Randomized controlled trials define shape of dose response for Pollinex Quattro Birch allergoid immunotherapy.

BACKGROUND: The Birch Allergoid, Tyrosine Adsorbate, Monophosphoryl Lipid A (POLLINEX® Quattro Plus 1.0 ml Birch 100%) is an effective, well-tolerated short course subcutaneous immunotherapy. We performed 2 phase II studies to determine its optimal cumulative dose. METHODS: The studies were conducted in Germany, Austria and Poland (EudraCT numbers: 2012-004336-28 PQBirch203 and 2015-000984-15 PQBirch204) using a wide range of cumulative doses. In both studies, subjects were administered 6 therapy injections weekly outside the pollen season. Conjunctival Provocation Tests were performed at screening, baseline and 3-4 weeks after completing treatment, to quantify the reduction in Total Symptom Scores (as the primary endpoint) with each cumulative dose. Multiple Comparison Procedure and Modeling analysis was used to test for the dose response, shape of the curve and estimation of the median effective dose (ED50 ), a measure of potency. RESULTS: Statistically significant dose responses (P < .01 & .001) were seen, respectively. The highest cumulative dose in PQBirch204 (27 300 standardized units [SU]) approached a plateau. Potency of the PQBirch was demonstrated by an ED50 2723 SU, just over half the current dose. Prevalence of treatment-emergent adverse events was similar for active doses, most being short-lived and mild. Compliance was over 85% in all groups. CONCLUSION: Increasing the cumulative dose of PQBirch 5.5-fold from 5100 to 27 300 SU achieved an absolute point difference from placebo of 1.91, a relative difference 32.3% and an increase in efficacy of 50%, without compromising safety. The cumulative dose response was confirmed to be curvilinear in shape.

Strong dose response after immunotherapy with PQ grass using conjunctival provocation testing.

BACKGROUND: Pollinex Quattro Grass (PQ Grass) is an effective, well-tolerated, short pre-seasonal subcutaneous immunotherapy to treat seasonal allergic rhinoconjunctivitis (SAR) due to grass pollen. In this Phase II study, 4 cumulative doses of PQ Grass and placebo were evaluated to determine its optimal cumulative dose. METHODS: Patients with grass pollen-induced SAR were randomised to either a cumulative dose of PQ Grass (5100, 14400, 27600 and 35600 SU) or placebo, administered as 6 weekly subcutaneous injections over 31-41 days (EudraCT number 2017-000333-31). Standardized conjunctival provocation tests (CPT) using grass pollen allergen extract were performed at screening, baseline and post-treatment to determine the total symptom score (TSS) assessed approximately 4 weeks after dosing. Three models were pre-defined (Emax, logistic, and linear in log-dose model) to evaluate a dose response relationship. RESULTS: In total, 95.5% of the 447 randomized patients received all 6 injections. A highly statistically significant (p < 0.0001), monotonic dose response was observed for all three pre-specified models. All treatment groups showed a statistically significant decrease from baseline in TSS compared to placebo, with the largest decrease observed after 27600 SU (p < 0.0001). The full course of 6 injections was completed by 95.5% of patients. Treatment-emergent adverse events were similar across PQ Grass groups, and mostly mild and transient in nature. CONCLUSIONS: PQ Grass demonstrated a strong curvilinear dose response in TSS following CPT without compromising its safety profile.

Hemodynamic effects of basal and stimulated release of endogenous nitric oxide in isolated human lungs.

Background-We compared the hemodynamic responses to inhibition or stimulation of endothelial nitric oxide (NO) release of isolated explanted lungs from transplantation recipients with pulmonary hypertension and in normotensive unallocated donor lungs. Methods and Results-Lungs from 10 patients with severe pulmonary hypertension (SPH) and from 16 patients with severe chronic obstructive lung disease (COLD) were studied. Fourteen normotensive lungs were studied as controls. The lungs were perfused at a constant flow. In protocol 1 N(G)-nitro-L-arginine methyl ester caused a similar rise in baseline pulmonary artery pressure (PAP) that was similar in SPH (+17.1+/-4.2 mm Hg; n=5), COLD (+15.5+/-4.8 mm Hg; n=8), and control lungs (+14.5+/-1.5 mm Hg; n=7). Arterial occlusion demonstrated that most of the changes with N(G)-nitro-L-arginine methyl ester were precapillary. The response to sodium nitroprusside (10(-8) to 10(-4) mol/L) was similar in all groups. In protocol 2, the lungs were preconstricted, and acetylcholine (10(-9) to 10(-5) mol/L) caused a lesser fall in PAP in both COLD and SPH lungs compared with control (-41.9+/-8.6%, -55. 7+/-7.6%, and -73.2+/-2.5%, respectively; P<0.05), whereas sodium nitroprusside (10(-5) mol/L) decreased PAP to initial levels in all lungs. Conclusions-Stimulated release of NO is impaired in arteries of lungs with plexogenic or hypoxemic pulmonary hypertension. In contrast, basal release of NO appears to be maintained.

