Role of CXC chemokine receptor-2 in a murine model of bronchopulmonary dysplasia.

The contribution of neutrophils and CXC chemokines to the pathogenesis of bronchopulmonary dysplasia is not well defined. The transgenic expression of IL-1ß in the pulmonary epithelium causes lung inflammation and disrupts alveolar development in infant mice. To study the hypothesis that CXC chemokine receptor-2 (CXCR2) is a mediator of inflammatory lung injury, we compared lung development in IL-1ß-expressing mice with wild-type (IL-1ß/CXCR2(+/+)) or null (IL-1ß/CXCR2(-/-)) CXCR2 loci. CXCR2 deficiency abolished the transmigration of neutrophils into the alveolar lumen in IL-1ß-expressing mice, but did not alter the number of neutrophils in the parenchyma. The deletion of CXCR2 increased the alveolar chord length and reduced the survival of mice when IL-1ß was expressed from the pseudoglandular to the alveolar stages. The capillary configuration was highly abnormal in both IL-1ß/CXCR2(+/+) and IL-1ß/CXCR2(-/-) lungs, but in very different ways. The cellular area of the parenchyma and the total capillary area of IL-1ß/CXCR2(+/+) and IL-1ß/CXCR2(-/-) mice were smaller than those of control/CXCR2(+/+) and control/CXCR2(-/-) mice, but the ratio of capillary area to cellular area was similar in all four genotypes. When IL-1ß was expressed during the saccular stage, IL-1ß/CXCR2(-/-) mice had smaller alveolar chord lengths and better survival than did IL-1ß/CXCR2(+/+) mice. Independent of the timing of IL-1ß expression, IL-1ß increased alveolar septal thickness in mice with wild-type CXCR2 loci, but not in CXCR2 null mice. Depending on the developmental stage at the time of the inflammatory insult, inhibition of the CXCR2 pathway may exert opposite effects on alveolar septation in the neonatal lung.

Developmental stage is a major determinant of lung injury in a murine model of bronchopulmonary dysplasia.

Bronchopulmonary dysplasia (BPD) is a common inflammatory lung disease in premature infants. To study the hypothesis that the sensitivity of the lung to inflammatory injury depends on the developmental stage, we studied postnatal lung development in transgenic mice expressing human IL-1ß (hIL-1ß) in the lungs during the late canalicular-early saccular, saccular, or late saccular-alveolar stage. Overexpression of hIL-1ß in the saccular stage caused arrest in alveolar development, airway remodeling, and goblet cell hyperplasia in the lungs as well as poor growth and survival of infant mice. Overexpression of hIL-1ß during the late canalicular-early saccular stage did not adversely affect lung development, growth, or survival of the pups. Mice expressing hIL-1ß from the late saccular to alveolar stage had smaller alveolar chord length, thinner septal walls, less airway remodeling and mucus metaplasia, and better survival than mice expressing hIL-1ß during the saccular stage. Human IL-1ß overexpression in the saccular stage was sufficient to cause a BPD-like illness in infant mice, whereas the lung was more resistant to hIL-1ß-induced injury at earlier and later developmental stages.

beta6 Integrin subunit deficiency alleviates lung injury in a mouse model of bronchopulmonary dysplasia.

Pulmonary inflammation is associated with the development of bronchopulmonary dysplasia in premature infants. We have previously shown that perinatal pulmonary expression of human IL-1beta is sufficient to cause a lung disease similar to bronchopulmonary dysplasia, characterized by inflammation, impaired alveolarization, poor postnatal growth, and increased mortality in infant mice. The alphavbeta6 integrin plays a critical role in regulating inflammation in the adult lung. To study the role of the beta6 integrin subunit in neonatal inflammatory lung disease, we compared the pulmonary development in IL-1beta-expressing infant mice with wild-type or null beta6 integrin loci. Absence of the beta6 integrin subunit decreased the mortality and improved the postnatal growth of IL-1beta-expressing pups. The disrupted alveolar development of IL-1beta-expressing mice was improved by beta6 integrin deficiency. IL-1beta-expressing beta6(-/-) pups had shorter alveolar chord length and thinner alveolar walls than IL-1beta-expressing beta6(+/+) pups. In addition, the absence of the beta6 integrin subunit reduced IL-1beta-induced neutrophil and macrophage infiltration into the alveolar spaces. beta6 integrin subunit deficiency suppressed inflammation and goblet cell hyperplasia in the airways and alleviated airway remodeling in IL-1beta-expressing mice. The expression of the chemoattractant proteins, keratinocyte-derived chemokine, macrophage-inflammatory protein-2, calgranulin A, and calgranulin B, of osteopontin, and of the chitinase-like lectins, Ym1 and Ym2, was lower in IL-1beta-expressing beta6(-/-) than in IL-1beta-expressing beta6(+/+) mice. We conclude that absence of the beta6 integrin subunit protects the infant murine lung against IL-1beta-induced inflammation and injury.

