Cleide Suguihara

Inhibition of the SDF-1/CXCR4 Axis Attenuates Neonatal Hypoxia-Induced Pulmonary Hypertension

Exposure of the neonatal lung to chronic hypoxia produces significant pulmonary vascular remodeling, right ventricular hypertrophy, and decreased lung alveolarization. Given recent data suggesting that stem cells could contribute to pulmonary vascular remodeling and right ventricular hypertrophy, we tested the hypothesis that blockade of SDF-1 (stromal cell–derived factor 1), a key stem cell mobilizer or its receptor, CXCR4 (CXC chemokine receptor 4), would attenuate and reverse hypoxia-induced cardiopulmonary remodeling in newborn mice. Neonatal mice exposed to normoxia or hypoxia were randomly assigned to receive daily intraperitoneal injections of normal saline, AMD3100, or anti–SDF-1 antibody from postnatal day 1 to 7 (preventive strategy) or postnatal day 7 to 14 (therapeutic strategy). As compared to normal saline, inhibition of the SDF-1/CXCR4 axis significantly improved lung alveolarization and decreased pulmonary hypertension, right ventricular hypertrophy, vascular remodeling, vascular cell proliferation, and lung or right ventricular stem cell expressions to near baseline values. We therefore conclude that the SDF-1/CXCR4 axis both prevents and reverses hypoxia-induced cardiopulmonary remodeling in neonatal mice, by decreasing progenitor cell recruitment to the pulmonary vasculature, as well as by decreasing pulmonary vascular cell proliferation. These data offer novel insights into the role of the SDF-1/CXCR4 axis in the pathogenesis of neonatal hypoxia-induced cardiopulmonary remodeling and have important therapeutic implications.

Long-term reparative effects of mesenchymal stem cell therapy following neonatal hyperoxia-induced lung injury.

Background:Mesenchymal stem cell (MSC) therapy may prevent neonatal hyperoxia-induced lung injury (HILI). There are, however, no clear data on the therapeutic efficacy of MSC therapy in established HILI, the duration of the reparative effects, and the exact mechanisms of repair. The main objective of this study was to evaluate whether the long-term reparative effects of a single intratracheal (IT) dose of MSCs or MSC-conditioned medium (CM) are comparable in established HILI.Methods:Newborn rats exposed to normoxia or hyperoxia from postnatal day (P)2)–P16 were randomized to receive IT MSCs, IT CM, or IT placebo (PL) on P9. Alveolarization and angiogenesis were evaluated at P16, P30, and P100.Results:At all time periods, there were marked improvements in alveolar and vascular development in hyperoxic pups treated with MSCs or CM as compared with PL. This was associated with decreased expression of inflammatory mediators and an upregulation of angiogenic factors. Of note, at P100, the improvements were more substantial with MSCs as compared with CM.Conclusion:These data suggest that acute effects of MSC therapy in HILI are mainly paracrine mediated; however, optimum long-term improvement following HILI requires treatment with the MSCs themselves or potentially repetitive administration of CM.

Effect of respiratory syncytial virus on apnea in weanling rats

Apnea is a common complication of respiratory syncytial virus (RSV) infection in young infants. The purpose of this study was to determine whether this infection affects apnea triggered by sensorineural stimulation in weanling rats. We also studied which neurotransmitters are involved in this response and whether passive prophylaxis with a specific neutralizing antibody (palivizumab) confers protection against it. Weanling rats were inoculated intranasally with RSV, adenovirus, or virus-free medium. Changes in respiratory rate and apnea in response to nerve stimulation with increasing doses of capsaicin were measured by plethysmography. Capsaicin-induced apnea was significantly longer in RSV-infected rats at postinoculation days 2 (upper airways infection) and 5 (lower airways infection), and apnea-related mortality occurred only in the RSV-infected group. Reduction in the duration of apnea was observed after selective inhibition of central γ-aminobutyric acid (GABA) type A receptors and neurokinin type 1 receptors for substance P. Prophylactic palivizumab protected against apnea and apnea-related mortality. These results suggest that sensorineural stimulation during RSV infection is associated with the development of apnea and apnea-related death in early life, whose mechanism involves the release of GABA acting on central GABA type A receptors and substance P acting on neurokinin type 1 receptors.

