Increased Expression of Long Noncoding RNA LOC100506314 in T cells from Patients with Nonsegmental Vitiligo and Its Contribution to Vitiligo Pathogenesis.

This study aimed to identify the abnormal expression of long noncoding RNAs (lncRNAs) in T cells from patients with vitiligo and to investigate their functional roles in the immune system. Using microarray analysis, the expression levels of RNA transcripts in T cells from patients with vitiligo and controls were compared. We identified several genes and validated their expression levels in T cells from 41 vitiligo patients and 41 controls. The biological functions of the lncRNAs were studied in a transfection study using an RNA pull-down assay, followed by proteomic analysis and western blotting. The expression levels of 134 genes were significantly increased, and those of 142 genes were significantly decreased in T cells from vitiligo patients. After validation, six genes had increased expression, and three genes had decreased expression in T cells from patients with vitiligo. T-cell expression of LOC100506314 was increased in vitiligo, especially CD4+, but not CD8+ T cells. The expression levels of LOC100506314 in CD4+ T cells was positively and significantly associated with the severity of vitiligo. LOC100506314 was bound to the signal transducer and activator of transcription 3 (STAT3) and macrophage migration inhibitory factor (MIF). Enhanced expression of LOC100506314 inhibited the phosphorylation of STAT3, protein kinase B (AKT), and extracellular signal-regulated protein kinases (ERK), as well as the levels of nuclear protein of p65 and the expression of IL-6 and IL-17 in Jurkat cells and T cells from patients with vitiligo. In conclusion, this study showed that the expression of LOC100506314 was elevated in CD4+ T cells from patients with vitiligo and associated the severity of vitiligo. LOC100506314 interacted with STAT3 and MIF and inhibited IL-6 and IL-17 expression by suppressing the STAT3, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), AKT, and ERK pathways. Enhanced expression of LOC100506314 in T cells may be a potential treatment strategy for vitiligo.

HLA-B*27 Heavy Chain Homo-Oligomers Promote the Cytotoxicity of NK Cells via Activation of PI3K/AKT Signaling.

Background and Objectives: Ankylosing spondylitis (AS) is a chronic inflammatory disease and is highly linked with the expression of the human leukocytic antigen-B*27 (HLA-B*27) genotype. HLA-B*27 heavy chain (B*27-HC) has an innate characteristic to slowly fold, resulting in the accumulation of the misfolded B*27-HC and the formation of homo-oligomeric B*27-HC molecules. The homo-oligomeric B*27-HC can act as a ligand of KIR3DL2. Interaction of the homo-oligomeric B*27-HC molecules with KIR3DL2 will trigger the survival and activation of KIR3DL2-positive NK cells. However, the effects of homo-oligomeric B*27-HC molecules associated with KIR3DL2 on the cytotoxic activity of NK cells and their cytokine expressions remain unknown. Materials and Methods: HLA-B*-2704-HC was overexpressed in the HMy2.C1R (C1R) cell line. Western blotting and quantitative RT-PCR were used to analyze the protein expression and cytokine expression, respectively, when C1R-B*-2704 cells that overexpress B*2704-HC were co-cultured with NK-92MI cells. Flow cytometry was used to analyze the cytotoxicity mediated by NK-92MI cells. Results: Our results revealed that NK-92MI cells up-regulated the expression of perforin and enhanced the cytotoxic activity via augmentation of PI3K/AKT signaling after co-culturing with C1R-B*2704 cells. Suppression of the dimerized B*27-HC formation or treatment with an inhibitor of PI3K, LY294002, or with an anti-B*27-HC monoclonal antibody can reduce the perforin expression of NK-92MI after co-culturing with C1R-B*-2704. Co-culturing with C1R-B*-2704 cells suppressed the TNF-α and IL6 expressions of NK-92MI cells. Conclusion: Stimulation of NK cell-mediated cytotoxicity by homo-oligomeric B*27-HC molecules may contribute to the pathogenesis of AS.

Brain-Derived Neurotrophic Factor Suppressed Proinflammatory Cytokines Secretion and Enhanced MicroRNA(miR)-3168 Expression in Macrophages.

