Fig. 7.

Connective tissue growth factor (CTGF) expression is decreased in pulmonary microvascular endothelial cells (PMVECs) isolated from transgenic HIF1/2 KO mice. Conversely, CTGF expression is increased in primary human PMVECs isolated from a patient with IPF exposed to hypoxia. A: by quantitative PCR on cell lysates, CTGF mRNA expression increased dramatically in control PMVECs with exposure to hypoxia (O₂ 1% for 24 h) compared with PMVECs from endothelium-targeted mice, confirmed at the protein level by immunoblot on whole cell lysates for CTGF (B), demonstrating a reduction in CTGF in HIF1/2 KO compared with the hypoxic control. β-actin shown as loading control. C: quantitative PCR of whole cell lysates of PMVECs from normal subjects and patients with IPF and PH exposed to hypoxia (O₂ 1%) for 6 h, demonstrating increased mRNA expression of CTGF in IPF PMVECs compared with normal control. D: by immunoblot of whole cell lysates collected after exposure to hypoxia, CTGF expression was increased in IPF PVMECs compared with normal PMVECs. β-actin shown for loading control. E: representative images from lung sections with immunohistochemical staining for CTGF from normal subjects with no immunostain observed, diffuse parenchymal staining in samples from patients with IPF alone, and in patients with IPF and PH, demonstrating immunostain-positive cells in the lung, including the vascular endothelium (arrowheads points to representative cell). Scale bars: 50 μm at ×60 magnification. n = 4 per group for in vitro experiments. *P < 0.05. n = 10 per group for immunohistochemistry, except in IPF + PH where n = 4.