Neonatal hypoxic hyperlipidemia in the rat: effects on aldosterone and corticosterone synthesis in vitro

Neonatal hypoxia increases aldosterone production and plasma lipids. Because fatty acids can inhibit aldosterone synthesis, we hypothesized that increases in plasma lipids restrain aldosteronogenesis in the hypoxic neonate. We exposed rats to 7 days of hypoxia from birth to 7 days of age (suckling) or from 28 to 35 days of age (weaned at day 21). Plasma was analyzed for lipid content, and steroidogenesis was studied in dispersed whole adrenal glands untreated and treated to wash away lipids. Hypoxia increased plasma cholesterol, triglycerides, and nonesterified fatty acids in the suckling neonatal rat only. Washing away lipids increased aldosterone production in cells from 7-day-old rats exposed to hypoxia, but not in cells from normoxic 7-day-old rats or from normoxic or hypoxic 35-day-old rats. Addition of oleic or linolenic acid to washed cells inhibited both aldosterone and corticosterone production, although cells from hypoxic 7-day-old rats were less sensitive. We conclude that hypoxia induces hyperlipidemia in the suckling neonate and that elevated nonesterified fatty acids inhibit aldosteronogenesis.     

Effects of hypoxia on the development of intestinal enzymes in neonatal and juvenile rats

Hypoxia in the neonate is known to alter the activity of hepatic and pancreatic enzymes involved in lipid and carbohydrate metabolism. The purpose of this study was to evaluate the effect of neonatal hypoxia on the activity of intestinal enzymes, and to determine whether the administration of glucocorticoids to neonates can mimic the effects of hypoxia. Hypoxia in neonatal rats (0-7 days) increased protein content, and lactase and maltase activity in the duodenal and the jejunal segments of the small intestine compared with normoxic controls. Hypoxia in juvenile rats (28-35 days) did not change these enzymes. Two weeks after returning hypoxic (0-7 days) pups to normoxia, their body weight remained lower than the age-matched controls. In the group recovering from hypoxia, sucrase, maltase, and leucine aminopeptidase activities were lower in the duodenal and the jejunal segment. Compared with controls, LDH activity was lower only in the jejunal intestine in the group recovering from hypoxia. All enzyme activities returned to control levels 3 weeks after recovery. Neonatal rats treated with dexamethasone had a decrease in body weight, but increases in sucrase and maltase activity in both the duodenal and the jejunal segment. Hypoxia in newborn rats caused a delayed maturation of small intestinal enzymes. Increases in serum glucocorticoids after hypoxic exposure probably do not play a major role in the delayed maturation of the disaccharidase activity in the small intestine.     

Effect of neonatal hypoxia on leptin, insulin, growth hormone and body composition in the rat.

The purpose of the present study was to evaluate the effect of exposure to hypoxia from birth to 7 days of age on leptin, insulin, growth hormone (GH), insulin-like growth factor-1 (IGF-1), glucose, corticosterone, body weight, and body composition in rats studied at 7 days of age and then after return to normoxia. Hypoxia for the first 7 days of life resulted in a significant decrease in plasma leptin, body weight, and an increase in corticosterone and insulin with no change in plasma glucose, GH or IGF-1. There was no significant effect of hypoxia on % lean body mass, but a small but significant increase in % body fat. Bone mineral density (BMD) was lower in 7-day-old hypoxic rats as compared to normoxic controls. All hormonal variables and BMD had normalized by 7 days after return to normoxia. However, body weight remained lower even 5 weeks after return to normoxia. We conclude that leptin is decreased during neonatal hypoxia despite no change in adiposity. Furthermore, insulin is increased probably to overcome the effects of increased counterregulatory hormones (such as corticosterone).     

Cardioprotection in chronically hypoxic rabbits persists on exposure to normoxia: role of NOS and KATP channels

