We postulated that the hypoxic response in sickle cell disease (SCD) contributes to altered gene expression and pulmonary hypertension, a complication associated with early mortality.
Methods and Results
To identify genes regulated by the hypoxic response and not other effects of chronic anemia, we compared expression variation in peripheral blood mononuclear cells from 13 SCD subjects with hemoglobin SS genotype and 15 Chuvash polycythemia subjects (VHLR200W homozygotes with constitutive up-regulation of hypoxia inducible factors in the absence of anemia or hypoxia). At 5% false discovery rate, 1040 genes exhibited >1.15 fold change in both conditions; 297 were up-regulated and 743 down-regulated including MAPK8 encoding a mitogen-activated protein kinase important for apoptosis, T-cell differentiation and inflammatory responses. Association mapping with a focus on local regulatory polymorphisms in 61 SCD patients identified expression quantitative trait loci (eQTL) for 103 of these hypoxia response genes. In a University of Illinois SCD cohort the A allele of a MAPK8 eQTL, rs10857560, was associated with pre-capillary pulmonary hypertension defined as mean pulmonary artery pressure ≥25 and pulmonary capillary wedge pressure ≤15 mm Hg at right heart catheterization (allele frequency=0.66; OR=13.8, P=0.00036, n=238). This association was confirmed in an independent Walk-PHaSST cohort (allele frequency=0.65; OR=11.3, P=0.0025, n=519). The homozygous AA genotype of rs10857560 was associated with decreased MAPK8 expression and present in all 14 identified pre-capillary pulmonary hypertension cases among the combined 757 patients.
Our study demonstrates a prominent hypoxic transcription component in SCD and a MAPK8 eQTL associated with pre-capillary pulmonary hypertension.
sickle cell disease; MAPK8; hypoxic response; expression quantitative trait loci; association mapping; pre-capillary pulmonary hypertension
In congenital Chuvash polycythemia (CP), VHLR200W homozygosity leads to elevated hypoxia inducible factor (HIF) levels at normoxia. CP is often treated by phlebotomy resulting in iron deficiency, permitting us to examine the separate and synergistic effects of iron deficiency and HIF signaling on gene expression. We compared peripheral blood mononuclear cell gene expression profiles of eight VHLR200W homozygotes with 17 wildtype individuals with normal iron status and found 812 up-regulated and 2120 down-regulated genes at false discovery rate 0.05. Among differential genes we identified three major gene regulation modules involving induction of innate immune responses, alteration of carbohydrate and lipid metabolism, and down-regulation of cell proliferation, stress-induced apoptosis and T-cell activation. These observations suggest molecular mechanisms for previous observations in CP of lower blood sugar without increased insulin and low oncogenic potential. Studies including 16 additional VHLR200W homozygotes with low ferritin indicated that iron deficiency enhanced the induction effect of VHLR200W for 50 genes including hemoglobin synthesis loci but suppressed the effect for 107 genes enriched for HIF-2 targets. This pattern is consistent with potentiation of HIF-1α protein stability by iron deficiency but a trend for down-regulation of HIF-2α translation by iron deficiency overriding an increase in HIF-2α protein stability.
We aimed to identify risk factors for acute pulmonary events in children and adolescents in the Pulmonary Hypertension and the Hypoxic Response in SCD (PUSH) study.
Patients with hemoglobin SS (n=376) and other sickle cell genotypes (n=127) aged 3-20 years were studied at four centers in a cross-sectional manner. A sub-group (n=293) was followed for a median of 21 months (range 9-35).
A patient-reported history of one or more acute pulmonary events, either acute chest syndrome (ACS) or pneumonia, was obtained in 195 hemoglobin SS patients (52%) and 51 patients with other genotypes (40%). By logistic regression, history of acute pulmonary events was independently associated with patient-reported history of asthma (p<0.0001), older age (p=0.001), >3 severe pain episodes in the preceding 12 months (p=0.002), higher tricuspid regurgitation velocity (TRV) (p=0.028), and higher white blood cell (WBC) count (p=0.043) among hemoglobin SS patients. History of acute pulmonary events was associated with >3 severe pain episodes (p=0.009) among patients with other genotypes. During follow-up, 43 patients (15%) had at least one new ACS episode including 11 without a baseline history of acute pulmonary events. History of acute pulmonary events (odds ratio 5.4; p<0.0001) and younger age (odds ratio 0.9; p=0.010) were independently associated with developing a new episode during follow-up.
