Sustained hypercapnia induces cerebral microvascular degeneration in the immature brain through induction of nitrative stress

摘要

Hypercapnia is regularly observed in chronic lung disease, such as bronchopulmonary dysplasia in preterm infants. Hypercapnia results in increased nitric oxide synthase activity and in vitro formation of nitrates. Neural vasculature of the immature subject is particularly sensitive to nitrative stress. We investigated whether exposure to clinically relevant sustained high CO2 causes microvascular degeneration in the newborn brain by inducing nitrative stress, and whether this microvascular degeneration has an impact on brain growth. Newborn rat pups were exposed to 10% CO2 as inspired gas (PaCO2 = 60–70 mmHg) starting within 24 h of birth until postnatal day 7 (P7). Brains were notably collected at different time points to measure vascular density, determine brain cortical nitrite/nitrate, and trans-arachidonic acids (TAAs; products of nitration) levels as effectors of vessel damage. Chronic exposure of rat pups to high CO2 (PaCO2 ≈ 65 mmHg) induced a 20% loss in cerebrovascular density at P3 and a 15% decrease in brain mass at P7; at P30, brain mass remained lower in CO2-exposed animals. Within 24 h of exposure to CO2, brain eNOS expression and production of nitrite/nitrate doubled, lipid nitration products (TAAs) increased, and protein nitration (3-nitrotyrosine immunoreactivity) was also coincidently augmented on brain microvessels (lectin positive). Intracerebroventricular injection of TAAs (10 μM) replicated cerebrovascular degeneration. Treatment of rat pups with NOS inhibitor (l-Nω-nitroarginine methyl ester) or a peroxynitrite decomposition catalyst (FeTPPS) prevented hypercapnia-induced microvascular degeneration and preserved brain mass. Cytotoxic effects of high CO2 were reproduced in vitro/ex vivo on cultured endothelial cells and sprouting microvessels. In summary, hypercapnia at values frequently observed in preterm infants with chronic lung disease results in increased nitrative stress, which leads to cerebral cortical microvascular degeneration and curtails brain growth.