Sporadic primary pulmonary hypertension is associated with germline mutations of the gene encoding BMPR-II, a receptor member of the TGF-beta family.

BACKGROUND: Primary pulmonary hypertension (PPH), resulting from occlusion of small pulmonary arteries, is a devastating condition. Mutations of the bone morphogenetic protein receptor type II gene (BMPR2), a component of the transforming growth factor beta (TGF-beta) family which plays a key role in cell growth, have recently been identified as causing familial PPH. We have searched for BMPR2 gene mutations in sporadic PPH patients to determine whether the same genetic defect underlies the more common form of the disorder. METHODS: We investigated 50 unrelated patients, with a clinical diagnosis of PPH and no identifiable family history of pulmonary hypertension, by direct sequencing of the entire coding region and intron/exon boundaries of the BMPR2 gene. DNA from available parent pairs (n=5) was used to assess the occurrence of spontaneous (de novo) mutations contributing to sporadic PPH. RESULTS: We found a total of 11 different heterozygous germline mutations of the BMPR2 gene in 13 of the 50 PPH patients studied, including missense (n=3), nonsense (n=3), and frameshift (n=5) mutations each predicted to alter the cell signalling response to specific ligands. Parental analysis showed three occurrences of paternal transmission and two of de novo mutation of the BMPR2 gene in sporadic PPH. CONCLUSION: The sporadic form of PPH is associated with germline mutations of the gene encoding the receptor protein BMPR-II in at least 26% of cases. A molecular classification of PPH, based upon the presence or absence of BMPR2 mutations, has important implications for patient management and screening of relatives.

Pharmacokinetics of cyclosporine in heart and lung transplant candidates and recipients with cystic fibrosis and Eisenmenger's syndrome.

OBJECTIVE: To compare the pharmacokinetics of cyclosporine in patients with either cystic fibrosis or Eisenmenger's syndrome. METHODS: Patients in the study were heart and lung transplant candidates with either cystic fibrosis (n = 6) or Eisenmenger's syndrome (n = 5), as well as patients who received heart and lung transplantation for either cystic fibrosis (n = 13) or Eisenmenger's syndrome (n = 7). This was no experimental pharmacokinetic study in transplant candidates and an exploratory population pharmacokinetic study in transplant recipients. RESULTS: Patients with cystic fibrosis showed higher blood cyclosporine clearance, higher apparent oral clearance, shorter mean residence time, and more erratic absorption. The coefficient of variation of pharmacokinetic parameters was higher in patients with cystic fibrosis. There were no significant differences in metabolite indexes between the two groups of patients after either oral or intravenous administration. A significant negative correlation was found between cyclosporine clearance and hematocrit (r = 0.81 [95% confidence interval, -0.95 to -0.4.1]). Dose-normalized predose blood concentration measurements were lower in patients with cystic fibrosis after transplantation. There was a significant correlation between hematocrit and log dose-normalized cyclosporine concentration (r = 0.40 [95% confidence interval, 0.30 to 0.49]). The total daily dose per 100 ng/ml trough blood concentration required was estimated to be 2.36 times (95% confidence interval, 1.96 to 2.84) higher in patients with cystic fibrosis. CONCLUSIONS: Cyclosporine pharmacokinetics is more variable in patients with cystic fibrosis. The difference in cyclosporine clearance between the two groups is accounted for by differences in binding in blood rather than metabolism. The findings suggest that patients with cystic fibrosis could be conservatively given initial oral doses that are 1.5 times higher than those for patients who receive transplants because of Eisenmenger's syndrome.

Heart-lung transplantation: better use of resources.