Asthma history, frequent pain and higher values for TRV and WBC count were independently associated with history of acute pulmonary events in hemoglobin SS patients and frequent pain was associated in those with other genotypes. Measures to reduce pain episodes and control asthma may help to decrease the incidence of acute pulmonary events in SCD.
sickle cell disease; acute chest syndrome; vaso-occlusive crisis; asthma; pain
In Chuvash polycythemia, a homozygous 598C>T mutation in the von Hippel-Lindau gene (VHL) leads to an R200W substitution in VHL protein, impaired degradation of α-subunits of hypoxia inducible factor (HIF)-1 and HIF-2, and augmented hypoxic responses during normoxia. Chronic hypoxia of high altitude is associated with decreased serum glucose and insulin concentrations. Other investigators reported that HIF-1 promotes cellular glucose uptake by increased expression of GLUT1 and increased glycolysis by increased expression of enzymes such as PDK. On the other hand, inactivation of Vhl in murine liver leads to hypoglycemia associated with a HIF-2-related decrease in the expression of the gluconeogenic enzymes genes Pepck, G6pc, and Glut2. We therefore hypothesized that glucose concentrations are decreased in individuals with Chuvash polycythemia. We found that 88 Chuvash VHLR200W homozygotes had lower random glucose and glycosylated hemoglobin A1c levels than 52 Chuvash subjects with wildtype VHL alleles. Serum metabolomics revealed higher glycerol and citrate levels in the VHLR200W homozygotes. We expanded these observations in VHLR200W homozygote mice and found that they had lower fasting glucose values and lower glucose excursions than wild-type control mice but no change in fasting insulin concentrations. Hepatic expression of Glut2 and G6pc but not Pdk2 was decreased and skeletal muscle expression of Glut1, Pdk1 and Pdk4 was increased. These results suggest that both decreased hepatic gluconeogenesis and increased skeletal uptake and glycolysis contribute to the decreased glucose concentrations. Further study is needed to determine whether pharmacologically manipulating HIF expression might be beneficial for treatment of diabetic patients.
VHL; hypoxia inducible factors; glucose; insulin; glycolysis; gluconeogenesis
How often elevated serum ferritin in primary-care patients reflects increased iron stores (normally 0.8 g in men, 0.4 g in women) is not known. The Hereditary Hemochromatosis and Iron Overload Screening (HEIRS) study screened 101,168 primary-care participants (44% Caucasians, 27% African-Americans, 14% Asians/Pacific Islanders, 13% Hispanics, 2% others). Follow-up clinical evaluation was performed in 302 of 333 HFE C282Y homozygotes regardless of iron measures and 1,375 of 1,920 nonhomozygotes with serum ferritin >300 μg/L (men), >200 μg/L (women) and transferrin saturation >50% (men), >45% (women). Quantitative phlebotomy was conducted in 122 of 175 C282Y homozygotes and 122 of 1,102 nonhomozygotes with non-transfusional serum ferritin elevation at evaluation. The estimated prevalence in the Caucasian population of C282Y homozygotes with serum ferritin >900 μg/L at evaluation was 20 per 10,000 men and 4 per 10,000 women; this constellation was predictive of iron stores >4 g in men and >2 g in women. The estimated prevalence per 10,000 of non-C282Y homozygotes with serum ferritin >900 μg/L at evaluation was 7 among Caucasians, 13 among Hispanics, 20 among African Americans, and 38 among Asians and Pacific Islanders, and this constellation was predictive of iron stores >2 g but <4 g. In conclusion, serum ferritin >900 μg/L after initial elevations of both serum ferritin and transferrin saturation is predictive of mildly increased iron stores in multiple ethnic populations regardless of HFE genotype. Serum ferritin >900 μg/L in male C282Y homozygotes is predictive of moderately increased iron stores.
Non-invasively assessed pulmonary pressure elevations and left ventricular diastolic dysfunction (LVDD) are associated with increased mortality in adults with sickle cell disease (SCD), but their relationship to exercise intolerance has not been evaluated prospectively.
