Hyperoxia disrupts pulmonary epithelial barrier in newborn rats via the deterioration of occludin and ZO-1

Abstract

BackgroundProlonged exposure to hyperoxia in neonates can cause hyperoxic acute lung injury (HALI), which is characterized by increased pulmonary permeability and diffuse infiltration of various inflammatory cells. Disruption of the epithelial barrier may lead to altered pulmonary permeability and maintenance of barrier properties requires intact epithelial tight junctions (TJs). However, in neonatal animals, relatively little is known about how the TJ proteins are expressed in the pulmonary epithelium, including whether expression of TJ proteins is regulated in response to hyperoxia exposure. This study determines whether changes in tight junctions play an important role in disruption of the pulmonary epithelial barrier during hyperoxic acute lung injury.MethodsNewborn rats, randomly divided into two groups, were exposed to hyperoxia (95% oxygen) or normoxia for 1–7 days, and the severity of lung injury was assessed; location and expression of key tight junction protein occludin and ZO-1 were examined by immunofluorescence staining and immunobloting; messenger RNA in lung tissue was studied by RT-PCR; transmission electron microscopy study was performed for the detection of tight junction morphology.ResultsWe found that different durations of hyperoxia exposure caused different degrees of lung injury in newborn rats. Treatment with hyperoxia for prolonged duration contributed to more serious lung injury, which was characterized by increased wet-to-dry ratio, extravascular lung water content, and bronchoalveolar lavage fluid (BALF):serum FD4 ratio. Transmission electron microscopy study demonstrated that hyperoxia destroyed the structure of tight junctions and prolonged hyperoxia exposure, enhancing the structure destruction. The results were compatible with pathohistologic findings. We found that hyperoxia markedly disrupted the membrane localization and downregulated the cytoplasm expression of the key tight junction proteins occludin and ZO-1 in the alveolar epithelium by immunofluorescence. The changes of messenger RNA and protein expression of occludin and ZO-1 in lung tissue detected by RT-PCR and immunoblotting were consistent with the degree of lung injury.ConclusionsThese data suggest that the disruption of the pulmonary epithelial barrier induced by hyperoxia is, at least in part, due to massive deterioration in the expression and localization of key TJ proteins.

Hyperoxia stimulates the transdifferentiation of type II alveolar epithelial cells in newborn rats

“…Pups were killed, and lungs harvested at the end of 1, 3, 7, 14, and 21 days of exposure as described previously (58,61). Briefly, lung samples from the right middle lobe were inflated at 18 cmH2O pressure with PBS and immersed in 4% paraformaldehyde (PFA) at 4°C, overnight.…”

“…All animal procedures were reviewed and approved by the Laboratory Animal Ethics Committee of China Medical University. Pregnant Wistar rats (200 -250 g) were purchased from the Animal Lab, Experimental Research Center, Shengjing Hospital, China Medical University, and the BPD model for the present study has been previously described (51,61). In brief, term pups of pregnant Wistar rats were randomly marked and assigned to two groups: model group rats were exposed to 85% oxygen in a sealed Plexiglas chamber for 1-21 days beginning on the day of birth, and the control group rats were exposed to room air.…”

“…After being dried in an oven at 80°C for 48 h, lung tissues were weighed again to yield a final dry weight. Finally, the pulmonary wet/dry (W/D) ratio was calculated to determine the presence of pulmonary edema (61).…”

“…Following lung injury, AEC II-to-AEC I transdifferentiation is an important mechanism in injury repair (16,24,39). Tight junctions between epithelial cells regulate diffusion of solutes through the paracellular pathway (47,61).…”

Regulation of endothelial permeability in the primate corpora lutea: implications for ovarian hyperstimulation syndrome

“…However, Occludin could not be detected in the theca lutein cells (Groten et al 2006). The presence of Occludin in epithelial and endothelial cells has also been discovered in other species tissues, such as rat lung, human liver, mouse brain, and others (Langbein et al 2002, Butt et al 2012, Errede et al 2012, You et al 2012.…”