Browsing by Author "Kunsevi Kilola, Carine"

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    Characterization of human alveolar macrophages and blood monocyte-derived macrophages responses to M.tb in TB close contacts with and without type 2 diabetes
    (Stellenbosch : Stellenbosch University, 2022-12) Kunsevi Kilola, Carine; Ronacher, Katharina; Kleynhans, Leanie; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Molecular Biology and Human Genetics.
    ENGLISH ABSTRACT: Tuberculosis (TB), caused by Mycobacterium tuberculosis (M.tb), remains a global threat to human health and a challenge for TB control. Type 2 diabetes (T2D) is a known risk factor for TB, as people living with diabetes have an increased risk of developing TB. The chronic low-grade inflammation associated with T2D is likely to contribute to compromised immune responses to M.tb in the lung and the periphery. We hypothesize that lung macrophages from T2D patients have an impaired response to M.tb and reduced capacity to contain M.tb growth in vitro. Human alveolar macrophages (HAMs) and peripheral monocyte-derived macrophages (MDMs) from 23 close contacts (CCs) of active TB patients with and without T2D were isolated and stimulated with live H37Rv M.tb to determine M.tb uptake and killing. Circulating numbers of white blood cells and neutrophils were elevated in T2D compared to no T2D. Moreover, neutrophil counts in the lung was lower in T2D patients. HAMs of T2D patients have a reduced ability to control M.tb growth one day post infection, and the colony forming units (CFUs) in these cells at day one post infection was inversely correlated to the neutrophil count in this compartment. HbA1c and HDL was associated with M.tb killing in HAMs and MDMs, respectively. CFU fold change at day one in HAMs was correlated with HbA1c suggesting a high bacterial load in the lung is attributable to poor glycemic control. Furthermore, cytokines in HAMs and MDMs supernatants were determined using the Luminex platform. M.tb infection induced unique cytokine responses in MDMs and HAMs. Granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-6, and IL-1RA concentrations were significantly higher in the MDM culture supernatant of no T2D controls compared to T2D patients. HAMs of T2D patients produced more MIP-1α and IL-1β, than no T2D controls. We further identified unique pathways associated with individual cytokine clusters in MDMs and HAMs. Finally, we determined the phenotypes of HAMs and MDMs using Flow cytometry and Time-of-flight mass cytometry (CyTOF), respectively. The flow cytometry data revealed three different MDM subsets (CD11b+CD11c+, CD11b-CD11c+ and CD11b+CD11c-), with the CD11b+CD11c+ cell population being the most abundant. Diabetes affected the frequency of CD36, CD32, CD16, HLA-DR and CD206 in the MDMs subsets before and after infection. The CyTOF data revealed two dominant CD11b lineages in HAMs (CD11b+CD11c+ and CD11b++CD11c+) with CD11b+CD11c+ cells being the most predominant subpopulation. A trend suggested that the frequency of uninfected CD11b++CD11c+ expressing CD36 was increased in T2D group. Furthermore, the median metal intensity of CD32 was significantly lower in the HAMs T2D patients. During T2D, the immune response is differentially regulated in the two compartments (periphery vs site of infection), which may influence the response to infections. Alterations in macrophages phenotypes and function during diabetes, pre- and post-exposure to M.tb may result in dysregulated host defense functions in both HAMs and MDMs.