Polygenic risk assessment reveals pleiotropy between sarcoidosis and inflammatory disorders in the context of genetic ancestry.

Sarcoidosis is a complex disease of unknown etiology characterized by the presence of granulomatous inflammation. Though various immune system pathways have been implicated in disease, the relationship between the genetic determinants of sarcoidosis and other inflammatory disorders has not been characterized. Herein, we examined the degree of genetic pleiotropy common to sarcoidosis and other inflammatory disorders to identify shared pathways and disease systems pertinent to sarcoidosis onset. To achieve this, we quantify the association of common variant polygenic risk scores from nine complex inflammatory disorders with sarcoidosis risk. Enrichment analyses of genes implicated in pleiotropic associations were further used to elucidate candidate pathways. In European-Americans, we identify significant pleiotropy between risk of sarcoidosis and risk of asthma (R2=2.03%; P=8.89 × 10-9), celiac disease (R2=2.03%; P=8.21 × 10-9), primary biliary cirrhosis (R2=2.43%; P=2.01 × 10-10) and rheumatoid arthritis (R2=4.32%; P=2.50 × 10-17). These associations validate in African Americans only after accounting for the proportion of genome-wide European ancestry, where we demonstrate similar effects of polygenic risk for African-Americans with the highest levels of European ancestry. Variants and genes implicated in European-American pleiotropic associations were enriched for pathways involving interleukin-12, interleukin-27 and cell adhesion molecules, corroborating the hypothesized immunopathogenesis of disease.

Association of ANXA11 genetic variation with sarcoidosis in African Americans and European Americans.

A recent genome-wide association study in a German population and two subsequent studies in European populations found that a non-synonymous single-nucleotide polymorphism (SNP), rs1049550, within the annexin A11 (ANXA11) gene was associated with susceptibility to sarcoidosis. We sought to identify additional ANXA11 variants independently associated with sarcoidosis, determine whether any sarcoidosis-associated ANXA11 variants were associated with chest radiographic phenotypes, and explore human leukocyte antigen (HLA) SNP-SNP interactions with ANXA11. A total of 209 SNPs spanning 100 kb including the 5' promoter, coding, and 3' untranslated regions of ANXA11 were genotyped for 1689 sarcoidosis cases and 1252 controls. After adjustment for rs1049550, two additional novel ANXA11 sarcoidosis associations were identified only in African Americans--rs61860052 (odds ratio (OR)=0.62; 95% confidence interval (CI)=0.40-0.97) and rs4377299 (OR=1.31; 95% CI=1.06-1.63). These associations were more pronounced in radiologically-classified Scadding stage IV sarcoidosis cases. We also identified a significant SNP-SNP interaction between rs1049550 and a sarcoidosis risk SNP (rs9268839) near the HLA-DRA locus. This further genetic dissection of ANXA11 may provide additional insight into the immune dysregulation characteristic of sarcoidosis pathophysiology.

A genome-wide admixture scan for ancestry-linked genes predisposing to sarcoidosis in African-Americans.

Genome-wide linkage and association studies have uncovered variants associated with sarcoidosis, a multiorgan granulomatous inflammatory disease. African ancestry may influence disease pathogenesis, as African-Americans are more commonly affected by sarcoidosis. Therefore, we conducted the first sarcoidosis genome-wide ancestry scan using a map of 1384 highly ancestry-informative single-nucleotide polymorphisms genotyped on 1357 sarcoidosis cases and 703 unaffected controls self-identified as African-American. The most significant ancestry association was at marker rs11966463 on chromosome 6p22.3 (ancestry association risk ratio (aRR)=1.90; P=0.0002). When we restricted the analysis to biopsy-confirmed cases, the aRR for this marker increased to 2.01; P=0.00007. Among the eight other markers that demonstrated suggestive ancestry associations with sarcoidosis were rs1462906 on chromosome 8p12, which had the most significant association with European ancestry (aRR=0.65; P=0.002), and markers on chromosomes 5p13 (aRR=1.46; P=0.005) and 5q31 (aRR=0.67; P=0.005), which correspond to regions we previously identified through sib-pair linkage analyses. Overall, the most significant ancestry association for Scadding stage IV cases was to marker rs7919137 on chromosome 10p11.22 (aRR=0.27; P=2 × 10(-5)), a region not associated with disease susceptibility. In summary, through admixture mapping of sarcoidosis we have confirmed previous genetic linkages and identified several novel putative candidate loci for sarcoidosis.

