Genome-wide DNA methylation profiling in differentiating Crohn's disease from intestinal tuberculosis.

BACKGROUND: Differential diagnosis of Crohn's disease (CD) and intestinal tuberculosis (ITB) is still difficult in clinical pratice. DNA methylation has been considered as a favorable area for biomarker exploration and identification. OBJECTIVE: The purpose of the current study was to evaluate DNA methylation changes between CD and ITB. METHODS: We performed a genome-wide association study to identify differentially methylated positions (DMPs), including 8 CD patients (before the initial of biologics or immunomodulators), 6 ITB patients, and 8 healthy controls (HCs), in whole blood DNA using the Infinium HumanMethylation850 BeadChip. RESULTS: Patients in the CD group and ITB group were all observed with hypo-methylated changes compared with HCs. However, the CD group overlaps with the ITB group in DNA methylation, suggesting a stable epigenetic profile between the two diseases. The pathway enrichment analysis showed the alternation in inflammation-related pathway, immune system, and signal transduction. Focused on the DMPs located in the promoter region, further analysis indicated hypermethylation of cg03122532 (5'UTR of KCNJ15) could be a potential CD-specific biomarker. CONCLUSIONS: We identified specific differential methylation loci related to CD and ITB in blood DNA. DNA metylation as a important epigenetic modification could contribute to the pathogenesis study and biomarker exploration of the diseases.

Differential prevalence of pathobionts and host gene polymorphisms in chronic inflammatory intestinal diseases: Crohn's disease and intestinal tuberculosis.

BACKGROUND AND OBJECTIVES: Crohn's disease (CD) and Intestinal tuberculosis (ITB) are chronic inflammatory ulcero-constrictive intestinal diseases with similar phenotype. Although both are disease models of chronic inflammation and their clinical presentations, imaging, histological and endoscopic findings are very similar, yet their etiologies are diverse. Hence, we aimed to look at differences in the prevalence of pathobionts like adherent-invasive Escherichia coli (AIEC), Listeria monocytogenes, Campylobacter jejuni and Yersinia enterocolitica in CD and ITB as well as their associations with host-associated genetic polymorphisms in genes majorly involved in pathways of microbial handling and immune responses. METHODS: The study cohort included 142 subjects (69 patients with CD, 32 with ITB and 41 controls). RT- PCR amplification was used to detect the presence of AIEC, L. monocytogenes, C. jejuni, and Y. enterocolitica DNA in colonic mucosal biopsies. Additionally, we tested three SNPs in IRGM (rs13361189, rs10065172, and rs4958847), one SNP in ATG16L1 (rs2241880) and one SNP in TNFRSF1A (rs4149570) by real-time PCR with SYBR green from peripheral blood samples in this cohort. RESULTS: In patients with CD, AIEC was most frequently present (16/ 69, 23.19%) followed by L. monocytogenes (14/69, 20.29%), C. jejuni (9/69, 13.04%), and Y. enterocolitica (7/69, 10.14%). Among them, L. monocytogenes and Y. enterocolitica were significantly associated with CD (p = 0.02). In addition, we identified all the three SNPs in IRGM (rs13361189, rs10065172, and rs4958847), one SNP in ATG16L1 (rs2241880) and TNFRSF1A (rs4149570) with a significant difference in frequency in patients with CD compared with ITB and controls (p<0.05). CONCLUSION: Higher prevalence of host gene polymorphisms, as well as the presence of pathobionts, was seen in the colonic mucosa of patients with CD as compared to ITB, although both are disease models of chronic inflammation.

Interleukin-22 receptor 1 is expressed in multinucleated giant cells: A study on intestinal tuberculosis and Crohn's disease.

