P < 0.001) and substantially higher among younger patients, nonpostmenopausal women, and patients receiving monotherapy. While potentially relevant improvements in the entire cohort were observed in regard to body weight (−2.1 ± 5.4 kg), fasting plasma glucose (from 6.8 to 6.2 mmol/L), and triglycerides (2.4 to 2.0 mmol/L), statistically significant changes in metabolic parameters could only be detected in subgroup analyses. Moxonidine therapy reduced blood pressure and improved rates of blood pressure control in this group of patients. While the observed trend towards improvement in various metabolic parameters merits further investigation, the overall effect of moxonidine treatment is consistent with a reduction of total cardiovascular risk in this hypertensive metabolic syndrome cohort.This study was designed to assess the effects of moxonidine on blood pressure and aspects of the metabolic syndrome in racially diverse population of patients encountered in routine medical practice. Physicians collected data on a minimum of three consecutive patients with uncontrolled essential hypertension and criteria for metabolic syndrome, eligible to receive moxonidine (0.2–0.4 mg once daily) for 6 months, either as monotherapy or as adjunct therapy to current antihypertensive treatment. Systolic and diastolic blood pressure (BP) declined by an average of 24.5 + 14.3 mmHg and 12.6 + 9.1 mmHg, respectively. BP responder rates defined as attaining BP < 140/90 mmHg were significantly ( Hypertension is a major contributor to cardiovascular disease (CVD) risk, but a patient's global CVD risk is determined by the interplay of multiple risk factors. In particular, the grouping of risk factors—elevated blood pressure, abdominal obesity, dyslipidaemia, and abnormalities of glucose and insulin metabolism, commonly referred to as metabolic syndrome has been associated with a substantial worsening of cardiovascular prognosis and all-cause mortality . PostmenAbdominal obesity is not only a cardinal feature of the metabolic syndrome but also an important contributor to the development and progression of cardiovascular and metabolic disturbances linked to the syndrome. Overactivity of the sympathetic nervous system (SNS) is of particular importance in this context. The interplay between obesity, elevated SNS activity, and hypertensive target organ damage is already demonstrable in very young overweight or obese adults , 8. ElevTargeting the SNS directly, therefore, provides a logical and attractive therapeutic target –12 in thMoxonidine is a widely approved antihypertensive drug that lowers BP primarily by reducing central SNS activity via activation of imidazoline type-1 receptors in the rostral ventrolateral medulla . In addiTo this end, we conducted a large, multinational study to appraise the effects of moxonidine on blood pressure, anthropometric, lipid, and metabolic parameters of the metabolic syndrome in patients encountered in routine  medical practice.MERSY was a multinational, open-label, observational study with a planned duration of 6 months. Participating physicians were asked to collect data on a minimum of three consecutive patients with uncontrolled essential hypertension and metabolic syndrome, for whom moxonidine might be prescribed.The primary objective was to evaluate the long-term safety and efficacy of moxonidine in hypertensive patients with metabolic syndrome. The secondary objective was to assess the effect of long-term treatment of moxonidine on laboratory parameters associated with the metabolic syndrome.Moxonidine was prescribed at a dose of 0.2–0.4 mg once daily, either as monotherapy or as adjunctive therapy when the current antihypertensive treatment (which was to the discretion of the treating physician) was insufficient to achieve individual blood pressure targets or if it was not tolerated. The preferred maintenance dose of moxonidine was 0.4 mg/day, but physicians were permitted to initiate therapy at 0.2 mg and titrate to 0.4 mg/day after 2 weeks. After the baseline visit, a first follow-up visit was scheduled for between 1 and 3 months, according to the treating physician's usual practice for such consultations. A final visit was planned 6 months after starting moxonidine therapy. The study had no formal mechanisms to monitor compliance.Adult patients (age ≥ 18 years) of either sex were eligible for enrolment if they had essential hypertension of any grade, as defined by the 2003 guidelines of the European Society of Hypertension. Patients were either newly diagnosed as hypertensive or had BP levels that were above target despite the use of other antihypertensive drugs measured according to the 2003 guidelines of the European Society of Hypertension or had failed to tolerate current antihypertensive treatment. For Australia only, supplementary inclusion criteria specified age not greater than 75 years and the persistence of hypertension despite concurrent antihypertensive therapy.Criteria for a diagnosis of metabolic syndrome were based on the 2005 definition proposed by the International Diabetes Federation and comprised central obesity  plus any two of the following: triglyceride (TG) levels ≥150 mg/dL (≥1.7 mmol/L) or specific treatment for this lipid abnormality; high-density lipoprotein (HDL) cholesterol <40 mg/dL (<1.03 mmol/L) in men or <50 mg/dL (<1.29 mmol/L) in women, or specific treatment for this lipid abnormality; systolic BP (SBP) ≥ 130 mmHg or diastolic BP (DBP) ≥ 85 mmHg, or treatment of previously diagnosed hypertension;fasting plasma glucose (FPG) ≥ 100 mg/dL (≥5.6 mmol/L) or previously diagnosed type 2 diabetes. If FPG was >5.6 mmol/L (>100 mg/dL), an oral glucose tolerance test was recommended, but this was not compulsory. Specific advice on lifestyle modification was not mandated by the protocol. The only criterion prohibiting patients from taking part in this study was the presence of contraindications to moxonidine, as identified in the relevant National Summary of Product Characteristics (SPC). From observations and experience in a previous postmarketing surveillance study , it was χ2 test or Fisher's exact test. Ordinal qualitative variables were compared using the Wilcoxon test or the Kruskal-Wallis test. Quantitative variables were compared using variance analysis.Nominal qualitative variables were compared using the BP, laboratory and weight parameters were compared between visits using covariance analysis, with the baseline value as the adjusted variable.t-test. All tests were two sided, with significance declared at the 5% level.The absolute changes in heart rate (HR) between the baseline and postbaseline visits were summarized and analyzed through a one-sample In addition to pooled results, subgroup analyses were undertaken according to menopause status (as determined by questioning), type of antihypertensive regimen (monotherapy or combination therapy), and age (<65 years and ≥65 years).Data management and statistical analysis were conducted by the FOVEA Group, Rueil-Malmaison, France. Data entry was performed using Access version 9.0. Double data entry was used. Entered data were verified against case record form data when a discrepancy was found during double data entry. Quality control was performed using SAS version 8.2.The primary efficacy variable was the percentage of patients responding to antihypertensive therapy during the study. A response was defined as attainment of systemic arterial BP < 140/90 mmHg from baseline to each follow-up visit. BP limits of <130/80 mmHg were set for patients with a diagnosis of diabetes at baseline.Secondary efficacy variables comprised the absolute change in BP from baseline to each follow-up visit, the absolute change in laboratory parameters for metabolic syndrome -cholesterol, and creatinine, urinary albumin) from baseline to final visit at 6 months, and the absolute change in weight parameters (body mass index (BMI), waist-hip circumferences) from baseline to each follow-up visit.Suspect adverse drug reactions (SADRs)—defined as a response to a drug that was noxious and unintended and that occurred at doses normally used in humans for prophylaxis or treatment of a disease or to modify physical function—were screened for via active enquiry during follow-up and final visits. Each SADR was evaluated for duration, severity , and seriousness. Physicians' assessment of the causal relationship to the investigational drug was documented, as was the action taken to address all SADRs and the outcome of each event. Special provisions were made for the reporting of all SADRs regarded as serious. Any pregnancies identified during the study were recorded and monitored as discrete events.The study was conducted in accordance with the ICH GCP (1997) and the Therapeutic Goods Administration (TGA) “Note for Guidance on Good Clinical Practice” (CPMP/ICH/135/95) annotated with TGA comments (July 2000). Patients were free to withdraw from the study at any time, for any reason, specified or unspecified, without prejudice to their medical care. Physicians were free to exclude any patient at any time if this was judged to be in the interests of the patient. N = 5603) and safety (n = 5879) populations. Two countries (Bahrain and Switzerland) did not recruit any patients. The MERSY study was conducted in 13 countries between December 2006 and March 2008. P < 0.001), ~1 mmHg lower in nonpostmenopausal women , and ~5 mmHg higher in patients receiving multiple antihypertensive medications than in those prescribed monotherapy ((159.2 ± 14.1)/(65.0 ± 13.4) mmHg versus (154.7 ± 11.7)/(60.9 ± 12.1) mmHg; P < 0.001 versus monotherapy). Differing national legislations precluded a complete audit of ethnicity data.Principal demographic details of the ITT population are depicted in Tables P = 0.001), these differences were numerically small (data not shown).There was statistical evidence of variations in various metabolic syndrome-related metabolic parameters according to age, therapeutic regimen, and menopause status, but with the exception of FPG according to menopause status  (n = 3506) had received multiagent combination therapy for BP control. Diuretics were the single most widely prescribed class of drugs recorded in this subset of patients (n = 2277). A further 1200 patients had received monotherapy, of which 605 had received either an angiotensin-converting enzyme (ACE) inhibitor or an angiotensin receptor blocker (ARB), 199 had received a calcium-channel blocker, 160 had been prescribed a beta-blocker, and 142 had received a diuretic. A further 719 patients had received no antihypertensive medication during that month.In the month preceding the baseline visit, most patients (n = 5567), with diabetes proportionately more often recorded in older patients (54.2% at age ≥ 65 years versus 44.8% at <65 years), those taking multiple antihypertensive drugs (51.5% versus 28.7% versus monotherapy), and postmenopausal women  .A baseline diagnosis of diabetes was present for 47.1% (2623) of the ITT population for whom data were available (n = 3885), intolerance to current antihypertensive medication (n = 286), or a new diagnosis of hypertension (n = 886); 138 cases were classified as “other reasons.” Lack of efficacy of current therapies was more likely to be the reason in older patients, postmenopausal patients, and patients taking multiple antihypertensive drugs, whereas a new diagnosis of hypertension was proportionately more common in younger nonpostmenopausal patients.Documented reasons for initiating moxonidine therapy comprised lack of efficacy of current antihypertensive medication (n = 823) were being prescribed moxonidine 0.2 mg/day at that time and 76.3% were prescribed doses up to 0.4 mg/day (n = 3143). The percentage of patients who received moxonidine as monotherapy (19-20%) or as part of multiple combination therapy (80-81%) remained constant throughout the study.Moxonidine 0.2 mg/day was prescribed to 1731 patients at the baseline visit. Doses up to 0.4 mg/day were prescribed to a further 3635 patients. Among the 4118 patients of the ITT cohort for whom medication data were available from the last study visit, 20.0% (n = 1345) at first in-study clinical visit (between 1 and 3 months) to 41.3% (n = 2314) at final assessment at 6 months.  