘z

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