花背蟾蜍胎肺发育中相关活性物质的表达及其调控意义
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摘要
本文以花背蟾蜍蝌蚪为对象,利用生物显微技术研究了花背蟾蜍蝌蚪肺的发生;采用免疫组织化学方法研究了表皮生长因子受体(EGFR)、转化生长因子β(TGF-β_1,TGF-β_2,TGF-β_3)在花背蟾蜍蝌蚪肺发育过程中的表达情况,利用美国Image-pro plus5.0专业图像分析软件对表皮生长因子受体(EGFR)、转化生长因子β(TGF-β_1,TGF-β_2,TGF-β_3)的表达强度进行定量分析。
1.花背蟾蜍的肺开始发生在幼虫时期的第21期,它起始于咽部后侧,向腹侧突起形成肺芽,肺芽经22、23、24期的缓慢发育,直到25期才开始分化出现微小的肺囊,但是里面充满液体,到30期开始形成明显的肺囊,且肺囊中已有气体,33期开始出现肺泡,36期时肺壁已充分伸展,肺泡增多,肺壁变薄,典型的气-血屏障开始形成,37期肺壁内表面的网状隔膜明显,到38期时肺壁进一步变薄,39期肺已接近幼蟾肺,成年花背蟾蜍的肺成囊状,并且在肺的内壁上有许多网状隔膜,将肺的内壁分隔形成许多小室,在每个小室内又有少数次级皱褶将小室分隔为数个近似半球形或“C”形小腔,肺内表面覆盖有一层肺上皮细胞。
2.花背蟾蜍胎肺发育的早期,EGFR主要表达于肺网状隔膜的上皮细胞,且较集中于上皮细胞的游离表面;37-39期,EGFR在肺泡囊及肺泡上皮细胞的阳性表达最活跃,EGFR在间充质细胞区域性分布更加明显;幼蟾期,EGFR阳性反应主要定位在肺泡上皮细胞,在肺网状隔膜处几乎不表达。
EGFR表达变化的定量分析显示在花背蟾蜍胎肺发育的全过程中,EGFR的表达水平呈逐渐上升状态,在30-35期变化比较平稳,至37期达到峰值,继而逐渐下降,39期和幼蟾期的表达水平又有所回升。
3.TGF-β_1,TGF-β_2,TGF-β_3三者均定位于细胞浆,阳性表达区域基本一致。30-36期,TGF-β在肺网状隔膜处和间充质细胞均呈现弱阳性,并且在网状隔膜处呈区域性分布特点;37-39期,TGF-β在肺网状隔膜处表达变弱,在平滑肌和肺间质细胞TGF-β呈强阳性表达;幼蟾期,血管周围的平滑肌和肺间质细胞处呈阳性表达。
TGF-β表达变化的定量分析显示在花背蟾蜍肺发育的全过程中,TGF-β(β_1,β_2,β_3)的表达呈间歇性变化,表明TGF-β在不同的生长时期,在肺组织中的表达均不同。
4.在花背蟾蜍胎肺发育的不同时期,EGFR,TGF-β_1,TGF-β_2,TGF-β_3因子在肺网状隔膜处和肺泡上皮细胞处的表达定位有迁移,免疫组化反应强弱也有差异变化,提示它们在胎肺发育的不同阶段发挥不同的功能。EGFR具有促进细胞发育分化的作用,而TGF-β因子的作用正相反,是一种选择的抑制作用,因此推测这类活性物质在花背蟾蜍胎肺发育过程中不仅对肺网状隔膜的形成,还对肺泡上皮细胞的成熟分化具有重要的调节作用。花背蟾蜍胎肺的正常发育有赖于这些活性物质之间的对立统一协调作用的共同调控。
Based on the histological study of the development of fetal lung in Bufo raddei,the distribution of the epidermal growth factor receptor(EGFR),transforming growth factor-β_1,β_2,β_3(TGF-β_1,TGF-β_2,TGF-β_3)was investigated in.the fetal lung during Bufo raddei tadpole development by the immunohistochemical method,the Image-pro-plus 5.0 professional image analysis was used in quantitatively studying.
