肺結核分枝桿菌 (Mycobacterium tuberculosis) 感染是很重大的一個傳染性疾病。每年全球約有一千萬人受到結核分枝桿菌的感染，並造成每年約三百萬人死亡。在肺結核的病人當中，第二型糖尿病 (type II diabetes mellitus, DM) 是很重要的危險因子。在台灣，肺結核病患當中有 14 ~ 19% 合併糖尿病 (DMTB)，且糖尿病病人罹患肺結核的機率是正常人的 5 ~ 6 倍。但是第二型糖尿病病人其免疫系統所遭受到的影響和其胰島素拮抗現象 (insulin resistance) 的關係，至今尚未有完整的研究。在先前的研究中，我們發現到當第二型糖尿病病患合併感染肺結核 (DMTB) 時，其疾病的嚴重程度和治療的完成率，相較於沒有糖尿病的肺結核病患 (TB) 有著顯著的惡化。並且在利用 M. bovis BCG (BCG) 或是 BCG + IL-12 及 BCG + IFN-γ 刺激病人周邊血液白血球後發現，肺結核病患合併有糖尿病時，其周邊血白血球所產生的 IL-12 可能較少。推測 DMTB 病患其免疫細胞當中的吞噬細胞 (phagocytes) 可能產生缺陷，使其產生較少的 IL-12 而影響了人體對抗肺結核分枝桿菌的免疫反應當中 IL-12/ IFN-γ 的平衡，使得 DM 病患對於肺結核分枝桿菌的感染有較高的感受性。因此在本實驗中，我們收集更多病人確認 DMTB 病患其白血球細胞激素 (cytokine) IL-12 及IFN-γ 產生的能力，並研究病人吞噬細胞自然免疫反應 (innate immune response) 的功能。首先我們發現，和單純感染肺結核分枝桿菌的病人比較之後，DMTB 病患的白血球產生 IL-12 的能力確實較差。而不管有無糖尿病，肺結核病患的白血球產生 IFN-γ 的能力都有比健康人差的趨勢。接著我們比較單純得到 TB 或是DM 及 DMTB 病患其周邊血液顆粒球 (granulocytes) 和單核球 (monocytes) 吞噬能力還有氧活性分子 (reactive oxygen species, ROS)的產生能力。發現到 DMTB 病患其 phagocytes 的吞噬能力都是正常的，但是其產生 ROS 的能力則比單純得到 TB 或是 DM 的病患來差。這些結果告訴我們 DMTB 病患其 phagocytes 的 innate immune responses 有不正常的現象，這些不正常包括 IL-12 及 ROS 的產生。IL-12 減少會影響後續免疫反應中 IFN-γ 的產生和 T 細胞及吞噬細胞本身的活化，而 ROS 產量減少則會影響吞噬細胞殺菌還有細胞趨向性等重要免疫功能。因此 DMTB 病患其免疫系統中 IL-12 及 ROS 生成的缺失，可能就是第二型糖尿病病患對於 M. tuberculosis 感染感受度及預後較差的原因。

Mycobacterium tuberculosis (M. tuberculosis) infection is the leading infectious disease in the world, results in 10 million new cases and at least 3 million deaths each year. M. tuberculosis multiplies intracellular, so the cell-mediated immune responses and T helper-1 (Th1) cytokines are crucial for limiting M. tuberculosis infection. Previous studies showed that type II diabetes mellitus (DM) is an important risk factor for worsening tuberculosis. However, the exact mechanism of how diabetes influences the immune responses to fight against tuberculosis is not fully understood. In this study, we investigated the innate immune responses mediated by phagocytes from TB patients and TB patients with type II DM (DMTB) by monitoring their phagocytosis ability and reactive oxygen species (ROS) production, and the amounts of innate immunity cytokines (IL-12, IFN-γ) production by peripheral leukocytes in the presence of M. bovis BCG and IL-12 or IFN-γ. First, we found that IL-12 production is decreased in DMTB patients compared with normal controls and TB patients. This finding suggests that there is truly an immune defect in DMTB patients, especially on phagocytes, which is responsible for IL-12 production. Second, phagocytes isolated from DMTB patients had normal phagocytosis ability, but their ROS production is lower than control group and TB patients. Although there is a correlation between IFN-γ production and ROS production in TB patients, this correlation was not found in DMTB patients. These findings suggest that phagocytes from DMTB patients may have abnormality in activation and cytokine secretion. These findings give us evidences that type II DM may influence the innate immunity of phagocyte responses to mycobacteria and lead to more serious infections

Babior,B.M. (1999). NADPH oxidase: an update. Blood?93, 1464-1476.