BACKGROUND: It is challenging to distinguish intestinal tuberculosis from Crohn's disease due to dynamic changes in epidemiology and similar clinical characteristics. Recent studies have shown that polymorphisms in genes involved in the interleukin (IL)-23/IL-17 axis may affect intestinal mucosal immunity by affecting the differentiation of Th17 cells. AIM: To investigate the specific single-nucleotide polymorphisms (SNPs) in genes involved in the IL-23/IL-17 axis and possible pathways that affect susceptibility to intestinal tuberculosis and Crohn's disease. METHODS: We analysed 133 patients with intestinal tuberculosis, 128 with Crohn's disease, and 500 normal controls. DNA was extracted from paraffin-embedded specimens or whole blood. Four SNPs in the IL23/Th17 axis (IL22 rs2227473, IL1ß rs1143627, TGFß rs4803455, and IL17 rs8193036) were genotyped with TaqMan assays. The transcriptional activity levels of different genotypes of rs2227473 were detected by dual luciferase reporter gene assay. The expression of IL-22R1 in different intestinal diseases was detected by immunohistochemistry. RESULTS: The A allele frequency of rs2227473 (P = 0.030, odds ratio = 0.60, 95% confidence interval: 0.37-0.95) showed an abnormal distribution between intestinal tuberculosis and healthy controls. The presence of the A allele was associated with a higher IL-22 transcriptional activity (P < 0.05). In addition, IL-22R1 was expressed in intestinal lymphoid tissues, especially under conditions of intestinal tuberculosis, and highly expressed in macrophage-derived Langhans giant cells. The results of immunohistochemistry showed that the expression of IL-22R1 in patients with Crohn's disease and intestinal tuberculosis was significantly higher than that in patients with intestinal polyps and colon cancer (P < 0.01). CONCLUSION: High IL-22 expression seems to be a protective factor for intestinal tuberculosis. IL-22R1 is expressed in Langhans giant cells, suggesting that the IL-22/IL-22R1 system links adaptive and innate immunity.

The Polymorphism rs17525495 of LTA4H Is Associated with Susceptibility of Crohn's Disease instead of Intestinal Tuberculosis in a Chinese Han Population.

BACKGROUND: Because of the similarity of intestinal tuberculosis and Crohn's disease in disease phenotype, differential diagnosis has always been a clinical problem. Arachidonic acid metabolites play an important role in the inflammatory response of intestinal tuberculosis and Crohn's disease. Recent studies have shown that the polymorphism locus in the promoter region of LTA4H gene affects LTB4 expression level and the susceptibility to extrapulmonary tuberculosis. Thus, we identified a total of 148 patients with intestinal tuberculosis, 145 with Crohn's disease, and 700 normal controls in this study. METHODS: All the study participants were local Han people from Jiangxi Province in the past eleven years. DNA was extracted from the paraffin-embedded specimens or the whole blood. The LTA4H promoter SNP (rs17525495) was genotyped with TaqMan assay. RESULTS: The T-alleles frequency was not significantly increased in patients with intestinal tuberculosis compared with healthy control group (p=0.630; OR=1.07; 95%CI=0.81-1.41), while patients with Crohn's disease have significantly increased T allele frequency compared with healthy population (p=0.032; OR=1.34; 95%CI=1.03-1.75). During treatment, the presence of the T allele significantly increased the proportion of Crohn's patients requiring glucocorticoids (p<0.05). CONCLUSIONS: The T allele of LTA4H gene SNP (rs17525495) is a risk factor for Crohn's disease instead of intestinal tuberculosis. More importantly, there may be a potential association of the different genotypes of rs17525495 with the treatment efficacy of 5-ASA and glucocorticoids in patients with Crohn's disease. The association between LTA4H polymorphism and drugs therapeutic effects might contribute to the practice of precision medicine and the prediction of clinical outcomes.

Cytokine gene expression in intestinal tuberculosis and Crohn's disease.