Methods and Results
Echocardiography, six-minute walk distance, hemolytic rate, and serum concentrations of ferritin and erythropoietin were evaluated in a cohort of 483 subjects with homozygous hemoglobin S in the US and UK Walk-PHaSST study. Tricuspid regurgitation velocity (TRV), which reflects systolic pulmonary artery pressure, was 2.7 to <3.0 m/sec (mean±SD 2.8±0.1) in 26% of the subjects and ≥3.0 m/sec (3.4±0.4) in 11%. LV lateral E/e′ ratio, which has been shown to reflect LV filling pressure in other conditions but has not been studied in SCD, was significantly higher in the groups with TRV ≥2.7 m/sec. Increased hemolysis (P<0.0001), LV lateral E/e′ ratio (P=0.0001), BUN (P=0.0002) and erythropoietin (P=0.002) were independently associated with an increased TRV. Further, female gender (P<0.0001), older age (P<0.0001), LV lateral E/e′ ratio (P=0.014), and TRV (P=0.019) were independent predictors of a shorter six-minute walk distance.
Echocardiography-estimated elevated pulmonary artery systolic pressure and LV lateral E/e′ ratio were independently associated with poor exercise capacity in a large cohort of patients with sickle cell anemia. Controlled trials investigating whether strategies to prevent or delay pulmonary hypertension and/or LVDD will improve exercise capacity and long-term outcomes in sickle cell anemia should be considered.
sickle cell anemia; pulmonary hypertension; left ventricular diastolic dysfunction; echocardiography; six-minute walk
We analyzed entry data from 163 adult hemoglobin SS and Sβ0 thalassemia patients enrolled in the prospective Sickle Cell Pulmonary Hypertension Screening Study and stratified their ECHO-determined tricuspid regurgitant jet velocity (TRV) and serum creatinine concentration according to three blood pressure categories. TRV was ≥2.5 m/sec in 27% of the patients with systolic blood pressure (SBP) <120 mm Hg and diastolic blood pressure (DBP) <70 mm Hg, in 37% with SBP 120–139 mm Hg or DBP 70–89 mm Hg, and in 93% with SBP 140 mm Hg or DBP 90 mm Hg or higher (P <0.0005 for trend). Serum creatinine concentration was 1.0 mg/dL or higher in 7% of patients with SBP <120 mm Hg and DBP <70 mm Hg, in 17% with SBP 120–139 mm Hg or DBP 70–89 mm Hg and 50% with SBP 140 mm Hg or DBP 90 mm Hg or higher (P <0.0005 for trend). Over two years of follow-up, there were trends for more frequent progression to elevated TRV (P = 0.073) or creatinine (P = 0.038) values according to the higher systemic blood pressure categories. Our findings suggest that systolic SBP 120–139 mm Hg or DBP 70–89 mm Hg defines a category of relative systemic hypertension in patients with sickle cell disease that is associated with increased risk for pulmonary hypertension and renal dysfunction. Whether antihypertensive and/or nitric oxide donor therapy in sickle cell disease patients with relative hypertension prevents these and other complications should be determined by clinical trials.
Alcohol consumption is associated with increased iron stores. In sub-Saharan Africa, high dietary ionic iron and the ferroportin Q248H allele have also been implicated in iron accumulation. We examined the associations of ferroportin Q248H, alcohol and dietary iron with serum ferritin, aspartate aminotransaminase (AST) and alanine aminotransaminase (ALT) concentrations in African Americans.
Inner-city African Americans (103 men, 40 women) were recruited from the community according to reported ingestion of >4 alcoholic drinks per day or <2 per week. Typical daily heme iron, non-heme iron and alcohol were estimated using University of Hawaii’s multiethnic dietary questionnaire. Based on dietary questionnaire estimates we established categories of < versus ≥56 g alcohol per day, equivalent to 4 alcoholic drinks per day assuming 14 g alcohol per drink.
Among 143 participants, 77% drank <56 g alcohol/day and 23% ≥56 g/d as estimated by the questionnaire. The prevalence of ferroportin Q248H was 23.3% with alcohol >56 g/d versus 7.5% with lower amounts (P=0.012). Among subjects with no history of HIV disease, serum ferritin concentration had positive relationships with male gender (P=0.041), alcohol consumption (P=0.021) and ALT concentration (P=0.0001) but not with dietary iron intake or ferroportin Q248H. Serum AST and ALT concentrations had significant positive associations with male gender and hepatitis C seropositivity but not with alcohol or dietary iron intake or ferroportin Q248H.
Our findings suggest a higher prevalence of ferroportin Q248H with greater alcohol consumption, and this higher prevalence raises the possibility that the allele might ameliorate the toxicity of alcohol. Our results suggest that alcohol but not dietary iron contributes to higher body iron stores in African Americans. Studies with larger numbers of participants are needed to further clarify the relationship of ferroportin Q248H with the toxicity of alcohol consumption.
