We characterized the functions of neutrophils in response to Mycobacterium tuberculosis (M. tb) infection, with particular reference to glutathione (GSH). We examined the effects of GSH in improving the ability of neutrophils to control intracellular M. tb infection. Our findings indicate that increasing the intracellular levels of GSH with a liposomal formulation of GSH (L-GSH) resulted in reduction in the levels of free radicals and increased acidification of M. tb containing phagosomes leading to the inhibition in the growth of M. tb. This inhibitory mechanism is dependent on the presence of TNF-α and IL-6. Our studies demonstrate a novel regulatory mechanism adapted by the neutrophils to control M. tb infection. 1. Introduction M. tb, a causative agent for tuberculosis (TB), is known to be a highly successful intracellular pathogen and is estimated to be harbored in one third of the world’s population [1]. Left untreated TB is known to have a 70% mortality rate [2]. Increasing drug resistance strains of M. tb and poor precautionary measurements for containing the spread, especially in “high burden” countries, have kept the disease from global eradication [1]. M. tb infection is mostly confined to lungs and can be transmitted by inhalation of droplets from individuals with active TB. M. tb has the ability to evade the host killing mechanisms and can successfully persist inside the phagocytic cells. Specifically, M. tb is capable of surviving within the host cells by preventing the fusion of the phagosomes containing bacteria with lysosomes, thereby avoiding exposure to the toxic lysosomal hydrolases [3]. The terminally differentiated polymorphonuclear leukocytes (PMN) or neutrophils are essential component of the human innate immune system and act as a first-line defense against invading microorganisms. Neutrophils are major inhabitants of the white blood cell population (WBC) since they constitute ~70% of the total WBC population [4]. Neutrophils form the bulk of early recruited leukocyte population that mediate and elicit immune responses against M. tb [4]. Neutrophils, after encountering the mycobacteria, exhibit innate immune responses such as phagocytosis which in turn triggers signaling events resulting in secretion of cytokines such as TNF-α and IL-1 that mediate early inflammatory responses [4–6]. The tripeptide, glutathione (GSH), protects all cells against oxidizing agents, free radicals, and reactive oxygen intermediates (ROI), either directly or through enzymatic action of GSH peroxidases and GSH-transferases [7, 8]. GSH is also important
References
[1]
World Health Organization, “Tuberculosis,” http://www.who.int/tb/publications/factsheet_global.pd.
[2]
E. W. Tiemersma, M. J. van der Werf, M. W. Borgdorff, B. G. Williams, and N. J. D. Nagelkerke, “Natural history of tuberculosis: duration and fatality of untreated pulmonary tuberculosis in HIV negative patients: a systematic review,” PLoS ONE, vol. 6, no. 4, Article ID e17601, 2011.
[3]
M. B. Goren, P. D'Arcy Hart, M. R. Young, and J. A. Armstrong, “Prevention of phagosome lysosome fusion in cultured macrophages by sulfatides of Mycobacterium tuberculosis,” Proceedings of the National Academy of Sciences of the United States of America, vol. 73, no. 7, pp. 2510–2514, 1976.
[4]
S.-Y. Eum, J.-H. Kong, M.-S. Hong et al., “Neutrophils are the predominant infected phagocytic cells in the airways of patients with active pulmonary TB,” Chest, vol. 137, no. 1, pp. 122–128, 2010.
[5]
I. Sugawara, T. Udagawa, and H. Yamada, “Rat neutrophils prevent the development of tuberculosis,” Infection and Immunity, vol. 72, no. 3, pp. 1804–1806, 2004.
[6]
P. Seiler, P. Aichele, B. Raupach, B. Odermatt, U. Steinhoff, and S. H. E. Kaufmann, “Rapid neutrophil response controls fast-replicating intracellular bacteria but not slow-replicating Mycobacterium tuberculosis,” Journal of Infectious Diseases, vol. 181, no. 2, pp. 671–680, 2000.
[7]
O. W. Griffith, “Biologic and pharmacologic regulation of mammalian glutathione synthesis,” Free Radical Biology and Medicine, vol. 27, no. 9-10, pp. 922–935, 1999.
[8]
R. Buhl, H. A. Jaffe, K. J. Holroyd et al., “Systemic glutathione deficiency in symptom-free HIV-seropositive individuals,” The Lancet, vol. 2, no. 8675, pp. 1294–1298, 1989.