Maternal IL-1beta production prevents lung injury in a mouse model of bronchopulmonary dysplasia.

Little is known about the influence of maternal inflammation on neonatal outcome. Production of IL-1beta in the lungs of newborn infants is associated with bronchopulmonary dysplasia. Using bitransgenic (bi-TG) mice in which human (h) IL-1beta is expressed with a doxycycline-inducible system controlled by the Clara cell secretory protein promoter, we have shown that hIL-1beta expression causes a bronchopulmonary dysplasia-like illness in infant mice. To study the hypothesis that maternal hIL-1beta production modifies the response of the newborn to hIL-1beta, doxycycline was administered to bi-TG and control dams from Embryonic Day 0, inducing production of hIL-1beta by the bi-TG dams before hIL-1beta production started in their bi-TG fetuses, or from Embryonic Day 15, inducing simultaneous production of hIL-1beta by both the bi-TG dams and their bi-TG fetuses. In addition to the lungs, hIL-1beta was expressed at low levels in the uteri of bi-TG dams. Maternal inflammation preceding fetal inflammation increased the survival and growth of hIL-1beta-expressing pups, enhanced alveolarization, and protected the airways against remodeling and goblet cell hyperplasia. Maternal hIL-1beta production preceding fetal hIL-1beta production caused silencing of several inflammatory genes, including CXC and CC chemokines, murine IL-1beta, serum amyloid A3, and Toll-like receptors 2 and 4, and suppressed the expression of chitinase-like lectins Ym1 and Ym2 in the lungs of infant mice. Maternal inflammation protects the newborn against subsequent hIL-1beta-induced lung inflammation and injury. In contrast, induction of hIL-1beta production simultaneously in bi-TG dams and their fetuses offered no protection against inflammatory lung disease in the neonate.

Matrix metalloproteinase-9 deficiency worsens lung injury in a model of bronchopulmonary dysplasia.

Increased activity of matrix metalloproteinase (MMP)-9 is associated with the development of bronchopulmonary dysplasia (BPD) in newborn infants, but the role of MMP-9 in the pathophysiology of BPD is unclear. We have shown that perinatal expression of interleukin-1 beta (IL-1 beta) in the lung is sufficient to cause a BPD-like illness in infant mice. To study the hypothesis that MMP-9 is an important downstream mediator in IL-1 beta-induced lung injury in the newborn, we compared the effects of IL-1 beta on fetal and postnatal lung inflammation and development in transgenic mice with regulatable pulmonary overexpression of human mature IL-1 beta with wild-type (IL-1 beta/MMP-9(+/+)) or null (IL-1 beta/MMP-9(-/-)) MMP-9 loci. IL-1 beta increased the expression of MMP-9 mRNA and amount of MMP-9 protein in the lungs of MMP-9(+/+) mice. IL-1 beta/MMP-9(-/-) mice had fewer neutrophils but more macrophages in the lungs than did IL-1 beta/MMP-9(+/+) mice. MMP-9 deficiency increased pulmonary cell death and macrophage clearance of dying cells in IL-1 beta-expressing mice. IL-1 beta/MMP-9(-/-) mice had more severe alveolar hypoplasia than IL-1 beta/MMP-9(+/+) mice, implying that IL-1 beta-induced lung disease was worsened in the absence of MMP-9. These results suggest that MMP-9 activity in the inflamed neonatal lung protects the lung against injury.

IL-1beta disrupts postnatal lung morphogenesis in the mouse.

Pulmonary inflammation and increased production of the inflammatory cytokine IL-1beta are associated with the development of bronchopulmonary dysplasia (BPD) in premature infants. To study the actions of IL-1beta in the fetal and newborn lung in vivo, we developed a bitransgenic mouse in which IL-1beta is expressed under conditional control in airway epithelial cells. Perinatal pulmonary expression of IL-1beta caused respiratory insufficiency that was associated with increased postnatal mortality. While intrauterine growth of IL-1beta-expressing mice was normal, their postnatal growth was impaired. IL-1beta disrupted alveolar septation and caused abnormalities in alpha-smooth muscle actin and elastin deposition in the septa of distal airspaces. IL-1beta disturbed capillary development and inhibited the production of vascular endothelial growth factor in the lungs of infant mice. IL-1beta induced the expression of CXC chemokines KC (CXCL1) and macrophage inflammatory protein-2 (CXCL2) and of CC chemokines monocyte chemotactic protein (MCP)-1 (CCL2) and MCP-3 (CCL7), consistent with neutrophilic and monocytic infiltration of the lungs. IL-1beta caused goblet cell metaplasia and bronchial smooth muscle hyperplasia. Perinatal expression of IL-1beta in epithelial cells of the lung caused a lung disease that was clinically and histologically similar to BPD.