PURPOSE: Our goal was to review the experience at Papworth Hospital, Cambridgeshire, England, with combined heart-lung transplantation. PATIENTS AND METHODS: Since April 1984, 31 patients have undergone heart-lung transplantation. Donors and recipients are carefully matched with regards to serology, morphology, and cytomegalovirus compatibility. A pulmonary preservation fluid has been developed that allows distant organ procurement with a single pulmonary artery flush technique. RESULTS: Acute cardiac rejection has not occurred in these patients. Twenty-three patients are alive between two months and over three years following transplantation. The actuarial survival rate at one year is 78 percent, and 70 percent at two years. Three patients died as a result of cytomegalovirus pneumonitis; in two patients, obliterative bronchiolitis developed, and both died, one after an opportunistic infection developed. Three patients died from other causes. The use of transbronchial biopsy of the lung has provided accurate, early, and safe diagnosis of pulmonary rejection. CONCLUSION: Developments in organ preservation and patient management, as well as careful selection of recipients and donors, have led to the effective use of resources and thereby to these good results. In particular, the incidence of obliterative bronchiolitis has been low, which is attributed to the early treatment of pulmonary rejection following diagnosis by transbronchial biopsy.

Transbronchial lung biopsy for the diagnosis of rejection in heart-lung transplant patients.

Long-term success of human lung transplantation has been hindered by the lack of an effective and repeatable method of obtained tissue from the transplanted lung for histology. Management of patients is complicated by the difficulty in distinguishing clinically between opportunistic infection of the lung and rejection. As a result, a large number of patients in recent reports develop chronic disabling obliterative bronchiolitis, believed to be the consequence of "chronic" rejection. Twenty-one patients have undergone heart-lung transplantation in our institute since 1984. During fiberoptic bronchoscopy, 43 transbronchial lung biopsies were performed in 15 patients. Twenty episodes of rejection occurred in 11 patients, from whom 16 sets of biopsies showed the typical changes of perivascular infiltrate and mucosal inflammation. Three biopsies were falsely negative; six routine biopsies performed when patients were well were all normal. Overall sensitivity was 84% and specificity 100%. By contrast, the sensitivity of the chest radiograph was only 40%. Opportunistic lung infection in 8 patients was diagnosed by transbronchial biopsy with a sensitivity of 38% and specificity of 100%. In no patient with opportunistic infection were the histologic features of rejection seen. Transbronchial lung biopsy offers a safe and repeatable method to obtain tissue from heart-lung transplants for histology. It has enabled the management of the lung transplant patient to be equivalent to that of the kidney, liver, and heart transplant patient.

Distant procurement of organs for clinical heart-lung transplantation using a single flush technique.

A more widespread application of combined heart-lung transplantation (HLT) for the treatment of pulmonary vascular disease has been limited by the lack of a simple and inexpensive method for preserving donor organs for transplantation from distant hospitals. Of the 12 HLT patients treated at Papworth Hospital, the last 7 involved distant procurement of donor organs. For this, a single flush hypothermic system was used, with a Ringer's solution containing albumin, mannitol, prostacyclin, and heparin. This was preceded by a prostacyclin infusion into the donor's pulmonary artery. A retrospective comparison has been made of the post-operative pulmonary function between the HLT patients with distant procurement, those with near procurement of organs, and a group of patients who underwent coronary artery bypass graft (CABG) over the same time span. The mean ischemic time for distant-procurement patients was 110 (+/- 10) min, compared with 50 (+/- 0.5) min for near procurement. During the initial 18 hr postoperatively, the alveolar-arterial oxygen gradients (A-aDO2) for both groups of HLT patients and the CABG patients were closely comparable. Two near-procurement patients died: 1 from tracheal dehiscence, and 1 from cytomegalovirus (CMV). One distant-procurement patient died of CMV. All the others recovered fully and are alive and well at home. No patient developed a classical reimplantation response, and pulmonary function in the distant-procurement group compared closely with the CABG patients. These observations suggest that a simple single flush preservation system can effectively extend the pulmonary ischemic time so facilitating distant organ procurement.

Development of bronchiolitis obliterans syndrome in recipients of heart-lung transplantation--early risk factors.