Hypoxia from birth increases resistance to myocardial ischemia in infant rabbits. We hypothesized that increased cardioprotection in hearts chronically hypoxic from birth persists following development in a normoxic environment and involves increased activation of nitric oxide synthase (NOS) and ATP-dependent K (K(ATP)) channels. Resistance to myocardial ischemia was determined in rabbits raised from birth to 10 days of age in a normoxic (Fi(O(2)) = 0.21) or hypoxic (Fi(O(2)) = 0.12) environment and subsequently exposed to normoxia for up to 60 days of age. Isolated hearts (n = 8/group) were subjected to 30 min of global ischemia followed by 35 min of reperfusion. At 10 days of age, resistance to myocardial ischemia (percent recovery postischemic recovery left ventricular developed pressure) was higher in chronically hypoxic hearts (68 +/- 4%) than normoxic controls (43 +/- 4%). At 10 days of age, N(G)-nitro-L-arginine methyl ester (200 microM) and glibenclamide (3 microM) abolished the cardioprotective effects of chronic hypoxia (45 +/- 4% and 46 +/- 5%, respectively) but had no effect on normoxic hearts. At 30 days of age resistance to ischemia in normoxic hearts declined (36 +/- 5%). However, in hearts subjected to chronic hypoxia from birth to 10 days and then exposed to normoxia until 30 days of age, resistance to ischemia persisted (63 +/- 4%). L-NAME or glibenclamide abolished cardioprotection in previously hypoxic hearts (37 +/- 4% and 39 +/- 5%, respectively) but had no effect on normoxic hearts. Increased cardioprotection was lost by 60 days. We conclude that cardioprotection conferred by adaptation to hypoxia from birth persists on subsequent exposure to normoxia and is associated with enhanced NOS activity and activation of K(ATP) channels.     

Adrenocortical responses to ACTH in neonatal rats: effect of hypoxia from birth on corticosterone,StAR, and PBR

The adrenocortical response to hypoxia may be a critical component of the adaptation to this common neonatal stress. Little is known about adrenal function in vivo in hypoxic neonates. The purpose of this study was to evaluate adrenocortical responses to ACTH in suckling rat pups exposed to hypoxia from birth to 5-7 days of age compared with normoxic controls. We also evaluated potential cellular controllers of steroidogenic function in situ. In 7-day-old pups at 0800, hypoxia from birth resulted in increased basal (12.2 +/- 1.4 ng/ml; n = 12) and ACTH-stimulated (94.0 +/- 9.4 ng/ml; n = 14) corticosterone levels compared with normoxic controls (basal = 8.3 +/- 0.5 ng/ml; n = 11; stimulated = 51.3 +/- 3.8 ng/ml; n = 8). This augmentation occurred despite no significant difference in plasma ACTH levels in normoxic vs. hypoxic pups before (85 +/- 4 vs. 78 +/- 8 pg/ml) or after (481 +/- 73 vs. 498 +/- 52 pg/ml) porcine ACTH injection (20 microg/kg). This effect was similar in the afternoon at 6 days of age and even greater at 5 days of age at 0800. The aldosterone response to ACTH was not augmented by exposure to hypoxia from birth. Adrenocortical hypoxia-inducible factor (HIF)-1alpha mRNA was undetectable by RT-PCR. Steroidogenic acute regulatory (StAR) protein in adrenal subcapsules (zona fasciculata/reticularis) was augmented by exposure to hypoxia; this effect was greatest at 5 days of age. Peripheral-type benzodiazepine receptor (PBR) protein was also increased at 6 and 7 days of age in pups exposed to hypoxia from birth. We conclude that hypoxia from birth results in an augmentation of the corticosterone but not aldosterone response to ACTH. This effect appears to be mediated at least in part by an increase in controllers of mitochondrial cholesterol transport (StAR and PBR) and to occur independently of measurable changes in endogenous plasma ACTH. The augmentation of the corticosterone response to acute increases in ACTH in hypoxic pups is likely to be an important component of the overall physiological adaptation to hypoxia in the neonate.     

Acid-base balance and lactatemia in acute hypoxia in normoxic or hypoxic albino rats

Albino rats Wistar family were raised since birth in normobaric hypoxic environment (10% O2 in N2). This hypoxic animal group and a normoxic control group were subjected to acute hypoxia in two spaced tests. The rats were exposed for 15 minutes to 7% O2 and later to 5% O2 gas mixture. At the end of the test with 7% O2 the hypoxic animals since birth showed a smaller quantity of blood lactate and their acid-base balance was more acid when compared to control animals. These differences were significant. In the considered metabolic parameters the differences between the 2 groups became not significant at the end of the test with 5% O2. We believe that the found differences in mentioned parameters between hypoxic and normoxic animals, also according to cellular adaptative processes, occurred during the rearing in hypoxic environment. In the test with 5% O2 the seriousness of the hypoxia overcomes the effects of adaptative mechanisms in hypoxic animals since birth. We believe that hypoxic rats since birth represent, limitedly to some aspects, different metabolic models compared to normoxic animals.     

