南方医科大学学报

南方医科大学学报 ›› 2020, Vol. 40 ›› Issue (12): 1703-1711.doi: 10.12122/j.issn.1673-4254.2020.12.02

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阳和平喘颗粒通过上调miR-139-5p和下调Notch1/Hes1通路促进哮喘大鼠BMSCs归巢

朱慧志，王 坤，杨 磊，徐晴雯，任冯春，刘向国   

  • 出版日期:2020-12-20 发布日期:2020-12-20

Yanghe Pingchuan granule promotes BMSCs homing in asthmatic rats by upregulating miR-139-5p and downregulating Notch1/Hes1 pathway

  • Online:2020-12-20 Published:2020-12-20

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摘要： 目的 观察阳和平喘颗粒对哮喘miR-139-5p、Notch1/Hes1信号通路及骨髓源性间充质干细胞（BMSCs）归巢的影响。方法 50 只 SD 大鼠随机均分为 5 组：正常对照（NC）组、模型对照（MC）组、BMSCs 移植（BMSCs）组、BMSCs+地塞米松（BMSCs+DXM）组[地塞米松 0.0625 mg/ （kg·d）]、BMSCs+阳和平喘组[阳和平喘颗粒3.5 g/ （kg·d）]。采用卵清蛋白致敏激发建立大鼠哮喘模型，BMSCs组、BMSCs+DXM组、BMSCs+阳和平喘组大鼠在激发最后1 d经尾静脉移植入1×106/mL BMSCs悬液。NC组、MC组以生理盐水灌胃，药物干预组分别予相应药物灌胃，灌胃量为1 mL/100 g，均持续1周。HE染色观察肺组织病理形态学，流式细胞术检测肺组织中CXCR4蛋白表达，酶联免疫吸附法检测肺组织γ干扰素（IFN-γ）、白介素-4（IL-4）表达。免疫荧光观察支气管上皮细胞CXCR4、Notch1、Hes1表达。RT-PCR法检测肺组织miR-139-5p、Notch1、Jagged1、RBP-J、Hes1基因表达，免疫印迹法检测肺组织 Notch1、Jagged1、Hes1 蛋白表达。结果 （1）与 NC 组比较，MC 组肺组织 CXCR4、IL-4、Notch1mRNA、Jagged1mRNA、RBP-JmRNA、Hes1mRNA及Notch1、Jagged1、Hes1蛋白表达升高，INF-γ、miR-139-5p mRNA表达降低（P<0.05）；（2）与MC组比较，BMSCs组、BMSCs+DXM组、BMSCs+阳和平喘组CXCR4、IFN-γ、miR-139-5pmRNA升高，IL-4、Notch1mRNA、Jagged1mRNA、RBP-JmRNA、Hes1mRNA及Notch1、Jagged1、Hes1蛋白表达降低（P<0.05）；（3）与BMSCs组比较，BMSCs+阳和平喘组CXCR4、IFN-γ、miR-139-5pmRNA表达升高，IL-4、Notch1mRNA表达量降低（P<0.05）。结论 阳和平喘颗粒通过上调miR-139-5p，下调Notch1/Hes1通路，强化BMSCs归巢对Th2炎症反应的抑制作用。

关键词: 哮喘；间充质干细胞；归巢；阳和平喘颗粒；Notch1/Hes1信号通路

Abstract: Objective To observe the effect of Yanghe Pingchuan (YHPC) granule on miR-139-5p, Notch1/Hes1 pathway and homing of bone marrow-derived mesenchymal stem cells (BMSCs) in asthmatic rats. Methods Fifty SD rats were randomized divided into normal control (NC) group, asthmatic model group, BMSCs transplantation group, BMSCs + dexamethasone (0.0625 mg/kg daily) group, and BMSCs+YHPC granule (3.5 g/kg daily) group. In all but the normal control group, asthmatic rat models were established by ovalbumin challenge, and BMSCs (1×106/mL) transplantation via the tail vein was performed in the latter 3 groups on last day of ovalbumin challenge. In all the groups, lung pathologies of the rats were evaluated using HE staining after the treatments. Flow cytometry was employed to detect pulmonary expression of CXCR4 protein, and ELISA was used to determine the expressions of interferon-γ (IFN-γ) and interleukin-4 (IL-4) in the lung tissue. The expressions of CXCR4, Notch1 and Hes1 in bronchial epithelial cells was examined using immunofluorescence assay. RT-PCR was used to detect the expressions of miR-139-5p, Notch1, Jagged1, RBP-J and Hes1 mRNAs, and the protein expressions of Notch1, Jagged1 and Hes1 were detected with Western blotting. Results Compared with the normal control rats, the asthmatic rats exhibited significantly increased expressions of CXCR4, IL-4, Notch1, Jagged1, RBP-J, and Hes1 mRNA and Notch1, Jagged1, and Hes1 proteins and lowered expressions of INF-γ mRNA and miR-139-5p in the lung tissues (P<0.05 or 0.01). Compared with those in the asthmatic model group, the mRNA expressions of CXCR4, IFN-γ, and miR-139-5p increased and the expressions of IL-4, Notch1, Jagged1, RBP-J, and Hes1 mRNA and Notch1, Jagged1, and Hes1 proteins decreased significantly in the 3 groups with BMSCs transplantation (P<0.05 or 0.01). The rats in BMSCs+YHPC granule group showed significantly higher CXCR4, IFN-γ, and miR-139-5p mRNA expressions and lower IL-4 and Notch1 mRNA expressions than those in
BMSCs transplantation group (P<0.05). Conclusion YHPC granule can enhance the inhibitory effect of BMSCs homing on Th2 inflammatory response in asthmatic rats by up-regulating miR-139-5p and down-regulating Notch1/Hes1 pathway.

Key words: asthma; bone marrow-derived mesenchymal stem cells; homing; Yanghe Pingchuan granule; Notch1/Hes1 signaling pathway