Screening And Identification Of Potenial Biomarkers For Multidrug-resistant Tuberculosis And Its Traditional Chinese Medicine Syndromes

At present,tuberculosis is still the most deady infectious disease in the world,especially multidrug-resistant tuberculosis（MDR-TB）is a key problem in global tuberculosis control Lack of effective early diagnosis methods,as well as long treatment cycles,large side effects,and patient intolerance,make the MDR-TB epidemic unable to be effectively controlled Traditional Chinese Medicine（TCM）treatment of MDR-TB has a good attenuating and synergistic effect.However,the lack of objective biological evidence for the classification and diagnosis of MDR-TB TCM syndromes has resulted in low efficacy of TCM treatment for MDR-TB.In first part study,we investigated long non-coding RNAs（IncRNAs）in plasma of MDR-TB patients,DS-TB（drug-sensitive tuberculosis）patients,and HC（healthy control）individuals and the potential diagnostic value for MDR-TB,using microarray,bioinformatics analysis and real-time quantitative polymerase chain reaction（RT-qPCR）methods.In second part study,isobaric tags for relative and absolute quantification-coupled twodimensional liquids chromatography-tandem mass spectrometry（iTRAQ-2DLC-MS/MS）combined with bioinformatics analysis method was applied to identify and screen the differentially expressed serum proteins（DEPs）,followed by the biological functional analysis.Additionally,DEPs among three different TCM syndromes of MDR-TB were verified and analyzed by enzyme-linked immune sorbent assay（ELISA）.Finally,a receiver operating characteristic（ROC）curve was performed to estimate the diagnostic ability of DEPs.The first part study found,compared with the DS-TB group,that a total of 543 IncRNAs（195 mRNAs）differentially expressed in the MDR-TB group,of which 181（63 mRNAs）were up-regulated and 362 were down-regulated（132 mRNAs）;Compared with the HC group,a total of 699 IncRNAs（231 mRNAs）were found to be differentially expressed in the MDR-TB group,including 345（123 mRNAs）up-regulated IncRNAs and 354 down-regulated IncRNAs（108 mRNAs）.Gene ontology（GO）,Kyoto Encyclopedia of Genes and Genomes（KEGG）and coding-noncoding co-expression（CNC）analyses showed that functions of differentially expressed IncRNAs were mainly enriched in the metabolism and regulation of substances,such as the fatty acid degradation pathway and the lysine degradation pathway.RT-qPCR confirmed that three IncRNAs were up-regulated in the MDR-TB group,including NR₁10750（P=0.036）,uc.212-（P=0.011）,and NR₁31237（P=0.008）.The second part study found,A total of 71 proteins were differentially expressed in the three different TCM syndrome groups of MDR-TB:pulmonary Yin deficiency（PYD）syndromegroup,Hyperactivity of Fire due to Yin deficiency（HFYD）syndrome group and deficiency of Qi and Yin（DQY）syndrome group.The expression level of transforming growth factor-beta-induced protein ig-h3（TGFBI）waslower in PYD syndrome group（P=0.002）.Pro-protein convertase subtilisin/kexin type 9（PCSK9）overexpressed in HFYD syndrome group（P<0.0001）.C-C motif chemokine 14（CCL14）expression level was reduced inDQY syndromegroup（P=0.004）.Therefore,this study demonstrated that serum TGFBI,PCSK9 and CCL14 are new potential biomarkers of PYD syndrome,HFYD syndrome and DQY syndrome for MDR-TB respectively.The first part study reveals the potential value of IncRNAs as biomarkers for early diagnosis of MDR-TB and the underlying mechanisms of these aberrantly expressed IncRNAs in the pathogenesis of MDR-TB.The first part study,this study provids biological basis for MDR-TB TCM syndrome classification and of great significance to take different treatment options for MDR-TB different TCM syndrome candidates.