To identify factors that affect manifestations of sickle cell anemia we compared patients 11-30 years of age from University of Ibadan, Nigeria (n = 214) and University of Illinois at Chicago, USA (n = 209). Paralleling findings in the general populations of the two countries, the Chicago patients were more often overweight or obese as defined by the Centers for Disease Control (Atlanta, GA, USA) guidelines, and more often had elevated blood pressure (BP) as defined by the National Heart, Lung, and Blood Institute (Bethesda, MD, USA) guidelines. The Ibadan sickle cell anemia patients less frequently received the pneumococcal vaccine or hydroxyurea (HU) therapy. Consistent with lower rates of elevated BP and increased body mass index (BMI), stroke history was less frequent in the Ibadan patients ≥18 years old. Furthermore, in combined analyses, systolic and diastolic BP directly correlated with BMI, and elevated weight status independently associated with history of stroke [odds ratio (OR) 2.7, p = 0.019]. Our findings are consistent with the possibility that higher values for BMI and BP in Chicago sickle cell anemia patients may contribute to an increased risk of stroke and highlights the need for measures to reduce these risk factors. On the other hand, lower pneumococcal vaccination and HU therapy rates in Ibadan patients highlights the need for more improved vaccination coverage and for studies to define the role of HU therapy in Africa.
Hydroxyurea (HU); hypertension; pneumococcal vaccination; sickle cell anemia; weight
Background. The mechanisms of severe malarial anemia and cerebral malaria, which are extreme manifestations of Plasmodium falciparum malaria, are not fully understood.
Methods. Children aged <6 years from southern Zambia presenting to the hospital with severe malarial anemia (n = 72), cerebral malaria (n = 28), or uncomplicated malaria (n = 66) were studied prospectively. Children with overlapping severe anemia and cerebral malaria were excluded.
Results. Low interleukin 10 concentrations had the strongest association with severe anemia (standard β = .61; P < .001) followed by high tumor necrosis factor α and sFas concentrations, low weight-for-age z scores, presence of stool parasites, and splenomegaly (standard β = .15–.25; P ≤ .031); most of these factors were also associated with lower reticulocytes. Greater parasitemia was associated with higher interleukin 10 and tumor necrosis factor α concentrations, whereas sulfadoxizole/pyrimethamine therapy and lower weight-for-age z scores were associated with lower interleukin 10 levels. Thrombocytopenia and elevated tissue plasminogen activator inhibitor 1 levels had the strongest associations with cerebral malaria (standard β = .37 or .36; P < .0001), followed by exposure to traditional herbal medicine and hemoglobinuria (standard β = .21–.31; P ≤ .006).
Conclusions. Predictors of severe malarial anemia (altered immune responses, poor nutrition, intestinal parasites, and impaired erythropoiesis) differed from those of cerebral malaria (thrombocytopenia, herbal medicine, and intravascular hemolysis). Improved preventive and therapeutic measures may need to consider these differences.
The genetic bases of the highly variable degrees of anaemia and haemolysis in persons with Hb SS are not fully known, but several studies have indicated that G6PD deficiency is not a factor. The G6PD202A and G6PD376G alleles and α-thalassaemia were determined by molecular genetic testing in 261 children and adolescents with Hb SS in a multicentre study. G6PD202A,376G (G6PD A-) was defined as hemizygosity for both alleles in males and homozygosity in females. Among the participants 41% were receiving hydroxycarbamide.
The prevalence of G6PD202A,376G was 13.6% in males and 3.3% in females with an overall prevalence of 8.7%. G6PD202A,376G was associated with a 10 g/l decrease in haemoglobin concentration (P=0.008) but not with increased haemolysis as measured by lactate dehydrogenase, bilirubin, aspartate-aminotransferase, reticulocyte count or a haemolytic component derived from these markers (P>0.09). Similar results were found within a sub-group of children who were not receiving hydroxycarbamide. By comparison, single and double α-globin deletions were associated with progressively higher haemoglobin concentrations (P=0.005 for trend), progressively lower values for haemolytic component (P=0.007), and increased severe pain episodes (P<0.001).