HLA and environmental interactions in sarcoidosis.

Sarcoidosis is a systemic granulomatosis of unknown etiology despite being described over 100 years ago. While both genetic predisposition and environmental exposures have been proposed as playing a role in this disease, there have not been any systematic investigations of gene-environmental interaction in this disease. In the ACCESS dataset, detailed environmental histories and high resolution HLA class II typing were performed on 476 cases of newly diagnosed sarcoidosis and 476 matched controls from the patients' community. We evaluated gene-environmental interactions in exposures or HLA class II alleles that were present in > 5% of the population and had an odd ratio of > 1.0. Four exposures and four HLA Class II alleles met these criteria and were evaluated. Significant interaction was observed between HLA DRB1*1101 and insecticide exposure at work (p < 0.10) and suggestive interaction was observed between HLA DRB1*1101 and exposure to mold and musty odors and DRB1*1501 and insecticide exposure at work (P < 0.15). In addition, HLA DRB1*1101 and insecticide exposure at work was associated with extrapulmonary sarcoidosis, specifically cardiac sarcoidosis and hypercalcemia (p<0.05) and HLA DRB1*1101 and exposure to molds and musty odors was associated with pulmonary only sarcoidosis (P < 0.05). These studies suggest that sarcoidosis is due to an interaction of genetic predisposition and environmental exposure in at least some cases of sarcoidosis. Future studies in defined phenotypes of sarcoidosis may be necessary to define environmental and genetic associations with sarcoidosis.

Genetic linkage analysis of sarcoidosis phenotypes: the sarcoidosis genetic analysis (SAGA) study.

The sarcoidosis genetic analysis (SAGA) study previously identified eight chromosomal regions with suggestive evidence for linkage to sarcoidosis susceptibility in African-American sib pairs. Since the clinical course of sarcoidosis is variable and likely under genetic control, we used the affected relative pair portion of the SAGA sample (n=344 pairs) to perform multipoint linkage analyses with covariates based on pulmonary and organ involvement phenotypes. Chest radiographic resolution was the pulmonary phenotype with the highest LOD (logarithm of the backward odds, or likelihood ratio) score of 5.11 at D1S3720 on chromosome 1p36 (P=4 x 10(-5)). In general, higher LOD scores were attained for covariates that modeled clustered organ system involvement rather than individual organ systems, with the cardiac/renal group having the highest LOD score of 6.65 at chromosome 18q22 (P=2 x 10(-5)). The highest LOD scores for the other three organ involvement groups of liver/spleen/bone marrow, neuro/lymph and ocular/skin/joint were 3.72 at 10p11 (P=0.0004), 5.16 at 7p22 (P=4 x 10(-5)) and 2.93 at 10q26 (P=0.001), respectively. Most of the phenotype linkages did not overlap with the regions previously found linked to susceptibility. Our results suggest that genes influencing clinical presentation of sarcoidosis in African Americans are likely to be different from those that underlie disease susceptibility.

Genome-wide search for sarcoidosis susceptibility genes in African Americans.

Sarcoidosis, a systemic granulomatous disease of unknown etiology, likely results from an environmental insult in a genetically susceptible host. In the US, African Americans are more commonly affected with sarcoidosis and suffer greater morbidity than Caucasians. We searched for sarcoidosis susceptibility loci by conducting a genome-wide, sib pair multipoint linkage analysis in 229 African-American families ascertained through two or more sibs with a history of sarcoidosis. Using the Haseman-Elston regression technique, linkage peaks with P-values less than 0.05 were identified on chromosomes 1p22, 2p25, 5p15-13, 5q11, 5q35, 9q34, 11p15 and 20q13 with the most prominent peak at D5S2500 on chromosome 5q11 (P=0.0005). We found agreement for linkage with the previously reported genome scan of a German population at chromosomes 1p and 9q. Based on the multiple suggestive regions for linkage found in our study population, it is likely that more than one gene influences sarcoidosis susceptibility in African Americans. Fine mapping of the linked regions, particularly on chromosome 5q, should help to refine linkage signals and guide further sarcoidosis candidate gene investigation.