Lipid and fatty acid profiles in the brain, liver, and stomach contents of neonatal rats: effects of hypoxia

Neonatal hypoxia leads to clinically significant fatty liver, presumably due to disturbances in lipid metabolism. To fully evaluate lipid metabolism, the present study analyzed the complete lipid profile of the brain, liver, and ingested stomach contents of 7-day-old rats exposed to hypoxia from birth. Hypoxia had negligible direct effects on lipid metabolism in the brain. Conversely, hypoxia exhibited direct effects on hepatic lipid metabolism that could not be fully explained by changes in dietary intake. Triacylglyceride concentration was significantly increased in the hypoxic liver but remained unchanged in the brain and stomach contents. Diacylglyceride concentration was increased in both the brain and liver, and this was associated with increased diacylglyceride in the stomach contents. Most n-3 and n-6 fatty acids were increased in the liver, but not in the brain, of hypoxic pups. These changes did not reflect those measured in the stomach contents. Saturated fatty acid concentrations were increased in both the hypoxic brain and liver, and these changes reflected those in the stomach contents. Hypoxia also increased total phospholipid concentration in the brain and stomach contents. We conclude that neonatal hypoxia indirectly affects specific lipid and fatty acid concentrations in the brain and liver through alterations in the absorbed stomach contents. Hypoxia also exhibits some direct affects through modulation of metabolic pathways in situ, mostly in the liver. In this respect, the neonatal brain exhibits tighter control on lipid homeostasis than the liver during neonatal hypoxia.     

Hypertriglyceridemia associated with decreased post-heparin plasma hepatic triglyceride lipase activity in hypoxic rats

Exposure of sated rats to 45% N2 in air for 5h increased serum triglyceride levels by 212% over the levels in normoxic rats. This increase in triglyceride levels was accompanied by a decrease in plasma triglyceride hydrolase activity after intravenous injection of heparin. Further fractionation of the activity by inhibition of lipoprotein lipase indicated that the low triglyceride hydrolase activity is mainly due to a reduction in hepatic triglyceride lipase, which is inversely correlated with the serum triglyceride level. The hypoxic exposure decreased the arterial blood [acetoacetate]/[beta-hydroxybutyrate] ratio in the sated rats, which is believed to reflect the oxidation-reduction state in hepatic mitochondria, but did not affect the level of serum enzymes indicative of tissue damage. On the other hand, triglyceride levels did not change during hypoxic exposure in fasted rats. Thus, hypertriglyceridemia in sated rats following exposure to hypoxia may result from impaired removal of circulating triglycerides by hepatic triglyceride lipase located in the sinusoidal surface of the liver.     

Effects of acute hypoxia and hyperoxia on respiratory rate in albino rats chronically exposed to hypoxia

Changes in respiratory frequencies with hypoxic or hyperoxic exposure were studied in: 12 normoxic control rats (N) born and raised in normoxic environment at sea level; 12 rats (A) born and raised in normoxic environment at sea level exposed to normobaric hypoxia (10% O2 in N2) as adults; 12 rats of first generation (G1) raised in the above mentioned hypoxic environment since a few hours after birth; 12 rats of third generation (G3) conceived and born in the hypoxic environment of hypoxic parents of second generation and maintained continuously under hypoxic conditions until their utilization. The response of A rats to 10% O2 and 7% O2 breathing was elevated (57% and 86% over air breathing). The mean respiratory frequency of A rats exposed to 7% O2 rose to a greater extent than did that of N rats. The G1 and G3 rats were less responsive to 7% O2 (64% and 37% over air breathing, respectively) than N and A rats; however, in G1 rats the exposure to 7% O2 produced a greater rise of frequency than in G3 rats. Furthermore A rats, G1 rats and G3 rats were less responsive to 97% O2 breathing (19%, 19% and 11% below air breathing, respectively). Comparing these data with previous findings we suggest that, with chronic exposure to hypoxia, changes in ventilatory response to hypoxia and hyperoxia occur in the following manner: I) loss of response to hypoxia if chronic exposure is begun in the immediate postnatal period; 2) degree of response to hypoxia or hyperoxia influenced by duration of chronic exposure.     