1.The histological study demonstrated that the development of the lung in larvae of Bufo raddei was studied at different developmental stages from the youngest forms to those entering the metamorphosis,the lung in larvae of Bufo raddei developed in 21 stage in the beginning,it is derivated from the back of the pharynx,next it formed the protuberant lung bud,the lung bud developed slowness during the 22,23,24 stage,while in the 25 stage,it is differentiated the slim lung sac,but the lung was filled with fluid,in the 30 stage,the lung formed the obvious lung sac,the lung was filled with air,in the 39 stage,the lung has approached to the young Bufo raddei fetal lung,the wall of the lung showed attenuation, the lung of the adult Bufo raddei becomed vesicle,the inside wall of the lung there are many net diaphragm and secondary septa so that it is separated to a lot of alveolar sac and pulmonary alveoli,in the inside of the lung surface was covered with a layer of epithelium.
2.In the early developmental phase,EGFR localized in epithelium of the reticular septa;37-39 stage,the positive reactions of EGFR were mainly detected in alveolar sac and alveoli epithelium.The localization patterns in the interstitial cells for EGFR were essentially identical;In young toad lung,rich EGFR was mainly localized in alveoli epithelium.
The quantitative analysis of EGFR showed:the changes of the EGFR expression level taked on the ascending trend in the development of the lung,at the 30-35 stage,the change was calm,but at 37 stage,the level was reached the peak,after the stage gradually descended,at the 39 stage and the young toad lung,it began rasied.
3.TGF-β_1,TGF-β_2,TGF-β_3 were placed in the cytoplasm,the localization patterns of TGF-βin the fetal lung were essentially identical,at the stage 30-35,the TGF-βweak reaction was present in the epithelium of the reticular septa and interstitial cells;at the stage 37-39,the TGF-βimmunoreactivity was reduced in epithelium of the reticular septa,but evident staining was observed in smooth muscle cells and interstitial cells;In young toad lung,the TGF-βwas mainly observed in smooth muscle cells and interstitial cells around blood vessel.
The quantitative analysis of TGF-βshowed:the changes of the TGF-βexpression level taked on the intermission trend in the development of the lung,it is showed at the different stages,the expression level of the TGF-βwere different.
4.The positive reactions of EGFR,TGF-β_1,TGF-β_2,TGF-β_3 were respectively different stages,which suggests that their functions are changing in individual developmental periods,the EGFR can promote the cells to develop and differentiating,but on the contrary,TGF-βinhibits lung development,They not only stimulate the process of epithelium of the reticular septa,but also modulate epithelium of alveolar sac and differention.The Bufo raddei normal fetal lung development depends on the unity of two opposites.
1.花背蟾蜍的肺开始发生在幼虫时期的第21期,它起始于咽部后侧,向腹侧突起形成肺芽,肺芽经22、23、24期的缓慢发育,直到25期才开始分化出现微小的肺囊,但是里面充满液体,到30期开始形成明显的肺囊,且肺囊中已有气体,33期开始出现肺泡,36期时肺壁已充分伸展,肺泡增多,肺壁变薄,典型的气-血屏障开始形成,37期肺壁内表面的网状隔膜明显,到38期时肺壁进一步变薄,39期肺已接近幼蟾肺,成年花背蟾蜍的肺成囊状,并且在肺的内壁上有许多网状隔膜,将肺的内壁分隔形成许多小室,在每个小室内又有少数次级皱褶将小室分隔为数个近似半球形或“C”形小腔,肺内表面覆盖有一层肺上皮细胞。
2.花背蟾蜍胎肺发育的早期,EGFR主要表达于肺网状隔膜的上皮细胞,且较集中于上皮细胞的游离表面;37-39期,EGFR在肺泡囊及肺泡上皮细胞的阳性表达最活跃,EGFR在间充质细胞区域性分布更加明显;幼蟾期,EGFR阳性反应主要定位在肺泡上皮细胞,在肺网状隔膜处几乎不表达。
EGFR表达变化的定量分析显示在花背蟾蜍胎肺发育的全过程中,EGFR的表达水平呈逐渐上升状态,在30-35期变化比较平稳,至37期达到峰值,继而逐渐下降,39期和幼蟾期的表达水平又有所回升。
3.TGF-β_1,TGF-β_2,TGF-β_3三者均定位于细胞浆,阳性表达区域基本一致。30-36期,TGF-β在肺网状隔膜处和间充质细胞均呈现弱阳性,并且在网状隔膜处呈区域性分布特点;37-39期,TGF-β在肺网状隔膜处表达变弱,在平滑肌和肺间质细胞TGF-β呈强阳性表达;幼蟾期,血管周围的平滑肌和肺间质细胞处呈阳性表达。
TGF-β表达变化的定量分析显示在花背蟾蜍肺发育的全过程中,TGF-β(β_1,β_2,β_3)的表达呈间歇性变化,表明TGF-β在不同的生长时期,在肺组织中的表达均不同。
