Shin-Seok Lee

Impaired differentiation and cytotoxicity of natural killer cells in systemic lupus erythematosus

OBJECTIVE To determine the cytotoxicity of natural killer (NK) cells and the level of differentiation of hematopoietic stem cells (HSCs) into NK cells in systemic lupus erythematosus (SLE). METHODS Patients with SLE (n=108), rheumatoid arthritis (RA; n=90), Behçets disease (n=39), or ankylosing spondylitis (n=41) and healthy control subjects (n=173) were enrolled in the study. NK cell levels, NK cell cytotoxicities, and lymphokine-activated killer (LAK) activities against K562 cells were measured by flow cytometry. Gene expression was assessed by reverse transcription-polymerase chain reaction. NK cells were differentiated from peripheral blood and bone marrow HSCs in vitro. RESULTS Percentages and absolute numbers of NK cells, cytotoxicities, and LAK activities were significantly lower in the peripheral blood of SLE and RA patients than in that of healthy controls. In particular, this NK cell deficiency was more prominent in patients with lupus nephritis and those with thrombocytopenia. Notably, purified NK cells derived from SLE patients, but not RA patients, were found to have lower cytotoxicities and LAK activities than those from healthy controls. This defect of NK cells in SLE patients was found to be related to lower numbers of NK precursors and to the down-regulation of perforin and granzyme in NK cells. The proliferative capacity of HSCs, the percentages of NK cells differentiated from HSCs, and NK cell cytotoxicities were significantly lower in SLE patients. CONCLUSION In SLE patients, circulating levels of NK cells were diminished and their cytotoxicities were impaired. Furthermore, the differentiation of HSCs into NK cells was found to be defective. These abnormalities possibly contribute to immune system dysregulation in SLE.

Mucosal-Associated Invariant T Cell Deficiency in Systemic Lupus Erythematosus

Mucosal-associated invariant T (MAIT) cells contribute to protection against certain microorganism infections and play an important role in mucosal immunity. However, the role of MAIT cells remains enigmatic in autoimmune diseases. In this study, we examined the level and function of MAIT cells in patients with rheumatic diseases. MAIT cell, cytokine, and programmed death-1 (PD-1) levels were measured by flow cytometry. Circulating MAIT cell levels were significantly reduced in systemic lupus erythematosus (SLE) and rheumatoid arthritis patients. In particular, this MAIT cell deficiency was more prominent in CD8+ and double-negative T cell subsets, and significantly correlated with disease activity, such as SLE disease activity index and 28-joint disease activity score. Interestingly, MAIT cell frequency was significantly correlated with NKT cell frequency in SLE patients. IFN-γ production in MAIT cells was impaired in SLE patients, which was due to an intrinsic defect in the Ca2+/calcineurin/NFAT1 signaling pathway. In SLE patients, MAIT cells were poorly activated by α-galactosylceramide–stimulated NKT cells, thereby showing the dysfunction between MAIT cells and NKT cells. Notably, an elevated expression of PD-1 in MAIT cells and NKT cells was associated with SLE. In rheumatoid arthritis patients, MAIT cell levels were significantly higher in synovial fluid than in peripheral blood. Our study primarily demonstrates that MAIT cells are numerically and functionally deficient in SLE. In addition, we report a novel finding that this MAIT cell deficiency is associated with NKT cell deficiency and elevated PD-1 expression. These abnormalities possibly contribute to dysregulated mucosal immunity in SLE.

High-density genotyping of immune-related loci identifies new SLE risk variants in individuals with Asian ancestry.

Systemic lupus erythematosus (SLE) has a strong but incompletely understood genetic architecture. We conducted an association study with replication in 4,478 SLE cases and 12,656 controls from six East Asian cohorts to identify new SLE susceptibility loci and better localize known loci. We identified ten new loci and confirmed 20 known loci with genome-wide significance. Among the new loci, the most significant locus was GTF2IRD1-GTF2I at 7q11.23 (rs73366469, Pmeta = 3.75 × 10−117, odds ratio (OR) = 2.38), followed by DEF6, IL12B, TCF7, TERT, CD226, PCNXL3, RASGRP1, SYNGR1 and SIGLEC6. We identified the most likely functional variants at each locus by analyzing epigenetic marks and gene expression data. Ten candidate variants are known to alter gene expression in cis or in trans. Enrichment analysis highlights the importance of these loci in B cell and T cell biology. The new loci, together with previously known loci, increase the explained heritability of SLE to 24%. The new loci share functional and ontological characteristics with previously reported loci and are possible drug targets for SLE therapeutics.