Brainstem Amino Acid Neurotransmitters and Ventilatory Response to Hypoxia in Piglets

The ventilatory response to hypoxia is influenced by the balance between inhibitory (GABA, glycine, and taurine) and excitatory (glutamate and aspartate) brainstem amino acid (AA) neurotransmitters. To assess the effects of AA in the nucleus tractus solitarius (NTS) on the ventilatory response to hypoxia at 1 and 2 wk of age, inhibitory and excitatory AA were sampled by microdialysis in unanesthetized and chronically instrumented piglets. Microdialysis samples from the NTS area were collected at 5-min intervals and minute ventilation (VE), arterial blood pressure (ABP), and arterial blood gases (ABG) were measured while the animals were in quiet sleep. A biphasic ventilatory response to hypoxia was observed in wk 1 and 2, but the decrease in VE at 10 and 15 min was more marked in wk 1. This was associated with an increase in inhibitory AA during hypoxia in wk 1. Excitatory AA levels were elevated during hypoxia in wk 1 and 2. Changes in ABP, pH, and ABG during hypoxia were not different between weeks. These data suggest that the larger depression in the ventilatory response to hypoxia observed in younger piglets is mediated by predominance of the inhibitory AA neurotransmitters, GABA, glycine, and taurine, in the NTS.

Effect of Cyclooxygenase Inhibition on Retinal and Choroidal Blood Flow during Hypercarbia in Newborn Piglets

The effect of the cyclooxygenase inhibitor, indomethacin, on choroidal (ChBF) and retinal (RBF) blood flow during hypercarbia was examined in 16 paralyzed and mechanically ventilated piglets less than 8 d old. The animals were randomly assigned to a control group (mean ± SEM: wt, 1.66 ± 0.1 kg; n = 8) that received a placebo infusion or to an indomethacin treatment group (wt, 1.68 ± 0.2 kg; n = 8) that received an infusion of indomethacin (5 mg/kg i.v. over 30 min). Baseline ChBF and RBF were measured using radiolabeled microspheres in room air before and 15 min after the administration of placebo or indomethacin. Animals were then exposed to 30 min of hypercarbia (6–7% CO2, arterial CO2 pressure 8–10 kPa) and measurements were repeated. There were no significant differences in RBF between control (40 ± 3 mL/ min/100 g) and indomethacin-treated animals (40 ± 3 mL/ min/100 g) before administration of placebo or indomethacin. However, RBF decreased significantly in the indomethacin-treated animals (28 ± 2 mL/min/100 g) compared to the control group (42 ± 4 mL/min/100 g) 15 min after administration of placebo or indomethacin. Furthermore, an increase in RBF occurred during hypercarbia in the control group (86 ± 6 mL/min/100 g), but this change was blunted in the indomethacin-treated animals (33 ± 5 mL/min/100 g) (p < 0.001). In contrast, ChBF did not differ significantly between the control and indomethacin groups during the periods studied. These results suggest that the increase in RBF during hypercarbia is at least partially mediated by cyclooxygenase by-products of ara-chidonic acid metabolism. The change in ChBF appears to be less influenced by these mediators.

Toll-like receptor 4–deficient mice are resistant to chronic hypoxia-induced pulmonary hypertension

ABSTRACT Current data suggest that Toll-like receptor 4 (TLR4), a key molecule in the innate immune response, may also be activated following tissue injury. Activation of this receptor is known to induce the production of several proinflammatory cytokines. Given that pulmonary inflammation has been shown to be a key contributor to chronic hypoxia-induced pulmonary vascular remodeling, the authors hypothesized that TLR4-deficient mice would be less susceptible to pulmonary hypertension (PH) as compared to mice with intact TLR4. TLR4-deficient and TLR4-intact strains of inbred mice were exposed to 4, 8, and 16 weeks of hypoxia (0.10 FiO2) or normoxia (0.21 FiO2) in a normobaric chamber. After chronic hypoxic exposure, TLR4-intact mice developed significant PH evidenced by increased right ventricular systolic pressure, right ventricular hypertrophy, and pulmonary artery medial thickening. In contrast, TLR4-deficient mice had no significant change in any of these parameters and this was associated with decreased pulmonary vascular inflammatory response as compared to the TLR4-intact mice. These results suggest that TLR4 deficiency may decrease the susceptibility to developing PH by attenuating the pulmonary vascular inflammatory response to chronic hypoxia.

High tidal volume ventilation activates Smad2 and upregulates expression of connective tissue growth factor in newborn rat lung.

High tidal volume (VT) ventilation plays a key role in ventilator induced lung injury and bronchopulmonary dysplasia. However, little is known about the effect of high VT on expression of growth factors that are critical to lung development. In a previous study, we demonstrated that connective tissue growth factor (CTGF) inhibits branching morphogenesis. In this study, we investigated the effect of high VT on CTGF expression in newborn rat lungs. Newborn rats were ventilated with normal VT (10 mL/kg) or high VT (25 mL/kg) for 6 h. Nonventilated animals served as controls. We found that high VT upregulated CTGF expression. To identify the potential signaling pathways mediating high VT induction of CTGF, newborn rats were ventilated with high VT for 1 or 3 h. Temporal expression of TGF-βs, p-Smad2, Smad7, and CTGF was analyzed. High VT ventilation did not change gene expression of TGF-βs and Smad7 but induced rapid and sustained expression of p-Smad2 that precedes increased CTGF expression. CTGF and p-Smad2 were localized in bronchiolar epithelial cells, alveolar walls and septa. These data suggest that high Vt ventilation activates the Smad2 pathway, which may be responsible for downstream induction of CTGF expression in newborn rat lungs.