Sarcoidosis and granuloma genes: a family-based study in African-Americans.

The evidence for a genetic component in the aetiology of sarcoidosis includes familial aggregation, associations with genetic polymorphisms, and linkage to the major histocompatibility complex class region on chromosome 6p. Unfortunately, the majority of genetic associations with sarcoidosis have not been consistently replicated. In the present study, using a family-based study design, which controls for population stratification, the authors attempted to replicate previously reported associations between sarcoidosis and three attractive candidate genes studied primarily in case-control samples. In 225 nuclear families, ascertained through African Americans with a history of sarcoidosis, no evidence was found for an association between sarcoidosis susceptibility and polymorphisms in the angiotensin converting enzyme, vitamin D receptor and tumour necrosis factor-alpha genes. Further analyses of chronic and acute disease phenotypes failed to reveal any notable associations. Assuming an underlying inheritance model with an additive allelic effect on disease risk, the current study had approximately 80-90% statistical power to detect a 3-fold increased risk associated with the putative risk allele of the polymorphisms under study. The present authors conclude that in African-Americans, the angiotensin converting enzyme, vitamin D receptor, and tumour necrosis factor-alpha genes are not significant risk factors for sarcoidosis susceptibility.

Clinical characteristics of patients in a case control study of sarcoidosis.

Sarcoidosis may be affected by sex, race, and age. A Case Control Etiologic Study of Sarcoidosis (ACCESS) enrolled 736 patients with sarcoidosis within 6 mo of diagnosis from 10 clinical centers in the United States. Using the ACCESS sarcoidosis assessment system, we determined organ involvement for the whole group and for subgroups differentiated by sex, race, and age (less than 40 yr or 40 yr and older). The study population was heterogeneous in terms of race (53% white, 44% black), sex (64% female, 36% male), and age (46% < 40 yr old, 54% > or = 40 yr old). Women were more likely to have eye and neurologic involvement (chi(2) = 4.74, p < 0.05 and chi(2) = 4.60, p < 0.05 respectively), have erythema nodosum (chi(2) = 7.28, p < 0.01), and to be age 40 yr or over (chi(2) = 6.07, p < 0.02) whereas men were more likely to be hypercalcemic (chi(2) = 7.38, p < 0.01). Black subjects were more likely to have skin involvement other than erythema nodosum (chi(2) = 5.47, p < 0.05), and eye (chi(2) = 13.8, p < 0.0001), liver (chi(2) = 23.3, p < 0.0001), bone marrow (chi(2) = 18.8, p < 0.001), and extrathoracic lymph node involvement (chi(2) = 7.21, p < 0.01). We conclude that the initial presentation of sarcoidosis is related to sex, race, and age.

Familial aggregation of sarcoidosis. A case-control etiologic study of sarcoidosis (ACCESS).

Despite reports of familial clustering of sarcoidosis, little empirical evidence exists that disease risk in family members of sarcoidosis cases is greater than that in the general population. To address this question, we estimated sarcoidosis familial relative risk using data on disease occurrence in 10,862 first- and 17,047 second-degree relatives of 706 age, sex, race, and geographically matched cases and controls who participated in the multicenter ACCESS (A Case-Control Etiology Study of Sarcoidosis) study from 1996 to 1999. Familial relative risk estimates were calculated using a logistic regression technique that accounted for the dependence between relatives. Sibs had the highest relative risk (odds ratio [OR] = 5.8; 95% confidence interval [CI] = 2.1-15.9), followed by avuncular relationships (OR = 5.7; 95% CI = 1.6-20.7), grandparents (OR = 5.2; 95% CI = 1.5-18.0), and then parents (OR = 3.8; 95% CI = 1.2-11.3). In a multivariate model fit to the parents and sibs data, the familial relative risk adjusted for age, sex, relative class, and shared environment was 4.7 (95% CI = 2.3-9.7). White cases had a markedly higher familial relative risk compared with African-American cases (18.0 versus 2.8; p = 0.098). In summary, a significant elevated risk of sarcoidosis was observed among first- and second-degree relatives of sarcoidosis cases compared with relatives of matched control subjects.

