Effects of hypercapnia and NO synthase inhibition in sustained hypoxic pulmonary vasoconstriction

Background

Acute respiratory disorders may lead to sustained alveolar hypoxia with hypercapnia resulting in impaired pulmonary gas exchange. Hypoxic pulmonary vasoconstriction (HPV) optimizes gas exchange during local acute (0-30 min), as well as sustained (> 30 min) hypoxia by matching blood perfusion to alveolar ventilation. Hypercapnia with acidosis improves pulmonary gas exchange in repetitive conditions of acute hypoxia by potentiating HPV and preventing pulmonary endothelial dysfunction. This study investigated, if the beneficial effects of hypercapnia with acidosis are preserved during sustained hypoxia as it occurs, e.g in permissive hypercapnic ventilation in intensive care units. Furthermore, the effects of NO synthase inhibitors under such conditions were examined.

Method

We employed isolated perfused and ventilated rabbit lungs to determine the influence of hypercapnia with or without acidosis (pH corrected with sodium bicarbonate), and inhibitors of endothelial as well as inducible NO synthase on acute or sustained HPV (180 min) and endothelial permeability.

Results

In hypercapnic acidosis, HPV was intensified in sustained hypoxia, in contrast to hypercapnia without acidosis when HPV was amplified during both phases. L-N^G-Nitroarginine (L-NNA), a non-selective NO synthase inhibitor, enhanced acute as well as sustained HPV under all conditions, however, the amplification of sustained HPV induced by hypercapnia with or without acidosis compared to normocapnia disappeared. In contrast 1400 W, a selective inhibitor of inducible NO synthase (iNOS), decreased HPV in normocapnia and hypercapnia without acidosis at late time points of sustained HPV and selectively reversed the amplification of sustained HPV during hypercapnia without acidosis. Hypoxic hypercapnia without acidosis increased capillary filtration coefficient (Kfc). This increase disappeared after administration of 1400 W.

Conclusion

Hypercapnia with and without acidosis increased HPV during conditions of sustained hypoxia. The increase of sustained HPV and endothelial permeability in hypoxic hypercapnia without acidosis was iNOS dependent.     

Inducible NOS inhibition reverses tobacco-smoke-induced emphysema and pulmonary hypertension in mice

Chronic obstructive pulmonary disease (COPD) is one of the most common causes of death worldwide. We report in an emphysema model of mice chronically exposed to tobacco smoke that pulmonary vascular dysfunction, vascular remodeling, and pulmonary hypertension (PH) precede development of alveolar destruction. We provide evidence for a causative role of inducible nitric oxide synthase (iNOS) and peroxynitrite in this context. Mice lacking iNOS were protected against emphysema and PH. Treatment of wild-type mice with the iNOS inhibitor N(6)-(1-iminoethyl)-L-lysine (L-NIL) prevented structural and functional alterations of both the lung vasculature and alveoli and also reversed established disease. In chimeric mice lacking iNOS in bone marrow (BM)-derived cells, PH was dependent on iNOS from BM-derived cells, whereas emphysema development was dependent on iNOS from non-BM-derived cells. Similar regulatory and structural alterations as seen in mouse lungs were found in lung tissue from humans with end-stage COPD.     

Detection of reactive oxygen species in isolated, perfused lungs by electron spin resonance spectroscopy

Background

Interleukin (IL)-9 is a Th2-derived cytokine with pleiotropic biological effects, which recently has been proposed as a candidate gene for asthma and allergy. We aimed to evaluate the therapeutic effect of a neutralizing anti-IL-9 antibody in a mouse model of airway eosinophilic inflammation and compared any such effect with anti-IL-5 treatment.

Methods

OVA-sensitized Balb/c mice were intraperitoneally pretreated with a single dose (100 μg) of an anti-mouse IL-9 monoclonal antibody (clone D9302C12) or its vehicle. A third group was given 50 μg of a monoclonal anti-mouse IL-5 antibody (TRFK-5) or its vehicle. Animals were subsequently exposed to OVA on five days via airways. Newly produced eosinophils were labelled using 5-bromo-2'-deoxyuridine (BrdU). BrdU⁺ eosinophils and CD34⁺ cell numbers were examined by immunocytochemistry. After culture and stimulation with OVA or PMA+IC, intracellular staining of IL-9 in bone marrow cells from OVA-exposed animals was measured by Flow Cytometry. The Mann-Whitney U-test was used to determine significant differences between groups.

Results

Anti-IL-9 significantly reduced bone marrow eosinophilia, primarily by decrease of newly produced (BrdU⁺) and mature eosinophils. Anti-IL-9 treatment also reduced blood neutrophil counts, but did not affect BAL neutrophils. Anti-IL-5 was able to reduce eosinophil numbers in all tissue compartments, as well as BrdU⁺ eosinophils and CD34⁺ progenitor cells, and in all instances to a greater extent than anti-IL-9. Also, FACS analysis showed that IL-9 is over-expressed in bone marrow CD4⁺ cells after allergen exposure.

Conclusions

Our data shows that a single dose of a neutralizing IL-9 antibody is not sufficient to reduce allergen-induced influx of newly produced cells from bone marrow to airways. However, in response to allergen, bone marrow cells over-express IL-9. This data suggest that IL-9 may participate in the regulation of granulocytopoiesis in allergic inflammation.     

Nebulization of the acidified sodium nitrite formulation attenuates acute hypoxic pulmonary vasoconstriction

Background

Generalized hypoxic pulmonary vasoconstriction (HPV) occurring during exposure to hypoxia is a detrimental process resulting in an increase in lung vascular resistance. Nebulization of sodium nitrite has been shown to inhibit HPV. The aim of this project was to investigate and compare the effects of nebulization of nitrite and different formulations of acidified sodium nitrite on acute HPV.

Methods

Ex vivo isolated rabbit lungs perfused with erythrocytes in Krebs-Henseleit buffer (adjusted to 10% hematocrit) and in vivo anesthetized catheterized rabbits were challenged with periods of hypoxic ventilation alternating with periods of normoxic ventilation. After baseline hypoxic challenges, vehicle, sodium nitrite or acidified sodium nitrite was delivered via nebulization. In the ex vivo model, pulmonary arterial pressure and nitric oxide concentrations in exhaled gas were monitored. Nitrite and nitrite/nitrate were measured in samples of perfusion buffer. Pulmonary arterial pressure, systemic arterial pressure, cardiac output and blood gases were monitored in the in vivo model.

Results

In the ex vivo model, nitrite nebulization attenuated HPV and increased nitric oxide concentrations in exhaled gas and nitrite concentrations in the perfusate. The acidified forms of sodium nitrite induced higher levels of nitric oxide in exhaled gas and had longer vasodilating effects compared to nitrite alone. All nitrite formulations increased concentrations of circulating nitrite to the same degree. In the in vivo model, inhaled nitrite inhibited HPV, while pulmonary arterial pressure, cardiac output and blood gases were not affected. All nitrite formulations had similar potency to inhibit HPV. The tested concentration of appeared tolerable.

Conclusion

Nitrite alone and in acidified forms effectively and similarly attenuates HPV. However, acidified nitrite formulations induce a more pronounced increase in nitric oxide exhalation.