[9]
B. de Quay, R. Malinverni, and B. H. Lauterburg, “Glutathione depletion in HIV-infected patients: role of cysteine deficiency and effect of oral N-acetylcysteine,” AIDS, vol. 6, no. 8, pp. 815–819, 1992.
[10]
H.-P. Eck, H. Gmunder, M. Hartmann, D. Petzoldt, V. Daniel, and W. Droge, “Low concentrations of acid-soluble thiol (cysteine) in the blood plasma of HIV-1-infected patients,” Biological Chemistry Hoppe-Seyler, vol. 370, no. 2, pp. 101–108, 1989.
[11]
B. Helbling, J. von Overbeck, and B. H. Lauterburg, “Decreased release of glutathione into the systemic circulation of patients with HIV infection,” European Journal of Clinical Investigation, vol. 26, no. 1, pp. 38–44, 1996.
[12]
M. Klein, K. Jarosova, S. Forejtova et al., “Gold and tuberculin skin tests for the detection of latent tuberculosis infection in patients treated with tumour necrosis factor alpha blocking agents,” Clinical and Experimental Rheumatology, vol. 31, no. 1, pp. 111–117, 2013.
[13]
C. Guerra, D. Morris, A. Sipin et al., “Glutathione and adaptive immune responses against Mycobacterium tuberculosis infection in healthy and HIV infected individuals,” PLoS ONE, vol. 6, no. 12, Article ID e28378, 2011.
[14]
C. Guerra, K. Johal, D. Morris et al., “Control of Mycobacterium tuberculosis growth by activated natural killer cells,” Clinical and Experimental Immunology, vol. 168, no. 1, pp. 142–152, 2012.
[15]
V. Venketaraman, Y. K. Dayaram, M. T. Talaue, and N. D. Connell, “Glutathione and nitrosoglutathione in macrophage defense against Mycobacterium tuberculosis,” Infection and Immunity, vol. 73, no. 3, pp. 1886–1889, 2005.
[16]
D. Morris, C. Guerra, M. Khurasany et al., “Glutathione supplementation improves macrophage functions in HIV,” Journal of Interferon & Cytokine Research, vol. 33, no. 5, pp. 270–279, 2013.
[17]
V. Venketaraman, Y. K. Dayaram, A. G. Amin et al., “Role of glutathione in macrophage control of mycobacteria,” Infection and Immunity, vol. 71, no. 4, pp. 1864–1871, 2003.
[18]
Y. K. Dayaram, M. T. Talaue, N. D. Connell, and V. Venketaraman, “Characterization of a glutathione metabolic mutant of Mycobacterium tuberculosis and its resistance to glutathione and nitrosoglutathione,” Journal of Bacteriology, vol. 188, no. 4, pp. 1364–1372, 2006.
[19]
D. Morris, C. Guerra, C. Donohue, H. Oh, M. Khurasany, and V. Venketaraman, “Unveiling the mechanisms for decreased glutathione in individuals with HIV infection,” Clinical and Developmental Immunology, vol. 2012, Article ID 734125, 2012.
[20]
V. Venketaraman, A. Millman, M. Salman et al., “Glutathione levels and immune responses in tuberculosis patients,” Microbial Pathogenesis, vol. 44, no. 3, pp. 255–261, 2008.
[21]
D. G. Russell, C. E. Barry III, and J. L. Flynn, “Tuberculosis: what we don't know can, and does, hurt us,” Science, vol. 328, no. 5980, pp. 852–856, 2010.
[22]
J. Keane, S. Gershon, R. P. Wise et al., “Tuberculosis associated with infliximab, a tumor necrosis factor α-neutralizing agent,” The New England Journal of Medicine, vol. 345, no. 15, pp. 1098–1104, 2001.
[23]
J. J. Gómez-Reino, L. Carmona, V. Rodríguez Valverde, E. M. Mola, and M. D. Montero, “Treatment of rheumatoid arthritis with tumor necrosis factor inhibitors may predispose to significant increase in tuberculosis risk: a multicenter active-surveillance report,” Arthritis and Rheumatism, vol. 48, no. 8, pp. 2122–2127, 2003.
[24]
T. Botha and B. Ryffel, “Reactivation of latent tuberculosis infection in TNF-deficient mice,” Journal of Immunology, vol. 171, no. 6, pp. 3110–3118, 2003.