Effects of respiratory mechanical unloading on thoracoabdominal motion in meconium-injured piglets and rabbits

Impaired pulmonary mechanics can cause chest wall distortion (CWD) so that work of breathing is dissipated in deforming the rib cage. We hypothesized that respiratory mechanical unloading as a technique of assisted mechanical ventilation would reduce CWD in animals with injured lungs. We studied five piglets and five adult rabbits to test across different ages and chest configurations. As a result of intratracheal meconium instillation, lung compliance decreased from 21 (median; range 17-35) to 9.5 (6.7-14) mL/kPa/kg in rabbits and from 26 (18-31) to 7.9 (4.9-11) in piglets. Airway resistance increased from 5.0 (4.6-6.1) to 6.9 (5.8-7.9) kPa/L/s in rabbits only. Respiratory inductive plethysmography was used to measure the phase shift between the rib cage and abdominal compartment movements and the total compartmental displacement ratio. We aimed at unloading at least three-fourths of lung elastance in all animals and 2.0 kPa/L/s of resistance in rabbits. Elastic unloading decreased the phase shift in all but one animal. It reduced the total compartmental displacement ratio from 1.27 (1.14-3.73) to 1.16(1.02-1.82) in piglets and from 1.77 (1.45-5.24) to 1.37 (1.11-4.78) in rabbits. The inspiratory rib cage expansion increased, whereas abdominal expansion did not. The tidal esophageal pressure deflection decreased. Tidal volume increased, whereas respiratory rate remained unaffected so that the partial pressure of arterial CO2 decreased. Resistive unloading as an adjunct to elastic unloading further reduced CWD and induced a more rapid, shallower breathing. We conclude that respiratory unloading as a mechanical support to spontaneous breathing reduces CWD. We speculate that the decrease in CWD increases ventilatory efficiency for a given diaphragmatic effort.

Hemodynamic and Ventilatory Effects of High-Frequency Jet and Conventional Ventilation in Piglets with Lung Lavage

The cardiovascular and ventilatory effects of high-frequency jet ventilation (HFJV) and conventional ventilation (CV) were evaluated in a saline lung lavage model in piglets. After saline lavage and stabilization on CV, animals were randomized to either mode of ventilation for 4 h. Arterial blood gases, cardiac output, mean pulmonary and arterial blood pressures, and pulmonary and systemic vascular resistances were compared between groups. Alveolar-arterial oxygen gradient (AaDO2) was lower in the HFJV than in the CV group after 3 h of ventilation (p less than 0.04). The peak inspiratory pressure necessary to maintain PaCO2 within the normal range was approximately half as much in the HFJV group as in the CV group (p less than 0.005) throughout the entire study period. Mean airway pressure, cardiac output, mean pulmonary and arterial blood pressures as well as pulmonary and systemic vascular resistances were not statistically different between groups. These results suggest that in this model, HFJV produces better oxygenation with lower peak airway pressures compared to CV without producing hemodynamic compromise.

The Effect of Escherichia coli Endotoxin Infusion on the Ventilatory Response to Hypoxia in Unanesthetized Newborn Piglets

To determine the effects of endotoxemia on the neonatal ventilatory response to hypoxia, 17 chronically instrumented and unanesthetized newborn piglets (≤7 d) were studied before and 30 min after the administration of Escherichia coli O55:B5 endotoxin (n = 8) or normal saline (n = 9). Minute ventilation, oxygen consumption, heart rate, arterial blood pressure, and blood gases were measured during normoxia and 10 min of hypoxia (fraction of inspired oxygen, 0.10). Basal ventilation was not modified by E. coli endotoxin infusion (mean ± SE, 516 ± 49 versus 539 ± 56 mL/min/kg), but the ventilatory response to hypoxia was markedly attenuated at 1 min (955 ± 57 versus 718 ± 97 mL/min/kg, p < 0.002, saline versus endotoxin) and at 10 min (788 ± 51 versus 624 ± 66 mL/min/kg, p < 0.002). A larger decrease in oxygen consumption was observed during hypoxia and endotoxemia (6.3 ± 2.8 versus 18.3 ± 2.7%, p < 0.03, pre-versus post-endotoxin). A significant correlation was demonstrated between the changes in minute ventilation and oxygen consumption with hypoxia during endotoxemia (r = 0.9, p < 0.002). The ventilatory response to hypoxia was not modified by the saline infusion. These data show a significant attenuation in the ventilatory response to hypoxia during E. coli endotoxemia. This decrease in ventilation was associated with a significant decrease in the metabolic rate during hypoxia and endotoxemia.