豚鼠肺脏上皮细胞形成及Toll样受体（TLR）信号因子的发育表达
发布时间：2018-10-10 16:09
【摘要】：豚鼠是研究人哮喘病、结核病等呼吸道疾病的理想动物模型之一。同时,豚鼠也是研究肺脏的先天免疫机制和呼吸道疾病的致病机理的常用模型。因此,本研究对豚鼠肺脏发育过程中上皮细胞的发育及其Toll样受体(TLRs)信号表达进行了初步的研究。 试验中,我们利用组织化学染色(HE)和免疫组织化学(IHC)染色方法对不同发育时期豚鼠呼吸道主要上皮细胞的类型与组成进行了形态学观察,并对TLRs信号分子的发育表达利用定量反转录PCR(qRT-PCR)和免疫印迹(WB)法进行了分析。结果显示,在胚胎(Embryonic)发育到45d和52d时,豚鼠肺脏分别处于假腺期和小管期；在胚胎发育到59d至出生前,肺脏呈现为肺泡增多的囊状期；在出生后7d至成年期(3个月),豚鼠肺脏发育处于肺泡期。IHC结果表明,在胚胎发育的第45d,肺脏存在具有多能性的基底细胞和肺泡Ⅱ型细胞,当胚胎发育到52d时,即肺脏发育的小管期,可以观察到纤毛细胞和Clara细胞。 荧光定量PCR分析结果表明,在出生后3个月(成年)、28d、14d、7d利0d,以及胚胎期E45d、E52d和E59d的不同发育阶段,豚鼠肺脏中TLR1、TLR2、TLR3、TLR4、TLR6、TLR7、 TLR8、TLR9、TLR10、MyD88和TRAF6,这11种TLR相关信号均有表达,但在出生前,各因子的表达量相对高于出生后。其中,在胚胎期E45d、E52d和E59d的豚鼠肺脏组织中,TLR1-10总体的表达水平为显著递增趋势,E52d时,TLR4表达量最高,E45d表达量次之；在E52d和E59d时,TRAF6和MyD88相对表达量最高；而在出生后到成年(3个月)期,各因子的表达量显著降低,且出生后的不同时期,同一因子间的表达差异不显著。同时,利用蛋白免疫印迹Western Blotting技术在蛋白质水平进一步对上述因子的表达进行了验证。上述结果表明,豚鼠肺脏在发育过程中,随着上皮细胞的增殖和分化,肺脏在形态学和功能上趋于成熟。TLRs在出生前的增高现象在分子水平提示豚鼠肺脏出生前的形态功能成熟性,并具备比较完善的免疫反应能力；而出生后这些因子的表达显著降低,将有利于避免机体诱导过度的免疫反应对组织器官造成免疫损伤,并处于随时识别免疫刺激源以激活先天免疫和获得性免疫反应的状态。这些结果都为进一步探讨豚鼠肺脏上皮细胞发育生物学的特征及其先天免疫机制的形成过程等提供了理论依据。
[Abstract]:Guinea pig is one of the ideal animal models to study respiratory diseases such as asthma, tuberculosis and so on. At the same time, guinea pig is also a common model to study the innate immune mechanism of lung and the pathogenic mechanism of respiratory diseases. Therefore, the development of epithelial cells and the expression of (TLRs) signal of Toll like receptor in guinea pig lung were studied in this study. In the experiment, the types and compositions of the main epithelial cells in the respiratory tract of guinea pigs at different developmental stages were observed by histochemical staining (HE) and immunohistochemical (IHC) staining. The developmental expression of TLRs signaling molecules was analyzed by quantitative reverse transcription PCR (qRT-PCR) and Western blot (WB). The results showed that the lungs of guinea pigs were in pseudoglandular phase and tubule stage at 45 days and 52 days of embryonic (Embryonic) development, and the lungs of guinea pigs showed a cystic phase of alveolar proliferation during the development of embryo from day 59 to pre-birth. From 7 days after birth to 3 months of adulthood, the lung development of guinea pigs was in alveolar phase. IHC results showed that there were pluripotent basal cells and alveolar type 鈪，
本文编号：2262415
[Abstract]:Guinea pig is one of the ideal animal models to study respiratory diseases such as asthma, tuberculosis and so on. At the same time, guinea pig is also a common model to study the innate immune mechanism of lung and the pathogenic mechanism of respiratory diseases. Therefore, the development of epithelial cells and the expression of (TLRs) signal of Toll like receptor in guinea pig lung were studied in this study. In the experiment, the types and compositions of the main epithelial cells in the respiratory tract of guinea pigs at different developmental stages were observed by histochemical staining (HE) and immunohistochemical (IHC) staining. The developmental expression of TLRs signaling molecules was analyzed by quantitative reverse transcription PCR (qRT-PCR) and Western blot (WB). The results showed that the lungs of guinea pigs were in pseudoglandular phase and tubule stage at 45 days and 52 days of embryonic (Embryonic) development, and the lungs of guinea pigs showed a cystic phase of alveolar proliferation during the development of embryo from day 59 to pre-birth. From 7 days after birth to 3 months of adulthood, the lung development of guinea pigs was in alveolar phase. IHC results showed that there were pluripotent basal cells and alveolar type 鈪，
本文编号：2262415
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