RESUMEN
The onset and degree of injury occurring in animals that develop hyperoxic acute lung injury (HALI) is dependent on age at exposure, suggesting that developmentally regulated pathways/factors must underlie initiation of the epithelial injury and subsequent repair. Type II TGFß receptor interacting protein-1 (TRIP-1) is a negative regulator of TGFß signaling, which we have previously shown is a developmentally regulated protein with modulatory effects on epithelial-fibroblastic signaling. The aim of this study was to assess if type II alveolar epithelial cells overexpressing TRIP-1 are protected against hyperoxia-induced epithelial injury, and in turn HALI. Rat lung epithelial cells (RLE) overexpressing TRIP-1 or LacZ were exposed to 85% oxygen for 4 days. A surfactant protein C (SPC)-driven TRIP-1 overexpression mouse (TRIP-1AECTg+ ) was generated and exposed to hyperoxia (>95% for 4 days) at 4 weeks of age to assess the effects TRIP-1 overexpression has on HALI. RLE overexpressing TRIP-1 resisted hyperoxia-induced apoptosis. Mice overexpressing TRIP-1 in their lung type II alveolar epithelial cells (TRIP-1AECTg+ ) showed normal lung development, increased phospho-AKT level and E-cadherin, along with resistance to HALI, as evidence by less TGFß activation, apoptosis, alveolar macrophage influx, KC expression. Taken together, these findings point to existence of a TRIP-1 mediated molecular pathway affording protection against epithelial/acute lung injury.
Asunto(s)
Lesión Pulmonar Aguda/metabolismo , Factores Eucarióticos de Iniciación/metabolismo , Hipoxia/complicaciones , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Lesión Pulmonar Aguda/etiología , Células Epiteliales Alveolares/metabolismo , Animales , Apoptosis , Línea Celular , Factores Eucarióticos de Iniciación/genética , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Pulmón/citología , Pulmón/metabolismo , Ratones , Ratones Endogámicos C57BL , RatasRESUMEN
Pulmonary lymphatic development in chronic lung disease (CLD) has not been investigated, and anatomy of lymphatics in human infant lungs is not well defined. Hypothesis. Pulmonary lymphatic hypoplasia is present in CLD. Method. Autopsy lung tissues of eighteen subjects gestational ages 22 to 40 weeks with and without history of respiratory morbidity were stained with monoclonal antipodoplanin and reviewed under light microscopy. Percentage of parenchyma podoplanin stained at the acinar level was determined using computerized image analysis; 9 CLD and 4 control subjects gestational ages 27 to 36 weeks were suitable for the analysis. Results. Distinct, lymphatic-specific staining with respect to other vascular structures was appreciated in all gestations. Infants with and without respiratory morbidity had comparable lymphatic distribution which extended to the alveolar ductal level. Podoplanin staining per parenchyma was increased and statistically significant in the CLD group versus controls at the alveolar ductal level (0.06% ± 0.02% versus 0.04% ± 0.01%, 95% CI -0.04% to -0.002%, P < 0.03). Conclusion. Contrary to our hypothesis, the findings show that there is an increase in alveolar lymphatics in CLD. It is suggested that the findings, by expanding current knowledge of CLD pathology, may offer insight into the development of more effective therapies to tackle CLD.
Asunto(s)
Enfermedad Crónica , Enfermedades Pulmonares/patología , Pulmón/patología , Anomalías Linfáticas/patología , Autopsia , Humanos , LactanteRESUMEN
BACKGROUND: Myofibroblasts are the critical effector cells in the pathogenesis of pulmonary fibrosis which carries a high degree of morbidity and mortality. We have previously identified Type II TGFß receptor interacting protein 1 (TRIP-1), through proteomic analysis, as a key regulator of collagen contraction in primary human lung fibroblasts--a functional characteristic of myofibroblasts, and the last, but critical step in the process of fibrosis. However, whether or not TRIP-1 modulates fibroblast trans-differentiation to myofibroblasts is not known. METHODS: TRIP-1 expression was altered in primary human lung fibroblasts by siRNA and plasmid transfection. Transfected fibroblasts were then analyzed for myofibroblast features and function such as α-SMA expression, collagen contraction ability, and resistance to apoptosis. RESULTS: The down-regulation of TRIP-1 expression in primary human lung fibroblasts induces α-SMA expression and enhances resistance to apoptosis and collagen contraction ability. In contrast, TRIP-1 over-expression inhibits α-SMA expression. Remarkably, the effects of the loss of TRIP-1 are not abrogated by blockage of TGFß ligand activation of the Smad3 pathway or by Smad3 knockdown. Rather, a TRIP-1 mediated enhancement of AKT phosphorylation is the implicated pathway. In TRIP-1 knockdown fibroblasts, AKT inhibition prevents α-SMA induction, and transfection with a constitutively active AKT construct drives collagen contraction and decreases apoptosis. CONCLUSIONS: TRIP-1 regulates fibroblast acquisition of phenotype and function associated with myofibroblasts. The importance of this finding is it suggests TRIP-1 expression could be a potential target in therapeutic strategy aimed against pathological fibrosis.