Elevated corticosterone and inhibition of ACTH responses to CRH and ether in the neonatal rat: effect of hypoxia from birth

Hypoxia is a common cause of neonatal morbidity and mortality. We have previously demonstrated a dramatic ACTH-independent activation of adrenal steroidogenesis in hypoxic neonatal rats, leading to increases in circulating corticosterone levels. The purpose of the present study was to determine if this ACTH-independent increase in corticosterone inhibits the ACTH response to acute stimuli. Neonatal rats were exposed to normoxia (control) or hypoxia from birth to 5 or 7 days of age. At the end of the exposure, plasma ACTH and corticosterone were measured before and after either ether vapors were administered for 3 min or CRH (10 microg/kg) was given intraperitoneally. Thyroid function, pituitary pro-opiomelanocortin (POMC) mRNA and ACTH content, and hypothalamic corticotropin-releasing hormone (CRH), neuropeptide Y (NPY), and AVP mRNA were also assessed. Hypoxia led to a significant increase in corticosterone without a large increase in ACTH, confirming previous studies. The ACTH responses to ether or CRH administration were almost completely inhibited in hypoxic pups. Hypoxia did not affect the established regulators of the neonatal hypothalamic-pituitary-adrenal axis, including pituitary POMC or ACTH content, hypothalamic CRH, NPY, or AVP mRNA (parvo- or magnocellular), or thyroid function. We conclude that hypoxia from birth to 5 or 7 days of age leads to an attenuated ACTH response to acute stimuli, most likely due to glucocorticoid negative feedback. The neural and biochemical mechanism of this effect has yet to be elucidated.     

Metabolomic analysis of adrenal lipids during hypoxia in the neonatal rat: implications in steroidogenesis

The nursing rat pup exposed to hypoxia from birth exhibits ACTH-independent increases in corticosterone and renin/ANG II-independent increases in aldosterone. These increases are accompanied by significant elevation of plasma lipid concentrations in the hypoxic neonates. The purpose of the present study was to compare changes in the concentrations of specific fatty acid metabolites and lipid classes in serum and adrenal tissue from normoxic and hypoxic rat pups. We hypothesized that lipid alterations resulting from hypoxia may partly explain increases in steroidogenesis. Rats were exposed to normoxia or hypoxia from birth, and pooled serum and adrenal tissue from 7-day-old pups were subjected to metabolomic analyses. Hypoxia resulted in specific and significant changes in a number of fatty acid metabolites in both serum and the adrenal. Hypoxia increased the concentrations of oleic (18:1 n-9), eicosapentaenoic (EPA; 20:5 n-3), and arachidonic (20:4 n-6) acids in the triacylglyceride fraction of serum and decreased oleic and EPA concentrations in the cholesterol ester fraction. In the adrenal, hypoxia caused an increase in several n-6 fatty acids in the triacylglyceride fraction, including linoleic (18:2 n-6) and arachidonic acid. There was also an increase in the concentration of alpha-linolenic acid (18:3 n-3) in the triacylglyceride fraction of the hypoxic adrenal, along with an increase in linoleic acid concentration in the diacylglyceride fraction. We propose that specific changes in lipid metabolism in the adrenal, as a result of hypoxia, may partly explain the increased steroidogenesis previously observed. The mechanism responsible may involve alterations in cellular signaling and/or mitochondrial function. These cellular changes may be a mechanism by which the neonate can increase circulating adrenal steroids necessary for survival, therefore bypassing a relative insensitivity to normal stimuli.     

Changes in the number of nitrergic neurons following kainic acid administration and repeated long-term hypoxia

Using histochemical analysis (NADPH-diaphorase) we have been investigating the influence of intraperitoneal administration of kainic acid (KA), hypoxia and combination of both these factors on neurons of the hippocampus and on the primary auditory cortex (PAC) in male rats of the Wistar strain. Kainic acid was administered to 18-day-old animals, which were exposed to long-lasting repeated hypoxia from the 2nd till the 17th day of age in a hypobaric chamber (for 8 h a day). At the age of 22 or 90 days, the animals were transcardially perfused with 4 % paraformaldehyde under deep thiopental anesthesia. Cryostate sections were stained to identify NADPH-diaphorase positive neurons that were then quantified in the hippocampus, in the dentate gyrus and in the PAC. In 22-day-old animals both hypoxia and KA increased the number of NADPH-diaphorase positive neurons in the hilus, CA1, CA3 areas of the hippocampus and in the PAC. On the contrary, KA given to hypoxic animals lowered the number of NADPH-diaphorase positive neurons in the dentate gyrus. In 90-day-old animals, hypoxia and KA given to both normoxic and hypoxic animals lowered the number of NADPH-diaphorase positive neurons in some areas of the central nervous system.     