Bashar,M., Alcabes,P., Rom,W.N., and Condos,R. (2001). Increased incidence of multidrug-resistant tuberculosis in diabetic patients on the Bellevue Chest Service, 1987 to 1997. Chest 120, 1514-1519.
Boldizsar,F., Berki,T., Miseta,A., and Nemeth,P. (2002). Effect of hyperglycemia on the basal cytosolic free calcium level, calcium signal and tyrosine-phosphorylation in human T-cells. Immuno Lett 82, 159-164.
Boleti,H., Ojcius,D.M., and Dautry-Varsat,A. (2000). Fluorescent labelling of intracellular bacteria in living host cells. Journal of Microbiological Methods 40, 265-274.
Brightbill,H.D., Libraty,D.H., Krutzik,S.R., Yang,R.B., Belisle,J.T., Bleharski,J.R., Maitland,M., Norgard,M.V., Plevy,S.E., Smale,S.T., Brennan,P.J., Bloom,B.R., Godowski,P.J., and Modlin,R.L. (1999). Host defense mechanisms triggered by microbial lipoproteins through toll-like receptors. Science 285, 732-736.
Casanova,J.L. (2001). Mendelian susceptibility to mycobacterial infection in man. Swiss. Med. Wkly. 131, 445-454.
Ceriello,A. and Motz,E. (2004). Is oxidative stress the pathogenic mechanism underlying insulin resistance, diabetes, and cardiovascular disease? The common soil hypothesis revisited. Arterioscler Thromb Vasc Biol. 24, 816-823.
Chan,J., Fan,X.D., Hunter,S.W., Brennan,P.J., and Bloom,B.R. (1991). Lipoarabinomannan, a possible virulence factor involved in persistence of Mycobacterium tuberculosis within macrophages. Infect. Immun. 59, 1755-1761.
Chang,F.Y. and Shaio,M.F. (1995). Respiratory burst activity of monocytes from patients with non-insulin-dependent diabetes mellitus. Diabetes Research and Clinical Practice 29, 121-127.
Cohen,A.W., Hnasko,R., Schubert,W., and Lisanti,M.P. (2004). Role of caveolae and caveolins in health and disease. Physiol Rev 84, 1341-1379.
Coker,R., McKee,M., Atun,R., Dimitrova,B., Dodonova,E., Kuznetsov,S., and Drobniewski,F. (2006). Risk factors for pulmonary tuberculosis in Russia: case-control study. Brit. Med. J. 332, 85-87.
Collison,K.S., Parhar,R.S., Saleh,S.S., Meyer,B.F., Kwaasi,A.A., Hammami,M.M., Schmidt,A.M., Stern,D.M., and Al Mohanna,F.A. (2002). RAGE-mediated neutrophil dysfunction is evoked by advanced glycation end products (AGEs). J Leukoc Biol 71, 433-444.
Cooper,A.M., Magram,J., Ferrante,J., and Orme,I.M. (1997). Interleukin 12 (IL-12) is crucial to the development of protective immunity in mice intravenously infected with mycobacterium tuberculosis. J. Exp. Med. 186, 39-45.
Cross,D.A., Alessi,D.R., Cohen,P., Andjelkovich,M., and Hemmings,B.A. (1995). Inhibition of glycogen synthase kinase-3 by insulin mediated by protein kinase B. Nature 378, 785-789.
DeLeo,F.R., Allen,L.A., Apicella,M., and Nauseef,W.M. (1999). - NADPH oxidase activation and assembly during phagocytosis. J. immunol. 163, 6732-6740.