Asunto(s)
Transdiferenciación Celular/fisiología , Factor 3 de Iniciación Eucariótica/fisiología , Fibroblastos/fisiología , Pulmón/fisiología , Miofibroblastos/fisiología , Proteínas Proto-Oncogénicas c-akt/fisiología , Animales , Células Cultivadas , Humanos , Pulmón/citología , RatasRESUMEN
Epithelial-mesenchymal transition (EMT) is a process by which epithelial cells undergo conversion to a mesenchymal phenotype contributing to wound repair by fibrosis and to cancer cell acquisition of invasive ability. Recently, we showed that type II TGF-ß receptor interacting protein-1 (TRIP-1), a protein identified as a phosphorylation target of the TGF-ß type II receptor kinase and as a functional component of eukaryotic translation initiator factor 3 (eiF3) multiprotein complex, is a novel modulator of fibroblast collagen contraction, an important step in wound repair stimulated by TGF-ß1 action. TGF-ß1 drives EMT, but it is not known whether TRIP-1 expression influences EMT induction. To investigate whether TRIP-1 plays a role in EMT induction we studied the effect of downregulating TRIP-1 expression in the well-characterized A549 model of TGF-ß1 induction of EMT. Here we report that short hairpin RNA (shRNA)-mediated depletion of TRIP-1 gene transcripts in A549 cells promotes EMT as assessed by changes in phenotypic markers, morphology, and migrative ability. Knockdown of TRIP-1 dramatically increased A549 responsiveness to TGF-ß1 induction of EMT. Mechanistically, a pathway involving increased TGF-ß type II receptor level, enhanced Smad3 phosphorylation, and the transcription factor SLUG is implicated. Altogether, the findings point to regulation of endogenous TRIP-1 protein expression as a potential strategy to target EMT, and related invasive behavior, in cancer cells.
Asunto(s)
Transición Epitelial-Mesenquimal , Factor 3 de Iniciación Eucariótica/metabolismo , Factor de Crecimiento Transformador beta1/farmacología , Línea Celular Tumoral , Regulación hacia Abajo , Células Epiteliales/citología , Transición Epitelial-Mesenquimal/efectos de los fármacos , Factor 3 de Iniciación Eucariótica/genética , Humanos , Pulmón , ARN Interferente Pequeño/farmacología , Proteína smad3/metabolismo , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
We have shown previously that T1α/podoplanin is required for capillary tube formation by human lung microvascular lymphatic endothelial cells (HMVEC-LLy) and that cells with decreased podoplanin expression fail to properly activate the small GTPase RhoA shortly after the beginning of the lymphangiogenic process. The objective of this study was to determine whether podoplanin regulates HMVEC-LLy migration and whether this regulation is via modulation of small GTPase activation. In analysis of scratch wound assays, we found that small interfering RNA (siRNA) depletion of podoplanin expression in HMVEC-LLy inhibits VEGF-induced microtubule-organizing center (MTOC) and Golgi polarization and causes a dramatic reduction in directional migration compared with control siRNA-transfected cells. In addition, a striking redistribution of cortical actin to fiber networks across the cell body is observed in these cells, and, remarkably, it returns to control levels if the cells are cotransfected with a dominant-negative mutant of Cdc42. Moreover, cotransfection of a dominant-negative construct of Cdc42 into podoplanin knockdown HMVEC-LLy completely abrogated the effect of podoplanin deficiency, rescuing MTOC and Golgi polarization and cell migration to control level. Importantly, expression of constitutively active Cdc42 construct, like podoplanin knockdown, decreased RhoA-GTP level in HMVEC-LLy, demonstrating cross talk between both GTPases. Taken together, the results indicate that polarized migration of lymphatic endothelial cells in response to VEGF is mediated via a pathway of podoplanin regulation of small GTPase activities, in particular Cdc42.
Asunto(s)
Células Endoteliales/fisiología , Pulmón/fisiología , Glicoproteínas de Membrana/fisiología , Microcirculación/fisiología , Proteína de Unión al GTP cdc42/fisiología , Movimiento Celular/fisiología , Activadores de GTP Fosfohidrolasa/metabolismo , GTP Fosfohidrolasas/metabolismo , Aparato de Golgi/metabolismo , Humanos , Glicoproteínas de Membrana/deficiencia , Glicoproteínas de Membrana/genética , Proteínas de Unión al GTP Monoméricas/metabolismo , Proteína de Unión al GTP rhoA/metabolismoRESUMEN
Acute lung injury involving extremely immature lungs often heals without excessive fibrosis unlike later in gestation and in adults. Several factors may be involved, but fibroblast contraction of collagen has been linked to the level of wound fibrosis. To assess whether human lung fibroblasts of fetal versus adult origin differ in ability to contract collagen and define the molecular underpinnings, we performed three-dimensional collagen contraction assay, analyzed their differential mRNA profile, specifically for transforming growth factor-beta (TGF-beta) signaling pathway and extracellular matrix components, studied the cell response to TGF-beta in culture, and used two-dimensional gel electrophoresis followed by mass spectrometry to identify differences in their overall proteomes. Human lung fetal fibroblasts contracted the collagen matrix less than the adults. Smooth muscle actin expression did not differ. TGF-beta stimulation resulted in greater Smad3 phosphorylation in fetal compared with adults. mRNA and proteomic profiling reveal a number of TGF-beta pathways, ECM components, and cytoskeletal regulatory molecules are differentially expressed between the cell types. Of note is TGF-beta receptor interacting protein 1 (TRIP-1), which we show inhibits fibroblast collagen contraction and is higher in fetal than adult fibroblasts. We conclude that human lung fetal fibroblasts are less able to contract collagen than adult lung fibroblasts. The diminished ability is not due to impediment of Smad3 activation but rather, at least in part, due to their higher level of TRIP-1 expression. TRIP-1 is a novel modulator of fibroblast collagen contraction.
