Gene expression and cytokine pattern of pulmonary tuberculosis patients and their contacts in Ethiopia

Bekele, Adane Mihret (2013-03)

Thesis (PhD)--Stellenbosch University, 2013.

Thesis

ENGLISH ABSTRACT: The immune response against M. tuberculosis is multifactorial, involving a network of innate and adaptive immune responses. Characterization of the immune response, a clear understanding of the dynamics and interplay of different arms of the immune response and the identification of infection-stage specific biomarkers are critical to allow the development of better tools for combating tuberculosis. In an attempt to identify such biomarkers, we studied pulmonary tuberculosis patients and their contacts in Addis Ababa, Ethiopia as part of EDCTP and BMGF funded tuberculosis projects by using multiplex techniques. We analysed 45 genes using the Multiplex Ligation Dependent Probe Amplification (MLPA) technique and the expression of IL-4δ2, BLR1, MARCO, CCL-19, IL7R, Bcl2, FcyR1A, MMP9, and LTF genes discriminate TB cases from their healthy contacts. FoxP3, TGFß1 and CCL-19 discriminate latently infected from uninfected contacts. Single genes predict with an area under the Receiver Operating Characteristic (ROC) curve of 0.68 to 0.85 while a combination of genes identified up to 95% of the different groups. Similarly, the multiplex analysis of cytokines and chemokines also showed that single or combinations of plasma cytokines and chemokines discriminate between different clinical groups accurately. The median plasma level of EGF, fractalkine, IFN-y, IL-4, MCP-3 and IP-10 is significantly different (p<0.05) in active tuberculosis and non active tuberculosis infection and the median plasma levels of IFN-y, IL-4, MCP-3, MIP-1ß and IP-10 were significantly different (p<0.05) before and after treatment. We also found a significant difference (p<0.05) in plasma levels of cytokines of patients infected with the different lineages and different families of the modern lineage. The plasma level of IL-4 was significantly higher in patients infected with lineage 3 (p<0.05) as compared to lineage 4 and the CAS familyinfected patients had a higher plasma level of IL-4 (P<0.05) as compared to patients infected with H and T families but there was no difference between H and T families. We identified genes and cytokines which had been reported from other studies in different settings and we believe that these molecules are very promising biomarkers for classifying active tuberculosis, latent infection, absence of infection and treated infection. These markers may be suitable for the development of clinically useful tools but require further validation and qualification in different populations and in larger studies.

AFRIKAANSE OPSOMMING: Die immuunrespons teen M. tuberculosis is multifaktoriaal en betrek ‘n netwerk van niespesifieke and spesifieke immuunresponse. Karakterisering van die immuunrespons, ‘n duidelike insig in die dinamika en tussenspel deur die verskillende arms van die immuunrespons en die identifikasie van spesifieke biomerkers is krities belangrik om die ontwikkeling van nuwe hulpmiddels teen tuberkulose te bevorder. In ‘n poging om sulke biomerkers te identifiseer het ons pulmonale tuberkulose pasiënte en hulle kontakte in Addis Ababa, Etiopië, as deel van die EDCTP en BMGF befondste tuberkulose projekte bestudeer met multipleks tegnieke. Ons het 45 gene analiseer met ‘Multiplex Ligation Dependent Probe Amplification (MLPA)’ en gevind dat die geenuitdrukking van IL-4•2, BLR1, MARCO, CCL-19, IL7R, Bcl2, Fc•R1A, MMP9, en LTF TB pasiënte van hulle kontakte onderskei. FoxP3, TGF•1 en CCL-19 onderskei tussen latent infekteerde en ongeïnfekteerde kontakte. Enkele gene voorspel met ‘n area onder die ‘Receiver Operating Characteristic (ROC)’ kurwe van 0.68 tot 0.85 terwyl die kombinasie van gene 95% van die verskillende groepe identifiseer. Soortgelyk het multipleks analise van sitokiene en chemokiene verskillende kliniese groepe akkuraat van mekaar onderskei. Die mediane plasmavlakke van EGF, fractalkine, IFN-•, IL-4, MCP-3 en IP-10 is beduidend verskillend (p<0.05) in aktiewe tuberkulose en nie-aktiewe tuberkulose infeksie en die mediane plasmavlak van IFN-•, IL-4, MCP-3, MIP-1• en IP-10 was beduidend verskillend voor en na behandeling. Ons het ook beduidende verskille (p<0.05) in plasmavlakke van sitokiene in pasiënte gevind wat infekteer is met verskillende stamme and verskillende families van die moderne stamme. Die plasmavlak van IL-4 was beduidend hoër in pasiënte wat infekteer is met stam 3 (p<0.05) teenoor stam 4 en die CAS familie-infekteerde pasiënte het ‘n hoër plasmavlak van IL-4 (p<0.05) teenoor pasiënte met H en T familie infeksie hoewel daar geen versikke was tussen die H en T families nie. Ons het gene en sitokiene identifiseer wat deur ander werkers onder verskillende omstandighede ook beskryf is en ons glo dat hierdie molekules baie belowende biomerkers is om aktiewe tuberkulose, latent tuberkulose, die afwesigheid van infeksie en behandelde infeksie van mekaar te onderskei. Hierdie merkers mag toepaslik wees vir die ontwikkeling van bruikbare kliniese hulpmiddele maar benodig verdere validasie en kwalifikasie in verskillende populasiegroepe en in groter studies.

Please refer to this item in SUNScholar by using the following persistent URL: http://hdl.handle.net/10019.1/71942
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