Dye,C., Scheele,S., Dolin,P., Pathania,V., and Raviglione,M.C. (1999). Consensus statement. Global burden of tuberculosis: estimated incidence, prevalence, and mortality by country. WHO Global Surveillance and Monitoring Project. JAMA 282, 677-686.
Dziatkowiak,H., Kowalska,M., and Denys,A. (/19). - Phagocytic and bactericidal activity of granulocytes in diabetic children. -3.
Feinberg,J., Fieschi,C., Doffinger,R., Feinberg,M., Leclerc,T., Boisson-Dupuis,S., Picard,C., Bustamante,J., Chapgier,A., Filipe-Santos,O., Ku,C.L., de Beaucoudrey,L., Reichenbach,J., Antoni,G., Balde,R., Alcais,A., and Casanova,J.L. (2004). Bacillus Calmette Guerin triggers the IL-12/IFN-gamma axis by an IRAK-4- and NEMO-dependent, non-cognate interaction between monocytes, NK, and T lymphocytes. Eur. J. Immunol. 34, 3276-3284.
Fenton,M.J., Vermeulen,M.W., Kim,S., Burdick,M., Strieter,R.M., and Kornfeld,H. (1997). Induction of gamma interferon production in human alveolar macrophages by Mycobacterium tuberculosis. Infect. Immun. 65, 5149-5156.
Ferebee,S.H. (1970). Controlled chemoprophylaxis trials in tuberculosis. A general review. Bibl. Tuberc. 26, 28-106.
Ferrari,G.F., Langen,H.F., Naito,M.F., and Pieters,J. (1999). A coat protein on phagosomes involved in the intracellular survival of mycobacteria. Cell 97, 435-447.
Fieschi,C., Dupuis,S., Catherinot,E., Feinberg,J., Bustamante,J., Breiman,A., Altare,F., Baretto,R., Le Deist,F., Kayal,S., Koch,H., Richter,D., Brezina,M., Aksu,G., Wood,P., Al Jumaah,S., Raspall,M., Silva Duarte,A.J., Tuerlinckx,D., Virelizier,J.L., Fischer,A., Enright,A., Bernhoft,J., Cleary,A.M., Vermylen,C., Rodriguez-Gallego,C., Davies,G., Blutters-Sawatzki,R., Siegrist,C.A., Ehlayel,M.S., Novelli,V., Haas,W.H., Levy,J., Freihorst,J., Al Hajjar,S., Nadal,D., De,M., V, Jeppsson,O., Kutukculer,N., Frecerova,K., Caragol,I., Lammas,D., Kumararatne,D.S., Abel,L., and Casanova,J.L. (2003). Low penetrance, broad resistance, and favorable outcome of interleukin 12 receptor beta1 deficiency: medical and immunological implications. J Exp Med 17, 527-545.
Filipe-Santos,O., Bustamante,J., Haverkamp,M.H., Vinolo,E., Ku,C.L., Puel,A., Frucht,D.M., Christel,K., von Bernuth,H., Jouanguy,E., Feinberg,J., Durandy,A., Senechal,B., Chapgier,A., Vogt,G., de Beaucoudrey,L., Fieschi,C., Picard,C., Garfa,M., Chemli,J., Bejaoui,M., Tsolia,M.N., Kutukculer,N., Plebani,A., Notarangelo,L., Bodemer,C., Geissmann,F., Israel,A., Veron,M., Knackstedt,M., Barbouche,R., Abel,L., Magdorf,K., Gendrel,D., Agou,F., Holland,S.M., and Casanova,J.L. (2006). X-linked susceptibility to mycobacteria is caused by mutations in NEMO impairing CD40-dependent IL-12 production. J. Exp. Med. 10, 1745-1759.
Flynn,J.L., Chan,J., Triebold,K.J., Dalton,D.K., Stewart,T.A., and Bloom,B.R. (1993). An essential role for interferon gamma in resistance to Mycobacterium tuberculosis infection. J. Exp. Med. 178, 2249-2254.
Fortune,S.M., Solache,A., Jaeger,A., Hill,P.J., Belisle,J.T., Bloom,B.R., Rubin,E.J., and Ernst,J.D. (2004). Mycobacterium tuberculosis inhibits macrophage responses to IFN-gamma through myeloid differentiation factor 88-dependent and -independent mechanisms. J. Immunol. 172, 6272-6280.