Asunto(s)
Colágeno Tipo I/fisiología , Factor 3 de Iniciación Eucariótica/genética , Factor 3 de Iniciación Eucariótica/metabolismo , Fibroblastos/fisiología , Pulmón/citología , Adulto , Células Cultivadas , Electroforesis en Gel Bidimensional , Feto/citología , Fibroblastos/citología , Fibroblastos/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica , Humanos , Pulmón/embriología , Pulmón/metabolismo , Proteómica , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/patología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína smad3/metabolismo , Factor de Crecimiento Transformador beta/farmacología , Cicatrización de Heridas/fisiologíaRESUMEN
The lymphatic vasculature functions to maintain tissue perfusion homeostasis. Defects in its formation or disruption of the vessels result in lymphedema, the effective treatment of which is hampered by limited understanding of factors regulating lymph vessel formation. Mice lacking T1alpha/podoplanin, a lymphatic endothelial cell transmembrane protein, have malformed lymphatic vasculature with lymphedema at birth, but the molecular mechanism for this phenotype is unknown. Here, we show, using primary human lung microvascular lymphatic endothelial cells (HMVEC-LLy), that small interfering RNA-mediated silence of podoplanin gene expression has the dramatic effect of blocking capillary tube formation in Matrigel. In addition, localization of phosphorylated ezrin/radixin/moesin proteins to plasma membrane extensions, an early event in the capillary morphogenic program in lymphatic endothelial cells, is impaired. We find that cells with decreased podoplanin expression fail to properly activate the small GTPase RhoA early (by 30 min) after plating on Matrigel, and Rac1 shows a delay in its activation. Further indication that podoplanin action is linked to RhoA activation is that use of a cell-permeable inhibitor of Rho inhibited lymphatic endothelial capillary tube formation in the same manner as did podoplanin gene silencing, which was not mimicked by treatment with a Rac1 inhibitor. These data clearly demonstrate that early activation of RhoA in the lymphangiogenic process, which is required for the successful establishment of the capillary network, is dependent on podoplanin expression. To our knowledge, this is the first time that a mechanism has been suggested to explain the role of podoplanin in lymphangiogenesis.
Asunto(s)
Capilares/fisiología , Pulmón/fisiología , Glicoproteínas de Membrana/genética , Microcirculación/fisiología , Circulación Pulmonar/fisiología , Supervivencia Celular , Silenciador del Gen , Humanos , Pulmón/citología , Glicoproteínas de Membrana/inmunología , ARN/genética , ARN Interferente Pequeño , TransfecciónRESUMEN
It is unclear how sublethal hypoxia affects lung development. To investigate the effects of chronic hypoxia on postnatal lung remodeling, we treated neonatal rats with FIO2 of 0.12 for 10 d and analyzed lung development by morphometry and gene expression by DNA microarray. Our results showed the neonatal rats exposed to hypoxia reduced body weight by 42% and wet lung weight by 32% compared with the neonatal rats exposed to normoxia. In the neonatal rats exposed to hypoxia, the radial alveolar counts were decreased to 5.6 from 7.9 and the mean linear intercepts were increased to 56.5 mum from 38.2 mum. In DNA microarray analysis, approximately half of probed genes were unknown. Chronic hypoxia significantly regulated expression of genes that are involved in pathogenesis of pulmonary hypertension and postnatal lung remodeling. Chemokine ligand 12, jagged 2 were among those upregulated; c-kit, ephrin A1, and Hif-2alpha were among those downregulated. The altered expression of those genes was correlated with the lung development and remodeling.