Responder rates were significantly (P < 0.001) and substantially higher among younger patients (44.3% versus 33.4% in older patients), nonpostmenopausal women , and patients receiving monotherapy (55.7% versus 37.8% in those receiving multidrug therapy).The proportion of patients classified as responding to hypertension therapy increased progressively during the study, from 24.2% , notwithstanding differences in absolute values. There were likewise no subgroup-specific variations in the trend for total cholesterol, creatinine, or urinary albumin. On-treatment changes were recorded for mean values of every nominated laboratory parameter except creatinine . The proP < 0.001) for all comparisons, except HDL-C (P = 0.004). The reduction in LDL-C during treatment was more marked in younger patients than older ones (P = 0.007).By contrast, the reduction in TG and the increase in HDL-C levels was more marked in younger (versus older) patients, and the reduction in TGs was significantly larger in nonpostmenopausal women  , fell from 79.6 ± 9.1 beats/min to 74.1 ± 7.0 beats/min, an average reduction of –5.7 ± 8.2.Average weight declined by –2.1 ± 5.4 kg during the study and BMI declined by –0.7 ± 2.0 kg/mn = 46) of cases, respectively. The distribution of investigators' impressions of treatment was similar.Patients' assessments of treatment were “excellent,” “good,” “tolerable,” or “bad" in 44.4%, 48.3%, 6.4%, and 0.9% (n = 15). Events contributing at least 5% of the total SADR count comprised gastrointestinal disorders ; nervous system disorders ; general disorders and administration site conditions ; and skin and subcutaneous tissue disorders . The most frequent SADRs linked with the gastrointestinal system were dry mouth (48 events in 47 subjects) while most of the SADRs linked with the nervous system included dizziness (16 events in 16 subjects) or headache (13 events in 13 subjects). Nervous system disorders were the single largest category of events associated with study termination and the single largest source of events rated as severe (n = 9).During the course of the study, 195 SADRs were recorded in 132 patients (2.2% of the study population) . Of thesOf the 12 serious SADRs, 2 each were classified as nervous system disorders; vascular disorders; infections and infestations; or respiratory, thoracic, and mediastinal disorders. The remaining four serious SADRs comprised one case each in the categories of psychiatric disorder; pregnancy, puerperium, and perinatal conditions; renal and urinary disorders; and cardiac disorders. No deaths were reported during the study.This open-label phase IV trial was designed to assess the effect of moxonidine on BP and laboratory parameters associated with the metabolic syndrome after 6 months of treatment in a general practice setting. We sought to enlarge on previous experience in a single country  by recruThe results of the MERSY study are consistent with the previous experience with moxonidine in the management of hypertension –24. The While blood pressure reduction is the primary goal of antihypertensive therapy, potential effects on metabolic parameters also need to be taken into account. Such considerations have led to widespread recommendations in national and international guidelines to avoid beta-blockers and diuretics in patients with metabolic disturbances or diabetes mellitus if not indicated for additional comorbidities. The reasoning for these recommendations relates to the well-described weight gain with beta-blockers and the adverse metabolic effects (such as insulin resistance and hyperuricemia) encountered with both beta-blockers and diuretics. In contrast, antihypertensive agents that exert no or even beneficial metabolic effects, such as calcium channel blockers  and inhibitors of the renin-angiotensin-system , which have been shown to reduce new onset of diabetes , are conMoxonidine clearly falls into the second category, with proven efficacy in regard to blood pressure lowering and beneficial effects in regard to diverse metabolic parameters. While the effects of moxonidine on individual metabolic indices in our study could be considered as modest, the trends of all the changes seen were towards a profile of lower overall CVD risk, which was particularly evident in the subgroup analyses.In this context, the average reduction in body weight of 2.1 kg in our patients is noteworthy and replicates earlier studies of moxonidine in populations with metabolic syndrome , 21. WeiP < 0.001) and substantially higher among younger compared to older patients (44.3% versus 33.4%). Interestingly, the reduction in LDL-C and TG (P < 0.001) and the increase in HDL-C levels (P = 0.004) was also more marked in younger than in older patients. These findings may indicate that younger subjects derive specific benefit from centrally sympatholytic agents.Several lines of evidence suggest that sympathetic activation is of particular relevance in the earlier stages of hypertension , 8, 11. Our observation of reductions in TG and body weight in conjunction with improvement in blood pressure control are compatible with the suggestion  that visAdditional, albeit indirect evidence for this concept comes from the ALMAZ study , in whicMoxonidine was well tolerated when used in combinations with the range of first-line antihypertensives in MERSY, as was the case in other studies. More generally, the safety profile of moxonidine in the MERSY study was fully in accordance with the known effects of the drug. The overall incidence of SADRs was very low (<2.5%) and the nature of the SADRs observed was consistent with previous experience. No previously unreported terms of SADR were encountered during our study. The incidence of SADRs with moxonidine is usually highest during the first weeks of treatment and thereafter declines to very low levels. The independent decision of many investigators to start therapy at doses <0.4 mg/day may have contributed to the excellent tolerability profile of moxonidine in the MERSY study and may be regarded as an example of skill in clinical practice.In summary, in this large sample of patients with hypertension and concomitant metabolic syndrome, moxonidine enhanced blood pressure control when used alone or in combination and was associated with improvement in several aspects of metabolic syndrome. Moxonidine was well-tolerated during 6 months of continuous use. Given current estimations suggesting that approximately 70% of all incident hypertension is associated with overweight or obesity, it appears justified to recommend antihypertensive treatment with an agent that targets the underlying pathophysiology including sympathetic activation, particularly so if additional benefits can be achieved in regard to control of body weight and other metabolic markers characterizing the metabolic syndrome.


To assess associations of the trans-lamina cribrosa pressure difference (TLCPD) with glaucomatous optic neuropathy.The population-based Central India Eye and Medical Study included 4711 subjects. Based on a previous study with lumbar cerebrospinal fluid pressure (CSFP) measurements, CSFP was calculated as CSFP[mmHg] = 0.44 Body Mass Index[kg/m2]+0.16 Diastolic Blood Pressure[mmHg]−0.18×Age[Years] −1.91. TLCPD was IOP–CSFP.P<0.001; standardized coefficient beta:0.53; regression coefficient B:0.18; 95% confidence interval (CI):0.17, 0.18), lower body mass index , lower diastolic blood pressure , higher pulse , lower body height , higher educational level , higher cholesterol blood concentrations , longer axial length , thicker central cornea , higher corneal refractive power  and presence of glaucomatous optic neuropathy . Differences between glaucomatous subjects and non-glaucomatous subjects in CSFP were more pronounced for open-angle glaucoma (OAG) than for angle-closure glaucoma (ACG) (3.0 mmHg versus 1.8 mmHg), while differences between glaucomatous subjects and non-glaucomatous subjects in IOP were higher for ACG than for OAG (8.5 mmHg versus 3.0 mmHg). Presence of OAG was significantly associated with TLCPD  but not with IOP . Prevalence of ACG was significantly associated with IOP  but not with TLCPD (P = 0.92).Mean TLCPD was 3.64±4.25 mm Hg in the non-glaucomatous population and 9.65±8.17 mmHg in the glaucomatous group. In multivariate analysis, TLCPD was associated with older age (In OAG, but not in ACG, calculated TLCPD versus IOP showed a better association with glaucoma presence and amount of glaucomatous optic neuropathy. It supports the notion of a potential role of low CSFP in the pathogenesis of open-angle glaucoma. The trans-lamina cribrosa pressure difference (TLCPD) has been defined as the difference between intraocular pressure (IOP) minus cerebrospinal fluid pressure (CSFP), while the IOP, in a strict physical sense, is the transcorneal pressure difference between the intraocular compartment and the surrounding external atmosphere The Central India Eye and Medical Study (CIEMS) is a population-based cross-sectional study performed in 8 remote villages close to so called tribal regions in Central India between 2006 and 2008 All examinations were carried out during a day-long stay at a hospital to which the study participants were transported by a bus. Trained social workers asked questions about regarding background, living conditions, tobacco use and alcohol consumption, and any known diagnosis of major systemic diseases. Pulse, arterial blood pressure, and body height and weight were recorded. One-and-a-half hours after a standardized lunch, blood and urine samples were obtained and biochemically analyzed. A detailed ophthalmologic examination included refractometry for visual acuity assessment, frequency-doubling perimetry, Goldmann applanation tonometry, slit lamp biomicroscopy, sonographic corneal pachymetry and biometry , gonioscopy, digital photography of the cornea, lens, optic disc and macula, and confocal laser scanning tomography  of the optic disc. Glaucoma was defined by a glaucomatous appearance of the optic disc. The optic nerve head was glaucomatous (1) if the inferior-superior-nasal-temporal (ISNT)-rule of the neuroretinal rim shape was not fulfilled in early glaucoma and in eyes with a normally shaped optic disc ; or (2) if an abnormally large cup was present in a small optic disc which normally would not show cupping. In all eyes with glaucoma, the visibility of the retinal nerve fiber layer was locally and / or segmentally reduced. The assessment of the optic disc photographs was carried in a masked manner without knowledge of intraocular pressure or the perimetric results. Each photograph of a glaucomatous optic disc was independently adjudicated by two senior graders (VN and JBJ). The whole glaucoma group was differentiated into subjects with open-angle glaucoma and with primary angle closure glaucoma. Open-angle glaucoma was characterized by an open anterior chamber angle, in addition to a normal depth of the anterior chamber as assessed by slit lamp biomicroscopy. In angle-closure glaucoma, the anterior chamber angle was occluded or occludable. The anterior chamber angle was defined as occludable, if ≥270° of the posterior trabecular meshwork could not be seen upon gonioscopy 2]+0.16×Diastolic Blood Pressure [mmHg]−0.18×Age [Years]−1.91. The