Identification of Tuberculosis Susceptibility Genes with Human Macrophage Gene Expression Profiles

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Abstract
Although host genetics influences susceptibility to tuberculosis (TB), few genes determining disease outcome have been identified. We hypothesized that macrophages from individuals with different clinical manifestations of Mycobacterium tuberculosis (Mtb) infection would have distinct gene expression profiles and that polymorphisms in these genes may also be associated with susceptibility to TB. We measured gene expression levels of >38,500 genes from ex vivo Mtb-stimulated macrophages in 12 subjects with 3 clinical phenotypes: latent, pulmonary, and meningeal TB (n = 4 per group). After identifying differentially expressed genes, we confirmed these results in 34 additional subjects by real-time PCR. We also used a case-control study design to examine whether polymorphisms in differentially regulated genes were associated with susceptibility to these different clinical forms of TB. We compared gene expression profiles in Mtb-stimulated and unstimulated macrophages and identified 1,608 and 199 genes that were differentially expressed by >2- and >5-fold, respectively. In an independent sample set of 34 individuals and a subset of highly regulated genes, 90% of the microarray results were confirmed by RT-PCR, including expression levels of CCL1, which distinguished the 3 clinical groups. Furthermore, 6 single nucleotide polymorphisms (SNPs) in CCL1 were found to be associated with TB in a case-control genetic association study with 273 TB cases and 188 controls. To our knowledge, this is the first identification of CCL1 as a gene involved in host susceptibility to TB and the first study to combine microarray and DNA polymorphism studies to identify genes associated with TB susceptibility. These results suggest that genome-wide studies can provide an unbiased method to identify critical macrophage response genes that are associated with different clinical outcomes and that variation in innate immune response genes regulate susceptibility to TB. Although TB is a leading cause of death worldwide, the vast majority of infected individuals are asymptomatic and contains the bacillus in a latent form. Among those with active disease, 80% have localized pulmonary disease and 20% have disseminated forms. TB meningitis (TBM) is the most severe form of TB with 20–25% of sufferers dying, and of the survivors, many have disability. We currently do not understand the host factors that regulate this diverse spectrum of clinical outcomes. We hypothesized that variation in innate immune gene function is an important regulator of TB clinical outcomes. We measured the mRNA expression levels of >38,500 genes in macrophages taken from people with a history of latent, pulmonary, or meningeal TB and found genes with unique activation patterns among the clinical groups. Furthermore, we studied one of these genes further and found that CCL1 polymorphisms were associated with pulmonary TB (PTB) but not other types of TB disease. To our knowledge, this is the first study to combine mRNA expression studies with genetic studies to discover a novel gene that is associated with different clinical outcomes in TB. We speculate that this approach can be used to discover novel strategies for modulating immune function to prevent adverse outcomes in TB.