Familial risk ratio of sarcoidosis in African-American sibs and parents.

While sarcoidosis is thought to aggregate in families, little is known about the risk to relatives of sarcoidosis patients. To estimate the familial risk ratio (lambda) of sarcoidosis in sibs and parents of cases, the authors studied 179 African-American families ascertained through an index sarcoidosis case diagnosed at Henry Ford Hospital in Detroit, Michigan. Among those relatives enrolled between 1997 and 1999, 12 of 327 (3.7%) sibs and 11 of 161 (6.8%) parents reported a history of sarcoidosis. The lambda in this sample of relatives, estimated by computing an age, sex, and race standardized incidence ratio, was 2.24 (95% confidence interval: 1.16, 3.92) for sibs and 2.82 (95% confidence interval: 1.41, 5.05) for parents. For sibs and parents combined, lambda was 2.49 (95% confidence interval: 1.58, 3.73). Results stratified by proband characteristics indicated that lambda was greater for relatives of younger (lambda = 2.93, 95% confidence interval: 1.52, 5.12) and male (lambda = 3.98, 95% confidence interval: 1.99, 7.12) probands. A higher lambda was also found for male family members and sibs born later in the birth order. A Monte Carlo method was also used to estimate lambda, with similar results obtained. Overall, these results indicate that, in African Americans, sibs and parents of sarcoidosis cases have about a 2.5-fold increased risk for sarcoidosis and that heterogeneity in disease risk may exist among family members.

The natural resistance-associated macrophage protein gene in African Americans with sarcoidosis.

The histologic and clinical similarities between tuberculosis and sarcoidosis suggest a shared underlying pathophysiology. Human natural resistance-associated macrophage protein (NRAMP1), which is closely related to the mouse gene, has been associated with susceptibility to tuberculosis in some human populations. Given the importance of the Nramp1 gene in animal models of granulomatous disorders, the association with human tuberculosis, and the possible role of NRAMP1 in macrophage activation and function, we hypothesized that human NRAMP1 plays a role in susceptibility to sarcoidosis. We analyzed several NRAMP1 gene polymorphisms in a case-control study of 157 African American patients with sarcoidosis and 111 African American control subjects. Our results, in contrast to those in tuberculosis patients, showed that the less common genotypes were found more often in control subjects than in case patients (odds ratio, 0.48; 95% confidence interval, 0.28-0.81). In particular, one polymorphism, a (CA)(n) repeat in the immediate 5' region of the gene, was found to have a protective effect (P = 0. 014). Whereas NRAMP1 polymorphisms have been associated with increased susceptibility to tuberculosis, our results suggest that at least one NRAMP1 polymorphism may decrease susceptibility in sarcoidosis.

The Blau syndrome gene is not a major risk factor for sarcoidosis.

BACKGROUND: A genetic predisposition to sarcoidosis has long been postulated, although no specific susceptibility genes are known. Candidate genes for the two granulomatous inflammatory disorders with clinical similarities to sarcoidosis, Blau syndrome and Crohn's disease, have been localized to a 40 centimorgan region spanning the chromosome 16 centromere. PATIENTS AND METHODS: Using a sample of 35 African-American sibling pairs, who both had clinically confirmed sarcoidosis, we tested for genetic linkage between the 16p12-q21 interval (the likely location of the Blau syndrome gene) and sarcoidosis. RESULTS: We found no evidence for linkage to any of the eight markers we tested in the 16p12-q21 interval. Ninety percent of the 16p12-q21 region had a LOD score < -2 for a dominant gene conferring a relative risk of 3 or greater for sarcoidosis. One hundred percent of the region had a LOD score < -2 for a dominant gene with a relative risk of 3.5 or greater or recessive gene with relative risk of 2.5 or greater. Based on simulation results we could not exclude a dominant gene with relative risk < 5 at the 0.05 significance level, nor a recessive gene with relative risk < 3, over the entire 16p12-q21 interval. CONCLUSIONS: While the clinical similarities between Blau Syndrome and sarcoidosis suggest genetic homogeneity between the disorders, we found no evidence for linkage of sarcoidosis to the Blau syndrome locus. Our exclusion results suggest that the Blau Syndrome gene does not have a major effect on sarcoidosis susceptibility.