Asunto(s)
Perfilación de la Expresión Génica/métodos , Regulación del Desarrollo de la Expresión Génica , Hipoxia/genética , Hipoxia/patología , Análisis de Secuencia por Matrices de Oligonucleótidos , Alveolos Pulmonares/patología , Animales , Animales Recién Nacidos , Peso Corporal , Enfermedad Crónica , Modelos Animales de Enfermedad , Hipoxia/fisiopatología , Neovascularización Fisiológica , Tamaño de los Órganos , Alveolos Pulmonares/irrigación sanguínea , Alveolos Pulmonares/crecimiento & desarrollo , Ratas , Reproducibilidad de los ResultadosRESUMEN
Hyperoxia is one of the major contributors to the development of bronchopulmonary dysplasia (BPD), a chronic lung disease in premature infants. Emerging evidence suggests that the arrested lung development of BPD is associated with pulmonary endothelial cell death and vascular dysfunction resulting from hyperoxia-induced lung injury. A better understanding of the mechanism of hyperoxia-induced endothelial cell death will provide critical information for the pathogenesis and therapeutic development of BPD. Epidermal growth factor-like domain 7 (EGFL7) is a protein secreted from endothelial cells. It plays an important role in vascular tubulogenesis. In the present study, we found that Egfl7 gene expression was significantly decreased in the neonatal rat lungs after hyperoxic exposure. The Egfl7 expression was returned to near normal level 2 wk after discounting oxygen exposure during recovery period. In cultured human endothelial cells, hyperoxia also significantly reduced Egfl7 expression. These observations suggest that diminished levels of Egfl7 expression might be associated with hyperoxia-induced endothelial cell death and lung injury. When we overexpressed human Egfl7 (hEgfl7) in EA.hy926 human endothelial cell line, we found that hEgfl7 overexpression could partially block cytochrome c release from mitochondria and decrease caspase-3 activation. Further Western blotting analyses showed that hEgfl7 overexpression could reduce expression of a proapoptotic protein, Bax, and increase expression of an antiapoptotic protein, Bcl-xL. Theses findings indicate that hEGFL7 may protect endothelial cell from hyperoxia-induced apoptosis by inhibition of mitochondria-dependent apoptosis pathway.
Asunto(s)
Factores de Crecimiento Endotelial/fisiología , Endotelio Vascular/fisiología , Hiperoxia/prevención & control , Animales , Animales Recién Nacidos , Displasia Broncopulmonar/prevención & control , Proteínas de Unión al Calcio , Supervivencia Celular/fisiología , Familia de Proteínas EGF , Factores de Crecimiento Endotelial/genética , Endotelio Vascular/fisiopatología , Regulación de la Expresión Génica , Humanos , Hiperoxia/fisiopatología , Recién Nacido , L-Lactato Deshidrogenasa/análisis , Ratas , TransfecciónRESUMEN
BACKGROUND: Hyperoxia and tidal volume mechanical ventilation are independent factors in the genesis of lung injury, but it remains unclear the extent to which each is responsible or contributes to this process in newborns. OBJECTIVES: To study the independent and combined effects of hyperoxia and tidal volume mechanical ventilation on the induction of lung inflammation in a newborn piglet model of ventilator-induced lung injury. METHODS: Following exposure to either ambient air or F(I)O2 = 1.0 for a period of 3 days, newborn piglets were randomized to receive mechanical ventilation with either high tidal volume (20 ml/kg) or low tidal volume (6 ml/kg) for 4 h while controlling for pH. RESULTS: Monocyte chemoattractant protein-1 level in the lungs of animals randomized to hyperoxia with high tidal volume ventilation was significantly elevated, compared to all other groups (p < 0.05). Myeloperoxidase assayed in lung homogenate was found to be significantly higher in nonventilated animals exposed to hyperoxia (p < 0.01). Only in animals previously exposed to hyperoxia did the addition of high tidal volume ventilation further increase the level of myeloperoxidase present (p < 0.05). Pulmonary vascular resistance was significantly elevated after 4 h of mechanical ventilation compared to 1 h (p < 0.001). CONCLUSIONS: We conclude that in neonatal piglets undergoing hyperoxic stress, superimposition of high tidal volume ventilation exacerbates the lung inflammation as assessed by lung monocyte chemoattractant protein-1 and level of myeloperoxidase.
Asunto(s)
Hiperoxia/fisiopatología , Neumonía/etiología , Neumonía/fisiopatología , Volumen de Ventilación Pulmonar/fisiología , Animales , Animales Recién Nacidos , Líquido del Lavado Bronquioalveolar/química , Quimiocina CCL2/análisis , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Regulación de la Expresión Génica , Hiperoxia/metabolismo , Interleucina-1beta/análisis , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Pulmón/irrigación sanguínea , Pulmón/metabolismo , Pulmón/patología , Masculino , Peroxidasa/genética , Peroxidasa/metabolismo , Neumonía/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Respiración Artificial/efectos adversos , Porcinos , Factor de Necrosis Tumoral alfa/análisis , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Resistencia Vascular/fisiologíaRESUMEN
The peptides platelet-derived growth factor-A (PDGF-A) and especially -B have important roles in lung development. The effect of hyperoxic exposure with and without inhaled nitric oxide (iNO) on lung expression of PDGF and its receptors is unknown. We hypothesized that hyperoxia exposure would suppress mRNA expression and protein production of these ligands and their receptors. The addition of iNO to hyperoxia may further aggravate the effects of hyperoxia. Thirteen-day-old piglets were randomized to breathe 1) room air (RA); 2) 0.96 fraction of inspired oxygen (O2), or 3) 0.96 fraction of inspired oxygen plus 50 ppm of NO (O2+NO), for 5 d. Lungs were preserved for mRNA, Western immunoblot, and immunohistochemical analyses for PDGF-A and -B and their receptors PDGFR-alpha and -beta. PDGF-B mRNA expression was greater than that of PDGF-A or PDGFR-alpha and -beta in RA piglet lungs (p<0.05). Hyperoxia with or without iNO reduced lung PDGF-B mRNA and protein expression relative to the RA group lungs (p<0.01). PDGF-B immunostain intensity was significantly increased in the alveolar macrophages, which were present in greater numbers in the hyperoxia-exposed piglet lungs, with or without NO (p<0.01). PDGFR-beta immunostaining was significantly increased in airway epithelial cells in O2- and O2+NO-exposed piglets. PDGF-A and PDGFR-alpha immunostain intensity and distribution pattern were unchanged relative to the RA group. Sublethal hyperoxia decreases PDGF-B mRNA and protein expression but not PDGF-A or their receptors in piglet lungs. iNO neither aggravates nor ameliorates this effect.