Furst,D.E., Breedveld,F.C., Kalden,J.R., Smolen,J.S., Burmester,G.R., Bijlsma,J.W., Dougados,M., Emery,P., Keystone,E.C., Klareskog,L., and Mease,P.J. (2005). Updated consensus statement on biological agents, specifically tumour necrosis factor {alpha} (TNF{alpha}) blocking agents and interleukin-1 receptor antagonist (IL-1ra), for the treatment of rheumatic diseases, 2005. Ann Rheum Dis. 64 Suppl 4, iv2-iv14.
Gloire,G., Legrand-Poels,S., and Piette,J. (2006). NF-kappaB activation by reactive oxygen species: fifteen years later. Biochem. Pharmacol. 72, 1493-1505.
Guha,M., Bai,W., Nadler,J.L., and Natarajan,R. (2000). Molecular mechanisms of tumor necrosis factor alpha gene expression in monocytic cells via hyperglycemia-induced oxidant stress-dependent and -independent pathways. J Biol Chem 275, 17728-17739.
Hand,W.L., Hand,D.L., and Vasquez,Y. (2007). Increased polymorphonuclear leukocyte respiratory burst function in type 2 diabetes. Diabetes Res Clin Pract 44-50.
Hensley,K., Robinson,K.A., Gabbita,S.P., Salsman,S., and Floyd,R.A. (2000). Reactive oxygen species, cell signaling, and cell injury. Free Radic Biol Med 28, 1456-1462.
Hsueh,P.R., Liu,Y.C., So,J., Liu,C.Y., Yang,P.C., and Luh,K.T. (2006). Mycobacterium tuberculosis in Taiwan. J. infect. 52, 77-85.
Huse,M., Lillemeier,B.F., Kuhns,M.S., Chen,D.S., and Davis,M.M. (2006). T cells use two directionally distinct pathways for cytokine secretion. Nat. Immunol. 7, 247-255.
Hussain,R., Kaleem,A., Shahid,F., Dojki,M., Jamil,B., Mehmood,H., Dawood,G., and Dockrell,H.M. (2002). Cytokine profiles using whole-blood assays can discriminate between tuberculosis patients and healthy endemic controls in a BCG-vaccinated population. J Immunol Methods 264, 95-108.
Ikeda,S., Yamaoka-Tojo,M., Hilenski,L., Patrushev,N.A., Anwar,G.M., Quinn,M.T., and Ushio-Fukai,M. (2005). IQGAP1 regulates reactive oxygen species-dependent endothelial cell migration through interacting with Nox2. Arterioscler Thromb Vasc Biol 25, 2295-2300.
Inokuchi,N., Sugahara,K., Soda,H., Usui,T., Hirakata,Y., Fukushima,K., Yamada,Y., Kohno,S., and Kamihira,S. (2003). Relationship between whole-blood interferon-gamma production and extent of radiographic disease in patients with pulmonary tuberculosis. Diagn Microbiol Infect Dis. 46, 109-114.
Inoue,S., Lan,Y., Muran,J., and Tsuji,M. (1996). Reduced hydrogen peroxide production in neutrophils from patients with diabetes. Diabetes Res. Clin. Pr. 33, 119-127.
Jackson,S.H., Miller,G.F., Segal,B.H., Mardiney,M., III, Domachowske,J.B., Gallin,J.I., and Holland,S.M. (2001). IFN-gamma is effective in reducing infections in the mouse model of chronic granulomatous disease (CGD). J Interferon Cytokine Res. 21, 567-573.
Jenn-Tyang Chang and Chi-Chang Shieh. Influence of diabetes on manifestations and treatment outcome of pulmonary tuberculosis patients: clinical and mechanism investigation. 2006. Department of Microbiology & Immunology of National Cheng Kung University.
