Cyclooxygenase-2 expression in lung in patients with congenital heart malformations and pulmonary arterial hypertension.

BACKGROUND: Pulmonary arterial hypertension (PAH) is a cause of morbidity in patients with congenital heart disease (CHD). It has been hypothesized that prostanoides participate in the development of PAH. The aim of this study was to show the potential expression of cyclooxygenase-2 (COX-2) in patients with CHD and PAH. PATIENTS AND METHODS: We included patients with isolated left-to-right shunts undergoing lung biopsy before or concomitantly with cardiac surgery between 2004 and 2009.For determination of COX-2 expression, histological and immunohistochemistry analyses as well as quantitative polymerase chain reaction (qPCR) were performed. RESULTS: We were able to show COX-2 protein overexpression in the lung tissue of children with CHD and PAH. Furthermore, we showed an increase in COX-1 gene expression and an even stronger induction of COX-2 by using qPCR and immunohistochemistry. CONCLUSIONS: We examined the expression of COX-2 in lung tissue from patients with CHD and PAH. We showed that COX-2 is expressed in diseased lung tissue, indicating a relationship between COX-2 and vascular remodeling in pulmonary arteries in CHD.

Endothelial nitric oxide synthase gene polymorphism (Glu298Asp) and acute pulmonary hypertension post cardiopulmonary bypass in children with congenital cardiac diseases.

BACKGROUND: Intra-cardiac repair of congenital cardiac diseases in children with left-right shunt is often associated with acute elevation of pulmonary artery pressure following cardiopulmonary bypass. We studied the correlation between the Glu298Asp polymorphism of the endothelial nitric oxide synthase gene and pulmonary hypertension in children with congenital cardiac diseases. METHODS AND RESULTS: A total of 80 children with congenital cardiac diseases at a median age of 3.8 years, ranged 0.1-36.2 years, and 136 controls were enrolled. Most patients presented with significant left-to-right shunt - pulmonary-to-systemic blood flow of 2.8, with a range from 0.6 to 7.5. In all, 40 out of 80 children showed pulmonary hypertension with mean pressure of 42, ranged 26-82, millimetres of mercury. Thirty-one out of 40 children underwent intra-cardiac repair and 15 out of 31 operated patients were found to have an acute elevation of pulmonary artery pressure after cardiopulmonary bypass. The Glu298Asp polymorphism was identified using polymerase chain reaction and restriction fragment length polymorphism. Both in patients and in controls, the genotype distribution corresponded to the Hardy-Weinberg equilibrium. The gene frequency for Glu298Glu, Glu298Asp and Asp298Asp was not different in the control group compared to the patients (Armitage trend test: p = 0.37). The endothelial nitric oxide synthase polymorphism was related to acute post-operative elevation of pulmonary artery pressure (genotypic frequency 53.3 versus 25%; Armitage trend test: p = 0.038). In addition, the allelic frequency of the Glu298Asp was related to post-operative pulmonary hypertension (Fischer's exact test: p = 0.048). The positive predictive value was 71.43%. CONCLUSION: Patients with left-to-right shunt are more likely to develop acute elevation of pulmonary artery pressure after cardiopulmonary bypass when presenting with the Glu298Asp polymorphism of the gene endothelial nitric oxide synthase. This could be used as a genetic marker for the predisposition for the development of pulmonary hypertension after intra-cardiac repair.

Endothelin-1 and asymmetric dimethylarginine in children with left-to-right shunt after intracardiac repair.

BACKGROUND: Endothelin-1 (ET-1) is an endogenous vasoconstrictive peptide hormone and asymmetric dimethylarginine (ADMA) acts as an endogenous inhibitor of nitric oxide synthase. We hypothesized that both could contribute to pulmonary hypertension in patients with left-to-right shunt after intracardiac repair. PATIENTS AND METHODS: We prospectively analyzed ET-1 and ADMA plasma levels in 31 patients (m = 16; f = 15) at an age of 0.6 [0.2-27] years (median [range]) with left-to-right shunt (ASD II: n = 12; VSD: n = 11; AVSD: n = 8) presenting with a Qp/Qs of 2.7 [1.4-6.3] and a pulmonary arterial mean pressure (PAP) of 23 [13-57] mmHg. Blood specimens were taken prior to cardiopulmonary bypass (CPB), after weaning from CPB and at 3, 6, 12 and 24 h after CPB. RESULTS: 12/31 patients were found to have pulmonary hypertension prior to intracardiac repair and 11/12 patients showed persistent pulmonary hypertension during the first 24 h after CPB. Patients with pulmonary hypertension at 12 h after CPB showed significant higher plasma ET-1 compared with patients with normal PAP (1.4 [0-7.9] versus 0.5 [0-2.5] pg/ml; P = 0.048 (Mann-Whitney)). Plasma ADMA decreased from 1.3 [0.75-2.3] micromol/l before CPB to 0.7 [0.4-2.1] micromol/l at 12 h (P < 0.05). However patients with pulmonary hypertension did not show different ADMA plasma levels. CONCLUSIONS: Increased plasma ET-1 but not inhibition of nitric oxide synthase by ADMA is associated with pulmonary hypertension after intracardiac repair.