Pathogenesis of bronchopulmonary dysplasia: the role of interleukin 1beta in the regulation of inflammation-mediated pulmonary retinoic acid pathways in transgenic mice.

BACKGROUND: Pulmonary inflammation, increased production of the inflammatory cytokine interleukin-1beta (IL-1beta), and vitamin A deficiency are risk factors for the development of bronchopulmonary dysplasia (BPD) in premature infants. To determine the mechanisms by which IL-1beta influences lung development, we have generated transgenic mice in which human IL-1beta is expressed in the lung epithelium with a doxycycline-inducible system controlled by the Clara cell secretory protein promoter. Perinatal IL-1beta production in these mice causes a phenotype that is strikingly similar to BPD. Pulmonary pathology in the mice shows inflammation, lack of alveolar septation, and impaired vascular development of the lung, similar to the histological characteristics of BPD. Retinoic acid (RA), one of the most biologically active derivatives of vitamin A, increases septation. Proteins involved in mediating the cellular responses to RA include the cellular retinoic acid binding proteins CRABP-I and CRABP-II and the nuclear retinoic acid receptors RAR-alpha, RAR-beta, and RAR-gamma. OBJECTIVE: To test the hypothesis that IL-1beta inhibits the expression of proteins involved in mediating the cellular response to RA. METHODS: The mRNA expression of CRABP-I, CRABP-II, RAR-alpha1, RAR-beta2, RAR-beta4, and RAR-gamma2 was studied with real-time RT-PCR on gestational day 18, and postnatal days 0, 1, 5, and 7 in IL-1beta-expressing mice and their control littermates. In addition, immunohistochemistry for CRABP-I was performed. RESULTS: IL-1beta decreased the mRNA expression and protein production of CRABP-I as well as the mRNA expression of RAR-gamma2. In contrast, no differences between IL-1beta-expressing and control mice were detected in the expression of CRABP-II, RAR-alpha1, RAR-beta2, or RAR-beta4. CONCLUSION: The present study demonstrates for the first time a link between inflammation and the retinoic acid pathway. Inhibition of CRABP-I and RAR-gamma2 expression may be one mechanism by which inflammation prevents alveolar septation. The therapeutic potential of RA in promoting septation in the setting of perinatal lung inflammation deserves further investigation.

Cimetidine does not prevent lung injury in newborn premature infants.

Animal studies have shown that induction of cytochrome P450 (CYP) in the lung by oxygen exposure may result in the release of free radical oxidants and arachidonic acid metabolites, which can cause lung injury that is reduced by treatment with cimetidine, a CYP inhibitor. To determine whether cimetidine would reduce lung injury in human infants at risk for chronic lung disease, we conducted a randomized clinical trial in which we administered either cimetidine or a placebo for 10 d beginning < 24 h after birth to 84 newborn infants weighing < or = 1250 g who were receiving O2 and mechanical ventilation. Cimetidine had no significant effect on severity of respiratory insufficiency assessed at 10 d postnatal age. F2-isoprostane levels (a marker of oxidant injury) in tracheal aspirates were significantly higher in the cimetidine group at 4 d and at 10 d. There were no significant differences between the groups in tracheal aspirate levels of inflammatory markers (leukotriene B4, IL-8, and nucleated cell count) or arachidonic acid metabolites. We conclude that cimetidine does not reduce lung injury in newborn premature infants receiving O2 and mechanical ventilation. It is possible that cimetidine was not an adequate CYP inhibitor in this context.

Interleukin-1beta causes pulmonary inflammation, emphysema, and airway remodeling in the adult murine lung.

The production of the inflammatory cytokine interleukin (IL)-1 is increased in lungs of patients with chronic obstructive pulmonary disease (COPD) or asthma. To characterize the in vivo actions of IL-1 in the lung, transgenic mice were generated in which human IL-1beta was expressed in the lung epithelium with a doxycycline-inducible system controlled by the rat Clara cell secretory protein (CCSP) promoter. Induction of IL-1beta expression in the lungs of adult mice caused pulmonary inflammation characterized by neutrophil and macrophage infiltrates. IL-1beta caused distal airspace enlargement, consistent with emphysema. IL-1beta caused disruption of elastin fibers in alveolar septa and fibrosis in airway walls and in the pleura. IL-1beta increased the thickness of conducting airways, enhanced mucin production, and caused lymphocytic aggregates in the airways. Decreased immunostaining for the winged helix transcription factor FOXA2 was associated with goblet cell hyperplasia in IL-1beta-expressing mice. The production of the neutrophil attractant CXC chemokines KC (CXCL1) and MIP-2 (CXCL2), and of matrix metalloproteases MMP-9 and MMP-12, was increased by IL-1beta. Chronic production of IL-1beta in respiratory epithelial cells of adult mice causes lung inflammation, enlargement of distal airspaces, mucus metaplasia, and airway fibrosis in the adult mouse.