[25]
S. Ehlers, “Why does tumor necrosis factor targeted therapy reactivate tuberculosis?” Journal of Rheumatology, vol. 32, no. 74, pp. 35–39, 2005.
[26]
J. L. Flynn, M. M. Goldstein, J. Chan et al., “Tumor necrosis factor-α is required in the protective immune response against Mycobacterium tuberculosis in mice,” Immunity, vol. 2, no. 6, pp. 561–572, 1995.
[27]
K. O. Kisich, M. Higgins, G. Diamond, and L. Heifets, “Tumor necrosis factor alpha stimulates killing of Mycobacterium tuberculosis by human neutrophils,” Infection and Immunity, vol. 70, no. 8, pp. 4591–4599, 2002.
[28]
J. Askling, C. M. Fored, L. Brandt et al., “Risk and case characteristics of tuberculosis in rheumatoid arthritis associated with tumor necrosis factor antagonists in Sweden,” Arthritis and Rheumatism, vol. 52, no. 7, pp. 1986–1992, 2005.
[29]
MMWR, “Tuberculosis associated with blocking agents against tumor necrosis factor-alpha-California, 2002-2003,” Morbidity and Mortality Weekly Report, vol. 53, p. 68, 2004.
[30]
R. S. Wallis, M. S. Broder, J. Y. Wong, M. E. Hanson, and D. O. Beenhouwer, “Granulomatous infectious diseases associated with tumor necrosis factor antagonists,” Clinical Infectious Diseases, vol. 38, no. 9, pp. 1261–1265, 2004.
[31]
I. S. Leal, M. Flórido, P. Andersen, and R. Appelberg, “Interleukin-6 regulates the phenotype of the immune response to a tuberculosis subunit vaccine,” Immunology, vol. 103, no. 3, pp. 375–381, 2001.
[32]
C. H. Ladel, C. Blum, A. Dreher, K. Reifenberg, M. Kopf, and S. H. E. Kaufmann, “Lethal tuberculosis in interleukin-6-deficient mutant mice,” Infection and Immunity, vol. 65, no. 11, pp. 4843–4849, 1997.
[33]
R. Appelberg, A. G. Castro, J. Pedrosa, and P. Minoprio, “Role of interleukin-6 in the induction of protective T cells during mycobacterial infections in mice,” Immunology, vol. 82, no. 3, pp. 361–364, 1994.
[34]
M. Okada, Y. Kita, N. Kanamaru et al., “Anti-IL-6 receptor antibody causes less promotion of tuberculosis infection than Anti-TNF-α antibody in mice,” Clinical and Developmental Immunology, vol. 2011, Article ID 404929, 2011.
[35]
J. L. Rothstein, T. F. Lint, and H. Schreiber, “Tumor necrosis factor/cachectin. Induction of hemorrhagic necrosis in normal tissue requires the fifth component of complement (C5),” Journal of Experimental Medicine, vol. 168, no. 6, pp. 2007–2021, 1988.
[36]
R. M. Strieter, S. L. Kunkel, and R. C. Bone, “Role of tumor necrosis factor-α in disease states and inflammation,” Critical Care Medicine, vol. 21, no. 10, pp. S447–S463, 1993.
[37]
R. Buhl, C. Vogelmeier, M. Critenden et al., “Augmentation of glutathione in the fluid lining the epithelium of the lower respiratory tract by directly administering glutathione aerosol,” Proceedings of the National Academy of Sciences of the United States of America, vol. 87, no. 11, pp. 4063–4067, 1990.
[38]
J. H. Roum, Z. Borok, N. G. McElvaney et al., “Glutathione aerosol suppresses lung epithelial surface inflammatory cell-derived oxidants in cystic fibrosis,” Journal of Applied Physiology, vol. 87, no. 1, pp. 438–443, 1999.
[39]
C. Skerry, J. Harper, M. Klunk, W. R. Bishai, and S. K. Jain, “Adjunctive TNF inhibition with standard treatment enhances bacterial clearance in a murine model of necrotic TB granulomas,” PLoS ONE, vol. 7, no. 6, Article ID e39680, 2012.
[40]
G. D. Zeevalk, L. P. Bernard, and F. T. Guilford, “Liposomal-glutathione provides maintenance of intracellular glutathione and neuroprotection in mesencephalic neuronal cells,” Neurochemical Research, vol. 35, no. 10, pp. 1575–1587, 2010.
