Chronic O2 exposure in the newborn rat results in decreased pulmonary arterial nitric oxide release and altered smooth muscle response to isoprostane.

Chronic oxygen exposure in the newborn rat results in lung isoprostane formation, which may contribute to the pulmonary hypertension evident in this animal model. The purpose of this study was to investigate the pulmonary arterial smooth muscle responses to 8-iso-prostaglandin F(2alpha) (8-iso-PGF(2a)) in newborn rats exposed to 60% O2 for 14 days. Because, in the adult rat, 8-iso-PGF(2alpha) may have a relaxant effect, mediated by nitric oxide (NO), we also sought to evaluate the pulmonary arterial NO synthase (NOS) protein content and NO release in the newborn exposed to chronic hyperoxia. Compared with air-exposed control animals, 8-iso-PGF(2a) induced a significantly greater force (P < 0.01) and reduced (P < 0.01) relaxation of precontracted pulmonary arteries in the 60% O2-treated animals. These changes were reproduced in control pulmonary arteries by NOS blockade by using NG-nitro-L-arginine methyl ester. Pulmonary arterial endothelial NOS was unaltered, but the inducible NOS protein content was significantly decreased (P < 0.01) in the experimental group. Pulmonary (P < 0.05) and aortic (P < 0.01) tissue ex vivo NO accumulation was significantly reduced in the 60% O2-treated animals. We speculate that impaired pulmonary vascular tissue NO metabolism after chronic O2 exposure potentiates 8-iso-PGF(2alpha)-induced vasoconstriction in the newborn rat, thus contributing to pulmonary hypertension.     

Reverse phase high performance liquid chromatography in the synthesis of leukotrienes A4 and C4

The chromatographic (RP HPLC) behaviour of leukotriene C4, its methyl ester, leukotriene A4 methyl ester and some chemicals involved in their synthesis have been investigated. Optimal conditions of separation were determined for the gradient and isocratic HPLC. Parameters of the interaction of the substances with hydrophobic surface are discussed in terms of solvophobic theory.     

Bronchial epithelium-associated pulmonary arterial muscle relaxation in the rat is absent in the fetus and suppressed by postnatal hypoxia

We recently reported the existence of a bronchial epithelium-derived relaxing factor (BrEpRF) capable of reducing pulmonary arterial smooth muscle force generation in the newborn rat. We reasoned in this study that BrEpRF has physiological significance in the control of pulmonary vascular tone. We hypothesized that the release and/or activity of this factor can be stimulated and is suppressed prenatally or under hypoxic conditions postnatally. Therefore, we evaluated the pathways stimulated by the BrEpRF in fetal and newborn rat intrapulmonary arteries mounted with their adjacent bronchi in a wire myograph under both normoxic and hypoxic conditions. Under normoxic conditions, BrEpRF release/activation was observed in newborn vessels following methacholine stimulation of M(2) muscarinic receptors, which was mediated via a nitric oxide (NO)-dependent mechanism involving the phosphatidylinositol 3-kinase pathway. Hypoxia suppressed the BrEpRF-dependent modulation of basal and methacholine-induced pulmonary arterial muscle tone in newborn vessels without altering endothelium-dependent or -independent NO-mediated relaxation. In fetal pulmonary arteries studied under normoxic conditions, BrEpRF neither was active under basal conditions nor could it be stimulated with methacholine. We conclude that release/activation of the BrEpRF occurs by an oxygen-dependent mechanism in the newborn and is suppressed during late fetal life. These results suggest that the BrEpRF may be involved in postnatal adaptation of the pulmonary circulation and that its suppression may contribute to hypoxic pulmonary vasoconstriction.     