Numerical and functional deficiencies of natural killer T cells in systemic lupus erythematosus: their deficiency related to disease activity

OBJECTIVE This study was designed to examine the frequency of natural killer T (NKT) cells and the response to α-galactosylceramide (α-GalCer) in SLE patients and to investigate the clinical relevance of NKT cell levels. METHODS Patients with SLE (n = 128) and age- and sex-matched healthy controls (HCs) (n = 92) were enrolled in the study. NKT cell and CD1d levels were measured by flow cytometry. Gene expression was determined by RT-PCR, and cytokine secretion by multiple cytokine assay. Peripheral blood mononuclear cells (PBMCs) were cultured in vitro with α-GalCer. Proliferation indices of NKT cells were estimated by flow cytometry. RESULTS Percentages and absolute numbers of NKT cells were significantly lower in the peripheral blood of SLE patients than in that of HCs, whereas CD1d levels in PBMCs were comparable between these two groups. Notably, this NKT cell deficiency was found to be correlated with SLEDAI. NKT cell proliferation was found to be impaired in SLE patients, and cytokine production by NKT cells in response to α-GalCer was diminished. This poor responsiveness to α-GalCer was found to be due to NKT cell dysfunction rather than to an abnormality in CD1d-expressing cells. CONCLUSIONS Our data show that NKT cell levels and functions are defective in SLE patients. Furthermore, these deficiencies were found to reflect disease activity. It would appear that these NKT cell abnormalities could contribute to immune system dysregulation in SLE.

Circulating mucosal-associated invariant T cell levels and their cytokine levels in healthy adults

Mucosal-associated invariant T (MAIT) cells have been reported to play an antimicrobial role in infectious diseases. However, little is known about age- and gender-related changes in circulating MAIT cell level and function in healthy population. The purposes of this study were to examine the level and cytokine production of circulating MAIT cells and their subsets in healthy adults and to investigate potential relationships between clinical parameters and MAIT cell levels or their subset levels. One hundred thirty-three healthy subjects were enrolled in this study. MAIT cells, their subset, and cytokine levels were measured by flow cytometry. Circulating MAIT cell levels were found to vary widely (0.19% to 21.7%) in the study subjects and to be significantly lower in elderly subjects (age, 61-92 years) than in young subjects (age, 21-40 years) (p<0.0005). No significant difference was found in the circulating MAIT cell levels between male and female subjects. A linear regression analysis revealed that circulating MAIT cell levels declined annually by 3.2% among men and 1.8% among women, respectively. Notably, the proportion of CD4+ MAIT cells increased with age, whereas that of CD8+ MAIT cells decreased with age. In addition, the production of interleukin (IL)-4 by MAIT cells was found to be significantly increased in elderly subjects and the ratio of interferon (IFN)-γ/IL-4 was lower as compared with young subjects, showing a Th1 to Th2 shift in cytokine profile in elderly subjects. Our data suggest that aging is associated with a reduction in circulating MAIT cells, accompanied with alterations in subset composition and cytokine profile.

High-density genotyping of immune loci in Koreans and Europeans identifies eight new rheumatoid arthritis risk loci.

OBJECTIVE A highly polygenic aetiology and high degree of allele-sharing between ancestries have been well elucidated in genetic studies of rheumatoid arthritis. Recently, the high-density genotyping array Immunochip for immune disease loci identified 14 new rheumatoid arthritis risk loci among individuals of European ancestry. Here, we aimed to identify new rheumatoid arthritis risk loci using Korean-specific Immunochip data. METHODS We analysed Korean rheumatoid arthritis case-control samples using the Immunochip and genome-wide association studies (GWAS) array to search for new risk alleles of rheumatoid arthritis with anticitrullinated peptide antibodies. To increase power, we performed a meta-analysis of Korean data with previously published European Immunochip and GWAS data for a total sample size of 9299 Korean and 45,790 European case-control samples. RESULTS We identified eight new rheumatoid arthritis susceptibility loci (TNFSF4, LBH, EOMES, ETS1-FLI1, COG6, RAD51B, UBASH3A and SYNGR1) that passed a genome-wide significance threshold (p<5×10(-8)), with evidence for three independent risk alleles at 1q25/TNFSF4. The risk alleles from the seven new loci except for the TNFSF4 locus (monomorphic in Koreans), together with risk alleles from previously established RA risk loci, exhibited a high correlation of effect sizes between ancestries. Further, we refined the number of single nucleotide polymorphisms (SNPs) that represent potentially causal variants through a trans-ethnic comparison of densely genotyped SNPs. CONCLUSIONS This study demonstrates the advantage of dense-mapping and trans-ancestral analysis for identification of potentially causal SNPs. In addition, our findings support the importance of T cells in the pathogenesis and the fact of frequent overlap of risk loci among diverse autoimmune diseases.