The influence of T cell receptor and cytokine genes on sarcoidosis susceptibility in African Americans.

The pathogenesis of sarcoidosis, a multisystem granulomatous disorder, is mediated through immunoregulatory pathways. While sarcoidosis clusters in families, inherited risk factors remain undefined. In search of possible sarcoidosis susceptibility genes, we examined anonymous polymorphic genetic markers tightly linked to six different candidate gene regions on chromosomes 2q13, 5q31, 6p23-25, 7p14-15, 14q11 and 22q11. These candidate regions contain T cell receptor, interleukin (IL) and interferon regulatory factor (IRF) genes. Our study population consisted of 105 African-American sarcoidosis cases and 95 unrelated healthy controls. The allelic frequency distribution of two out of the six markers, IL-1 alpha marker (p = 0.010) on 2q13 and the F13A marker (p = 0.0006) on 6p23-25, was statistically significantly different in cases compared with controls. The two alleles most strongly associated with sarcoidosis were IL-1 alpha*137 (Odds Ratio (OR) = 2.60; 95% confidence interval (CI) = 1.36-4.98) and F13A*188 (OR = 2.42; 95% CI = 1.37-4.30). Individuals that had both of these alleles were at a six-fold increased risk for sarcoidosis (OR = 6.19; 95% CI = 2.54-15.10). Restricting the analysis to cases with at least one first or second-degree relative affected with sarcoidosis increased the OR to 15.38. IL-1 levels are elevated in sarcoidosis and the F13A marker is tightly linked to a gene that codes for a newly identified interferon regulatory factor protein (IRF-4), which is thought to play a role in T cell effector functions. Our results suggest genetic susceptibility to sarcoidosis may be conferred by more than one immune-related gene that act synergistically on disease risk.

Single versus multiple doses of acetazolamide for metabolic alkalosis in critically ill medical patients: a randomized, double-blind trial.

OBJECTIVE: To compare two dosing regimens of acetazolamide for the reversal of metabolic alkalosis in mechanically ventilated patients with asthma or chronic obstructive pulmonary disease. DESIGN: A randomized, double-blind, placebo-controlled trial. SETTING: A 35-bed medical intensive care unit in a tertiary care teaching hospital. PATIENTS: Forty mechanically ventilated patients with a metabolic alkalosis (arterial pH > or = 7.48 and serum bicarbonate concentration > or = 26 mEq/L) resistant to fluid or potassium therapy (serum potassium concentration, > or = 4 mEq/L) not receiving acetazolamide or sodium bicarbonate in the previous 72 hrs. INTERVENTIONS: Stratified by previous diuretic use and randomized to receive intravenous administration of acetazolamide, one dose of 500 mg or 250 mg every 6 hrs for a total of four doses. MEASUREMENTS AND MAIN RESULTS: Serum bicarbonate and potassium concentrations were drawn every 6 hrs for 72 hrs, arterial blood gases were drawn every 12 hrs for 72 hrs, and both urine chloride and pH were drawn at hours 0, 6, 12, 18, 24, 48, and 72. By using generalized estimating equation techniques, no difference was found between the two dosing regimens at any point over the study period for serum bicarbonate, serum potassium, or urine chloride end points. Results did not differ between diuretic- and nondiuretic-treated patients. Serum bicarbonate concentrations remained significantly decreased in both treatment groups 72 hrs after administration of the first acetazolamide dose (31.8 +/- 4.9-25.3 +/- 3.8 mEq/L, p < .0001 [250 mg x 4]; 31.9 +/- 25.4-25.4 +/- 3.6 mEq/L, p < .0001 [500 mg x 1]). CONCLUSIONS: We conclude that a single 500-mg dose of acetazolamide reverses nonchloride responsive metabolic alkaloses in medical intensive care unit patients as effectively as multiple doses of 250 mg. Studies to examine the prolonged duration of action of acetazolamide observed in this study as well as the effect of acetazolamide on clinical end points, such as duration of mechanical ventilation, are warranted.