Given the internationally recognized definition of bronchiolitis obliterans syndrome (BOS) and longer follow up of heart-lung transplant recipients, it is possible to establish some of the major risk factors for development and progression of BOS. Between April 1984 and 31 December 1993, 157 patients underwent heart-lung transplantation; 126 survived at least six months after operation and so were at risk of developing BOS. The following early risk factors were assessed for development of BOS grade 1 (21-35% decline in FEV1) and progression from grade 1 to grade 2 (36-50% decline in FEV1): age, gender and underlying diagnosis of the recipient, evidence of acute rejection and cytomegalovirus (CMV) infection within 6 months of operation, peak FEV1 achieved, age and gender of the donor, cold ischemic time of the graft, and matching of CMV serological status and HLA antigens of donor and recipient. The number of acute rejection episodes observed remained the single most important determinant of development of BOS grade 1 (relative risk 1.17 (1.06, 1.29), P=0.002) and progression to BOS grade 2 (relative risk 1.58 (1.02, 2.46), P=0.03). No other factors were significantly related to development or progression of BOS, although both evidence of CMV infection and disease and the number of HLA mismatches increased the risk. Bronchiolitis obliterans syndrome is a major problem for medium-to-long-term survivors of cardiothoracic transplantation. Acute rejection early after transplantation is a sensitive prognostic indicator of subsequent functional decline and requires prompt attention.

The effect of recipient lung size on lung physiology after heart-lung transplantation.

We studied the postoperative course of lung volumes in 32 heart-lung transplant recipients relative to the predicted total lung capacity of the individual donors, to assess the degree of inaccuracy likely to result from the radiological method of matching of donor and recipient lung sizes. There was a tendency for recipients with large preoperative lung volumes--from, for example, emphysema--to receive smaller lungs, while those with smaller volumes from pulmonary vascular disease received bigger donor lungs, but no immediate problems were incurred. After an initial fall in total lung capacity, the postoperative value of the total lung capacity approached the recipients' pretransplant value about one year after the operation irrespective of the size of the donor lungs. This suggests that chest wall compliance is the major determinant of postoperative lung volume and not the donor lung size or compliance. Exact matching of donors' and recipients' lung sizes may not be necessary, and if required can be simply achieved by comparing the measured total lung capacity in the recipient with the predicted value of the donor based on sex, age, and height.

Risk factors for survival following combined heart-lung transplantation. The first 100 patients.

As the numbers of heart and lung transplant recipients have increased it has become possible to identify major risk factors for early (within 3 months) and later (after 3 months) death after this procedure. For 100 patients receiving organs between April 1984 and February 1991, and followed up until February 1992, patient characteristics, operative details, and early morbidity were assessed for their effects on early and later deaths. Recipient age, sex, and preoperative diagnosis did not have a significant effect on early (within 3 months) or later death. Positive cytomegalovirus antibody status of donor or recipient conferred greater risk of death within 90 days (odds ratio [OR] = 3.24, P = 0.06). Greater than 2 L blood in the first 24 hr after operation (OR = 6.00, P = 0.05), and ventilation for greater than 24 hr (OR = 4.87, P = 0.006) were significant prognostic indicators of early death. After the first 3 months, the main risk factor for death was rejection in the first 3 months (OR = 1.38 per episode, P = 0.008). Early infection in general and CMV infection in particular were associated with a small increase in risk. This study confirms the importance of matching donor and recipient for CMV and shows that difficulties during operation, reflected in postoperative bleeding and ventilation times increased the chance of early death. Later death was associated with early acute rejection. A detrimental effect of infection, including CMV infection, either does not exist, or is too small to be detected in a study of this size.

Herpes simplex virus infection in heart-lung transplant recipients.

We report our experience of herpes simplex virus infection in a series of 51 recipients of heart lung transplantation (HLT). Nine patients, all of whom were seropositive for the virus preoperatively, developed HSV infection. Seven episodes of culture-proved mucocutaneous HSV infection without evidence of pulmonary involvement occurred in four patients. Six episodes of HSV pneumonia were seen in a further five patients, one of whom died. Diagnosis of HSV pneumonia was by histological appearances on transbronchial biopsy, together with culture of lung tissue or bronchoalveolar lavage. Concomitant cytomegalovirus infection occurred in four patients. All patients who developed HSV pneumonia did so within the first two postoperative months; in four patients following augmented immunosuppression. We now suggest that HLT recipients who are HSV antibody-positive should receive prophylactic acyclovir for the first two months after surgery and at times of augmented immunosuppression.

Infection and reactivation with cytomegalovirus strains in lung transplant recipients.