Asunto(s)
Pulmón/metabolismo , Óxido Nítrico/metabolismo , Oxígeno/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Proteínas Proto-Oncogénicas c-sis/metabolismo , Receptores del Factor de Crecimiento Derivado de Plaquetas/metabolismo , Secuencia de Aminoácidos , Animales , Humanos , Pulmón/citología , Ratones , Datos de Secuencia Molecular , Factor de Crecimiento Derivado de Plaquetas/genética , Proteínas Proto-Oncogénicas c-sis/genética , ARN Mensajero/metabolismo , Distribución Aleatoria , Alineación de Secuencia , PorcinosRESUMEN
In this retrospective study, we tested the following hypotheses: rates of severe intraventricular hemorrhage (SIVH) and early neonatal survival are similar among extremely low birth weight (ELBW) infants treated with combination prophylaxis of phenobarbital and indomethacin compared with phenobarbital alone or no prophylaxis; and rates of patent ductus arteriosus (PDA) and necrotizing enterocolitis (NEC) are similar among indomethacin-exposed and nonexposed ELBW infants. Data were abstracted on 265 ELBW infants admitted into a level 3 neonatal intensive care unit from 1994 through 2002. Combination prophylaxis neither reduced the odds ratio (OR) of SIVH (OR = 1.53; 95% confidence interval [CI], 0.43 to 1.16) versus phenobarbital (OR = 2.91; 95% CI, 0.91 to 9.27 versus none (OR = 1; 95% CI, reference) nor increased the odds of early neonatal survival (OR = 0.72; 95% CI, 0.17 to 3.09 for combination prophylaxis versus OR = 0.66; 95% CI, 0.16 to 2.67 for phenobarbital versus OR = 1; 95% CI, reference for none). Indomethacin exposure reduced the odds of PDA (OR = 0.35; 95% CI, 0.17 to 0.75) without increasing the risk of NEC (OR = 1.37; 95% CI, 0.60 to 3.12). In conclusion, combination prophylaxis does not improve SIVH and early neonatal survival outcomes. Early exposure to indomethacin offers some benefits without any added risks.
Asunto(s)
Anticonvulsivantes/uso terapéutico , Indometacina/uso terapéutico , Recién Nacido de muy Bajo Peso , Hemorragias Intracraneales/prevención & control , Fenobarbital/uso terapéutico , Resultado del Embarazo , Antiinflamatorios no Esteroideos , Quimioterapia Combinada , Enterocolitis Necrotizante/prevención & control , Femenino , Humanos , Recién Nacido , Modelos Logísticos , Embarazo , Estudios RetrospectivosRESUMEN
OBJECTIVE: The pathogenesis of chronic lung disease (CLD) involves inflammation with proteolytic damage to lung extracellular matrix. Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases that, acting in concert with their tissue inhibitors, tightly orchestrate extracellular matrix morphogenesis and repair after injury. Imbalances in their levels relative to that of their inhibitors have been implicated in diseases characterized by matrix disruption and remodeling. We investigated the possibility that imbalances in MMP-9 and MMP-2 relative to their tissue inhibitor of metalloproteinase-1 (TIMP-1) and TIMP-2, respectively, in tracheal aspirates of preterm infants may be involved in the development of CLD. METHODS: Serial tracheal aspirates collected from birth until extubation in 49 ventilated preterm infants (24-32 weeks' gestations) were analyzed for MMP-2, MMP-9, TIMP-1, and TIMP-2. Data normalized by TA values of free secretory component of immunoglobulin A were compared for CLD (n = 22) versus no CLD (n = 27). Also, known clinical predictors of CLD (gestational age, birth weight, and sex) were assessed for both groups. Association of predictors with the outcome CLD was assessed by logistic regression. RESULTS: Mean gestational age was lower in CLD infants, but birth weight and gender were comparable for both groups. CLD infants had significantly lower TIMP-1 level with higher MMP-9/TIMP-1 ratio during the first 2 weeks of life and low TIMP-2 and MMP-2 levels during the first 3 days of life compared with no-CLD infants. Logistic regression analysis indicated that the findings are predictive of CLD. CONCLUSIONS: We conclude that low tracheal aspirate levels of TIMPs, with a high MMP-9/TIMP-1 ratio early in life, are associated with subsequent development of CLD.