BACKGROUND: Intestinal tuberculosis (TB) and Crohn's disease closely resemble each other clinically and morphologically. Little is known of cytokine regulation in intestinal TB. OBJECTIVE: To compare cytokine gene expression in colonic mucosa and peripheral blood mononuclear cells (PBMC) in TB with that in Crohn's disease. METHODS: Biopsies were obtained from normal and ulcerated colonic mucosa of 12 intestinal TB and 11 Crohn's disease patients, and PBMC from 15 intestinal TB and 12 Crohn's disease patients and 11 healthy volunteers. RNA was extracted, and the expression of selected cytokines, chemokines and pattern recognition receptors quantified by reverse transcriptase real-time polymerase chain reaction using SYBR green. RESULTS: The mRNA expression of interleukin-8 (IL-8), induced protein-10, tumour necrosis factor-alpha, IL-23 p19 and IL-12 p40, and Toll-like receptors (TLR) 1 and 2 in the ulcerated mucosa was increased in both intestinal TB and Crohn's disease. Expression of growth-related oncogene-alpha was increased in intestinal TB, while expression of interferon-gamma (IFN-) and TLR 4, 5 and 9 was increased in Crohn's disease. Expression of RANTES (regulated upon activation, normal T-cell expressed and secreted) was decreased in Crohn's disease. Secretion of IFN- or IL-10 from PBMC was not significantly altered in either disease. PBMC mRNA expression of IL-1, IL-6 and IL-8 mRNA was upregulated in Crohn's disease, while that of IL-17 was upregulated in intestinal TB. CONCLUSIONS: Cytokine gene expression patterns in intestinal mucosa and PBMC of intestinal TB were remarkably similar to Crohn's disease, and demonstrated innate immune activation and T-helper 1 polarisation.

LMP2/LMP7 gene variant: a risk factor for intestinal Mycobacterium tuberculosis infection in the Chinese population.

BACKGROUND AND AIMS: Low molecular mass protein-2 (LMP2) and low molecular mass protein-7 (LMP7) genes play a critical role in foreign antigen processing on the major histocompatibility complex-I CD8(+) cytotoxic T-lymphocyte pathway. This study was designed to investigate whether the sequence variants in the LMP2/LMP7 coding region were associated with intestinal Mycobacterium tuberculosis (M. tuberculosis) infection or with the co-infection of pulmonary tuberculosis. METHODS: A total of 168 patients with intestinal tuberculosis and 235 normal controls were recruited for this study. Two polymorphisms of LMP2 (Arg60-His) and LMP7 (Gln145-Lys) were identified by polymerase chain reaction-restriction fragment length polymorphism method. The associations of the LMP2/LMP7 genotype and haplotype with intestinal M. tuberculosis infection were assessed by using logistic regression analysis. RESULTS: The results revealed that LMP7 position codon 145 Lys/Lys and Gln/Lys alleles in the coding region were associated with the infection of intestinal M. tuberculosis (P=0.003, odds ratio [OR]= 3.86 and P < 0.001, OR = 2.28, respectively). Meanwhile, the Arg-Lys and Cys-Lys haplotypes exhibited significant relation to the intestinal M. tuberculosis infection (P= 0.006, OR=1.87; P=0.021, OR=1.83, respectively). No significant associations were observed for any of the single-nucleotide polymorphism genotypes or haplotypes with the co-infection of pulmonary tuberculosis (P > 0.05). CONCLUSIONS: The results indicated that the genetic variant within the LMP2/LMP7 gene would increase the risk of intestinal M. tuberculosis infection.

Use of polymerase chain reaction in the diagnosis of abdominal tuberculosis.

BACKGROUND: Early diagnosis and prompt treatment of abdominal tuberculosis is vitally important as it greatly reduces disease and treatment related morbidity and even mortality in extreme cases. A polymerase chain reaction (PCR) test was evaluated for its feasibility as a diagnostic tool in abdominal tuberculosis (TB) in the Indian scenario. METHODS: PCR for the identification of M. tuberculosis amplified a 340 bp nucleotide sequence located within the 38 kDa protein gene of M. tuberculosis. Tissues for processing were obtained from patients suspected to have abdominal TB. These were from various sources such as abdominal lymph nodes, segments of intestine and bowel obtained at various times and in different ways such as laparoscopy, colectomy, bowel and lymph node resection. Fifty such patients had their tissues sent for PCR. RESULTS: PCR results were compared with histopathology (HP). Of the 50 samples, 31 were positive for abdominal TB by HP whereas 30 were positive by PCR. Twenty-four of these were positive for both HP and PCR while of the seven samples positive for HP, five were negative and two gave inhibition by PCR. Six samples negative by HP were positive by PCR. CONCLUSION: This study demonstrates that PCR can be used as an effective tool to diagnose abdominal TB.