Upregulated mRNA expression of major histocompatibility complex class I chain-related gene A in colon and activated natural killer cells of Chinese patients with ulcerative colitis.

OBJECTIVE: To explore the expression of major histocompatibility complex class I chain-related gene A (MICA) and its ligand in colonic mucosa and the role of MICA-natural killer (NK) group 2D (NKG2D) interaction in activating NK cells in ulcerative colitis (UC) patients. METHODS: Intestinal mucosal biopsies were obtained from patients with UC and the controls. The expression of major histocompatibility complex class I-related gene (MIC) genes was determined by a reverse transcription polymerase chain reaction (RT-PCR) and the imaging of MICA expressed on colonic mucosa was measured by confocal microscopy resonance scanning. NKG2D and intracellular interferon (IFN)-γ expressions on NK cells were assayed by flow cytometry. RESULTS: The relative amount of MICA mRNA in the colonic mucosa of UC patients was significantly higher than in that of the controls (3.5408 ± 2.6658 vs 1.0477 ± 0.7201, P = 0.001), as were the major histocompatibility complex class I chain-related gene B (MICB) (8.9879 ± 3.2893 vs 4.6293 ± 1.2616, P < 0.001) and NKG2D mRNA expression (2.4395 ± 0.8147 vs 1.1624 ± 0.3954, P < 0.001). Confocal microscopy resonance scanning had shown that MICA was localized predominantly on the basolateral membranes of the epithelium. Further flow cytometry confirmed that the percentage of IFN-γ producer NK cells that expressed NKG2D in peripheral blood lymphocytes was higher in UC patients than in the healthy controls (45.36% ± 12.47% vs 27.45% ± 9.30%, P < 0.001). CONCLUSION: MICA, MICB and NKG2D were upregulated in the colonic mucosa of UC and were associated with activating NK cells with promoted NKG2D and IFN-γ production.

[CTLA-4 gene polymorphism regulates CTLA-4 mRNA stability and protein level in patients with ulcerative colitis].

OBJECTIVE: To investigate the effect of (AT)n repeat polymorphism of the 3'untranslated region in cytotoxic T-lymphocyte associated antigen 4 (CTLA-4) gene on CTLA-4 mRNA stability and full length (flCTLA-4) and soluble CTLA4 (sCTLA-4) expression in ulcerative colitis (UC). METHODS: flCTLA-4 mRNA in colonic biopsies and sCTLA-4 mRNA stability in peripheral blood mononuclear cells of UC patients were measured by quantitative PCR and half-life, respectively. The protein expression of flCTLA-4 in colonic biopsies and sCTLA-4 in sera of UC patients were determined by immunohistochemistry and enzyme-linked immunosorbent assay, respectively. The polymorphism of CTLA-4 (AT)n repeats in 300 UC and 700 age and sex matched healthy controls was genotyped by fluorescent PCR. RESULTS: Among the UC patients, sCTLA-4 mRNA expression levels were decreased in active disease compared to non-active disease (P= 0.004). Carriers of the longer alleles of the (AT)n repeats expressed lower levels of flCTLA-4 and sCTLA-4 mRNA and sCTLA-4 protein than those of the shorter alleles in UC (all P< 0.01), and mRNA with long (AT)n repeat alleles has shorter half-life than mRNA with short alleles and, hence, are unstable. The frequency of long allele carriers of CTLA-4 (AT)n repeats was significantly higher in UC patients than in the healthy controls (22.0% vs. 6.3%, P< 0.01, OR= 4.21, 95% CI: 2.79-6.33), and associated with extensive colitis (P= 0.008). CONCLUSION: CTLA-4 gene expression levels were associated with (AT)n repeat polymorphisms in UC patients. The expression of CTLA-4 mRNA and protein were decreased in carriers of the longer alleles of the (AT)n repeats of CTLA-4 gene. This study suggests that CTLA-4 plays an important role in genetic risk and pathophysiology for UC in central China.

Effect of cytotoxic T lymphocyte-associated molecule 4 1661 gene polymorphism on its expression and transcription in ulcerative colitis.