Jouanguy,E., Altare,F., Lamhamedi,S., Revy,P., Emile,J.F., Newport,M., Levin,M., Blanche,S., Seboun,E., Fischer,A., and Casanova,J.L. (1996). Interferon-gamma-receptor deficiency in an infant with fatal bacille Calmette-Guerin infection. N Engl J Med 335, 1956-1961.
Kamata,H., Honda,S., Maeda,S., Chang,L., Hirata,H., and Karin,M. (2005). Reactive oxygen species promote TNFalpha-induced death and sustained JNK activation by inhibiting MAP kinase phosphatases. Cell 120, 649-661.
Ladel,C.H., Szalay,G., Riedel,D., and Kaufmann,S.H. (1997). Interleukin-12 secretion by Mycobacterium tuberculosis-infected macrophages. Infect. Immun. 65, 1936-1938.
Lau,Y.L., Chan,G.C., Ha,S.Y., Hui,Y.F., and Yuen,K.Y. (1998). The role of phagocytic respiratory burst in host defense against Mycobacterium tuberculosis. Clin InfecT Dis. 26, 226-7.
Li,N. and Karin,M. (1999). - Is NF-kappaB the sensor of oxidative stress? FASEB J 13, 1137-1143.
Lienhardt,C. (2001). From exposure to disease: the role of environmental factors in susceptibility to and development of tuberculosis. Epidemiol Rev 23, 288-301.
Lin,J.C., Siu,L.K., Fung,C.P., Tsou,H.H., Wang,J.J., Chen,C.T., Wang,S.C., and Chang,F.Y. (2006). Impaired phagocytosis of capsular serotypes K1 or K2 Klebsiella pneumoniae in type 2 diabetes mellitus patients with poor glycemic control. J. Clin. Endocr. Metab. 91, 3084-3087.
Lin,Y., Zhang,M., Hofman,F.M., Gong,J., and Barnes,P.F. (1996). Absence of a prominent Th2 cytokine response in human tuberculosis. Infec Immun. 64, 1351-1356.
Marhoffer,W., Stein,M., Maeser E, and Federlin,K. (1992). Impairment of polymorphonuclear leukocyte function and metabolic control of diabetes. Diabetes Care 15, 256-260.
Martin,M., Rehani,K., Jope,R.S., and Michalek,S.M. (2005). Toll-like receptor-mediated cytokine production is differentially regulated by glycogen synthase kinase 3. Nat. Immunol. 6, 777-784.
Mazaki,Y., Hashimoto,S., Tsujimura,T., Morishige,M., Hashimoto,A., Aritake,K., Yamada,A., Nam,J.M., Kiyonari,H., Nakao,K., and Sabe,H. (2006). Neutrophil direction sensing and superoxide production linked by the GTPase-activating protein GIT2. Nat. Immunol. 7, 724-731.
Mohan,A.K., Cote,T.R., Block,J.A., Manadan,A.M., Siegel,J.N., and Braun,M.M. (2004). Tuberculosis following the use of etanercept, a tumor necrosis factor inhibitor. Clin Infect Dis. 39, 295-299.
Nabi,A.H., Islam,L.N., Rahman,M.M., and Biswas,K.B. (2005). Polymorphonuclear neutrophil dysfunctions in streptozotocin-induced type 1 diabetic rats. Journal of Biochemistry and Molecular Biology 38, 661-667.
Nakano,H., Nakajima,A., Sakon-Komazawa,S., Piao,J.H., Xue,X., and Okumura,K. (2006). Reactive oxygen species mediate crosstalk between NF-kappaB and JNK. Cell Death Differ. 13, 730-737.
Nikoulina,S.E., Ciaraldi,T.P., Mudaliar,S., Carter,L., Johnson,K., and Henry,R.R. (2002). Inhibition of glycogen synthase kinase 3 improves insulin action and glucose metabolism in human skeletal muscle. Diabetes 51, 2190-2198.
Ottenhoff,T.H., Kumararatne,D., and Casanova,J.L. (1998). Novel human immunodeficiencies reveal the essential role of type-I cytokines in immunity to intracellular bacteria. Immunol Today. 19, 491-494.
