Tuberculosis Trends in Saudis and Non-Saudis in the Kingdom of Saudi Arabia – A 10 Year Retrospective Study (2000–2009)

Tuberculosis (TB) remains a public health problem in the Kingdom of Saudi Arabia (KSA), which has a very large labour force from high TB endemic countries. Understanding the epidemiological and clinical features of the TB problem, and the TB burden in the immigrant workforce, is necessary for improved planning and implementation of TB services and prevention measures

The Effects of Macrophage Polarization and Granuloma Microenvironment on the Macrophage Response to Mycobacterium tuberculosis

Macrophages display considerable functional and phenotypic plasticity because the cytokine and biochemical environment affects their differentiation and maturation. Macrophages are the primary niche for Mycobacterium tuberculosis inside granulomas. This thesis examines the different macrophage subsets M1-like, M2-like, Mox macrophages and lipid loaded foamy macrophages and how they interact with Mycobacterium tuberculosis complex infection. Human monocyte-derived macrophages were differentiated in the presence of either granulocyte-macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF). By examining production of the pro-inflammatory cytokine TNF-α and the anti-inflammatory cytokine IL-10 in response to LPS it was established that these GM- and M-macrophages formed distinct subsets with characteristics similar to M1-like and M2-like macrophages. GM-macrophages and M-macrophages were also matured in the presence of the lipid OxPAPC; these macrophages termed “Mox” have been previously characterized in lipid rich murine atherosclerosis plaques that resemble lipid rich tuberculosis granuloma. The characterization of the generated macrophages subsets was examined and it was found in the presence of Mycobacterium bovis BCG infection, Mox M-macrophages became extremely elongated and spindle shaped. CD14 expression was significantly higher in Mycobacterium bovis BCG infected and uninfected M-macrophages and Mox M-macrophages compared to all GM-macrophages. There were trends towards lower expression of CD14 in M and Mox M macrophages after BCG infection. Also there was a trend towards higher expression of CD209 in M-macrophages and also reduced expression in cells infected with BCG. M-macrophages and Mox M-macrophages displayed significantly higher phagocytic abilities against latex beads and Mycobacterium bovis BCG. To examine lipid loaded foamy macrophages, U937 monocytes were used and differentiated into macrophages using phorbol 13-myristate 13-acetate (PMA). In vitro generated foamy macrophages with oleic acid-BSA were able to retain their lipid droplets up to 7 days. OA-BSA foamy macrophages were infected with Mycobacterium bovis BCG, Mycobacterium tuberculosis H37Rv and Mycobacterium tuberculosis GC1237 and it was observed that Mycobacterium tuberculosis GC1237 caused the highest level of host cell death. Both Mycobacterium tuberculosis H37Rv and Mycobacterium tuberculosis GC1237 induced more apoptosis in foamy macrophages compared to non-foamy macrophages. Mycobacterium bovis BCG infection of OA-BSA foamy cells decreased lipid content significantly after 48 hours infection. While looking at the ability of OA-BSA treated foamy macrophages to take up latex beads, dextran and Mycobacterium bovis BCG it was found that foamy macrophages had similar phagocytic capability to non-foamy cells but had a reduced ability to take up dextran. The induction of necrosis in the TB granuloma is an essential component for the pathogenesis and transmission of TB. While investigating the relative ability of macrophage subsets to undergo programmed necrosis in response to inflammatory stimuli, TNF-α or IFN-γ, it was found that programmed necrosis, necroptosis, could be induced in GM-macrophages and M-macrophages by TNF-α and to a lesser extent by IFN-γ. Necroptosis could be induced as well using TNF-α in OA-BSA treated and untreated macrophages. Necroptosis was more prominent in OA-BSA treated foamy macrophages compared to non-foamy macrophages. These data give and understanding and insight into the roles of macrophage polarization and lipid milieu in tuberculosis infection

Total number and incidence rate of TB cases in Saudi Arabia (2000–2009) by year and age group.

*<p>Chi-Square for linear trend.</p

(A) Annual TB incidence rate trends among Saudis (2000–2009); (B) Annual TB incidence rate trends among non-Saudis (2000–2009).

<p>(A) Annual TB incidence rate trends among Saudis (2000–2009); (B) Annual TB incidence rate trends among non-Saudis (2000–2009).</p

Total number and incidence rate of TB cases in Saudi Arabia (2000–2009) by Province.

*<p>Chi square for trend.</p

Number and percentage of TB cases in Saudi Arabia (2000–2010) by form of TB and nationality.

*<p>Pearson Chi-square.</p

Susceptibility of M. tuberculosis-infected host cells to phospho-MLKL driven necroptosis is dependent on cell type and presence of TNFα

An important feature of Mycobacterium tuberculosis pathogenesis is the ability to control cell death in infected host cells, including inhibition of apoptosis and stimulation of necrosis. Recently an alternative form of programmed cell death, necroptosis, has been described where necrotic cell death is induced by apoptotic stimuli under conditions where apoptotic execution is inhibited. We show for the first time that M. tuberculosis and TNFα synergise to induce necroptosis in murine fibroblasts via RIPK1-dependent mechanisms and characterized by phosphorylation of Ser345 of the MLKL necroptosis death effector. However, in murine macrophages M. tuberculosis and TNFα induce non-necroptotic cell death that is RIPK1-dependent but independent of MLKL phosphorylation. Instead, M. tuberculosis-infected macrophages undergo RIPK3-dependent cell death which occurs both in the presence and absence of TNFα and involves the production of mitochondrial ROS. Immunocytochemical staining for MLKL phosphorylation further demonstrated the occurrence of necroptosis in vivo in murine M. tuberculosis granulomas. Phosphorylated- MLKL immunoreactivity was observed associated with the cytoplasm and nucleus of fusiform cells in M. tuberculosis lesions but not in proximal macrophages. Thus whereas pMLKL-driven necroptosis does not appear to be a feature of M. tuberculosis-infected macrophage cell death, it may contribute to TNFα-induced cytotoxicity of the lung stroma and therefore contribute to necrotic cavitation and bacterial dissemination

Cumulative proportion of non-Saudi TB patients by country of origin (2000–2009).

<p>Cumulative proportion of non-Saudi TB patients by country of origin (2000–2009).</p

Total number of TB and incidence rate/100,000 in Saudi Arabia (2000–2009) by nationality.

*<p>Chi-Square for linear trend.</p