In conclusion, G6PD202A,376G may be associated with lower haemoglobin concentration in sickle cell anaemia by a mechanism other than increased haemolysis.
sickle cell anaemia; G6PD; haemolysis; alpha-thalassaemia; haemoglobin concentration
N-terminal (NT) pro-brain natriuretic peptide (proBNP) ≥160 ng/l has a 78% positive predictive value for pulmonary hypertension and is associated with increased mortality in US sickle cell disease patients, but the importance in sickle cell disease patients in Africa is not known. In a cross-sectional study at Ahmadu Bello University Teaching Hospital, Shika-Zaria, Nigeria, we studied 133 hydroxycarbamide-naïve Nigerian sickle cell anaemia patients aged 18-52 years at steady-state and 65 healthy controls. Twenty-six percent of patients versus 5% of controls had NT-proBNP ≥160 ng/l (P=0.0006). By logistic regression among the patients, human immunodeficiency virus seropositivity, higher serum ferritin and lower haemoglobin or higher lactate dehydrogenase independently predicted elevated NT-proBNP. After adjustment for haemoglobin concentration, elevated NT-proBNP concentration was associated with an estimated 7.8-fold increase in the odds of severe functional impairment, defined as an inability to walk more than 300 m in six min (95% confidence interval 1.5-32.6; P=0.005). Similarly, elevated tricuspid regurgitation velocity was associated with an estimated 5.6-fold increase in the odds of functional impairment (95% confidence interval 1.5-21.0; P=0.011). In conclusion, NT-proBNP elevation is common and is associated with markers of anaemia, inflammation and iron status and with severe functional impairment among sickle cell anaemia patients in Nigeria.
sickle cell disease; N-terminal pro-brain natriuretic peptide; six-minute walk; haemolysis; Africa
Hepatitis C virus (HCV) infection may be associated with thrombocytopenia and increased iron stores in patients receiving medical care. We aimed to determine how often changes in hematologic, iron metabolic and inflammatory markers occur in individuals with undiagnosed HCV in the community.
Inner-city African Americans (n=143) were recruited from the community according to reported ingestion of alcohol. They were divided broadly into those who drank more or less than 56 g alcohol/day as assessed by dietary questionnaire. HCV serology was determined and laboratory values were compared according to HCV seropositivity in analyses that adjusted for alcohol consumption.
The prevalence of HCV seropositivity was 23% among men and 29% among women. Levels of hepatocellular enzymes were higher with HCV seropositivity (P <0.0001) but hemoglobin concentrations, white blood cell and platelet counts and serum ferritin concentrations did not differ. The globulin fraction of the serum protein concentration (P=0.002) was increased with HCV seropositivity as expected with chronic inflammation. However, erythrocyte sedimentation rate and serum iron and haptoglobin levels did not differ significantly according to HCV status. Furthermore, multivariate analysis revealed that C-reactive protein was decreased and transferrin concentration was increased with both HCV and alcohol consumption (P<0.014).
Previously undiagnosed HCV seropositivity has little effect on the complete blood count and body iron stores but appears to perturb the response to an inflammatory stimulus, causing reduced rather than increased circulating CRP concentrations and increased rather than decreased transferrin concentrations.
HCV infection; African American; CRP; Iron metabolism
Fetal hemoglobin (HbF) is the major modifier of the clinical course of sickle cell anemia. Its levels are highly heritable and its interpersonal variability is modulated in part by three quantitative trait loci (QTL) that effect HbF gene expression. Genome-wide association studies (GWAS) have identified single nucleotide polymorphisms (SNPs) in these QTLs that are highly associated with HbF but explain only 10 to 12% of the variance of HbF. Combining SNPs into a genetic risk score (GRS) can help to explain a larger amount of the variability of HbF level but the challenge of this approach is to select the optimal number of SNPs to be included in the GRS.
Methods and Results
We develop a collection of 14 models with GRS composed of different numbers of SNPs, and use the ensemble of these models to predict HbF in sickle cell anemia patients. The models were trained in 841 sickle cell anemia patients and were tested in three independent cohorts. The ensemble of 14 models explained 23.4% of the variability in HbF in the discovery cohort, while the correlation between predicted and observed HbF in the 3 independent cohorts ranged between 0.28 and 0.44. The models included SNPs in BCL11A, the HBS1L-MYB intergenic region and the site of the HBB gene cluster, QTL previously associated with HbF.