Increased lipoprotein lipase content in the adipose tissue of suckling and weaning obese Zucker rats.

The aim of this study was to determine whether the increase in lipoprotein lipase activity displayed by the adipose tissue of obese (fa/fa) rats as compared with that of lean (Fa/fa) rats could be ascribed to a change in the content or in the catalytic properties of the enzyme. The question was addressed in rats of two ages: in 7-day-old suckling and in 30-day-old post-weaning pups. Inguinal fat-pads were removed surgically (7 days of age) or after killing (30 days of age), and acetone-extract powders were prepared. The relative quantity of enzyme was assessed by immunotitration using an antiserum raised in goat against purified lipoprotein lipase from rat adipose tissue. The results indicate that increases in enzyme activity in obese animals were strictly paralleled by increases in the amount of enzyme in suckling as well as in post-weaning pups. Moreover, the apparent Km values of lipoprotein lipase for its substrate triacylglycerol were identical in the two genotypes. In conclusion, the genotype-mediated increase in lipoprotein lipase activity in adipose tissue of obese Zucker rats was fully accounted for by an increase in the content of the enzyme. In addition, this work documents the mechanism of the increase in lipoprotein lipase activity during weaning, which is mediated mainly through changes in the adipose-tissue enzyme content.     

Interaction of hypoxic and hypercapnic stimuli on breathing pattern in the newborn rat

We aimed to investigate whether newborn rats respond to acute hypoxia with a biphasic pattern as other newborn species, the characteristics of their ventilatory response to hypercapnia, and the ventilatory response to combined hypoxic and hypercapnic stimuli. First, we established that newborn unanesthetized rats (2-4 days old) exposed to 10% O2 respond as other species. Their ventilation (VE), measured by flow plethysmography, immediately increased by 30%, then dropped and remained around normoxic values within 5 min. The drop was due to a decrease in tidal volume, while frequency remained elevated. Hence, alveolar ventilation was about 10% below normoxic value. At the same time O2 consumption, measured manometrically, dropped (-23%), possibly indicating a mechanism to protect vital organs. Ten percent CO2 in O2 breathing determined a substantial increase in VE (+47%), indicating that the respiratory pump is capable of a marked sustained hyperventilation. When CO2 was added to the hypoxic mixture, VE increased by about 85%, significantly more than without the concurrent hypoxic stimulus. Thus, even during the drop in VE of the biphasic response to hypoxia, the respiratory control system can respond with excitation to a further increase in chemical drive. Analysis of the breathing patterns suggests that in the newborn rat in hypoxia the inspiratory drive is decreased but the inspiratory on-switch mechanism is stimulated, hypercapnia increases ventilation mainly through an increase in respiratory drive, and moderate asphyxia induces the most powerful ventilatory response by combining the stimulatory action of hypercapnia and hypoxia.     

Hypoxia-induced caspase-3 activation and DNA fragmentation in cortical neurons of newborn piglets: role of nitric oxide

Hypoxia results in generation of nitric oxide (NO) free radicals, activation of caspase-3, and genomic DNA fragmentation. The present study tests the hypothesis that hypoxia-induced caspase-3 activation and DNA fragmentation are nitric oxide mediated. Studies were conducted in newborn piglets, divided into normoxic (n = 5), hypoxic (n = 5), and hypoxic-7-NINA (n = 6). Hypoxic-7-NINA group received the neuronal nitric oxide synthase inhibitor, 7-Nitroindazole (7-NINA). Caspase-3 activity was determined spectrofluorometrically using enzyme-specific substrates. Sections from the neocortex were stained with an antiserum recognizing active caspase-3. Purified DNA was separated by gel electrophoresis. Administration of 7-NINA resulted in decreased immunoreactivity of caspase-3 (mean LI: 20.2%) as compared to the untreated hypoxia group (mean LI: 57.5%) (P < 0.05). 7-NINA attenuated caspase-3 enzymatic activity as well in comparison to the untreated hypoxia group (P < 0.05). Furthermore, multiple low molecular weight bands corresponding to DNA fragments were present in the hypoxic but not in the normoxic or hypoxic-7-NINA groups. Inhibition of nNOS abates the hypoxia-induced increase in active caspase-3 immunoreactivity, as well as enzymatic activity in cortical neurons, and DNA fragmentation in brain homogenates. We conclude that the coordinate increase of capase-3 activity and fragmentation of nuclear DNA in the hypoxic newborn piglet brain are NO mediated.     

