Background: Human umbilical cord blood (UCB)-derived mesenchymal stem cells (MSCs) attenuate hyperoxic neonatal lung injury primarily through anti-inflammatory effects. We hypothesized that intratracheal transplantation of human UCB-derived MSCs could attenuate Escherichia coli (E. coli)-induced acute lung injury (ALI) in mice by suppressing the inflammatory response. Methods: Eight-week-old male ICR mice were randomized to control or ALI groups. ALI was induced by intratracheal E. coli instillation. Three-hours after E. coli instillation, MSCs, fibroblasts or phosphate-buffered saline were intratracheally administered randomly and survival was analyzed for 7 days post-injury. Lung histology including injury scores, myeloperoxidase (MPO) activity, and protein levels of interleukin (IL)-1α, IL-1β, IL-6, tumor necrosis factor (TNF)-α, and macrophage inflammatory protein (MIP)-2 as well as the wet-dry lung ratio and bacterial counts from blood and bronchoalveolar lavage (BAL) were evaluated at 1, 3, and 7 days post-injury. Levels of inflammatory cytokines in the lung were also profiled using protein macroarrays at day 3 post-injury which showed peak inflammation. Results: MSC transplantation increased survival and attenuated lung injuries in ALI mice, as evidenced by decreased injury scores on day 3 post-injury and reduced lung inflammation including increased MPO activity and protein levels of IL-1α, IL-1β, IL-6, TNF-α, and MIP-2 on day 3 and 7 post-injury. Inflammatory cytokine profiles in the lungs at day 3 post-injury were attenuated by MSC transplantation. MSCs also reduced the elevated lung water content at day 3 post-injury and bacterial counts in blood and BAL on day 7 post-injury. Conclusions: Intratracheal transplantation of UCB-derived MSCs attenuates E. coli-induced ALI primarily by down-modulating the inflammatory process and enhancing bacterial clearance.

Intratracheal transplantation of human umbilical cord blood (UCB)–derived mesenchymal stem cells (MSCs) protects against neonatal hyperoxic lung injury by a paracrine rather than a regenerative mechanism. However, the role of paracrine factors produced by the MSCs, such as vascular endothelial growth factor (VEGF), has not been delineated. This study examined whether VEGF secreted by MSCs plays a pivotal role in protecting against neonatal hyperoxic lung injury. VEGF was knocked down in human UCB–derived MSCs by transfection with small interfering RNA specific for human VEGF. The in vitro effects of MSCs with or without VEGF knockdown or neutralizing antibody were evaluated in a rat lung epithelial (L2) cell line challenged with H2O2. To confirm these results in vivo, newborn Sprague-Dawley rats were exposed to hyperoxia (90% O2) for 14 days. MSCs (1 × 105 cells) with or without VEGF knockdown were administered intratracheally at postnatal Day 5. Lungs were serially harvested for biochemical and histologic analyses. VEGF knockdown and antibody abolished the in vitro benefits of MSCs on H2O2–induced cell death and the up-regulation of inflammatory cytokines in L2 cells. VEGF knockdown also abolished the in vivo protective effects of MSCs in hyperoxic lung injury, such as the attenuation of impaired alveolarization and angiogenesis, reduction in the number of terminal deoxynucleotidyl transferase dUTP nick end labeling–positive and ED-1–positive cells, and down-regulation of proinflammatory cytokine levels. Our data indicate that VEGF secreted by transplanted MSCs is one of the critical paracrine factors that play seminal roles in attenuating hyperoxic lung injuries in neonatal rats.

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