Atrasentan treatment of pulmonary vascular disease in piglets with increased pulmonary blood flow.

We studied the effect of chronic endothelin A receptor blockade by atrasentan on the pulmonary endothelin-1 system and vascular endothelial growth factor (VEGF) expression in piglets with high pulmonary blood flow. Twenty-five 4-week-old piglets with high pulmonary blood flow were randomized to three groups: sham operated (n = 8), placebo (water) (n = 7), or treatment with atrasentan (2 mg/kg per day) (n = 10). After 3 months, mean pulmonary arterial pressure (PAP) was higher in the placebo group than in the sham group [18 +/- 2 mm Hg versus 14 +/- 1 mm Hg; P < 0.05 (ANOVA)]. Atrasentan treatment was associated with lower cardiac output, PAP (14 +/- 1 mm Hg), and medial wall thickness of pulmonary arteries (diameter: 50-150 microM) compared with placebo [13.6 +/- 3.0% versus 18.1 +/- 4.2%; P < 0.05 (ANOVA)]. Quantitative real-time polymerase chain reaction for endothelin-1, endothelin B receptor, and endothelin-converting enzyme-1 mRNA in lung tissue did not differ. However, immunostaining as well as mRNA for VEGF were lower in atrasentan-treated animals (relative gene expression: atrasentan versus placebo: 0.8 +/- 0.3 versus 1.5 +/- 0.3; P = 0.009). Atrasentan treatment effectively reduces medial hypertrophy in piglets with chronic pulmonary hyperperfusion. Chronic endothelin A receptor blockade by atrasentan may interfere with the expression of VEGF.

Pulmonary vascular changes in piglets with increased pulmonary blood flow and pressure.

In this model of pulmonary vascular disease, high pulmonary blood flow was created by an anastomosis between the left subclavian artery and the main pulmonary artery [Blalock-Taussig (BT) shunt] in 4-week-old piglets (n = 6). Additional ligation of the left pulmonary artery (LPA) was used to increase pulmonary artery pressure (n = 6). Seven piglets were sham-operated. After 3 months, mean pulmonary artery pressure was higher in animals with BT shunt and LPA ligation (22 +/- 5; mean+/-SD) compared to sham-operated animals (15 +/- 2). In addition, thickening of the medial coat (20.1 +/- 2.8% versus 13.6 +/- 3.1% wall thickness) and increased immunostaining for vascular endothelial growth factor A (VEGF-A) were observed. Relative gene expression for endothelin-converting enzyme-1 (ECE-1) mRNA was 1.8 times higher, and VEGF-A mRNA was 2.5 times higher in pigs with BT shunt and LPA ligation compared with sham-operated animals. VEGF receptor-1 and VEGF receptor-2 mRNA was lower in shunted animals and in animals with additional ligation of LPA. Upregulation of ECE-1 and VEGF-A, as well as changes in VEGFR expression in the pulmonary hypertensive lung, may contribute to pulmonary vascular changes.

Comparison of the pharmacokinetic profiles of two locally administered doxycycline gels in crevicular fluid and saliva.

BACKGROUND: Controlled-release delivery systems enable the clinician to extend the half-life period of locally administered antibiotics in gingival crevicular fluid (GCF) significantly. The aim of this split-mouth study was to compare the pharmacokinetic profile of two different doxycycline gels (DOXY and ATRI) for topical subgingival application. Pharmacokinetics of both doxycycline gels were analyzed in GCF and saliva. METHODS: In 10 patients suffering from severe periodontitis, 10 pairs of contralateral defects (pocket depth > or =5 mm/bleeding on probing or > or =6 mm) were randomly assigned either to the first application of DOXY or ATRI. Fourteen days after the topical application of the first antibiotic gel, the application of the second gel in the contralateral defect took place. Clinical examinations at baseline showed no significant differences between sites treated with DOXY and ATRI. Samples of GCF and saliva were drawn baseline, 2, 5 and 24 h after application, 2, 3, 4, 7, 9 and 11 days after application. Separation and quantitative measurement of both doxycycline-gels was performed with HPLC- and UV-detection at lambda=260 nm. RESULTS: In saliva specimens, time-dependent changes of mean doxycycline concentration were almost identical for both doxycycline-gels and declined from a maximum 2 h after application (ATRI: 6653.90+/-3096.14 microg/ml; DOXY: 5386.60+/-1542.02 microg/ml [arithmetic mean+/-SEM]) to zero values 9 days after application. In crevicular fluid specimens, sites treated with ATRI exhibited a faster decrease of mean doxycycline concentration (1085.30, 264.00, 273.94, and 258.00 microg/ml measured 2, 5, 24, and 48 h after application) than sites treated with DOXY (1388.38, 1300.40, 803.73, and 235.10 microg/ml). The faster decrease of ATRI compared with DOXY could not be proved to be statistically significantly different. CONCLUSIONS: Both doxycycline gels showed pharmacokinetics of controlled-release delivery systems.