Identification of Tuberculosis Susceptibility Genes with Human Macrophage Gene Expression Profiles

Nguyen Thuy Thuong Thuong,Jeremy J Farrar,Martin Hibberd,Cameron P Simmons,Mark Seielstad,Sarah J Dunstan,Alan Aderem,Vesteinn Thorsson,Guy E Thwaites,Nguyen Thi Ngoc Lan,Thomas R Hawn,Nguyen Than Ha Quyen,Yik Y Teo,Tran Thi Hong Chau,Jeffery S Cox

doi:10.1371/journal.ppat.1000229

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.

Highlights

TB, a leading cause of death worldwide, is characterized by different clinical forms including latent TB (LTB), localized pulmonary infection, and various forms of extrapulmonary TB including TB meningitis (TBM). 90% of people infected with Mycobacterium tuberculosis (Mtb) have latent infection with no symptoms and an immune response that contains the bacilli
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

Summary

Introduction

TB, a leading cause of death worldwide, is characterized by different clinical forms including latent TB (LTB), localized pulmonary infection, and various forms of extrapulmonary TB including TBM. 90% of people infected with Mtb have latent infection with no symptoms and an immune response that contains the bacilli. TB, a leading cause of death worldwide, is characterized by different clinical forms including latent TB (LTB), localized pulmonary infection, and various forms of extrapulmonary TB including TBM. In 10% of infected individuals, symptoms develop and most commonly manifest as pulmonary disease, which accounts for 80% of all forms of TB disease [1]. Only 10% of individuals who are infected with Mtb develop active disease, it is not known which immune responses are associated with susceptibility or resistance. It is not known why some individuals have disseminated TB that spreads to the meninges and central nervous system, while most people have localized disease in the lungs. Pathogen virulence traits, and host genetics have the potential to influence the different clinical manifestations of TB, it is not currently understood which factors are the most important [4]

Methods

Results

Discussion

Conclusion