association between higher CSFP and younger age, higher body mass index and higher blood pressure had also been found in other previous investigations P = 0.29) from the calculated CSFP (13.3±3.2 mm Hg). The Durbin-Watson value was 2.08. Values falling into the acceptable range of 1.5 to 2.5 indicate a non-significant autocorrelation for the residuals in the multiple regression models 2]+0.27×Diastolic Blood Pressure [mmHg]−0.08×Age [Years]−24.8.For the calculation of a formula to estimate the CSFP, we used the lumbar CSFP measurements obtained in a previous study on 72 patients (mean age: 42.0±13.4 years) who consecutively underwent lumbar puncture for diagnosis and treatment of neurological diseases P-values were 2-sided and were considered statistically significant when the values were less than 0.05.Only those subjects with assessable optic disc photographs and measurements of blood pressure, body height and weight, and IOP were included into the study. Statistical analysis was performed using a commercially available statistical software package . Continuous data were presented as mean ± standard deviation. In a first step, we examined the mean values. In a second step, we searched for associations between TLCPD, CSFP and other systemic and ocular parameters, first in univariate analysis, and then in multivariate analysis. In a third step, we compared TLCPD and CSFP between glaucomatous subjects and non-glaucomatous subjects and searched for associations between TLCP and CSFP on one side and parameters indicating the amount of glaucomatous optic nerve damage on the other side. In a fourth step, logistic regression analyses were performed to investigate associations of the presence of glaucoma and TLCPD. Odds ratios (OR) and 95% confidence intervals (CI) were presented. All P<0.001) and more myopic , while both groups did not differ significantly in axial length , IOP  and gender .Out of the 9422 eyes (4711 subjects), optic disc photographs and values for TLCPD were available for 8815 (93.6%) eyes of 4546 (96.5%) subjects. The other 165 participants were excluded since assessable photographs of the optic nerve head or values of blood pressure, body mass index or IOP were not available. The mean age of the 4546 participants (53.7% women) was 48.5±12.9 years , mean refractive error was −0.13±1.77 diopters , and mean axial length was 22.64±0.88 mm . The group of excluded subjects as compared with the group of subjects included into this study was significantly older  of 122 subjects  (2.7%). According to the gonioscopic finding, the whole glaucoma group was differentiated into an OAG group (n = 173 eyes) and subjects with ACG (n = 20 eyes). Using the criteria of the International Society of Geographical and Epidemiological Ophthalmology, glaucoma was detected in 129 eyes of 121 subjects, with 121 eyes fulfilling the criteria of open-angle glaucoma.P<0.001; correlation coefficient r: 0.52), lower body height , lower body weight , lower body mass index , lower level of education , lower diastolic blood pressure , lower systolic blood pressure , lower pulse , and higher blood concentration of cholesterol  and lower hemoglobin concentration ; and with the ocular parameters of shorter axial length , myopic refractive error , shallower anterior chamber , thicker lens , thicker central cornea , and higher corneal refractive power . It was not significantly associated with blood concentration of glucose (P = 0.13), high-density lipoproteins (P = 0.17), and of glycosylated hemoglobin (P = 0.91).In the non-glaucomatous population, mean TLCPD was 3.64±4.25 mm Hg (median: 3.42 mm Hg) . In univP-values. It resulted in a final model in which TLCPD was associated with the systemic parameters of older age (P<0.001), lower body mass index (P<0.001), lower diastolic blood pressure (P<0.001), higher pulse (P<0.001), lower body height (P<0.001), higher level of education (P<0.001), higher blood concentrations of cholesterol (P<0.001), and with the ocular parameters of longer axial length (P = 0.001), thicker central cornea (P<0.001), and higher corneal refractive power P<0.001). If we then added the presence of glaucoma to the list of independent variables, TLCPD was additionally associated with glaucomatous optic neuropathy  in this multivariate analysis model.The multivariate analysis within the non-glaucomatous group included TLCPD as dependent variable and all those parameters which were significantly associated with TLCPD in the univariate analysis. In a first step, we removed all parameters for which the variance inflation factor as part of the analysis of collinearity was higher than 2.0. We then dropped out of the list of independent parameters those variables, which were no longer significantly associated with TLCPD, starting with the parameters with the highest uropathy . In a siP<0.001; beta: 0.17; B: 5.22; 95%CI: 4.60, 5.84) after adjusting for lower pulse , shorter body height , lower educational level , longer axial length , thicker central cornea , higher corneal refractive power , and higher blood concentration of cholesterol .If the parameters of body mass index, age and diastolic blood pressure were dropped from the multivariate model, since these parameters were included in the formula to calculate the CSFP, TLCPD was significantly associated with a higher prevalence of glaucoma , female gender (P = 0.049), taller body height , higher body weight , higher body mass index , higher diastolic blood pressure  and systolic blood pressure , higher pulse , higher level of education , and higher blood concentration of cholesterol  and glucose , and with the ocular parameters of higher intraocular pressure  , lower corneal refractive power , refractive error , longer axial length , deeper anterior chamber , and thinner lens . It was not significantly associated with blood concentration of glycosylated hemoglobin (P = 0.20) and high-density lipoproteins (P = 0.65).In the non-glaucomatous population, mean estimated CSFP was 10.0±3.6 mm Hg (median: 10.2 mm Hg) . In univr: 0.22) , thickerP-values. We eventually arrived at a model in which CSFP was significantly associated with younger age (P<0.001), higher body mass index (P<0.001), higher systolic blood pressure (P<0.001), higher pulse (P<0.001), taller body height (P<0.001), and higher IOP (P = 0.03)  .P<0.001; beta: −0.08; B: −2.11; 95%CI: −2.57, −1.64) after adjusting for higher pulse , longer body height , female gender , higher educational level , higher blood concentration of cholesterol  and glucose , thicker central cornea , less corneal refractive power , and higher intraocular pressure . If the total glaucoma group was differentiated into OAG and ACG, the absolute value of the standardized coefficient beta in the model was higher for OAG (beta: −0.08) than for ACG (beta: −0.03). If the OAG group was further subdivided into a subgroup with an IOP measurement ≤21 mmHg  and a subgroup with an IOP>21 mmHg , the association between higher CSFP and lower glaucoma prevalence in the model described above was better for the normal-IOP OAG group  than for the high-IOP OAG group .If the parameters of body mass index, age and diastolic blood pressure were dropped from the multivariate model , higher CSFP was significantly associated with a lower prevalence of glaucoma (P<0.001), the estimated CSFP was lower  and the mean TLCPD was significantly (P<0.001) higher (9.8±8.2 mmHg versus 3.6±4.2 mmHg in the glaucoma group. The differences between both groups were highest for TLCPD (6.2 mmHg) followed by IOP (3.5 mmHg) and the estimated CSFP (2.4 mmHg). If the whole glaucoma group was divided into OAG and ACG, the difference between the glaucomatous group and the non-glaucomatous group in IOP was higher for ACG than for OAG (8.5 mmHg versus 3.0 mmHg), while the difference in CSFP between the glaucomatous group and the non-glaucomatous group was more pronounced for the OAG group than for the ACG group (3.0 mm Hg versus 1.8 mmHg). Correspondingly, the correlation coefficients between the pressure parameters and the presence of glaucomatous optic neuropathy were higher for TLCPD than for IOP in the OAG group (r = 0.17 versus r = 0.12), while the correlation coefficients were higher for IOP than for TLCPS in the ACG group (r = 0.11 versus r = 0.12). As a corollary, the association between presence of glaucoma and lower CSFP in binary regression analysis was stronger for OAG  than for ACG .Comparing the whole glaucoma group with the non-glaucomatous group showed that IOP was significantly higher , lower body mass index (P = 0.008), higher mean blood pressure (P = 0.02), prevalence of myopic retinopathy (P<0.001), lower level of education (P = 0.006), longer axial length (P = 0.002), and higher intraocular pressure  P<0.001; B =  0.16; OR: 1.17; 95%CI: 1.12, 1.22). If only IOP and TLCPD were included into the analysis, the presence of OAG was significantly associated with TLCPD  but not with IOP .A previous study had revealed that the prevalence of OAG was significantly associated with higher age (P = 0.02), higher IOP (P<0.001), more hyperopic refractive error (P<0.001), shallower anterior chamber (P = 0.01), and shorter axial length (P<0.001). If TLCPD was added to the model, it was not significantly associated with the presence of ACG (P = 0.55). If only IOP and TLCPD were included into the analysis with age as additional independent parameter, the presence of ACG was significantly associated with IOP  but not with TLCPD (P = 0.92).The prevalence of ACG was significantly correlated with older age  was stronger than the association between rim area and IOP  after adjusting for gender (P = 0.03), optic disc area (P<0.001) and axial length (P <0.001). This difference was more pronounced for the OAG than for the ACG group.If the amount of glaucomatous damage was quantified by neuroretinal area, the association between rim area and smaller TLCPD  CSFP was associated with the presence and severity of OAG Interestingly, using the formula described above resulted in a mean value of CSFP of 10.0±3.6 mm Hg (median: 10.2 mm Hg) which is similar to the reported mean CSFP as measured by lumbar puncture P<0.001) correlated with IOP  the range in which 95% of the measurements are to be expected (assuming a Gaussian distribution), the range will go from −4.9 mmHg to 12.1 mmHg. This range in relative terms is even larger than the normal range for CSFP. If one takes into account that TLCPD is more important than CSFP for the pressure conditions in the optic nerve head, the large normal range of TLCPD may exemplify the role alterations of the TLCPD may potentially play in the pathophysiology of the optic nerve head.In multivariate analysis in the non-glaucomatous study population, higher TLCPD was associated with among other parameters, with older age, lower body mass index, lower diastolic blood pressure, and more myopic refractive error . These rPotential limitations of our study should be mentioned. First, the whole statistical analysis depended on the formula to calculate the CSFP. The study, in which the basis parameter for that formula were assessed included a relatively small number of subjects, and these subjects had a clinical reason to undergo lumbar puncture In conclusion, in OAG but not in ACG, calculated TLCPD versus IOP showed a better association with presence of glaucoma and amount of glaucomatous optic neuropathy. It supports the notion of a potential role of low CSFP in the pathogenesis of OAG. In contrast, IOP as compared with TLCPD was better correlated with presence and amount of glaucomatous damage in ACG, fitting with the notion that the intraocular reason for the elevation of IOP, and not the retro-laminar pressure, was the major driving cause for glaucomatous optic neuropathy in ACG.