Cytomegalovirus pneumonia is a major cause of morbidity and death following lung transplantation (LT) (1). The case fatality rate is highest in the CMV-seronegative recipients (R-) of organs from seropositive donors (D+), which suggests that transmission of CMV may occur with the graft (1), but in seropositive recipients (R+) the comparative importance of reactivation of endogenous virus and reinfection with donor virus is poorly understood.

Risk factors for obliterative bronchiolitis in heart-lung transplant recipients.

Obliterative bronchiolitis is the major cause of death of long-term survivors of heart-lung transplantation. Of our first 75 patients who have received heart-lung transplantation, 38 have been followed for a year or longer. Eight patients developed clinical evidence of obliterative bronchiolitis within 15 months of transplantation, of whom four died with postmortem confirmation of extensive obliterative bronchiolitis, interstitial and pleural fibrosis, and vascular sclerosis in the heart and lungs. One further patient died before one year after chronic rejection. All nine patients had evidence on transbronchial biopsy of submucosal fibrosis and vascular sclerosis. Twelve of our remaining patients have shown similar areas of lung fibrosis on transbronchial biopsy, and the other eighteen are well and without fibrosis on transbronchial biopsy. Studies of the 274 biopsies obtained from 38 patients confirmed rejection on 182 occasions with more frequent, more persistent, and more severe rejection in the chronic rejection group than in the without-fibrosis or lung fibrosis groups. Opportunistic infection resulted in pneumonia on 19 occasions, and were most commonly found in lung fibrosis patients. We conclude that obliterative bronchiolitis is the likely outcome in patients with early, poorly controlled, severe rejection.

Transbronchial biopsies in children after heart-lung transplantation.

Sixty transbronchial biopsies have been performed in eight children after heart-lung transplantation. The selection of fiber-optic bronchoscope or a small (4 mm; 30 cm) rigid bronchoscope was made according to the size of endotracheal tube required at surgery. If the endotracheal tube was size 7.5 or greater, a fiber-optic bronchoscope was used, whereas if the endotracheal tube size was below 7, a rigid bronchoscope was used. For the diagnosis of lung rejection, the histology of biopsies revealed a sensitivity of 91% and specificity of 69% (similar to the result in adults). The histology also distinguished lung infection from rejection. Complications included three pneumothoraces and two clinically significant episodes of hemorrhage, one of which led to a cardiorespiratory arrest, which may have been caused by hypoxia. As a result, arterial oxygen saturation is now monitored during the procedure using a pulse oximeter.

Changes in factor VIII proteins after cardiopulmonary bypass in man suggest endothelial damage.

16 patients undergoing coronary artery bypass grafting using cardiopulmonary bypass (CPB) had blood samples taken at various times before, during and up to 1 week after surgery for estimation of beta-thromboglobulin (BTG), alpha-1-antichymotrypsin (ACT), factor VIII procoagulant protein (VIII:C), von Willebrand factor antigen (vWF:Ag) and ristocetin co-factor (vWF:RiCoF). vWF:Ag and vWF:RiCoF rose during and following surgery in a different manner to ACT. At 1 week there was a significantly disproportionate rise in vWF:Ag compared to vWF:RiCoF which suggested a degree of pulmonary endothelial damage. Prostacyclin, which was administered to 8 of the patients during CPB, reduced platelet activation as measured by a reduction in the release of BTG and also attenuated the consumption of VIII:C. It had no effect on pulmonary endothelial damage as measured by the ratio of vWF:Ag to vWF:RiCoF.

Role of prostacyclin in the treatment of primary pulmonary hypertension.

Over the last decade, new medical and surgical treatments have become available for primary pulmonary hypertension that have influenced the natural history of the disease. Vasodilator therapy is aimed at overcoming pulmonary vasoconstriction with consequent decrease in afterload and improved right ventricular output. The endpoint of vasodilator therapy is therefore to reduce pulmonary vascular resistance, without causing significant systemic hypotension. Prostacyclin, a potent vasodilator and antiaggregating agent, has been successfully used by continuous infusion in the treatment of severe pulmonary hypertension. The marked improvement in quality of life observed with the use of prostacyclin in these patients, as well as the increased survival time on the waiting list for transplantation, has influenced us to adopt prostacyclin infusion as a fundamental element of our therapeutic strategy for the management of this rare but fatal condition.