Asunto(s)
Displasia Broncopulmonar/etiología , Metaloproteinasa 2 de la Matriz/análisis , Metaloproteinasa 9 de la Matriz/análisis , Inhibidor Tisular de Metaloproteinasa-1/análisis , Inhibidor Tisular de Metaloproteinasa-2/análisis , Tráquea/química , Femenino , Humanos , Recién Nacido , Recien Nacido Prematuro , Modelos Logísticos , Masculino , Respiración Artificial , Factores de RiesgoRESUMEN
Microvascular development is critical for normal lung maturation. The aims of this study were (1) to quantitatively and qualitatively assess lung microvascular growth in the human fetus, from 22 to 40 weeks' gestation, and (2) to compare development in these infants to those with mild, moderate and severe chronic lung disease (CLD). Using 1- and 4-microm thick sections and electron microscopy, lungs were morphometrically assessed for surface density of distal air spaces; volume density of parenchymal vessels having an air-blood barrier (ABB); percent of distal air space wall having an ABB, and capillary loading, defined as ABB/mm2 of epithelial surface area. The percent of vessels with ABB increased in controls during development in parallel with increasing lung parenchyma. Infants with severe CLD had fewer ABBs and less capillary loading than controls up to 34 weeks' post-conceptional age (PCA), but by 36-40 weeks, showed catch-up growth. Microvasculature vessel diameter, septal thickness, and air sac diameter at 36-40 weeks' PCA were increased with severe CLD, and vessels were more distant from the air surface. We conclude that infants with severe CLD have both stunted secondary septation and microvascular development, but over time, the primary septal wall adapts by thinning and increasing the number of ABBs, thereby taking on the function of secondary septa.
Asunto(s)
Adaptación Fisiológica , Enfermedades Pulmonares/fisiopatología , Pulmón/irrigación sanguínea , Microcirculación/embriología , Microcirculación/crecimiento & desarrollo , Capilares/ultraestructura , Enfermedad Crónica , Edad Gestacional , Humanos , Recién Nacido , Recien Nacido Prematuro , Pulmón/crecimiento & desarrollo , Pulmón/patología , Enfermedades Pulmonares/patología , Microcirculación/patología , Microscopía ElectrónicaRESUMEN
Because echocardiographic studies on infants with chronic lung disease (CLD) suggest that pulmonary hypertension (PH) may contribute to its severity, we studied acinar arterial walls in the following surfactant-era infants: controls (n=38): 22-41 weeks of gestational age (GA), exposed briefly to oxygen and positive pressure ventilation, died within 48 hr of birth; prolonged rupture of fetal membranes (PROM) and persistent pulmonary hypertension (PPHN) (n=17); and SCORE (integrated area under curve of average daily FiO2 x average daily MAP) groups (<20, 20-69, and 70-500; mild, moderate, and severe clinical lung disease, respectively, n=35): 23-30 weeks GA, lived 7-79 days. Lungs were stained for elastic tissue and smooth muscle actin. Vessels were assessed for percent of vessel circumference with smooth muscle, extent of elastic laminae in the walls, and percent arterial wall thickness (%AWT) at three levels: terminal to respiratory bronchiole transition (TRB), alveolar duct, and saccule. At the alveolar ductal and saccular levels, percent arterial wall thickness (%AWT) in mild CLD (SCORE < 20) was less than controls (P < 0.05) and those with more severe CLD (SCORE 70-500), indicating that normal postnatal arterial wall thinning may be delayed, or there is remodeling associated with increased %AWT. Severe CLD infants also had a significantly higher percent of circumferential actin than those with milder disease (SCORE < or = 69) and controls. In moderate and severe CLD, there was an increase in extent of the elastic laminae compared to controls and mild CLD. These changes were also significantly greater in PROM and PPHN infants compared to even severe CLD. We conclude that PH is a real possibility in severe CLD infants after discharge at 36 weeks. Grading the severity of CLD at discharge, and echocardiographic studies, may guide subsequent oxygen therapy.
Asunto(s)
Arterias/patología , Enfermedades del Prematuro/patología , Enfermedades Pulmonares/patología , Pulmón/irrigación sanguínea , Pulmón/patología , Actinas/análisis , Displasia Broncopulmonar/tratamiento farmacológico , Displasia Broncopulmonar/patología , Enfermedad Crónica , Endotelio Vascular/patología , Femenino , Humanos , Recién Nacido , Posmaduro , Enfermedades del Prematuro/tratamiento farmacológico , Enfermedades Pulmonares/tratamiento farmacológico , Músculo Liso Vascular/patología , Tamaño de los Órganos , Síndrome de Circulación Fetal Persistente/patología , Surfactantes Pulmonares/uso terapéutico , Respiración ArtificialRESUMEN
Inflammation is strongly linked to the pathogenesis of chronic lung disease of prematurity (CLD). Premature gas-breathing of ambient or supplemental oxygen in a host with relatively deficient and poorly inducible antioxidant defenses may itself be injurious, and further amplified by mechanical stretch injury in the surfactant-insufficient lung.1 Cellular injury provokes an inflammatory response.Since inflammation is often detected at birth in the lungs of newborns who later develop CLD,2 it has been an attractive strategy to abrogate inflammation, but the arsenal is limited. Glucocorticoids have been widely used but are acknowledged to be potentially harmful to neurologic and somatic development, and are not recommended outside controlled trials.3 The number that benefit is comparable to the number harmed, according to meta-analysis.4 More specific blockade of harmful inflammation could overcome this obstacle. Examination of the inflammatory pathways that initiate and propagate lung injury and subsequent abnormal development points to promising new strategies that may one day be tailored to individual patients.