DOT1L, MLLT3, SIRT1, and SGK1 encode genes in a pathway that controls methylation of the histone H3 globular domain at lysine 79 (H3K79), thereby modulating expression of the ENaCα subunit. This study aimed to determine the role of variation in these regulatory genes on blood pressure response to HCTZ, and secondarily, untreated blood pressure.Nearly one-third of the United States adult population suffers from hypertension. Hydrochlorothiazide (HCTZ), one of the most commonly used medications to treat hypertension, has variable efficacy. The renal epithelial sodium channel (ENaC) provides a mechanism for fine-tuning sodium excretion, and is a major regulator of blood pressure homeostasis. P ≤ 0.01 in one cohort and replication by P ≤ 0.05 in the other cohort considered significant.We investigated associations between genetic variations in this candidate pathway and HCTZ blood pressure response in two separate hypertensive cohorts . In a secondary, exploratory analysis, we measured associations between these same genetic variations and untreated blood pressure. Associations were measured by linear regression, with only associations with DOT1L (rs2269879) was strongly associated with greater systolic (P = 0.0002) and diastolic (P = 0.0016) blood pressure response to hydrochlorothiazide in Caucasians. However, this association was not replicated in the other cohort. When untreated blood pressure levels were analyzed, we found directionally similar associations between a polymorphism in MLLT3 (rs12350051) and greater untreated systolic (P < 0.01 in both cohorts) and diastolic (P < 0.05 in both cohorts) blood pressure levels in both cohorts. However, when further replication was attempted in a third hypertensive cohort and in smaller, normotensive samples, significant associations were not observed.In one cohort, a polymorphism in DOT1L, MLLT3, SIRT1, and SGK1 are not likely associated with blood pressure response to HCTZ. However, a possibility exists that rs2269879 in DOT1L could be associated with HCTZ response in Caucasians. Additionally, exploratory analyses suggest rs12350051 in MLLT3 may be associated with untreated blood pressure in African-Americans. Replication efforts are needed to verify roles for these polymorphisms in human blood pressure regulation.Our data suggest polymorphisms in  Hydrochlorothiazide (HCTZ) is one of the most commonly prescribed antihypertensive drug in the US, with approximately 118 million prescriptions dispensed in 2010, either alone or combined with another antihypertensive .Patient response to thiazides varies widely, with differential responses between and within races . Because+/Cl- cotransporter (NCC) in the distal convoluted tubule [NEDD4L, a gene involved in ENaC regulation, and blood pressure response to diuretics [SCNN1G (which encodes the ENaCγ subunit) have been associated with HCTZ response [Thiazides achieve their initial diuretic action by preventing renal sodium reabsorption via inhibition of the Nad tubule -7. Howevd tubule . Becauseiuretics . In addiresponse ,11. In aresponse .MLLT3) binds to the ENaCα promoter and localizes Dot1 for di- and tri-methylation at H3 Lys79, which is associated with ENaCα gene repression [SGK1), which disrupts the assembly of the Af9/Dot1 complex [SIRT1) can also form a complex with Dot1 to decrease ENaCα expression. However, the mechanism for this interaction is still unclear [An epigenetic pathway was recently discovered that regulates ENaCα expression in the kidney by methylation of histone protein H3 at lysine 79 (H3K79) -15. At tpression . This re complex . Evidenc unclear .DOT1L, MLLT3, SIRT1, and SGK1 affect clinical blood pressure response to HCTZ in well-defined clinical cohorts. To test the second, we assessed associations of these polymorphisms with untreated clinical blood pressures as an exploratory analysis.We hypothesized that genetic variation in this epigenetic regulatory pathway plays a role in the antihypertensive effects of thiazides, through its regulation of ENaC. Secondarily, we hypothesized that variation in this pathway affects human blood pressure homeostasis. To test the first hypothesis, we evaluated whether polymorphisms in Study participants arose from the Pharmacogenomic Evaluation of Antihypertensive Responses  and the Genetic Epidemiology of Responses to Antihypertensives . Both studies were approved by the institutional review boards at each center where they were conducted, and all subjects provided informed, written consent before being screened for enrollment.PEAR was a multi-center clinical trial examining the role of genetic variability on blood pressure response to HCTZ and/or atenolol . Men andGERA was a two-center clinical trial designed to determine whether polymorphisms in renin-angiotensin-aldosterone system genes were predictive of the blood pressure response to HCTZ . BrieflyTwo normotensive samples were also used in an attempt to replicate untreated blood pressure associations found in GERA and PEAR African-Americans. The first sample drawn from was the Ethnic Pain Sensitivity trial, a single center study designed to examine ethnic differences in pain sensitivity . 206 heahttp://gvs.gs.washington.edu/GVS/ was queried for the genetic regions of DOT1L, MLLT3, SIRT1, and SGK1 in the HapMap YRI (African Ancestry) and CEPH (European Ancestry) populations. SNPs with a minor allele frequency less than 5% were excluded. The Genome Variation Server provided a comprehensive list of 144 SNPs meeting these criteria which tagged all four gene regions.A tagSNP approach was used to assure maximum coverage of common SNPs in each candidate gene region. A tagSNP selection tool using the Multipop-TagSelect algorithm  providedin silico by two separate programs, Pupasuite [To more precisely investigate the strongest candidate SNPs, putative functional SNPs (pfSNPs) were added to the lists generated by the Genome Variation Server. These were computed upasuite  and Fastupasuite . From thGenotypes in HCTZ-treated GERA and PEAR participants were determined using a custom GoldenGate Assay for the BeadXpress Reader System . Genotyping was carried out according to the manufacturer's protocol. Raw data conversion and quality control were completed in GenomeStudio software . Samples were excluded if their genotype call rate was below 90%. Individual SNPs were excluded from analysis if they were monomorphic in our cohorts, their call frequencies were below 75%, or their GenTrain scores were less than 0.3.For untreated blood pressure replication analyses, genotypes were determined using Taqman SNP Genotyping Assays and the Taqman 7900HT Real Time PCR System  according to the manufacturer's protocol.Associations between genotype and blood pressure responses to HCTZ were tested by linear regression after adjustment for covariates, including gender, age, and untreated blood pressure. Associations between untreated systolic blood pressure and diastolic blood pressure were tested in the same manner as described for HCTZ response, except covariate adjustments included only gender and age. Statistical analyses were completed in JMP Genomics 4 and SAS 9.2 .P ≤ 0.00028 for significance (0.05/180 = 0.00028). However, the SNPs analyzed are strong biological candidates, and do not represent independent tests due to linkage disequilibrium. Additionally, replication of findings is essential in genetic association studies. In our analyses, SNPs were considered significant if they associated with HCTZ blood pressure response in either GERA or PEAR with P ≤ 0.01, replicated with P ≤ 0.05 in the other study group, and had matching directions of effect. With this replication strategy, the overall P-value threshold for significance is 0.01 × 0.05 × 0.5 (for matching direction of effect) = 0.00025, which therefore meets the Bonferroni criterion, as similarly justified previously [Because of the large number of SNPs tested, adjustments for multiple comparisons were necessary to avoid false positives. A Bonferroni correction, assuming 180 independent tests, would require a eviously .2 analysis separately by race and study group. Those SNPs with Hardy-Weinberg equilibrium P-values less than a Bonferroni-corrected 0.00028 were flagged and analyzed under suspicion of genotyping error.As a quality control procedure, Hardy-Weinberg equilibrium was tested via χBaseline characteristics were similar among both study cohorts Table . Both noGERA - Genetic Epidemiology of Responses to Antihypertensives trial, PEAR HCTZ - Randomized to HCTZ in the Pharmacogenomic Evaluation of Antihypertensive Responses trial, PEAR ATEN - Randomized to atenolol in the Pharmacogenomic Evaluation of Antihypertensive Responses trial, EPS AA population - African Americans from the Ethic Pain Sensitivity trial, HTNDB AA Normotensives - Normotensive African Americans from a University of Florida hypertension database, BP - blood pressure, N/A - Not Available.Out of a total 180 SNPs genotyped with the GoldenGate assay, six SNPs were excluded for having a SNP call frequency less than 75% or a GenTrain score less than 0.3. An additional two SNPs failed these quality controls in GERA only, so were analyzed exclusively in PEAR. In GERA, 19 participant samples were excluded from analysis because of low genotype call rates (< 90%). In PEAR, one participant sample was excluded from analyses because of low call rates. Taqman genotyping of rs12350051 was completed in replication samples with approximately 7% duplication, revealing 98% concordance. Both rs2269879 and rs12350051 were in Hardy-Weinberg equilibrium in all groups analyzed.P-values for genetic associations with blood pressure response in GERA and PEAR HCTZ  and a 3.5 mmHg greater mean diastolic (P = 0.0016) response to HCTZ in PEAR HCTZ Caucasians . Because GERA P-values did not meet our predetermined criteria for significance, this SNP was not considered a replicated association.A Manhattan plot of -log Z Figure  indicateMLLT3) genotype in both PEAR HCTZ  and GERA African-Americans  at the pre-defined thresholds for association and replication , in intron 3. FastSNP showed the variant A allele at rs8113528 creates a possible binding site for p300, transcriptional co-activator that functions as a histone acetyltransferase. Additionally, UCSC Genome Browser http://genome.ucsc.edu/ indicates rs8113528 exists in an area surrounded by moderate histone acetylation.The strongest pharmacogenetic association with HCTZ response was with rs2269879 in DOT1L, and was only observed in PEAR Caucasians. Results with systolic and diastolic response in GERA were directionally consistent, but nonsignificant. Located in intron 7, rs2269879 was chosen for genotyping as a tagSNP. Upon review in the HapMap CEPH population, the SNP was found to be in perfect linkage with rs8113528 . Office measurements, the only blood pressure phenotypes available in GERA, may not possess high enough fidelity to detect this association with rs2269879. Supporting this theory, we observed similar, but much weaker associations with office blood pressure response in PEAR. Perhaps if another large hypertensive cohort, prospectively treated with HCTZ, becomes obtainable for analysis of home blood pressure responses, the association we found could be tested again for replication.Because the association only met the significance threshold in PEAR, and did not replicate in GERA, we cannot rule out that this is a chance finding. One reason for a lack of replication in GERA may be that the effect of this SNP can only be detected using home blood pressure. PEAR was the only study with the home blood pressure phenotype available. We decided pressure ,26. In apressure . PEAR hoDOT1L, MLLT3, SIRT1, SGK1 and blood pressure associations is low. TagSNPs within 5000 bases of each candidate gene were selected to try to detect any possible cis-regulatory regions. Great effort was spent on identifying pfSNPs in silico for each candidate gene, which were not required to be in the pre-defined gene region for tagSNP development. However, only