Antisense inhibition of epidermal growth factor receptor decreases expression of human surfactant protein A.
Antisense inhibition of epidermal growth factor receptor decreases expression of human surfactant protein A.
复制标题
表皮生长因子受体的反义抑制可降低人表面活性剂蛋白 A 的表达。
DOI：
10.1165/ajrcmb.22.6.3726
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发表时间：
2000
期刊：
影响因子：
--
通讯作者：
Snyder,JM
中科院分区：
文献类型：
--
作者：
Klein,JM;McCarthy,TA;Dagle,JM;Snyder,JM
表皮生长因子（EGF）通过与EGF受体的配体结合刺激胎肺组织中表面活性蛋白A（SP-A）的合成。我们假设抑制EGF受体信使RNA（mRNA）可以阻断人胎肺组织中SP-A在肺泡Ⅱ型细胞体外分化过程中的表达，将中期人胎肺外植体在无血清Waymouth培养基中培养3 ~ 5d，加入或不加入与EGF受体mRNA起始密码子互补的反义18-mer硫代磷酸寡核苷酸（ON）。使用类似修饰的有义和乱序ON作为额外的对照。 逆转录聚合酶链反应（RT-PCR）半定量检测EGF受体mRNA的表达。我们发现，与对照组相比，反义处理组的EGF受体mRNA水平显着降低3倍，在有义条件下没有影响。免疫组化染色显示，与对照组或正义组相比，反义治疗组远端肺上皮细胞中EGF受体蛋白的染色量减少。与对照组相比，反义EGF受体ON治疗降低了SP-A mRNA和蛋白质，在正义条件下没有影响。通过乳酸脱氢酶的释放测定，ON不影响组织活力。我们的结论是EGF受体mRNA的选择性降解与反义ON治疗的结果在SP-A在人胎肺表达下降。这些发现支持EGF受体在人肺泡II型细胞分化过程中调节SP-A基因表达的至关重要性。
Epidermal growth factor (EGF) stimulates surfactant protein A (SP-A) synthesis in fetal lung tissue through ligand binding to the EGF receptor. We hypothesized that inhibition of EGF receptor messenger RNA (mRNA) would block SP-A expression in human fetal lung tissue during alveolar type II cell differentiationin vitro.Midtrimester human fetal lung explants were maintained in serum-free Waymouth's medium for 3 to 5 d in the presence or absence of an antisense 18-mer phosphorothioate oligonucleotide (ON) complementary to the initiation codon region of EGF receptor mRNA. Sense and scrambled ONs similarly modified were used as additional controls. The concentration of EGF receptor mRNA was semiquantitatively determined by reverse transcriptase/polymerase chain reaction (RT-PCR). We found a significant 3-fold decrease in EGF receptor mRNA levels in the antisense-treated groups compared with the control group with no effect in the sense condition. Immunohistochemical staining revealed a decrease in the amount of staining for EGF receptor protein in distal pulmonary epithelial cells in the antisense-treated groups compared with either control or sense conditions. Treatment with antisense EGF receptor ON decreased both SP-A mRNA and protein compared with controls with no effect in the sense condition. The ONs did not affect tissue viability as measured by the release of lactate dehydrogenase. We conclude that selective degradation of EGF receptor mRNA with antisense ON treatment results in a decrease in SP-A expression in human fetal lung. These findings support the critical importance of the EGF receptor for the regulation of SP-A gene expression during human alveolar type II cell differentiation.