Asunto(s)
Movimiento Celular/fisiología , Recien Nacido Prematuro , Leucocitos/fisiología , Enfermedades Pulmonares/prevención & control , Enfermedades Pulmonares/fisiopatología , Neumonía/fisiopatología , Neumonía/terapia , Enfermedad Crónica , Humanos , Recién Nacido , Enfermedades Pulmonares/etiología , Neumonía/complicacionesRESUMEN
OBJECTIVE: Infants with chronic lung disease (CLD) have an arrest of primary and secondary septation. We hypothesized that this may be related to damage or abnormal development of lung collagen secondary to positive pressure ventilation. Our aims were to identify the sites and quantity of collagen in control infants 22 to 72 weeks' postconceptional age and compare these with infants with various degrees of severity of CLD. METHODS: The controls were 22 to 42 weeks' gestation (n = 30), received minimal ventilator care, and died within 48 hours of birth, plus 5 term infants who died at 43 to 72 weeks' postconceptional age from nonpulmonary causes. Infants who were 23 to 30 weeks' gestation, were at risk for CLD, and lived 5 to 94 days (n = 33) were separated into 3 groups on the basis of respiratory score (score group; the integrated area under the curve of the average daily fraction of inspired oxygen x mean airway pressure [cm H2O] over the number of days lived). The score groups, <20, 20 to 69, and 70 to 500, related clinically to mild to moderate and severe lung disease. The lungs were tracheally perfused and formalin fixed. Total lung volume was determined by water displacement. The paraffin-embedded lung blocks were sectioned 5 micro m thick, stained with Gomori's reticulum stain, hematoxylin and eosin, and immunohistochemically for collagen IV. The parenchyma was point-counted, and the volume density of collagen was measured. The chord diameter of the peripheral airway saccules and alveoli was measured. Descriptive collagen data were assessed on en face 40- micro m-thick sections through the alveolar or saccular walls on all infants at risk for CLD and in selected controls. RESULTS: In the controls, the volume density of collagen decreased from a maximum of 9% at 22 weeks to 5% at term and 72 weeks. With Scores < or =69, the fraction of collagen was similar to controls, but in infants with scores 70 to 500, it was increased relative to controls. However, when collagen was expressed as the volume density of interstitial tissue, ie, excluding parenchymal air space, it increased from a low of 5% at 22 weeks to 25% at 72 weeks. In infants with scores 70 to 500, 79% of infants had collagens greater than controls. Saccular and alveolar diameter increased from 40 micro m at 23 weeks to 100 microm at 72 weeks. Most infants with severe CLD (scores > or =70) had diameters more than twice that of controls at the same age. The total lung parenchymal collagen had a similar pattern as the volume density of collagen in interstitial tissue, increasing from 0.4 cm3 at 23 weeks to 9.7 cm3 at 72 weeks in the controls. Eighty-five percent of infants with scores 70 to 500 had total parenchymal collagen greater than the 95% confidence interval of the controls. With en face sections, a fine collagen mesh was seen at 23 weeks, which progressively increased in fiber size and quantity until 72 weeks. With severe CLD, the secondary collagen fibers in the saccular wall were thickened, tortuous, and disorganized relative to same-aged controls. Under 30 weeks, in the controls, the interstitium contained a wide, delicate network of interconnected collagen fibers. After positive pressure ventilation, some saccules markedly increased their diameter, which compressed and obliterated the interstitial network. In contrast with severe CLD, the interstitium was wide, with coarse wavy collagen fibers. CONCLUSIONS: Parenchymal collagen increases throughout development. Before 30 weeks, there is a delicate complex interstitial collagen network, which may be important for primary septation and subsequent normal development. Positive pressure ventilation, if excessive, and depending on lung maturity and disease state, over a short time can severely compress the interstitium and damage this collagen network and prevent normal primary septation and arrest or distort future lung development. With severe CLD, distal air space diameter increases. There is a failure of primary and secondary septation, arrested lung development and remodeling, with thickened cnt and remodeling, with thickened collagenous saccular walls, and a wide interstitium with increased quantity and size of collagen fibers that can affect the mechanics of ventilation. We conclude that normal lung development is dependent on a normal interstitium and, perhaps, collagen architecture and that origins of CLD begin early in the course of positive pressure ventilation.