An ensemble of 14 genetic risk models can predict HbF levels with accuracy between 0.28 and 0.44 and the approach may prove useful in other applications.
sickle cell disease; hemoglobin; genetics; association studies; risk prediction; risk factor
Chuvash polycythemia results from a homozygous 598C>T mutation in exon 3 of the von Hippel-Lindau (VHL) gene. This disrupts the normoxia pathway for degrading hypoxia inducible factor (HIF)-1α and HIF-2α causing altered expression of HIF-1 and HIF-2 inducible genes. As hypoxia induces expression of proinflammatory cytokines, we hypothesized that there might be an elevation of Th1 cytokines in the setting of Chuvash polycythemia. We analyzed plasma concentrations of Th1 (interleukins-2 and 12, interferon-γ, granulocyte-monocyte colony-stimulating factor, tumor necrosis factor-α) and Th2 cytokines (interleukins-4, 5, 10, and 13) using the Bio-Plex multiplex suspension array system in 34 VHL598C>T homozygotes and 32 VHL wild-type participants from Chuvashia. Concentrations of all the Th1 and Th2 cytokines measured were elevated in the VHL598C>T homozygotes compared with the control wild-type participants, but the ratios of Th1 to Th2 cytokines did not differ by genotype. In parallel, peripheral blood concentrations of CD4 positive T-helper cells and CD4/CD8 ratio were lower in the VHL598C>T homozygotes. In conclusion, the up-regulated hypoxic response in Chuvash polycythemia is associated with increased plasma products of both the Th1 and Th2 pathways, but the balance between the two pathways seems to be preserved.
In Chuvash polycythemia, homozygous von Hippel-Lindau (VHL) 598C>T leads to increased hypoxia inducible factor-1α and 2α, thromboses and lower systemic blood pressures. Circulating homocysteine, glutathione, γ-glutamyltransferase and cysteinylglycine concentrations were higher in 34 VHL598C>T homozygotes than in 37 normal controls and cysteine was lower. Multivariate analysis showed elevated homocysteine independently associated with higher mean systemic blood pressures and elevated glutathione was associated with lower pressures to a similar degree. Among VHL598C>T homozygotes, homocysteine was elevated with low and normal folate concentrations, consistent with a possible defect in the remethylation pathway. The elevated glutathione and γ-glutamyltranserase levels correlated positively with cysteinylglycine, consistent with possible upregulation of a glutathione synthetic enzyme and γ-glutamyltransferase. Cysteinylglycine correlated inversely with cysteine, consistent with possible reduced cysteinyldipeptidase activity. We conclude that up-regulated hypoxia-sensing may influence multiple steps in thiol metabolism. The effects of the resultant elevated levels of homocysteine and glutathione on systemic blood pressure may largely balance each other out.
VHL; polycythemia; homocysteine; folate; glutathione
Background: In adults with sickle cell disease (SCD), an increased
tricuspid regurgitant velocity (TRV) measured by Doppler echocardiography, an
increased serum N-terminal pro–brain natriuretic peptide (NT-pro-BNP) level,
and pulmonary hypertension (PH) diagnosed by right heart catheterization (RHC) are
independent risk factors for mortality.
Methods: A multidisciplinary committee was formed by
clinician-investigators experienced in the management of patients with PH and/or SCD.
Clinically important questions were posed, related evidence was appraised, and
questions were answered with evidence-based recommendations. Target audiences include
all clinicians who take care of patients with SCD.
Results: Mortality risk stratification guides decision making. An
increased risk for mortality is defined as a TRV equal to or greater than 2.5
m/second, an NT-pro-BNP level equal to or greater than 160 pg/ml, or RHC-confirmed
PH. For patients identified as having increased mortality risk, we make a strong
recommendation for hydroxyurea as first-line therapy and a weak recommendation for
chronic transfusions as an alternative therapy. For all patients with SCD with
elevated TRV alone or elevated NT-pro-BNP alone, and for patients with SCD with
RHC-confirmed PH with elevated pulmonary artery wedge pressure and low pulmonary
vascular resistance, we make a strong recommendation against PAH-specific therapy.
However, for select patients with SCD with RHC-confirmed PH who have elevated
pulmonary vascular resistance and normal pulmonary capillary wedge pressure, we make
a weak recommendation for either prostacyclin agonist or endothelin receptor
antagonist therapy and a strong recommendation against phosphodiesterase-5 inhibitor
Conclusions: Evidence-based recommendations for the management of
patients with SCD with increased mortality risk are provided, but will require
frequent reassessment and updating.
sickle cell disease; mortality; pulmonary hypertension; dyspnea; hemolysis
Sickle cell disease (SCD), caused by a mutation in the β-globin gene HBB, is widely distributed in malaria endemic regions. Cardiopulmonary complications are major causes of morbidity and mortality. Hemoglobin SS (Hb SS) represents a large proportion of SCD in the Americas, United Kingdom, and certain regions of Africa while higher proportions of hemoglobin SC are observed in Burkina Faso and hemoglobin Sβ-thalassemia in Greece and India. Coinheritance of α-thalassemia and persistence of hemoglobin F production are observed in highest frequency in certain regions of India and the Middle East. As confirmed in the PUSH and Walk-PHaSST studies, Hb SS, absence of co-inheriting alpha-thalassemia, and low hemoglobin F levels tend to be associated with more hemolysis, lower hemoglobin oxygen saturations, greater proportions of elevated tricuspid regurgitant jet velocity and brain natriuretic peptide, and increased left ventricular mass index. Identification of additional genetic modifiers will improve prediction of cardiopulmonary complications in SCD.