Adrenal lipid profiles of chemically sympathectomized normoxic and hypoxic neonatal rats.

Neonatal hypoxia is a common condition that elicits a coordinated endocrine response. In the neonatal rat, hypoxia induces an ACTH-independent increase in corticosterone which can be partially blocked by chemical sympathectomy. The present study sought to characterize the effects of sympathectomy on the adrenal lipid profile, since previous work suggested that augmented plasma corticosterone during hypoxia may be due to changes in adrenal lipid metabolism. Newborn rats were exposed to normoxia or hypoxia from birth to seven days of age, and guanethidine was used to produce the sympathectomy. Plasma epinephrine and norepinephrine were not significantly affected by hypoxia, while guanethidine decreased plasma norepinephrine in normoxic and hypoxic pups. Hypoxia alone increased the concentration of cholesterol esters in the adrenal gland; this increase was due to increases in cholesterol ester-associated oleic (18:1n9), docosahexaenoic (22:6n3), arachidonic (20:4n6), and adrenic (22:4n6) acids. Hypoxia also increased diglyceride-associated adrenic acid. Guanethidine treatment attenuated the hypoxia-induced increase in cholesterol ester-bound arachidonic and adrenic acids. Guanethidine also decreased saturated fatty acid concentrations and increased n3 fatty acid-enriched triglycerides. The results support the idea that the ACTH-independent corticosterone response to hypoxia in the neonatal rat is mediated by specific, sympathetically driven alterations in the adrenal lipid profile.     

Effect of 7-Nitroindazole Sodium on the Cellular Distribution of Neuronal Nitric Oxide Synthase in the Cerebral Cortex of Hypoxic Newborn Piglets

Cerebral hypoxia results in generation of nitric oxide (NO) free radicals by Ca⁺⁺-dependent activation of neuronal nitric oxide synthase (nNOS). The present study tests the hypothesis that the hypoxia-induced increased expression of nNOS in cortical neurons is mediated by NO. To test this hypothesis the cellular distribution of nNOS was determined immunohistochemically in the cerebral cortex of hypoxic newborn piglets with and without prior exposure to the selective nNOS inhibitor 7-nitroindazole sodium (7-NINA). Studies were conducted in newborn piglets, divided into normoxic (n = 6), normoxic treated with 7-NINA (n = 6), hypoxic (n = 6) and hypoxic pretreated with 7-NINA (n = 6). Hypoxia was induced by lowering the FiO₂ to 0.05–0.07 for 1 h. Cerebral tissue hypoxia was documented by decrease of ATP and phosphocreatine levels in both the hypoxic and 7-NINA pretreated hypoxic groups (P < 0.01). An increase in the number of nNOS immunoreactive neurons was observed in the frontal and parietal cortex of the hypoxic as compared to the normoxic groups (P < 0.05) which was attenuated by pretreatment with 7-NINA (P < 0.05 versus hypoxic). 7-NINA affected neither the cerebral energy metabolism nor the cellular distribution of nNOS in the cerebral cortex of normoxic animals. We conclude that nNOS expression in cortical neurons of hypoxic newborn piglets is NO-mediated. We speculate that nNOS inhibition by 7-NINA will protect against hypoxia-induced NO-mediated neuronal death.     

N-acetylcysteine inhibits hypoxic pulmonary hypertension most effectively in the initial phase of chronic hypoxia