Asunto(s)
Colágeno/metabolismo , Enfermedades Pulmonares/embriología , Enfermedades Pulmonares/metabolismo , Displasia Broncopulmonar/embriología , Displasia Broncopulmonar/metabolismo , Displasia Broncopulmonar/mortalidad , Niño , Preescolar , Enfermedad Crónica , Colágeno/química , Colágeno/inmunología , Colágeno Tipo IV/inmunología , Colágeno Tipo IV/metabolismo , Humanos , Inmunohistoquímica , Lactante , Recién Nacido , Recien Nacido Prematuro , Pulmón/anomalías , Pulmón/embriología , Pulmón/metabolismo , Pulmón/patología , Enfermedades Pulmonares/mortalidad , Enfermedades Pulmonares/patología , Mediciones del Volumen Pulmonar/métodos , Respiración con Presión Positiva/efectos adversos , Respiración con Presión Positiva/métodos , Síndrome de Dificultad Respiratoria del Recién Nacido/terapiaRESUMEN
Pulmonary hyperoxic injury manifests as widespread alveolar-epithelial and microvascular endothelial cell necrosis, resolution of which requires angiogenesis. We investigated the hypothesis that inhaled nitric oxide (iNO) and hyperoxia each decreases lung vascular endothelial growth factor (VEGF) expression but increases endostatin and that concurrent administration of both gases will show a greater effect. Piglets were randomized to breathe for 5 d room air (RA); RA + NO (RA + 50 ppm NO), O(2) (hyperoxia, F(I)O(2) >0.96), O(2) + NO, or O(2) + NO + REC (O(2) + NO plus recovery in 50% O(2) for 72 h. After the piglets were killed, we measured lung capillary leak, VEGF mRNA, VEGF, and endostatin protein in homogenates, plasma, and lavage. VEGF mRNA decreased significantly with O(2) and O(2) + NO compared with breathing RA (p < or = 0.05). VEGF protein declined in the experimental groups with a significant reduction in the recovery group compared with the RA group (p < or = 0.05). Similar but more dramatic, endostatin declined in all groups relative to the RA group (p < 0.001). Lavage fluid VEGF protein and lung capillary leak rose significantly with O(2) and O(2) + NO compared with RA, but endostatin was unchanged. At 72 h of recovery from hyperoxia, VEGF mRNA and lavage fluid VEGF but not lung VEGF protein had normalized. Hyperoxia and iNO suppresses lung endostatin expression, but iNO unlike hyperoxia alone does not alter lung VEGF production. Hyperoxia paradoxically raises lavageable VEGF levels. This latter effect and that on VEGF mRNA level but not protein is abrogated by recovery in reduced F(I)O(2) for 72 h.
Asunto(s)
Colágeno/metabolismo , Factores de Crecimiento Endotelial/sangre , Hiperoxia/metabolismo , Péptidos y Proteínas de Señalización Intercelular/sangre , Pulmón/metabolismo , Linfocinas/sangre , Óxido Nítrico/farmacología , Fragmentos de Péptidos/metabolismo , Administración por Inhalación , Animales , Apoptosis , Líquido del Lavado Bronquioalveolar , Permeabilidad Capilar/efectos de los fármacos , Endostatinas , Factores de Crecimiento Endotelial/genética , Expresión Génica/efectos de los fármacos , Hiperoxia/patología , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intercelular/genética , Pulmón/efectos de los fármacos , Pulmón/patología , Linfocinas/genética , Masculino , Oxígeno/farmacología , ARN Mensajero/análisis , Porcinos , Factor A de Crecimiento Endotelial Vascular , Factores de Crecimiento Endotelial VascularRESUMEN
The peptide endothelin-1 (ET-1) plays an unknown role in the pathogenesis and progression of two important neonatal pulmonary disorders, chronic lung disease (CLD) of prematurity and persistent pulmonary hypertension of the newborn (PPHN). Inhaled nitric oxide (INO) is a proven vasodilator therapy in PPHN and is an experimental therapy in CLD. We sought to determine the effects, if any, of the interaction of inhaled INO with ET-1 in these two separate disorders. Infants (n=21) with PPHN (mean gestation age, 39.4 weeks; mean birth weight, 3470 g) were treated with INO. All infants were <72 h of age at baseline. Plasma obtained at baseline and after 24 h of INO therapy was assessed for ET-1. The change in ET-1 levels with INO was inversely correlated with change in arterial partial pressure of O(2) (r=-0.71, P=0.0003). A separate group of 33 patients with CLD (mean gestational age, 27 weeks; mean birth weight, 740 g; mean age, 19 days) had tracheal aspirate levels of ET-1 obtained before, during, and after 7 days' administration of INO. Values were normalized by soluble secretory component of IgA. Tracheal aspirate ET-1 levels were detectable before INO therapy. There was no significant change during or after treatment with INO. There was not a significant correlation between baseline fractional inspired O(2) and ET-1 levels. There was a non-significant trend in the correlation between the change in ET-1 and the change in interleukin-8 levels in tracheal aspirate. This report confirms the presence of ET-1 in tracheal aspirate of premature infants who are developing CLD and reaffirms the presence of ET-1 in plasma of infants with PPHN. Short-term INO therapy was associated with a decrease in plasma ET-1 levels in PPHN, but did not affect tracheal aspirate ET-1 in CLD. Given the vasconstrictive, profibrotic, and proinflammatory properties of ET-1, specific ET-1 receptor antagonists could be considered as candidates for trials as adjunct therapy in either or both of these disorders.