Sickle Cell Disease; Echocardiogram; Pulmonary Function Test; Hemoglobin F; Alpha thalassemia
To evaluate the association between haemoglobinuria and chronic kidney disease (CKD) in sickle cell anaemia (SCA), we analysed 356 adult haemoglobin SS or Sβ° thalassaemia patients from the University of Illinois at Chicago (UIC) and 439 from the multi-centre Walk-Treatment of Pulmonary Hypertension and Sickle Cell Disease with Sildenafil Therapy (PHaSST) cohort. CKD was classified according to National Kidney Foundation Kidney Disease Outcomes Quality Initiatives guidelines. Haemoglobinuria, defined as positive haem on urine dipstick with absent red blood cells on microscopy, was confirmed by enzyme-linked immunosorbent assay in a subset of patients. The prevalence of CKD was 58% in the UIC cohort and 54% in the Walk-PHaSST cohort, and haemoglobinuria was observed in 36% and 20% of the patients, respectively. Pathway analysis in both cohorts indicated an independent association of lactate dehydrogenase with haemoglobinuria and, in turn, independent associations of haemoglobinuria and age with CKD (P<0.0001). After a median of 32 months of follow-up in the UIC cohort, haemoglobinuria was associated with progression of CKD (halving of estimated glomerular filtration rate or requirement for dialysis; Hazard ratio [HR] 13.9, 95% confidence interval [CI] 1.7-113.2, P=0.0012) and increasing albuminuria (HR 3.1, 95% CI: 1.3-7.7; logrank P=0.0035). In conclusion haemoglobinuria is common in SCA and is associated with CKD, consistent with a role for intravascular haemolysis in the pathogenesis of renal dysfunction in SCA.
Sickle Cell; Haemoglobinuria; Haemolysis; Kidney Disease; Albuminuria
Low steady state haemoglobin oxygen saturation in patients with sickle cell anaemia has been associated with the degree of anaemia and haemolysis. How much pulmonary dysfunction contributes to low saturation is not clear. In a prospective study of children and adolescents with sickle cell disease aged 3–20 years at steady state and matched controls, 52% of 391 patients versus 24% of 63 controls had steady state oxygen saturation <99% (P < 0·0001), 9% of patients versus no controls had saturation <95% (P = 0·008) and 8% of patients versus no controls had exercise-induced reduction in saturation ≥3%. Decreasing haemoglobin concentration (P ≤ 0·001) and increasing haemolysis (P ≤ 0·003) but not pulmonary function tests were independent predictors of both lower steady-state saturation and exercise-induced reduction in saturation. Neither history of stroke nor history of acute chest syndrome was significantly associated with lower steady-state oxygen saturation or exercise-induced reduction in saturation. Tricuspid regurgitation velocity was higher in patients with lower steady state haemoglobin oxygen saturation (P = 0·003) and with greater decline in oxygen saturation during the six-minute walk (P = 0·022). In conclusion, lower haemoglobin oxygen saturation is independently associated with increasing degrees of anaemia and haemolysis but not pulmonary function abnormalities among children and adolescents with sickle cell disease.
sickle cell disease; paediatric; oxygen saturation; six-minute walk; pulmonary hypertension
Haemolytic anaemia is variable among patients with sickle cell anaemia and can be estimated by reticulocyte count, lactate dehydrogenase, aspartate aminotransferase and bilirubin levels. Using principal component analysis of these measurements we computed a haemolytic score that we used as a subphenotype in a genome-wide association study. We identified in one cohort and replicated in two additional cohorts the association of a single nucleotide polymorphism in NPRL3 (rs7203560; chr16p13·3) (P = 6·04 × 10−07). This association was validated by targeted genotyping in a fourth independent cohort. The HBA1/HBA2 regulatory elements, hypersensitive sites (HS)-33, HS-40 and HS-48 are located in introns of NPRL3. Rs7203560 was in perfect linkage disequilibrium (LD) with rs9926112 (r2 = 1) and in strong LD with rs7197554 (r2 = 0·75) and rs13336641 (r2 = 0·77); the latter is located between HS-33 and HS-40 sites and next to a CTCF binding site. The minor allele for rs7203560 was associated with the −∝3·7thalassaemia gene deletion. When adjusting for HbF and ∝ thalassaemia, the association of NPRL3 with the haemolytic score was significant (P = 0·00375) and remained significant when examining only cases without gene deletion∝ thalassaemia (P = 0·02463). Perhaps by independently down-regulating expression of the HBA1/HBA2 genes, variants of the HBA1/HBA2 gene regulatory loci, tagged by rs7203560, reduce haemolysis in sickle cell anaemia.