Antibodies against modified low-density lipoproteins and their complexes in blood of patients with various manifestations of atherosclerosis

The study included 79 patients with coronary artery disease (CAD), 25 individuals with preclinical atherosclerosis and 59 healthy individuals. Key lipid parameters were examined in all the participants. Levels of antibodies (Abs) (IgG and IgM) against low density lipoproteins (LDL) modified by malondialdehyde (MDA), acetic anhydride and hypochlorite, were determined by the enzyme-linked immunosorbent assay (ELISA). Abs specificity was tested by competitive ELISA. Circulating immune complexes (CIC) were isolated by polyethylene glycol precipitation followed by determination of their cholesterol by the enzymatic method. Abs to hypochlorite-modified LDL (hypochlorite-LDL) detected in the serum samples did not demonstrate cross-reactivity with MDA-modified LDL (MDA-LDL) and acetylated LDL (acetyl-LDL). Patients with CAD had increased levels of CIC (p < 0.0001) and decreased levels of Abs (IgM) to hypochlorite- LDL, compared with healthy controls and patients with preclinical atherosclerosis (p = 0.006). A correlation between the levels of Abs (IgG) to the hypochlorite-LDL and Abs to MDA-LDL and acetyl-LDL was found. The content of the Abs (IgM) to MDA-LDL and acetyl-LDL correlated with CIC-cholesterol concentrations, while lipid parameters did not correlate with Abs levels.     

Effect of increased afterload on right ventricular function in newborn pigs

The effect of a progressive increase in right ventricular (RV) afterload was studied in pigs less than 24 h (group I) and 3-5 days old (group III). RV load was applied to increase mean pulmonary arterial pressure (Ppa) until right to left shunt was observed. Initially, pigs in group I had a significantly lower systemic arterial pressure (Psa = 63 +/- 2 vs. 82 +/- 5 mmHg) and higher Ppa (30 +/- 1 vs. 23 +/- 2 mmHg) even though the RV stroke work (RVSW) was similar (54.3 +/- 10.8 vs. 32.4 +/- 2.1 mmHg/ml) to group II. After a progressive rise in afterload, pigs in group I could maintain a higher RV stroke volume than those in group II (1.3 +/- 0.3 vs. 0.4 +/- 0.1 ml; P less than 0.05). At shunt condition, the RVSW was increased by 21 +/- 14% of the initial value in group I vs. a 32 +/- 8% decrease in group II (P less than 0.05). The ductus arteriosus was constricted and right-to-left shunt was observed in all animals at the foramen ovale level even though Ppa exceeded Psa before the rise in the right atrial pressure in group I. Thus, as RV afterload is increased in the pig, the older animals' right ventricle is progressively less capable of maintaining pulmonary blood flow than animals within 24 h of birth.     

A bronchial epithelium-derived factor reduces pulmonary vascular tone in the newborn rat.

The factors accounting for the maintenance of a low pulmonary vascular resistance postnatally are not completely understood. The aim of this study was to test the hypothesis that bronchial epithelium produces a factor capable of relaxing adjacent pulmonary arterial smooth muscle. We studied fourth-generation intralobar pulmonary arteries and bronchi of 4- to 8-day-old rats. Arteries were mounted on a wire myograph, alone or with the adjacent bronchus. The presence of the attached bronchus significantly reduced pulmonary artery force generation induced by the thromboxane analog (U-46619) or KCl whether the endothelium was present or absent (P < 0.01). The converse was not true in that bronchial force generation was not affected when studied with the adjacent pulmonary artery. Mechanical removal of the bronchial epithelium or addition of the nitric oxide (NO) synthase (NOS) nonspecific (N(G)-monomethyl-l-arginine) or the specific neuronal NOS (7-nitroindazole) inhibitors increased arterial force generation to levels comparable to the isolated artery preparation. Wortmannin, a phosphatidylinositol 3-kinase inhibitor, significantly decreased (P < 0.01) NO release of pulmonary arteries only when the adjacent bronchus was present. We conclude that bronchial epithelium in the newborn rat produces a factor capable of lowering pulmonary vascular muscle tone. This relaxant effect can be suppressed by NOS and phosphatidylinositol 3-kinase kinase inhibition, suggesting an action via NOS phosphorylation and NO release. We speculate that such a mechanism may be operative in vivo and plays an important role in control of pulmonary vascular resistance in the early postnatal period.     