SNPs with a minor allele frequency of 0.05 were considered for genotyping, so our study cannot rule out very rare SNPs in the candidate genes with large effect sizes affecting blood pressure response. Additionally, our data do not rule out whether or not this pathway plays any role thiazide response. If HCTZ did have some small effect on H3K79 methylation, redundancy in ENaCα regulation [The lack of association we found with HCTZ response suggests that genotyping polymorphisms in this pathway would likely not help predict patient response to thiazide diuretics. The likelihood that common SNP associations were missed with gulation  could coMLLT3. It was chosen as a tagSNP, and is located in intron 2, with no linkage to any known functional SNPs. In silico, rs12350051 was not observed in any known miRNA sequences, transcription factor binding sites, exonic splice sites, splice enhancer, or silencer sequences. Because the same blood pressure association was not seen in Caucasians, one possibility could be that this SNP is in high linkage disequilibrium with an undiscovered functional polymorphism in African-Americans.Little is known about the effect of this histone H3K79 methylation pathway on blood pressure regulation in humans, so exploratory analyses testing associations in untreated blood pressure phenotypes could also provide valuable information. The SNP that associated and replicated with untreated blood pressure was rs12350051 in One would expect using a patient population with a wide range of blood pressures to be the best method to detect genetic associations with untreated blood pressure. So the fact that untreated blood pressure associations were seen in PEAR and GERA is somewhat surprising, as these studies enrolled hypertensives spanning a relatively small blood pressure range. This was one of the reasons we attempted to replicate these findings in normotensive blood pressure ranges not represented in PEAR and GERA. However, the normotensive groups also had a narrow blood pressure range. The fact that no replication was observed in normotensives could be because of this narrow blood pressure range, the fact that they were younger, the differences in study protocols, leading to differences in blood pressure measurement precision, or the sample sizes were too small and lacked the power to detect the effect we observed in the larger hypertensive cohorts. Another possibility is that perhaps the effect of this SNP is easier to detect or only exerts an effect with higher blood pressures.Dot1 conditional knockout mice were shown to exhibit salt sensitive hypertension [Af17, which has been shown to compete with Af9 for Dot1 binding at the ENaCa promoter, [Our findings are not the first to detect associations between this H3K79 methylation pathway and blood pressure regulation. rtension . Conversromoter,  exhibit romoter, . Also, Sromoter,  has beenromoter, .MLLT3, SIRT1, or SGK1 and blood pressure responses to HCTZ in hypertensives. One SNP in DOT1L (rs2269879) could play a role in HCTZ response, but requires further investigation to replicate the association found in PEAR. Additionally, rs12350051 in MLLT3 was associated with untreated blood pressure in African-American hypertensive individuals. Because this was an exploratory analysis, and the association was not replicated in smaller normotensive samples, questions remain as to whether this polymorphism is involved in the blood pressure regulation of normotensives, and the mechanism by which rs12350051 exerts an effect on blood pressure. Further study in clinical populations with broader blood pressure ranges would help answer these questions.In conclusion, associations likely do not exist between common variation in ATEN: atenolol; HCTZ: hydrochlorothiazide; H3K79: histone H3 lysine 79; NCC: sodium - chloride cotransporter; ENaC: epithelial sodium channel; ENaCα: epithelial sodium channel α subunit; SNP: single nucleotide polymorphism; pfSNP: putative functional SNP; GERA: Genetic Epidemiology of Responses to Antihypertensives; PEAR: Pharmacogenomic Evaluation of Antihypertensive Responses.The authors declare that they have no competing interests.JDD participated in the study design, participated in genotyping, performed statistical analyses and drafted the manuscript. IZ participated in the design of the study and helped to draft the manuscript. BB participated in sample processing and genotyping and helped to draft the manuscript. KRB assisted with statistical analysis, STT and ALB participated in study coordination and helped to draft the manuscript. YG, TYL, ABC, EB, JGG, RMC, RBF participated in study coordination. BCK conceived the study, participated in its design, and helped to draft the manuscript. JAJ conceived the study, participated in its design, and helped to draft the manuscript. All authors read and approved the final manuscript.Supplementary Table 1. SNPs genotyped in SGK1, DOT1L, SIRT1, and MLLT3 gene regions.Click here for file


Background. Inflammation and remodeling are integral parts of asthma pathophysiology. We sought to describe the clinical and pathologic features of near fatal asthma exacerbation (NFE). Methods. Bronchial biopsies were collected prospectively from NFE I subjects. Another NFE II group and a moderate severity exacerbation control group (ME II) were retrospectively identified—no biopsies obtained. Results. All NFE II (n = 9) subjects exhibited remodeling and significant inflammation . NFE II group (n = 37) had a significant history of prior intubation and inhaled corticosteroids usage compared to ME II group (n = 41). They also exhibited leukocytosis, eosinophilia, and longer hospitalization days. Conclusions. Remodeling, eosinophilic, and neutrophilic inflammation were observed in NFE. NFE is associated with prior intubation and inhaled corticosteroids usage. Remodeling in asthma refers to structural changes in large and small airways, consisting of subepithelial fibrosis, increased vascularity, increased airway smooth muscle mass, and goblet cell hyperplasia of proximal and distal airways , 2. RemoRemodeling and persistent inflammation are present in relatively mild and “benign” asthma, but not many data exist regarding the pathologic features in severe asthmatic exacerbations or in near fatal asthma . This stThis observation would particularly help us how to optimally manage these severely compromised patients; studies in humans but also in animal models yield contradictory results about the effectiveness of anti-inflammatory medication against remodeling , 8. The We defined two subgroups of asthma subjects who presented with an acute exacerbation at the ED. One group consisted of severe asthmatics with an NFE who required intubation and mechanical ventilation and were admitted to the ICU (NFE group). The second group consisted of asthma patients with a less severe exacerbation who were admitted either to the ICU or to a medical ward and preserved spontaneous breathing. The latter group responded to conventional therapy and was designated as the moderate exacerbation (ME) group (controls). The study comprised of a prospective and a retrospective arm.Between 2004 and 2009, asthma patients admitted to Los Angeles County and University of Southern California (LAC and USC) Medical Center  and to the USC University Hospital (USCUH)  with an NFE were included prospectively in the study. Inclusion criteria were an NFE as defined previously and a diagnosis of asthma reported by the admitting and the ICU physician. Exclusion criteria were age below 16th year and history of Chronic Obstructive Pulmonary Disease (COPD). Patients underwent bronchoscopy and had mucosal biopsies of the large proximal airways. These patients comprised NFE I group. Unfortunately, no ME patients were prospectively included in the study; as a result, control biopsies were not available in this study.Because of the lack of a prospective control group, we retrieved in a retrospective fashion the medical records of adult asthmatics which presented at the ED and admitted to LAC and USC Medical Center or to USCUH between 1999 and 2004 with an asthma exacerbation. As defined previously, they were divided to those with an NFE  and those with an ME . No biopsies were available for these retrospectively collected groups. The same inclusion and exclusion criteria with the prospective data were applied. The purpose of the retrospective arm was therefore to provide an analysis of NFE patients  with a control ME group, unavailable in the prospective arm.We reviewed the charts for recording of (i) medication used at time of admission , (ii) smoking habit (in pack/years), and (iii) respiratory symptoms and signs, blood eosinophilia, and evidence of infection. Pulmonary function testing was not recorded in the charts. We believe that the explanation is twofold: the discomfort of the emergency setting at which admissions took place and the mainly low socioeconomic status of most patients which prevented them from regular and proper follow-up of their disease.The study was approved by the University of Southern California Institutional Review Board  for the medical chart review (retrospective group) as well as for the patients who underwent bronchoscopy and airway biopsies (prospective group). Informed consent was obtained from the patient directly or from the patient's legal surrogate.Bronchoscopy was performed through the endotracheal tube and mechanical ventilation was performed appropriately and according to the patient's best care and management. After each subject was stabilized with maximum conventional treatment and under sedation, bronchoscopy was performed at the bedside within 12 hours after intubation (using a Pentax bronchoscope). Five mucosal biopsies were taken from the right main bronchus, the bronchus intermedius, and from a lobar bronchus (middle lobe or upper lobe). The biopsies were superficial in order to prevent significant hemorrhage. Biopsy specimens were processed in the Pathology Department to be examined by one pathologist (MK).The airway tissues submitted in 10% formalin fixative were embedded in paraffin after appropriate dehydration and 5-micron-thick sections cut and stained as follows: one hematoxylin and eosin-stained slide, one PAS- (periodic acid-schiff-) stained slide, and one trichrome-stained slide with at least 2 sections per slide. The numbers of eosinophils, neutrophils, lymphocytes, and plasma cells in the highest area of cellularity of the bronchial wall were scored semiquantitatively as follows: 1–5 cells/HPF (high-power field) = 1+; 6–10 cells/HPF = 2+; >10 cells/HPF = 3+. Next, the respiratory basal lamina was graded as normal or thickened. The presence of mucinous and squamous metaplasia was determined by counting 30 contiguous respiratory epithelial cells and noting the percentage of mucinous or squamous cells among them. Bronchial wall edema was judged as mild, moderate, or severe. Finally, to determine the vascularity of the bronchial wall, the maximum number of capillaries in the airway wall was counted under a high-power microscopic field (×400).t test or a nonparametric equivalent (Mann-Whitney test). χ2 or Fisher's exact test was performed for qualitative variables. Relative risk and its 95% confidence interval were calculated. Correlation analysis was performed in all groups. A Roc analysis in the retrospective arm was done to find the features with the largest area under the curve for subsequent intubation. Multiple logistic regression analysis was also performed to identify independent factors associated with intubation in the retrospective arm, although the large number of missing values for some key variables limited the utility of this approach. The threshold of statistical significance was P < 0.05.Values are presented as means ± standard deviation or frequency and percent. Measured values between groups NFE II and ME II were analyzed with two-tailed Patients' classification in groups is shown in The histopathological findings of the NFE I