Effects of the immunosuppressant cyclosporine on the circulation of heart transplant recipients.

The effect of cyclosporine on the systemic circulation and on heart rate is unknown for heart transplant recipients. Thirty-four heart transplant recipients were studied by right-sided cardiac catheterization after endomyocardial biopsy. A direct linear relation was found between systemic and pulmonary vascular resistance and cyclosporine trough blood levels, which were negatively related to heart rate. The effect of cyclosporine on pulmonary vascular resistance, however, was not statistically significant by multivariate analysis when patient age was considered. In contrast, renal function appeared unrelated to systemic vascular resistance or heart rate. It appears that cyclosporine trough blood levels may have a direct effect on systemic vascular resistance as well as an unexplained negative chronotropic effect on heart rate.

Impairment of pulmonary-artery endothelium-dependent relaxation in chronic obstructive lung disease is not due to dysfunction of endothelial cell membrane receptors nor to L-arginine deficiency.

1. Endothelium-dependent relaxation mediated by endothelium-derived relaxing factor (EDRF) or nitric oxide (NO), is impaired in pulmonary arteries (PA) of hypoxic patients with chronic obstructive lung disease (COLD). To determine the mechanisms responsible for this impairment, we compared the response of rings of isolated PA from 12 COLD patients and 8 controls to the endothelium-dependent vasodilators acetylcholine (ACh), adenosine diphosphate (ADP), and the calcium ionophore, A23187. The response of PA rings to the endothelium-independent nitro-vasodilator sodium nitroprusside (SNP) was also studied in both groups. The PA rings had been pre-contracted by the alpha-adrenoceptor agonist phenylephrine (PE). 2. Endothelium-dependent relaxation was significantly reduced in PA rings from COLD patients as compared with controls when tested with ACh (37.8 +/- 8.8% vs 73.4 +/- 7.9%), ADP (38.4 +/- 6.7% vs 80 +/- 5.6%), and the calcium ionophore, A23187 (35.8 +/- 6.1% vs 87 +/- 6.6%). Relaxation with SNP was, however, significantly greater in PA rings from COLD patients (99.4 +/- 0.6% vs 90.3 +/- 3.1%), as was the contractile response to PE (1.91 +/- 0.21 g vs 1.33 +/- 0.15 g). Pretreatment with the specific inhibitor of NO formation, NG-monomethyl-L-arginine (L-NMMA; 10(-4) M) significantly reduced the relaxation to ACh in all PA rings. This inhibition could be reversed by L-arginine (10(-3) M), the substrate for NO synthesis. Pretreatment with L-arginine alone, however, did not restore the impaired endothelium-dependent relaxation of PA rings from COLD patients. 3. We conclude that EDRF (NO) production is impaired in PA rings from COLD patients and that this impairment is neither due to endothelial receptors dysfunction nor a defect of L-arginine availability and/or transport. Our hypothesis is that the abnormality must lie within the biosynthesis pathway of NO from L-arginine, possibly involving the endothelial enzyme cell, NO synthase, the normal function of which might be altered by chronic hypoxia.

Impairment of pulmonary endothelium-dependent relaxation in patients with Eisenmenger's syndrome.

A comparison has been made between the endothelium-dependent relaxation of pulmonary arteries (PA) obtained at heart-lung transplantation from 4 patients with Eisenmenger's syndrome and secondary pulmonary hypertension, and PA obtained at lobectomy from 4 patients with lung carcinoma, the controls. All vascular rings were studied immediately after lung excision. PA rings from control patients dose-dependently relaxed to cumulative doses of acetylcholine (ACh, 10(-10) to 10(-5) M), achieving a maximal relaxation of 80 +/- 5% (mean +/- s.e. mean) from precontraction with phenylephrine. By contrast, PA rings from Eisenmenger's syndrome patients achieved a maximal relaxation of only 34 +/- 12% (P less than 0.05, unpaired t test), with even paradoxical contraction at high doses of ACh (10(-6) to 10(-5) M). Sodium nitroprusside (10(-4) M) relaxed all PA rings, with and without endothelium (carefully removed before study), obtained from both control and Eisenmenger's syndrome patients. These results provide the first evidence that endothelium-dependent relaxation of PA mediated by endothelium-derived relaxing factors is impaired in Eisenmenger's syndrome patients with secondary pulmonary hypertension.