Exposure to chronic hypoxia results in hypoxic pulmonary hypertension (HPH). In rats HPH develops during the first two weeks of exposure to hypoxia, then it stabilizes and does not increase in severity. We hypothesize that free radical injury to pulmonary vascular wall is an important mechanism in the early days of the hypoxic exposure. Thus antioxidant treatment just before and at the beginning of hypoxia should be more effective in reducing HPH than antioxidant therapy of developed pulmonary hypertension. We studied adult male rats exposed for 4 weeks to isobaric hypoxia (F(iO2) = 0.1) and treated with the antioxidant, N-acetylcysteine (NAC, 20 g/l in drinking water). NAC was given "early" (7 days before and the first 7 days of hypoxia) or "late" (last two weeks of hypoxic exposure). These experimental groups were compared with normoxic controls and untreated hypoxic rats (3-4 weeks hypoxia). All animals kept in hypoxia had significantly higher mean pulmonary arterial blood pressure (PAP) than normoxic animals. PAP was significantly lower in hypoxic animals with early (27.1 +/- 0.9 mmHg) than late NAC treatment (30.5 +/- 1.0 mmHg, P < 0.05; hypoxic without NAC 32.6 +/- 1.2 mmHg, normoxic controls 14.9 +/- 0.7 mmHg). Early but not late NAC treatment inhibited hypoxia-induced increase in right ventricle weight and muscularization of distal pulmonary arteries assessed by quantitative histology. We conclude that release of free oxygen radicals in early phases of exposure to hypoxia induces injury to pulmonary vessels that contributes to their structural remodeling and development of HPH.     

Induction of matrix metalloproteinase-2 enhances systemic arterial contraction after hypoxia

This study was carried out to determine the role of increased vascular matrix metalloproteinase-2 (MMP-2) expression in the changes in systemic arterial contraction after prolonged hypoxia. Rats and mice were exposed to hypoxia (10% and 8% O(2), respectively) or normoxia (21% O(2)) for 16 h, 48 h, or 7 days. Aortae and mesenteric arteries were either mounted in organ bath myographs or frozen in liquid nitrogen. MMP-2 inhibition with cyclic CTTHWGFTLC (CTT) reduced contraction to phenylephrine (PE) in aortae and mesenteric arteries from rats exposed to hypoxia for 7 days but not in vessels from normoxic rats. Similarly, CTT reduced contraction to Big endothelin-1 (Big ET-1) in aortae from rats exposed to hypoxia for 7 days. Responses to PE were reduced in hypoxic MMP-2(-/-) mice compared with MMP-2(+/+) mice. Increased contraction to Big ET-1 after hypoxia was observed in MMP-2(+/+) mice but not in MMP-2(-/-) mice. Rat aortic MMP-2 and membrane type 1 (MT1)-MMP protein levels and MMP activity were increased after 7 days of hypoxia. Rat aortic MMP-2 and MT1-MMP mRNA levels were increased in the deep medial vascular smooth muscle. We conclude that hypoxic induction of MMP-2 expression potentiates contraction in systemic conduit and resistance arteries. This may preserve the capacity to regulate the systemic circulation in the transition between the alterations in vascular tone and structural remodeling that occurs during prolonged hypoxic epochs.     

Neonatal dexamethasone treatment increases the risk for pulmonary hypertension in adult rats

Dexamethasone (Dex) treatment during a critical period of lung development causes lung hypoplasia in infant rats. However, the effects of Dex on the pulmonary circulation are unknown. To determine whether Dex increases the risk for development of pulmonary hypertension, we treated newborn Sprague-Dawley rats with Dex (0.25 microg/day, days 3-13). Litters were divided equally between Dex-treated and vehicle control (ethanol) rats. Rats were raised in either room air until 10 wk of age (normoxic groups) or room air until 7 wk of age and then in a hypoxia chamber (inspired O(2) fraction = 0.10; hypoxic groups) for 3 wk to induce pulmonary hypertension. Compared with vehicle control rats, Dex treatment of neonatal rats reduced alveolarization (by 42%; P < 0.05) and barium-filled pulmonary artery counts (by 37%; P < 0.05) in 10-wk-old adults. Pulmonary arterial pressure and the ratio of right ventricle to left ventricle plus septum weights (RV/LV+S) were higher in 10-wk-old Dex-treated normoxic rats compared with those in normoxic control rats (by 16 and 16% respectively; P < 0.05). Small pulmonary arteries of adult normoxic Dex-treated rats showed increased vessel wall thickness compared with that in control rats (by 15%; P < 0.05). After 3 wk of hypoxia, RV/LV+S values were 36% higher in rats treated with Dex in the neonatal period compared with those in hypoxic control rats (P < 0.05). RV/LV+S was 42% higher in hypoxic control rats compared with those in normoxic control rats (P < 0.05). We conclude that Dex treatment of neonatal rats caused sustained lung hypoplasia and increased pulmonary arterial pressures and augmented the severity of hypoxia-induced pulmonary hypertension in adult rats.