subjects are shown in The retrospective arm consisted of groups NFE II and ME II. 2 patients had 2 severe exacerbations each. Each exacerbation was considered a separate event . Patient2 administered, higher arterial PO2 and PCO2, and lower pH than the ME II group. They also had profound leukocytosis, higher peripheral blood eosinophils (percentage and absolute counts), as well as longer hospitalization days . In addition, these patients exhibited higher alveolar-arterial oxygen gradient (A-aO2) but did not reach statistical significance (P = 0.09). Upper respiratory tract infection was common but not different between study subjects and controls.The NFE II group exhibited higher heart rate (HR), respiratory rate (RR), and diastolic blood pressure (DBP). They had higher FiOP < 0.05). Doses of corticosteroids have not been sufficiently recorded into the charts and are missing from our study. Multiple logistic regression conducted with covariates that had no more that 25% missing data identified DBP (P = 0.0041) and RR (P = 0.0024) as independent clinical risk factors for intubation. The Roc analysis determined that the length of hospitalization, absolute eosinophils, and per cent eosinophilia had the larger area under the curve for intubation .NFE II group had a significant history of prior intubation and prior usage of inhaled corticosteroids comparing to controls . Although they may also exhibit remodeling, our primary goal was to investigate the pathology of the most severe asthma population. No conclusions can be made regarding the clinical characteristics associated with remodeling in those cases, which was the secondary purpose of this study. Retrospectively collected NFE II group exhibited, however, very similar characteristics with NFE I group in terms of age, race, sex, history, and medication used. They also had a similar mainly low socioeconomic status as most of them were underinsured and the study derived from a public hospital. As a result, we believe that clinical characteristics of NFE II patients, which are compatible with the currently known phenotype of Near Fatal Asthma (NFA) , may not be extrapolated to NFE I patients and therefore associated with remodeling. Retrospective population provides, however, a very good reflection of the prospective arm of the study.A second limitation is the lack of pulmonary function testing and of other missing data . We attribute the former to the poor monitoring of asthma that these patients showed, an element explained in detail here in after. Regarding the latter, these data have been collected retrospectively from charts frequently over a decade old and were unable to be retrieved. A third limitation of our study is the only semiquantitative description of tissue specimens. We acknowledge that limitation and hope to be addressed in a future study.Remodeling in near fatal asthma is a reasonable finding, along with evidence of acute and chronic inflammation. Epithelial basal lamina thickening, increased smooth muscle mass, vascularity, and goblet cell hyperplasia are all observed in various asthma stages  –12. Althβ (transforming growth factor-β), conversion of fibroblasts to myofibroblasts, and excessive extracellular matrix expression (ECM). Phipps et al. and Flood-Page and colleagues found that patients with atopic asthma who received mepolizumab, an anti-IL5 antibody, exhibited a decrease in lung eosinophilia along with a reduction of markers of remodeling, suggesting a close association [A major finding in the tissue specimens of near fatal asthmatics in our study is the combination of predominant eosinophilic inflammation with remodeling. While eosinophils have been associated, as found in our study, with persistent, severe asthma, their role in the pathogenesis of remodeling is not so clear. Eosinophils have been shown to induce remodeling through the production of a strong fibrogenic cytokine, TGF-ociation , 14. Micociation , 16. UnfThe use of inhaled corticosteroids (CSs) was rather low in our study subjects (17% in ME II group and 48% in NFE II group) and signifies nonadherence to current treatment guidelines , 19. TheAlthough cases of COPD have been excluded from the study, a significant proportion of study subjects were current or former smokers. Thus, certain of the study subjects might have coexistent asthma and COPD or may have been misdiagnosed as asthma instead of COPD. In the absence of pulmonary function testing and scheduled posthospitalization visit, this issue may not be addressed adequately.In conclusion, this is a descriptive study showing that eosinophilic inflammation and remodeling are the predominant features of near fatal asthma exacerbations. These exacerbations are associated with a history of prior endotracheal intubation and use of inhaled CS. The lack of a moderate severity exacerbation control group, pulmonary function testing, and the semiquantitative pathological description is the limitations of our study.


Pulmonary arterial hypertension is a rare disease, which requires a high index of suspicion to diagnose when patients initially present. Initial symptoms can be nonspecific and include complaints such as fatigue and mild dyspnea. Once the disease is suspected, echocardiography is used to estimate the pulmonary arterial (PA) pressure and to exclude secondary causes of elevated PA pressures such as left heart disease. Right heart catheterization with vasodilator challenge is critical to the proper assessment of pulmonary hemodynamics and to determine whether patients are likely to benefit from vasodilator therapy. Pathologically, the disease is characterized by deleterious remodeling of the distal pulmonary arterial and arteriolar circulation, which results in increased pulmonary vascular resistance. In the last fifteen years, medications from three different classes have been approved for the treatment of pulmonary arterial hypertension. These include the prostanoids, endothelin receptor antagonists, and phosphodiesterase-5 inhibitors. Pulmonary arterial hypertension (PAH) is a progressive incurable disease that is characterized by extensive remodeling of the pulmonary circulation predominantly in the distal pulmonary arteries and arterioles. Proliferation of pulmonary vascular endothelial and smooth muscle cells leads to intimal and medial thickening of the pulmonary resistance vessels . In someAlthough PAH is a rare disease, it often strikes patients who are otherwise healthy in the middle of their life. Its devastating impact on the lives of thousands of people has led to an intense focus of research in pulmonary vascular biology over the last quarter century that has resulted in the development of numerous new therapies that have improved the prognosis considerably. Unfortunately, a cure for PAH has not yet been forthcoming and long-term survival remains poor. Even with modern medical therapy, most patient experience progression of their disease and many are referred for lung transplantation. The discovery this decade of a genetic defect that is associated with a substantial number of cases has raised the hope that a cure for PAH may eventually be found , 4.The pathogenesis of PAH remains unclear. However, several important imbalances in mediators of pulmonary vascular cell growth and apoptosis have been described. Patients with PAH demonstrate a decrease in the synthesis of prostacyclin. This potent pulmonary vasodilator also has important inhibitory effects on platelet aggregation and cellular proliferation. Patients with PAH have decreased pulmonary expression of prostaglandin synthase, the major enzyme responsible for its synthesis from the arachidonic acid pathway. They also have a reduction in the circulating levels of PGI2 relative to circulating levels of thromboxane . These cEndothelin is a potent vasoconstrictor and smooth muscle mitogen secreted by the pulmonary endothelium. Immunohistochemistry studies have demonstrated increased expression of endothelin in the obliterative vascular lesions found in the lungs of patients with PAH . CirculaIn the healthy person, the pulmonary vasculature dilates in response to increased flow allowing the lung to accommodate a marked increase in blood flow during exercise without much of a rise in PA pressure. Synthesis and release of nitric oxide (NO) by endothelial nitric oxide synthase (eNOS) in the pulmonary vascular endothelium plays an important role in flow-mediated vasodilation in the pulmonary circulation. Patients with PAH have decreased pulmonary expression of eNOS and decreased levels of NO in exhaled air, raising the possibility that decreased NO synthesis contributes to the rise in PVR , 9. The The current accepted definition of PAH is a mean PA pressure of greater than 25 mm Hg with a mean pulmonary capillary wedge pressure of less than 15 mm Hg and a pulmonary vascular resistance (PVR) of >3 Woods units. Elevation in peak PA pressure reported on transthoracic echocardiogram is not uncommon, but in the great majority of cases, it is not caused by PAH. A number of pathological states can elevate PA pressures, most notably chronic heart and lung diseases. During the past several decades, the World Health Organization (WHO) has proposed a new terminology to describe the various forms of pulmonary hypertension. The most recent update to this classification system was the 4th World Symposium on Pulmonary Hypertension in Dana Point, California, in 2008  Table 1Table 1.WHO group 1 is termed PAH and is characterized by a diffuse pulmonary vasculopathy of the pulmonary arteries and arterioles that is progressive in nature and results in marked increases in the transpulmonary pressure gradient. This group is further subdivided into idiopathic PAH  and heritable PAH. The latter is used to describe PAH associated with a number of genetic mutations that have been linked to the development of PAH , 4. PAH WHO group 2 refers to elevation of PA pressure that is caused by an increase in pulmonary venous pressure from left-sided heart disease. This type of pulmonary hypertension is termed pulmonary venous hypertension and is characterized by a nearly normal pulmonary vascular resistance (<3 Woods Units). WHO group 3 includes pulmonary hypertension that occurs with chronic lung diseases such as emphysema, interstitial lung disease, and sleep disordered breathing. PA pressure elevation is usually moderate in this group and often reflects the severity of the underlying lung disease. WHO group 4 refers to pulmonary hypertension that occurs in association with venous thromboembolic disease. The last WHO group is reserved for a variety of diseases known to be associated with pulmonary hypertension for unclear reasons such as pulmonary sarcoidosis.In addition to the etiological grouping, the WHO also uses a grading score for functional class based on the model developed for heart failure by the New York Heart Association. Patients are designated as one of four functional classifications based on the severity of their dyspnea and physical limitations . The WHOIdiopathic PAH is the most common form of the disease affecting 40%–50% of the patient in WHO diagnosis group 1. The estimated incidence of IPAH ranges from 7–50 per million , 13. AppPrompt diagnosis of PAH in an early stage is important so that treatment may be started before right heart failure occurs. However, time to diagnosis once symptoms develop is typically long and is impacted by the lack of specific disease symptoms and the rarity of the disease that results in a low level of clinical suspicion. Thirty years ago, the mean time between symptom onset and diagnosis was 2 years , 14. PatWhen PAH is suspected from the patient's symptoms or detected by echocardiogram performed for other reasons, further evaluation should start with a complete history and physical exam. A thorough interview may reveal a family history of early death due to heart failure or other events suggestive of pulmonary hypertension. A history of other heart and lung diseases, connective tissue disease, venous