Park,H.S., Lee,S.H., Park,D., Lee,J.S., Ryu,S.H., Lee,W.J., Rhee,S.G., and Bae,Y.S. (2004). Sequential activation of phosphatidylinositol 3-kinase, beta Pix, Rac1, and Nox1 in growth factor-induced production of H2O2. Mol. Cell Biol. 24, 4384-94.
Perez,A., Brown,H.S., III, and Restrepo,B.I. (2006). Association between tuberculosis and diabetes in the Mexican border and non-border regions of Texas. Am J Trop Med Hyg 74, 604-611.
Raja,A. (2004). Immunology of tuberculosis. Indian J Med Res. 120, 213-232.
Rajagopalan,S. (2005). Serious infections in elderly patients with diabetes mellitus. Clinical Infectious Diseases 40, 990-996.
Schlesinger,L.S., Bellinger-Kawahara,C.G., Payne,N.R., and Horwitz,M.A. (1990). Phagocytosis of Mycobacterium tuberculosis is mediated by human monocyte complement receptors and complement component C3. J. Immunol. 144, 2771-2780.
Schlesinger,L.S., Hull,S.R., and Kaufman,T.M. (1994). - Binding of the terminal mannosyl units of lipoarabinomannan from a virulent strain of Mycobacterium tuberculosis to human macrophages. J. Immunol. 152, 4070-4079.
Schluger,N.W. (2001). Recent advances in our understanding of human host responses to tuberculosis. Respir Res 2, 157-163.
Shaikh,M.A., Singla,R., Khan,N.B., Sharif,N.S., and Saigh,M.O. (2003). Does diabetes alter the radiological presentation of pulmonary tuberculosis. Saudi. Med. J. 24, 278-281.
Smith,I. (2003). Mycobacterium tuberculosis pathogenesis and molecular determinants of virulence. Clin Microbiol Rev 16, 463-496.
Steinbrecher,K.A., Wilson,W., III, Cogswell,P.C., and Baldwin,A.S. (2005). Glycogen synthase kinase 3beta functions to specify gene-specific, NF-kappaB-dependent transcription. Mol Cell Biol 25, 8444-8455.
Tse,H.M., Milton,M.J., Schreiner,S., Profozich,J.L., Trucco,M., and Piganelli,J.D. (2007). Disruption of innate-mediated proinflammatory cytokine and reactive oxygen species third signal leads to antigen-specific hyporesponsiveness. J Immunol. 178, 908-917.
Ushio-Fukai,M. (2006). Localizing NADPH oxidase-derived ROS. Sci STKE 22.
Vilhardt,F. and van Deurs,B. (2004). The phagocyte NADPH oxidase depends on cholesterol-enriched membrane microdomains for assembly. EMBO J 23, 739-748.
Wallgren A. (1948). The time-tabel of tuberculosis. Tubercle 29, 245-251.
Wang,J., Wakeham,J., Harkness,R., and Xing,Z. (1999). Macrophages are a significant source of type 1 cytokines during mycobacterial infection. J. Clin. Invest 103, 1023-1029.
Wu,H.P., Hua,C.C., and Chuang,D.Y. (2007). Decreased in vitro interferon-gamma production in patients with cavitary tuberculosis on chest radiography. Respir Med 101, 48-52.
Yamashiro,S., Kawakami,K., Uezu,K., Kinjo,T., Miyagi,K., Nakamura,K., and Saito,A. (2005). Lower expression of Th1-related cytokines and inducible nitric oxide synthase in mice with streptozotocin-induced diabetes mellitus infected with Mycobacterium tuberculosis. Clin. Exp. Immunol. 139, 57-64.
Zhang,M., Gately,M.K., Wang,E., Gong,J., Wolf,S.F., Lu,S., Modlin,R.L., and Barnes,P.F. (1994). Interleukin 12 at the site of disease in tuberculosis. J. Clin Invest. 93, 1733-1739.
Zhang,Y., Heym,B., Allen,B., Young,D., and Cole,S. (19920). The catalase-peroxidase gene and isoniazid resistance of Mycobacterium tuberculosis. Nature 358, 591-593.
Zimmet,P., Alberti,K.G., and Shaw,J. (2001). Global and societal implications of the diabetes epidemic. Nature 414, 782-787.