Volutrauma activates the clotting cascade in the newborn but not adult rat

Coagulopathy and alveolar fibrin deposition are common in sick neonates and attributed to the primary disease, as opposed to their ventilatory support. Hypothesizing that high tidal volume ventilation activates the extrinsic coagulation pathway, we air ventilated newborn and adult rats at low (10 ml/kg) or high (30 ml/kg) tidal volume and compared them with age-matched nonventilated controls. Blood was collected at the end of the experiment for measurement of clot time, tissue factor, and other coagulation factor content. Similar measurements were obtained from lung lavage material. The newborn clot time (44+/-1) was lower and plasma tissue factor content higher (103.4+/-0.4) than adults (88+/-4 s and 26.6+/-1.4 units; P<0.01). High, but not low, tidal volume ventilation of newborns for as little as 15 min significantly reduced clot time and increased plasma tissue factor content (P<0.01). High volume ventilation increased plasma factor Xa (0.1+/-0.1 to 1.6+/-0.4 nM; P<0.01) and thrombin (1.3+/-0.2 to 2.2+/-0.4 nM; P<0.05) and decreased antithrombin (0.12+/-0.01 to 0.05+/-0.01; P<0.01) in the newborn. Lung lavage material of high volume-ventilated newborns showed increased (P<0.01) factor Xa and thrombin. No changes in these parameters were observed in adult rats that were high volume ventilated for up to 90 min. Compared with adults, newborn rats have a greater propensity for volutrauma-activated intravascular coagulation. These data suggest that mechanical ventilation promotes neonatal thrombosis via lung tissue factor release.     

Catalytic properties and amino acid sequence of endo-1→3-β-D-glucanase from the marine mollusk <Emphasis Type="Italic">Tapes literata</Emphasis>

A specific 1→3-β-D-glucanase with molecular mass 37 kDa was isolated in homogeneous state from crystalline style of the commercial marine mollusk Tapes literata. It exhibits maximal activity within the pH range from 4.5 to 7.5 at 45dgC. The 1→3-β-D-glucanase catalyzes hydrolysis of β-1→3 bonds in glucans as an endoenzyme with retention of bond configuration, and it has transglycosylating activity. The K _m for hydrolysis of laminaran is 0.25 mg/ml. The enzyme is classified as a glucan endo-(1→3)-β-D-glucosidase (EC 3.2.1.39). The cDNA encoding this 1→3-β-D-glucanase from T. literata was sequenced, and the amino acid sequence of the enzyme was determined. The endo-1→3-β-D-glucanase from T. literata was assigned to the 16th structural family (GHF 16) of O-glycoside hydrolases.     

Pulmonary hypertension in the newborn GTP cyclohydrolase I-deficient mouse

Tetrahydrobiopterin (BH4) is a regulator of endothelial nitric oxide synthase (eNOS) activity. Deficient levels result in eNOS uncoupling, with a shift from nitric oxide to superoxide generation. The hph-1 mutant mouse has deficient GTP cyclohydrolase I (GTPCH1) activity, resulting in low BH4 tissue content. The adult hph-1 mouse has pulmonary hypertension, but whether such condition is present from birth is not known. Thus, we evaluated newborn animals’ pulmonary arterial medial thickness, biopterin content (BH4 + BH2), H₂O₂ and eNOS, right ventricle-to-left ventricle + septum (RV/LV + septum) ratio, near-resistance pulmonary artery agonist-induced force, and endothelium-dependent and -independent relaxation. The lung biopterin content was inversely related to age for both types, but significantly lower in hph-1 mice, compared to wild-type animals. As judged by the RV/LV + septum ratio, newborn hph-1 mice have pulmonary hypertension and, after a 2-week 13% oxygen exposure, the ratios were similar in both types. The pulmonary arterial agonist-induced force was reduced (P < 0.01) in hph-1 animals and no type-dependent difference in endothelium-dependent or -independent vasorelaxation was observed. Compared to wild-type mice, the lung H₂O₂ content was increased, whereas the eNOS expression was decreased (P < 0.01) in hph-1 animals. The pulmonary arterial medial thickness, a surrogate marker of vascular remodeling, was increased (P < 0.01) in hph-1 compared to wild-type mice. In conclusion, our data suggest that pulmonary hypertension is present from birth in the GTPCH1-deficient mice, not as a result of impaired vasodilation, but secondary to vascular remodeling.