thrombosis, pulmonary embolism, or hypercoagulopathy should be sought. Patients should be questioned about risk factors for HIV infection, hepatitis, or cirrhosis and asked if they are aware of any congenital or rheumatic heart disease and an assessment should be made regarding the likelihood of sleep disordered breathing. Finally, patients should be questioned about previous use of amphetamines or other appetite suppressants and travel to or immigration from countries known to be endemic for schistosomiasis.Physical signs of elevated PA pressure are few and may be subtle. Pulmonary hypertension often results in enough tricuspid regurgitation to produce a systolic murmur best heard at the right sternal border. The second heart sound is often split due to enlargement of the right ventricle and the pulmonic component may be accentuated . Signs oThe examiner should also look for signs of connective tissue diseases such as malar rash, telangiectasias, calcinosis, sclerodactyly, and Raynaud's disease. Evidence of liver cirrhosis or portal hypertension and other diseases that can contribute to the development of pulmonary hypertension such as thyroid disease, obstructive sleep apnea, and hemolytic anemia should be sought. Signs of chronic heart disease, pulmonary fibrosis, or emphysema suggest that patients may have pulmonary venous hypertension or pulmonary hypertension associated with chronic lung disease rather than PAH. Patients are most often referred for evaluation of pulmonary hypertension because of progressive dyspnea that is not readily explained by other diseases or because elevated PA pressure was found during an echocardiogram. The approach to diagnosis is similar in both scenarios. An ECG and chest film are helpful in excluding subclinical heart or lung disease and may provide additional information about the severity of the patient's PAH. Signs of elevated right-sided pressure and right heart strain such as right axis deviation, increased p-wave amplitude in lead 2 and t-wave inversion in the precordial leads suggest more severe pulmonary hypertension. Enlarged right ventricle and proximal pulmonary arteries can be seen on chest film in some patients and peripheral pruning of the pulmonary vessels may be apparent. Increased filling of the retrosternal air space on lateral chest film suggests right ventricular enlargement. Hilar adenopathy may suggest sarcoidosis.Pulmonary function tests should be done to help exclude obstructive or restrictive lung disease and obtain a baseline measure of diffusion capacity. Adequate oxygenation should be confirmed by pulse oximetry or arterial blood gas analysis. When suspected, hypercarbia should be excluded by arterial or venous blood gas analysis. The 6 minute walk test has become a mainstay of evaluating PAH patients and the 6 minute walking distance is the most frequent primary efficacy outcome measured in clinical trials of presently available PAH therapies. The baseline 6 minute walking distance should be established ideally before initiating treatment. Change in blood pressure, pulse and oxygen saturation should be recorded along with the Borg dyspnea index. Patients who have a fall in oxygenation saturation are more likely to have pulmonary vascular disease. Inability to increase blood pressure suggests impaired cardiac function. A formal sleep study may not be necessary in all patients but should be considered in those with symptoms suggestive of obstructive sleep apnea or other forms of sleep disordered breathing.Laboratory blood tests important in the initial evaluation of PAH include HIV test, liver function tests, thyroid function tests, and serologies to rule out lupus, scleroderma, and rheumatoid arthritis. A baseline BNP and serum sodium may be helpful in monitoring disease progression , 17. SexA good quality transthoracic echocardiogram is extremely useful in evaluating pulmonary hypertension , 19. RigThe echocardiogram can also help look for left-sided heart disease that may elevate PA pressure by increasing pulmonary venous pressures. Enlargement of the left atrium suggests pulmonary venous hypertension or mitral valve disease. Decreased left ventricular systolic function is readily apparent when present. Left ventricular hypertrophy raises the concern of pulmonary venous hypertension from diastolic dysfunction and some echocardiographers will report impaired diastolic filling of the left ventricle by examining the pattern of blood flow through the mitral valve during diastole. Findings such as these should alert the clinician to the possibility of left sided heart disease and consideration of additional cardiac studies.Definitive diagnosis of PAH cannot be made without accurate measurement of PA pressure and an assessment of pulmonary venous pressure and cardiac output. At the present time, this essentially requires right heart catheterization. Right heart catheterization for the initial evaluation of PAH has become the standard of care as described by the recommendations in the Dana Point 2009 WHO consensus as well as by guidelines written and adopted by the American College of Cardiologists, American Thoracic Society, and the American College of Chest Physicians . In addiSome clinicians rely on echocardiogram to make the diagnosis of PAH because it is less invasive, but because of the rarity of PAH relative to that of other diseases that increase peak PA pressure, it is necessary to demonstrate that the elevation in PA pressure is due to increased PVR. This can only be done by direct catheterization of the pulmonary artery, measurement of PA pressure and downstream pulmonary venous pressure using the pulmonary artery occlusion technique, and accurate measurement of cardiac output. It is also important to realize the PA pressure is dependent on cardiac output and that a moderate elevation of PA pressure on echocardiogram can be seen in severe PAH when right ventricular failure occurs. Right heart catheterization also allows for evaluation of left to right intracardiac shunts and acute reactivity to pulmonary vasodilators. Medications used for vasodilator testing while the right heart catheter is in place include inhaled nitric oxide, intravenous epoprostenol, or intravenous adenosine. The definition of a positive vasodilator response is a greater than 10% decrease in pulmonary artery mean pressure to an absolute mean value of less than 40 mm/Hg with no decrease in cardiac output. In addition to acute vasodilator testing, patients with mild or moderate pulmonary hypertension are sometimes exercised with the right heart catheter in place to determine if PA pressure rises during exercise.Over the last 15 years, 9 different drugs representing 3 new classes of medication have been developed for the treatment of PAH . This exAs mentioned previously, patients with pulmonary hypertension have reduced expression of endothelial nitric oxide synthase in the pulmonary vasculature . While nSildenafil was the first PDE5 inhibitor approved for the treatment of PAH. It had previously demonstrated potential in the treatment of pulmonary hypertension in animal models  and as aP < 0.001 for all treatment groups compared to placebo). Analysis of hemodynamic data demonstrated a significant improvement in pulmonary vascular resistance with sildenafil, and there was a dose-response effect with the 80 mg group demonstrating the greatest reduction (−261 dyn*sec*cm−5 with 80 mg as compared with an increase of 49 dyn*sec*cm−5 in the placebo group). The improvement in cardiac index also exhibited a dose response relationship although the improvement in the 20 mg group compared with placebo did not reach statistical significance. The incidence of clinical worsening did not differ between the two treatment groups. Two hundred fifty nine of the patients enrolled in a long-term extension study of 80 mg of sildenafil three times daily. During a one year follow up, 15 of those patients withdrew, and 14 died. Of the remaining 230 patients, eight were receiving additional therapy. Analysis of the other 222 patients revealed that the mean change from baseline in the six-minute walking distance was 51 meters (95% CI of 41 to 60). Analysis of the primary endpoint demonstrated an increase in walk distance for all groups randomized to sildenafil. At week 12, the mean placebo-corrected effects were 45 m, 46 m, and 50 m for those in the 20 mg, 40 mg, and 80 mg groups respectively  Study Group randomized 405 treatment-naive or bosentan treated patients with PAH to placebo or to tadalafil 2.5, 10, 20, or 40 mg orally once daily . The stuP < 0.01. In addition, time to clinical worsening was significantly improved in the tadalafil 40 mg group (P = 0.041). Fifty-three percent of the patients were on background therapy with bosentan during the trial. Bosentan-naïve patients given 40 mg of tadalafil had a greater increase in walking distance (44 meters) than patients on background bosentan (23 meters). Further supporting the idea of a blunted improvement in exercise capacity for patients already receiving PAH therapy, bosentan-naïve patients tended to have better results in all secondary end points. The improvement in 6-minute walking distance was sustained for 10 months in a subgroup of patients who had been enrolled in a long-term extension study of 20 or 40 mg tadalafil. Of the 213 patients who had completed 44 weeks of tadalafil therapy at the time of publication, the mean change in 6-minute walking distance was 37 meters at 16 weeks and 38 m after 44 weeks.Tadalafil at doses of 10, 20, and 40 mg, compared with placebo, improved the six-minute walk distance in a dose-dependent manner at week 16 . However, the 40 mg dose of tadalafil was the only one that reached the prespecified value of statistical significance of P < 0.002 for the comparison of the treatment groups). With adjustment for baseline differences in 6-minute walk distance and vasodilator use, the epoprostenol treated group still exhibited a median increase of 31 meters, while the conventional therapy group exhibited a median decrease of 29 meters (P < 0.02). Quality of life as measured by the Chronic Heart Failure Questionnaire improved only in the epoprostenol-treated group. The mean PA pressure decreased 4.8 ± 1.3 mm Hg in the epoprostenol treated group, while it increased 1.9 ± 1.6 mm Hg in the conventional therapy group . Eight patients randomized to the conventional therapy group died during the 12 weeks of the study, but none of those assign to treatment with epoprostenol died (P < 0.05).Prostacyclin was discovered in 1976 and was found to have vasodilating, antiplatelet, and antiproliferative properties. The chemical analogue epoprostenol was synthesized and tested in the same year . The finSide effects are common at higher doses of prostacyclin infusion and occur as a result of systemic vasodilation. The most common side effects include facial flushing, headache, and jaw pain, especially when chewing. Other adverse events include diarrhea and bone pain in the legs and bottom of the feet. The side effects were reported more frequently in the group that received epoprostenol. Serious complications included four episodes of nonfatal catheter-related sepsis and one episode of nonfatal catheter related thrombosis resulting in a paradoxical embolism.P < 0.001). Hemodynamic assessment demonstrated a decrease in mean PA pressure of 5.0 ± 1 mm Hg in the intervention group, while the conventional therapy arm demonstrated an increase in mean PA pressure of 0.94  ±  1.1 mm Hg (difference between groups of −5.97 mm Hg with 95% CI of −8.98 to −2.96 mm Hg). Cardiac index increased 0.5 ± 0.08 L/min/m2 with the addition of epoprostenol, while it decreased 0.1 ± 0.08 L/min/m2 in the control arm (difference between groups of 0.60 L/min/m2 with 95% CI of 0.39 to 0.81 L/min/m2). Adverse events related to the use of a continuous intravenous infusion system for epoprostenol included 4% incidences of sepsis, cellulitis, hemorrhage, and pneumothorax. Similar numbers of patients died in