4.在花背蟾蜍胎肺发育的不同时期,EGFR,TGF-β_1,TGF-β_2,TGF-β_3因子在肺网状隔膜处和肺泡上皮细胞处的表达定位有迁移,免疫组化反应强弱也有差异变化,提示它们在胎肺发育的不同阶段发挥不同的功能。EGFR具有促进细胞发育分化的作用,而TGF-β因子的作用正相反,是一种选择的抑制作用,因此推测这类活性物质在花背蟾蜍胎肺发育过程中不仅对肺网状隔膜的形成,还对肺泡上皮细胞的成熟分化具有重要的调节作用。花背蟾蜍胎肺的正常发育有赖于这些活性物质之间的对立统一协调作用的共同调控。
Based on the histological study of the development of fetal lung in Bufo raddei,the distribution of the epidermal growth factor receptor(EGFR),transforming growth factor-β_1,β_2,β_3(TGF-β_1,TGF-β_2,TGF-β_3)was investigated in.the fetal lung during Bufo raddei tadpole development by the immunohistochemical method,the Image-pro-plus 5.0 professional image analysis was used in quantitatively studying.
1.The histological study demonstrated that the development of the lung in larvae of Bufo raddei was studied at different developmental stages from the youngest forms to those entering the metamorphosis,the lung in larvae of Bufo raddei developed in 21 stage in the beginning,it is derivated from the back of the pharynx,next it formed the protuberant lung bud,the lung bud developed slowness during the 22,23,24 stage,while in the 25 stage,it is differentiated the slim lung sac,but the lung was filled with fluid,in the 30 stage,the lung formed the obvious lung sac,the lung was filled with air,in the 39 stage,the lung has approached to the young Bufo raddei fetal lung,the wall of the lung showed attenuation, the lung of the adult Bufo raddei becomed vesicle,the inside wall of the lung there are many net diaphragm and secondary septa so that it is separated to a lot of alveolar sac and pulmonary alveoli,in the inside of the lung surface was covered with a layer of epithelium.
2.In the early developmental phase,EGFR localized in epithelium of the reticular septa;37-39 stage,the positive reactions of EGFR were mainly detected in alveolar sac and alveoli epithelium.The localization patterns in the interstitial cells for EGFR were essentially identical;In young toad lung,rich EGFR was mainly localized in alveoli epithelium.
The quantitative analysis of EGFR showed:the changes of the EGFR expression level taked on the ascending trend in the development of the lung,at the 30-35 stage,the change was calm,but at 37 stage,the level was reached the peak,after the stage gradually descended,at the 39 stage and the young toad lung,it began rasied.
3.TGF-β_1,TGF-β_2,TGF-β_3 were placed in the cytoplasm,the localization patterns of TGF-βin the fetal lung were essentially identical,at the stage 30-35,the TGF-βweak reaction was present in the epithelium of the reticular septa and interstitial cells;at the stage 37-39,the TGF-βimmunoreactivity was reduced in epithelium of the reticular septa,but evident staining was observed in smooth muscle cells and interstitial cells;In young toad lung,the TGF-βwas mainly observed in smooth muscle cells and interstitial cells around blood vessel.
The quantitative analysis of TGF-βshowed:the changes of the TGF-βexpression level taked on the intermission trend in the development of the lung,it is showed at the different stages,the expression level of the TGF-βwere different.
4.The positive reactions of EGFR,TGF-β_1,TGF-β_2,TGF-β_3 were respectively different stages,which suggests that their functions are changing in individual developmental periods,the EGFR can promote the cells to develop and differentiating,but on the contrary,TGF-βinhibits lung development,They not only stimulate the process of epithelium of the reticular septa,but also modulate epithelium of alveolar sac and differention.The Bufo raddei normal fetal lung development depends on the unity of two opposites.
引文
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