OBJECTIVE: Our aim was to investigate the expression of cytotoxic T lymphocyte-associated molecule 4 (CTLA-4) in ulcerative colitis (UC) and to evaluate the effect of CTLA-4 gene -1661A/G polymorphism on CTLA-4 expression and transcription. METHODS: A total of 20 UC patients and 22 healthy controls matched by age and sex were enrolled at Zhongnan Hospital of Wuhan University in central China. The CTLA-4 -1661A/G polymorphism was genotyped by the polymerase chain reaction-restriction fragment length polymorphism method. A Western blot analysis was performed to determine the full length CTLA-4 (flCTLA-4) protein expression in the peripheral blood of the UC patients. Serum-soluble CTLA-4 (sCTLA-4) levels were measured by enzyme-linked immunosorbent assay. CTLA-4-1661G mutant promoter transcription function was analyzed by site-directed PCR-based mutagenesis. RESULTS: CTLA-4 protein expression on CD4(+) T cells in UC patients was lower than that in the healthy controls (P < 0.001) while serum sCTLA-4 in the UC patients was significantly higher than that in the healthy controls (P < 0.001). No correlation was found between flCTLA-4 and sCTLA-4 expression levels and the -1661 A/G polymorphism of the CTLA-4 gene. Meanwhile, CTLA-4 -1661 allele A had no significant impact on the promoter activity compared with allele G (P > 0.05). CONCLUSION: CTLA-4 expressions were aberrant in UC patients compared with the healthy controls. CTLA-4 -1661A/G polymorphism had no significant impact on CTLA-4 expression and transcription in the peripheral CD4 T cells of UC patients.

[Association of cytotoxic T lymphocyte antigen-4 promoter C-658T polymorphism with ulcerative colitis in Chinese].

OBJECTIVE: To investigate the association of gene polymorphism of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) with ulcerative colitis (UC) in Chinese. METHODS: One hundred and seventeen patients with UC and 246 healthy controls were genotyped for the polymorphisms of C-658T in the promoter and C61T at the 3' untranslated region of the CTLA-4 gene using polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) and polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP), respectively. The genotype and allele frequencies of the two groups were calculated and compared by chi square test. RESULTS: The frequency of TT+CT genotype at the CTLA-4 gene C-658T in the promoter was significantly higher in UC patients than that in healthy controls (P=0.015). The frequency of the T allele at this locus was also significantly higher in UC patients than that in the controls (P=0.033). The frequencies of TT genotype and T allele at the C-658T locus were highly associated with extensive colitis in UC patients (P=0.037, and P=0.0067, respectively). CONCLUSION: The T allele of CTLA-4 promoter C-658T locus was highly associated with UC in Chinese Han of central China.

[An association between MICB 0106 allele and ulcerative colitis in Chinese Han in Hubei province].

OBJECTIVE: To investigate the association between the exon 2, 3, 4 of MHC class I chain-related gene-B (MICB) and ulcerative colitis (UC) in Chinese Han. METHODS: Using polymerase chain reaction single-stranded conformation polymorphism, allele frequency of MICB exon 2, 3 and 4 in 105 patients with UC and 213 age and sex matched healthy controls were genotyped. All of the studied individuals were Chinese Han. RESULTS: Allele frequency of MICB 0106 was increased in patients with UC as compared with normal controls (19.0% vs 8.9%, P = 0.000, Pc < 0.001, OR = 2.402, 95% CI: 1.488-3.879). The frequency of MICB 0106 was increased significantly in patients with extensive colitis (24.4% vs 8.9%, P = 0.000, Pc < 0.001, OR = 3.294, 95% CI: 1.800-6.027), moderate and severe disease (24.1% vs 8.9%, P = 0.000, Pc < 0.001, OR = 3.294, 95% CI: 1.893-5.576) and in those with extra intestinal manifestations (20.5% vs 8.9%, P = 0.002, Pc = 0.012, OR = 2.626, 95% CI: 1. 18 4. 61). Furthermore, MICB 0106 allele was higher in frequency in the male patients with UC (22. % vs 8. % P = 0. 01, Pc =0 . 06, OR =3 . 76, 95% CI:1 . 37 6. 78) and the patients more than4 0 years old (28.8% vs 8.3% P = 0.000, Pc <0 .001, OR= 4 .500, 95% I:2 . 81 8.504) as compared with healthy controls. CONCLUSION: MICB 0106 allele is positively associated with UC, especially with extensive colitis, moderate and severe disease, presence of extra intestinal manifestations, male gender and age of more than 40 years in Chinese Han in Hubei province.