each group, and minor side effects related to the use of epoprostenol were in keeping with other studies.Epoprostenol has also been shown to be effective in the treatment of PAH associated with scleroderma. A short-term hemodynamic study of the effect of intravenous epoprostenol in patients with PAH associated with scleroderma demonstrated reductions in pulmonary vascular resistance and an increase in cardiac output . AnotherP = 0.006). While the overall treatment effect was small, patients in the highest quartile dose of treprostinil (>13.8 ng/kg/min) had a placebo adjusted increase in walking distance of 36.1 ± 10 meters. Interestingly, the patients with the lowest baseline 6-minute walking distance in the treprostinil group experienced the greatest benefit. Patients who walked less than 150 meters at baseline experienced a treatment effect of 51 ± 16 meters (P = 0.002), while patients who walked more than 351 meters at baseline experienced no substantial treatment effect. These findings suggest that treprostinil infusion may be more effective when used at higher doses in patients with more advanced PAH.Epoprostenol's short plasma half-life of approximately 3 minutes mandates continuous intravenous infusion that is both cumbersome for the patient and associated with potentially serious catheter-related complications. Treprostinil is a stable prostacyclin analogue that is chemically stable at room temperature and has a half-life of three to four hours. These properties permit continuous subcutaneous infusion via a needle that can be placed under the skin by the patient. Initial data, published in abstract form, demonstrated the safety and beneficial acute hemodynamic effects of this approach to prostacyclin therapy , 37. The2 compared to a decrement of 0.06 L/min/m2 in the placebo group (P = 0.0001 for comparison of change from baseline), and the mean PA pressure decreased 2.3 ± 0.5 mm Hg as compared to an increase of 0.7 ± 0.6 mm Hg in the placebo group (P = 0.0003 for comparison of change from baseline). The treprostinil-treated group had a higher incidence of infusion site pain (85% versus 27%) and also had a higher incidence of classic prostacyclin related side effects. Three patients in the treprostinil-treated group experienced an episode of gastrointestinal bleeding. Two of these patients had a supratherapeutic international normalized ratio (INR), and all of the episodes resolved without significant consequences.Analysis of cardiopulmonary hemodynamics in this study demonstrated statistically significant improvements in all variables with treprostinil treatment. In particular, cardiac index improved by 0.12 L/min/mSubcutaneous treprostinil infusion allows for the delivery of continuous prostacyclin infusion without the need for a central catheter, and thereby eliminates the danger of central venous catheter infection. At the same time, its longer half-life provides a greater cushion of time for restarting the infusion in the event that it is interrupted because of pump failure or loss of intravenous access. These advantages increase the number of patients who are eligible for continuous prostacyclin infusion therapy. Unfortunately, most patients experience some degree of discomfort at the infusion site, and in approximately a quarter of patients, the degree of discomfort is too great to continue therapy. In addition to subcutaneous infusion, treprostinil can also be used as a continuous intravenous infusion .Prostacyclin analogues also act as effective pulmonary vasodilators when inhaled. Aerosolized prostacyclin was developed to mitigate the incidence of systemic side effects that occur with prostacyclin therapy and to obviate the need for subcutaneous or central venous access and tethering the patient to a battery operated infusion pump. Another potential advantage of inhaled prostacyclin is the improvement of ventilation perfusion matching. This may occur because the greatest pulmonary vasodilator effect occurs in areas of the lung that are well ventilated and thereby reduces blood flow to areas that are not. Several early trials demonstrated the feasibility of this approach in the treatment of PAH. The pivotal trial was performed in Europe and randomized 203 patients with WHO functional class III or IV pulmonary hypertension to inhaled iloprost or placebo . Most ofP = 0.007). There were no significant differences amongst groups with different types of pulmonary hypertension. Adverse effects were similar between the two groups although there was a statistically significant higher incidence of syncope in the treatment group. These events were not associated with clinical deterioration, and the authors concluded that the higher incidence of syncope may have resulted from treated patients increasing their activity too quickly. Given the concern that intermittent inhalations (as opposed to a continuous intravenous infusion of a prostanoid) may not have a durable effect throughout the day, the 12-week hemodynamic assessment included measurement of preinhalation values. While measurements done after iloprost inhalation showed the most improvement from baseline compared to the placebo group, the pre-inhalation change in PVR from baseline of −9 ± 275 dyn·sec·cm−5 was still significantly better than the 96 ± 322 dyn·sec·cm−5 increase from baseline in the placebo group (P < 0.01).Given their desire for a robust measure of efficacy and the fact that continuous intravenous epoprostenol had previously demonstrated a survival advantage, these investigators chose a combined primary endpoint of an improvement of at least one NYHA functional class and a 10 percent improvement in 6-minute walking distance. This combined endpoint was met by 16.8 percent of the patients who received iloprost as compared with 4.9 percent of the patients who received placebo  and 20 meters at week 12 (P = 0.0004). Importantly, there was evidence that benefit was maintained throughout the 6-hour interval between treatments. The median improvement in 6-minute walking distance measured just before inhalation of treprostinil was 14 meters at week 12 (P = 0.0066). Amongst the secondary end points, only quality-of-life measures and NT-proBNP demonstrated improvement with inhaled treprostinil compared to placebo.Of the 235 patients randomized, 23 withdrew from the study prematurely. Eight patients withdrew consent, while the rest discontinued due to adverse events or worsening disease. The between treatment median difference in change from baseline in peak six-minute walk distance was 19 meters at week 6  tested for the treatment of PAH was bosentan, which is a nonselective agent that has similar binding affinity for ET-A and ET-B. After a small initial double-blind placebo-controlled study  demonstrP < 0.001). There was a nonsignificant trend towards further improvement with the 250 mg dose as compared to 125 mg. Bosentan also significantly increased the time to clinical worsening as compared to placebo. The overall incidence of adverse events between the two groups was similar with the exception of a dose-dependent increase in the incidence of liver transaminase elevation with bosentan therapy.At the conclusion of sixteen weeks, 6 minute walking distance decreased an average of 8 meters in the placebo group and increased 36 m in the combined bosentan groups  were concurrent randomized, placebo-controlled, double-blind studies which differed in terms of the investigative sites and doses of ambrisentan. ARIES-1 randomized patients in North or South America to placebo or 5 or 10 mg per day of ambrisentan. ARIES-2 randomized patients in Europe, the middle east, or Australia to placebo or 2.5 or 5 mg of ambrisentan. In both studies, patients had idiopathic PAH or PAH associated with connective tissue disease, HIV infection, or anorexigen use and the primary endpoint was the change in 6-minute walk distance from baseline to week 12. The results of both studies were published together .P = 0.008) and 51 meters (P < 0.001) for the 5 and 10 mg doses, respectively, in ARIES 1 and 32 meters (P = 0.022) and 59 meters (P < 0.001) for the 2.5 and 5 mg doses in ARIES-2. Improvements in secondary end points such as time to clinical worsening, functional class, symptom assessments, and B-type natriuretic peptide measurements were also seen for the 5 and 10 mg doses. In order to assess the effects of more sustained therapy, these trials also included a long-term study that was open to all patients who completed the initial study and all placebo patients who discontinued because of early escape. Of the 280 patients who completed 48 weeks of ambrisentan monotherapy, the improvement from baseline in 6-minute walk distance was 39 meters.At 12 weeks, the six-minute walk distance increased in all ambrisentan groups. The mean placebo-corrected treatment effects were 31 meters (A third endothelin antagonist, sitaxsentan, demonstrated promise in PAH based on its selectivity for the ET-A receptor, and on a lack of significant interaction with sildenafil. However, the manufacturer withdrew it from all available markets and halted further clinical trials in December 2010 due to problems with hepatotoxicity.Both bosentan and Ambrisentan have been shown to cause transient elevation of liver transaminases, and these laboratory tests should be monitored at least monthly for as long as the patient is taking them. The ERAs have also been associated with small decreases in hemoglobin and are highly teratogenic. Sexually active patients on these medications should use a double-barrier method of contraception, and fertile woman should be monitored closely for pregnancy.Few clinical trials have compared the safety and efficacy of one PAH drug or one class of PAH drugs to another in a head-to-head trial. Wilkins et al.  found noThe use of a combination of drugs from different PAH classes is an area of great interest. Several randomized placebo controlled trials have examined the benefit of adding a second PAH medication to patients who failed to respond to or have deteriorated on their initial therapy using a randomized placebo-controlled design , 51–53. Continued long-term care is an important aspect of patient survival once diagnosis and PAH therapy has started. Frequent office visit are a necessity for patients starting any continuous infusion or inhaled therapy to ensure accuracy and compliance to therapy. Patients starting on parenteral therapy are seen biweekly to monthly when first beginning therapy. Once a patient's condition has stabilized, they are typically followed in the office every 1–3 months. Along with frequent office visits, 6-minute walk tests, echocardiogram, and plasma BNP levels are performed 2-3 times a year or more often if the patient requires frequent titration of parenteral therapy or addition or change in therapy due to suboptimal treatment response.Concurrent medications used for PAH patients include diuretics to control peripheral edema as necessary, digoxin, warfarin, and supplemental oxygen. Digoxin may improve right ventricular contractility, but its clinical effect is small . Some stInpatient hospitalization may be required for patient with worsening right heart failure and fluid overload for IV diuresis and stabilization of disease. If a patient's condition does not improve despite an aggressive course of therapy, a repeat right heart catheterization may be warranted to reassess pulmonary hemodynamics.Referral for lung transplantation should be considered soon after initial diagnosis and initiation of therapy, especially if there is a delay in adequate response to therapy. Lung transplant evaluation should be completed early in the course of therapy, as the evaluation process and wait for suitable donor organ is lengthy, and the patients who fail to respond to medical therapy can deteriorate suddenly. Timing of lung transplant is important in that a patient must be stable enough to tolerate transplant surgery but have a worsening prognosis with a decreased likelihood of surviving without transplant.