Angiotensin-converting enzyme gene polymorphism and risk of sarcoidosis.

The angiotensin-converting enzyme (ACE) has been implicated in the pathophysiology of sarcoidosis. Serum ACE levels in normal and sarcoidosis patients are influenced by an insertion (I)/deletion (D) polymorphism in the ACE gene. To elucidate the role of this ACE gene polymorphism in sarcoidosis, we conducted a case-control study in African Americans and Caucasians. The ACE gene (I/D) polymorphism did not differ between 60 Caucasian cases and 48 control subjects (p = 0.577). In contrast, a comparison of 183 African-American cases and 111 control subjects resulted in a marked difference in genotypic distributions (p = 0.005). In African Americans, the risk for sarcoidosis was 1.30 (95% confidence interval [CI] = 0.72 to 2. 36) for ID heterozygotes, and 3.17 (95% CI = 1.50 to 6.71) for deletion/deletion (DD) homozygotes. The risk associated with the DD homozygotes was even greater in African Americans when cases were restricted to those with a positive family history (odds ratio = 4. 83; 95% CI = 1.86 to 12.59). Further analyses of African-American cases showed that the ACE genotype was not associated with disease severity, extrathoracic involvement, or overall radiographic change 2 to 4 yr after diagnosis. We did find a moderate association between the II genotype and radiographic progression (OR = 2.97; 95% CI = 1.01 to 8.76). Our results suggest the ACE genotype may play a more important role in sarcoidosis susceptibility and progression in African Americans than Caucasians.

Epidemiology, demographics, and genetics of sarcoidosis.

Epidemiological knowledge of sarcoidosis is based mainly on studies performed more than 30 years ago. These early case-control studies produced some interesting risk factor-disease associations, but a clear causative mechanism in sarcoidosis remains unknown. Studies in military and veteran populations showed a clear preponderance of sarcoidosis in African Americans compared with Caucasians. Our recent sarcoidosis incidence study in a racially heterogeneous population found African Americans at three- to fourfold greater risk, which was less than the 10 to 17 times greater risk previously reported. Females are consistently found at greater risk than males, although the relative risk difference generally does not exceed two. The striking racial differences and numerous reports of familial clustering suggest genetic susceptibility. We have found that familial sarcoidosis is almost three times more common in African-American (17%) than Caucasian cases (6%). Future genetic studies can benefit from the extensive catalog of candidate genes that is emerging from the human genome project. The epidemiological evidence to date strongly suggests that studies seeking causes for sarcoidosis need to consider both environmental and genetic risk factors to be successful because the two likely interact with each other to produce disease.

Analysis of HLA-DPB1 polymorphisms in African-Americans with sarcoidosis.

Several studies have found weak associations between certain human leukocyte antigen (HLA) alleles and sarcoidosis, but none have been conclusive. Glutamic acid at position 69 in HLA-DPB1 has been reported to be strongly associated with chronic beryllium disease. The immunopathologic and clinical similarities between chronic beryllium disease (CBD) and sarcoidosis suggest that similar immune-response genes may be involved in susceptibility in both diseases. We analyzed the DNA sequence of HLA-DPB1 exon 2, which contains the hypervariable regions involved in binding antigens, in blood samples from African-American sarcoidosis patients and healthy controls. Results indicate that Val36 (odds ratio [OR] = 2.30) and Asp55 (OR = 2.03) are associated with increased risk for sarcoidosis, but no association with Glu69 was found. These results suggest that although HLA-DPB1 Glu69 is not associated with sarcoidosis, other alleles may make some contribution to susceptibility to sarcoidosis in African-Americans.

Genetics of sarcoidosis.

Hereditary susceptibility to sarcoidosis is suggested by ethnic preponderance, familial clustering, and multigenerational involvement. The genetics of sarcoidosis cannot be adequately addressed in small samples of patients; a large-scale study with stratification for patient phenotypic differences is necessary. A study that uses both genetic marker and environmental data would be able to control for and examine different causative mechanisms. Until such a well-designed, comprehensive study is carried out, we are left with interesting patterns of disease in families and uncertain allelic associations.