Wang Mingwu, Huang Jianyan, Lu Michael, Zhang Shunhua, Ding Chuanqing
Formerly Doheny Eye Institute (M.W.), Keck School of Medicine, University of Southern California, Los Angeles, CA; currently Department of Ophthalmology and Vision Science (M.W.), University of Arizona College of Medicine, Tucson, AZ; Department of Ophthalmology (C.D.), Doheny Eye Institute, Keck School of Medicine, University of Southern California, Los Angeles, CA; Department of Cell and Neurobiology (J.H., M.L., C.D.), Keck School of Medicine, University of Southern California, Los Angeles, CA; and Department of Ophthalmology (S.Z.), Peking Union Medical College, Beijing, China.
Eye Contact Lens. 2015 Sep;41(5):297-303. doi: 10.1097/ICL.0000000000000123.
Epithelial sodium channel (ENaC) plays a critical role in the control of Na(+) balance and the development and progression of exocrine gland pathologic condition. The aim of the present study was to investigate the presence of ENaC in the rabbit lacrimal gland (LG) and its potential changes during induced autoimmune dacryoadenitis (IAD) and pregnancy.
Total messenger RNA (mRNA) of α, β, and γ subunits was extracted from whole LG, acinar cells, and ductal cells by laser capture microdissection (LCM) for real-time reverse-transcriptase polymerase chain reaction. Lacrimal glands were processed for Western blot and immunofluorescence.
Messenger RNA for both α and γ was expressed in whole LG lysates, whereas β was undetectable. In rabbits with IAD, the levels of mRNA for α and γ were 20.9% and 58.9% lower (P<0.05), whereas no significant changes were observed in term-pregnant rabbits (P=0.152). However, we were unable to detect mRNA of any subunit in LCM specimens of ductal cells because of their low levels. Western blot demonstrated bands for both α (90 kDa) and γ (85 kDa) but β was undetectable. In rabbits with IAD, densitometry analysis showed that expression of α decreased 22%, whereas γ decreased 26% (P<0.05). In pregnant rabbits, however, α expression was 31% lower, whereas γ expression was 34% lower (P<0.05). From immunofluorescence studies, all subunits were present in ductal cells, whereas virtually no immunoreactivity was detected in acini. No noticeable changes of their distribution pattern and intensity were found in rabbits with IAD or during pregnancy.
The present study demonstrated the presence of ENaC in the rabbit LG and its alterations in IAD and pregnancy, suggesting that ENaC may contribute to the pathogenesis of altered LG secretion and ocular surface symptoms in these animals.
上皮钠通道（ENaC）在钠平衡的控制以及外分泌腺病理状况的发生和发展中起关键作用。本研究的目的是调查兔泪腺（LG）中ENaC的存在情况及其在诱导性自身免疫性泪腺炎（IAD）和妊娠期间的潜在变化。
通过激光捕获显微切割（LCM）从整个泪腺、腺泡细胞和导管细胞中提取α、β和γ亚基的总信使核糖核酸（mRNA），用于实时逆转录聚合酶链反应。对泪腺进行蛋白质免疫印迹和免疫荧光检测。
α和γ的信使核糖核酸在整个泪腺裂解物中均有表达，而β未检测到。在患有IAD的兔子中，α和γ的信使核糖核酸水平分别降低了20.9%和58.9%（P<0.05），而足月妊娠的兔子中未观察到显著变化（P=0.152）。然而，由于导管细胞中mRNA水平较低，我们在其LCM标本中未能检测到任何亚基的mRNA。蛋白质免疫印迹显示α（90 kDa）和γ（85 kDa）的条带，但未检测到β。在患有IAD的兔子中，光密度分析显示α的表达下降了22%，而γ下降了26%（P<0.05）。然而，在妊娠兔子中，α的表达降低了31%，而γ的表达降低了34%（P<0.05）。免疫荧光研究表明，所有亚基均存在于导管细胞中，而在腺泡中几乎未检测到免疫反应性。在患有IAD的兔子或妊娠期间，未发现其分布模式和强度有明显变化。
本研究证明兔泪腺中存在ENaC及其在IAD和妊娠中的改变，表明ENaC可能参与了这些动物泪腺分泌改变和眼表症状的发病机制。