haemolysis; sickle cell anaemia; haemolytic anaemia; genetic analysis; thalassaemia
The role of pulmonary hypertension as a cause of mortality in sickle cell disease (SCD) is controversial.
Methods and Results
We evaluated the relationship between an elevated estimated pulmonary artery systolic pressure and mortality in patients with SCD. We followed patients from the walk-PHaSST screening cohort for a median of 29 months. A tricuspid regurgitation velocity (TRV)≥3.0 m/s cuttof, which has a 67–75% positive predictive value for mean pulmonary artery pressure ≥25 mm Hg was used. Among 572 subjects, 11.2% had TRV≥3.0 m/sec. Among 582 with a measured NT-proBNP, 24.1% had values ≥160 pg/mL. Of 22 deaths during follow-up, 50% had a TRV≥3.0 m/sec. At 24 months the cumulative survival was 83% with TRV≥3.0 m/sec and 98% with TRV<3.0 m/sec (p<0.0001). The hazard ratios for death were 11.1 (95% CI 4.1–30.1; p<0.0001) for TRV≥3.0 m/sec, 4.6 (1.8–11.3; p = 0.001) for NT-proBNP≥160 pg/mL, and 14.9 (5.5–39.9; p<0.0001) for both TRV≥3.0 m/sec and NT-proBNP≥160 pg/mL. Age >47 years, male gender, chronic transfusions, WHO class III–IV, increased hemolytic markers, ferritin and creatinine were also associated with increased risk of death.
A TRV≥3.0 m/sec occurs in approximately 10% of individuals and has the highest risk for death of any measured variable.
The study is registered in ClinicalTrials.gov with identifier
Background & Aims
Celiac disease is an increasingly recognized disorder in Caucasian populations of European origin. Little is known about its prevalence in non-Caucasians. Although it is thought to be a cause of iron deficiency anemia, little is known about the extent to which celiac disease contributes to iron deficiency in Caucasians, and especially non-Caucasians. We analyzed samples collected from participants in the Hemochromatosis and Iron Overload Screening (HEIRS) study to identify individuals with iron deficiency and assess the frequency of celiac disease.
We analyzed serum samples from white men (25 y old or older) and women (50 y old or older) who participated the HEIRS study; cases were defined as individuals with iron deficiency (serum level of ferritin ≤12 mg/L) and controls were those without (serum level of ferritin >100 mg/L in men and >50 mg/L in women). All samples were also analyzed for human recombinant tissue transglutaminase immunoglobulin A; positive results were confirmed by an assay for endomysial antibodies. Patients with positive results from both celiac disease tests were presumed to have untreated celiac disease, and those with a positive result from only 1 test were excluded from analysis. We analyzed HLA genotypes and frequencies of celiac disease between Caucasians and non-Caucasians with iron deficiency.
Celiac disease occurred in 14 of 567 of cases (2.5%) and in only 1 of 1136 controls (0.1%; Fisher’s exact test, P=1.92 × 10−6). Celiac disease was more common in Caucasian cases (14/363, 4%) than non-Caucasian cases (0/204; P=.003). Only 1 Caucasian control and no non-Caucasian controls had celiac disease. The odds of celiac disease in individuals with iron deficiency was 28-fold (95% confidence interval, 3.7–212.8) that of controls; 13/14 cases with celiac disease carried the DQ2.5 variant of the HLA genotype.
Celiac disease is associated with iron deficiency of Caucasians. Celiac disease is rare among non-Caucasians—even among individuals with features of celiac disease, such as iron deficiency. Celiac disease is also rare among individuals without iron deficiency. Men and post-menopausal women with iron deficiency should be tested for celiac disease.
SNP; risk factor; gluten allergy; intestine; absorption