McCray P B, Reenstra W W, Louie E, Johnson J, Bettencourt J D, Bastacky J
Department of Pulmonary Medicine, Children's Hospital Research Institute, Oakland 94609.
Am J Physiol. 1992 Apr;262(4 Pt 1):L472-81. doi: 10.1152/ajplung.1992.262.4.L472.
We studied the developmental expression of the cystic fibrosis (CF) gene in human lung tissue from normal and CF-affected fetuses. Two unrelated CF fetuses, both homozygous for the delta F508 deletion, were examined. Cystic fibrosis transmembrane conductance regulator (CFTR) mRNA was present in second-trimester CF lung and in first- and second-trimester normal lung as assessed by amplification of reverse transcribed total RNA with the use of the polymerase chain reaction. CFTR protein was identified by immunoprecipitation in normal second-trimester fetal lung explants. To evaluate possible functional consequences of CF in the fetus, lung tissue explants were grown in submersion organ culture. By light and electron microscopy, the CF fetal lung explants appeared normal. When explants from normal fetal lung were exposed to 8-(4-chlorophenylthio) adenosine 3',-5'cyclic monophosphate (CPT-cAMP), and 3-isobutyl-1-methylxanthine (IBMX) for 24 h, the intraluminal fluid content increased, as assessed by a 40 +/- 4% increase in cross-sectional diameter. In contrast, identically treated CF explants showed no significant change in explant diameter (3 +/- 1.6%). The transepithelial potential (psi t) across fetal lung explants was measured with microelectrodes. In normal second-trimester explants, CPT-cAMP and IBMX caused hyperpolarization of psi t (-0.93 +/- 14 mV to -4.3 +/- 1.2 mV); in contrast, CF fetal lung explants showed no significant change in psi t with CPT-cAMP and IBMX (-0.84 +/- 0.07 mV to -1.21 +/- 0.26 mV). This study confirms the presence of CFTR mRNA and protein in human fetal lung and suggests that although the CF fetal lung appears normal morphologically, there is a defect in cAMP-mediated fluid secretion in the lung of the CF fetus.
我们研究了囊性纤维化（CF）基因在正常胎儿和患CF胎儿的人肺组织中的发育表达情况。检查了两名无关的CF胎儿，二者均为ΔF508缺失的纯合子。通过使用聚合酶链反应对逆转录的总RNA进行扩增评估，CF跨膜传导调节因子（CFTR）mRNA存在于妊娠中期的CF肺以及妊娠早期和中期的正常肺中。通过免疫沉淀在正常妊娠中期胎儿肺外植体中鉴定出CFTR蛋白。为了评估CF对胎儿可能产生的功能影响，将肺组织外植体在浸没器官培养中培养。通过光学和电子显微镜观察，CF胎儿肺外植体看起来正常。当将正常胎儿肺的外植体暴露于8-（4-氯苯硫基）腺苷3'，5'-环磷酸（CPT-cAMP）和3-异丁基-1-甲基黄嘌呤（IBMX）24小时时，通过横截面直径增加40±4%评估，管腔内液体含量增加。相比之下，经过相同处理的CF外植体的直径没有显著变化（3±1.6%）。用微电极测量跨胎儿肺外植体的跨上皮电位（ψt）。在正常妊娠中期外植体中，CPT-cAMP和IBMX导致ψt超极化（从-0.93±14 mV到-4.3±1.2 mV）；相比之下，CF胎儿肺外植体在CPT-cAMP和IBMX作用下ψt没有显著变化（从-0.84±0.07 mV到-1.21±0.26 mV）。这项研究证实了CFTR mRNA和蛋白在人胎儿肺中的存在，并表明尽管CF胎儿肺在形态上看起来正常，但CF胎儿的肺中存在cAMP介导的液体分泌缺陷。