Dysfunction of Natural Killer T Cells in Patients with Active Mycobacterium tuberculosis Infection

ABSTRACT Natural killer T (NKT) cells are known to play a protective role in the immune responses of mice against a variety of infectious pathogens. However, little is known about the detailed information of NKT cells in patients with Mycobacterium tuberculosis infection. The aims of this study were to examine NKT cell levels and functions in patients with active M. tuberculosis infection, to investigate relationships between NKT cell levels and clinical parameters, and to determine the mechanism responsible for the poor response to α-galactosylceramide (α-GalCer). NKT cell levels were significantly lower in the peripheral blood of pulmonary tuberculosis and extrapulmonary tuberculosis patients, and the proliferative responses of NKT cells to α-GalCer were also lower in patients, whereas NKT cell levels and responses were comparable in latent tuberculosis infection subjects and healthy controls. Furthermore, this NKT cell deficiency was found to be correlated with serum C-reactive protein levels. In addition, the poor response to α-GalCer in M. tuberculosis-infected patients was found to be due to increased NKT cell apoptosis, reduced CD1d expression, and a defect in NKT cells. Notably, M. tuberculosis infection was associated with an elevated expression of the inhibitory programmed death-1 (PD-1) receptor on NKT cells, and blockade of PD-1 signaling enhanced the response to α-GalCer. This study shows that NKT cell levels and functions are reduced in M. tuberculosis-infected patients and these deficiencies were found to reflect the presence of active tuberculosis.

Bone destruction by receptor activator of nuclear factor κB ligand-expressing T cells in chronic gouty arthritis.

IntroductionThe purpose of this study was to analyze the cellular expressions of pro-resorptive cytokines in gouty tophus tissues, to determine the capacity of monosodium urate monohydrate (MSU) crystals to induce these cytokines, and to understand the mechanisms of bone destruction in chronic gout.MethodsFourteen fixed, paraffin-embedded, uninfected tophus samples were analyzed immunohistochemically. Peripheral blood mononuclear cells (PBMCs) were cultured in vitro with MSU crystals, and gene expression was assessed by reverse transcription-polymerase chain reaction. In vitro osteoclastogenesis was performed using PBMCs and synovial fluid mononuclear cells (SFMCs).ResultsCD4+ T cells, CD8+ T cells, CD20+ B cells and mast cells infiltrated tophus tissues. Tartrate-resistant acid phosphatase (TRAP)+ osteoclasts were present around tophi and in osteolytic lesions. Interleukin (IL)-1, IL-6 and tumor necrosis factor (TNF)-alpha were produced from infiltrated mononuclear cells, whereas receptor activator of nuclear factor κB ligand (RANKL) was strongly expressed in T cells. However, osteoprotegerin (OPG) was not or was weakly expressed in tophus tissues. MSU crystals induced the expressions of IL-1, IL-6, TNF-alpha and RANKL in PBMCs, but inhibited OPG expression. In addition, the pro-resorptive cytokines were highly expressed in SFMCs of gouty arthritis patients. Furthermore, in vitro osteoclastogenesis was enhanced in SFMC cultures, but inhibited in T cell-depleted SFMC cultures.ConclusionsOur study demonstrates that RANKL-expressing T cells and TRAP+ osteoclasts are present within gouty tophus tissues, and that infiltrating cells express pro-resorptive cytokines. Furthermore, our data show that MSU crystals have the potential to induce pro-resorptive cytokines, and T cells are involved in osteoclastogenesis in chronic gout.

Natural killer T cell deficiency in active adult-onset Still's Disease: correlation of deficiency of natural killer T cells with dysfunction of natural killer cells.

OBJECTIVE To examine the levels and functions of natural killer (NK) and natural killer T (NKT) cells, investigate relationships between NK and NKT cells, and determine the clinical relevance of NKT cell levels in patients with adult-onset Stills disease (AOSD). METHODS Patients with active untreated AOSD (n = 20) and age- and sex-matched healthy controls (n = 20) were studied. NK and NKT cell levels were measured by flow cytometry. Peripheral blood mononuclear cells were cultured in vitro with α-galactosylceramide (αGalCer). NK cytotoxicity against K562 cells and proliferation indices of NKT cells were estimated by flow cytometry. RESULTS Percentages and absolute numbers of NKT cells were significantly lower in the peripheral blood of AOSD patients than in that of healthy controls. Proliferative responses of NKT cells to αGalCer were also lower in patients, and this was found to be due to proinflammatory cytokines and NKT cell apoptosis. In addition, NK cytotoxicity was found to be significantly lower in patients than in healthy controls, but NK cell levels were comparable in the 2 groups. Notably, this NKT cell deficiency was found to be correlated with NK cell dysfunction and to reflect active disease status. Furthermore, αGalCer-mediated NK cytotoxicity, showing the interaction between NK and NKT cells, was significantly lower in AOSD patients than in healthy controls. CONCLUSION These findings demonstrate that NK and NKT cell functions are defective in AOSD patients and suggest that these abnormalities contribute to innate immune dysfunction in AOSD.

Appearance of Systemic Lupus Erythematosus in Patients with Myasthenia Gravis following Thymectomy : Two Case Reports

We report two cases of systemic lupus erythematosus (SLE) in myasthenia gravis (MG) patients who had undergone thymectomy. SLE developed in the patients 3 months or 13 yr after thymectomy, and polyarthritis was the main clinical manifestation of SLE. Both patients fulfilled at least four of the revised criteria for the classification of SLE. In this report, we describe two postthymectomy lupus patients and perform a comparative review of previous cases.