We investigated the role of brain-derived neurotrophic factor (BDNF) and its signaling pathway in the proinflammatory cytokines production of macrophages. The effects of different concentrations of BDNF on proinflammatory cytokines expression and secretion in U937 cell-differentiated macrophages, and human monocyte-derived macrophages were analyzed using enzyme-linked immunosorbent assay and real-time polymerase chain reaction. The CRISPR-Cas9 system was used to knockout p75 neurotrophin receptor (p75NTR), one of the BDNF receptors. Next-generation sequencing (NGS) was conducted to search for BDNF-regulated microRNA. A very low concentration of BDNF (1 ng/mL) could suppress the secretion of interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, and IL-6 in lipopolysaccharide (LPS)-stimulated macrophages but did not change their mRNA expression. BDNF suppressed IL-1ß and IL-6 secretion in human monocyte-derived macrophages. In U937 cells, BDNF suppressed the phosphorylation of JNK and c-Jun. The p75NTR knockout strongly suppressed IL-1ß, IL-6, and TNF-α secretion in macrophages and LPS-stimulated macrophages. BDNF regulated the expression of miR-3168 with Ras-related protein Rab-11A as its target. In conclusion, BDNF suppressed proinflammatory cytokines secretion in macrophages and inhibited the phosphorylation of JNK. Knockout of p75NTR suppressed proinflammatory cytokines expression and secretion. BDNF upregulated the expression of miR-3168. The inhibition of p75NTR could be a potential strategy to control inflammation.

Down-Regulation of LOC645166 in T Cells of Ankylosing Spondylitis Patients Promotes the NF-κB Signaling via Decreasingly Blocking Recruitment of the IKK Complex to K63-Linked Polyubiquitin Chains.

Ankylosing spondylitis (AS) is a chronic inflammatory disease that mainly affects the spine. AS is highly associated with the expression of HLA-B27. Up to 95% AS patients are HLA-B27-positive. However, only 1%-2% of the HLA-B27-positive carriers suffer from AS, implying that other factors may also govern the development of AS. Long non-coding RNAs (lncRNAs) can regulate the immune response via their binding proteins. In the present study, we have identified that the levels of lncRNA, LOC645166, in T cells of AS patients were reduced. Overexpression of LOC645166 in Jurkat cells down-regulated the IL-23p19 expression and suppressed the JAK2/STAT3 signaling in response to stimulation by phorbol 12-myristate 13-acetate. Suppression of STAT3 activation by LOC645166 was also observed when Jurkat cells or T cells of AS patient were treated with anti-CD3/CD28 antibodies. In order to explore the role of LOC645166 in the pathogenesis of AS, RNA pull-down assay plus proteomic approach and western blotting were performed and identified that LOC645166 prefers binding the K63-linked polyubiquitin chains. LOC645166 can suppress recruitment of the IKK complex to K63-linked polyubiquitin chains and diminish IKK2 activation, leading to down-regulation of NF-κB activation. Down-regulation of LOC645166 expression in T cells of AS patients up-regulates NF-kB activation via decreasingly impeding recruitment of the IKK complex to K63-linked polyubiquitin chains, allowing AS patients to exhibit more sensitivity to stimulation by the proinflammatory cytokines or by TLR ligands.

Increased Serum Levels of Brain-Derived Neurotrophic Factor Contribute to Inflammatory Responses in Patients with Rheumatoid Arthritis.

The aim of this study is to investigate the role of brain-derived neurotrophic factor (BDNF) in the inflammatory responses in patients with rheumatoid arthritis (RA). Serum levels of BDNF and the precursor form of BDNF (proBDNF) from 625 RA patients and 40 controls were analyzed using enzyme-linked immunosorbent assay. Effects of BDNF on the mitogen-activated protein kinase pathway were analyzed by Western blotting. Microarray analysis was conducted to search BDNF regulated gene expression in Jurkat cells, and the differentially expressed genes were validated using T cells from patients with RA and controls. Serum BDNF levels were significantly elevated in patients with RA compared with the controls. Low serum BDNF levels were found in RA patients with anxiety or receiving biologics treatment. BDNF (20 ng/mL) enhanced the phosphorylation of ERK, JNK, and c-Jun, but suppressed the phosphorylation of p38, whereas BDNF (200 ng/mL) enhanced the phosphorylation of ERK and p38. After validation, the expression of CAMK2A, MASP2, GNG13, and MUC5AC, regulated by BDNF and one of its receptors, NGFR, was increased in RA T cells. BDNF increased the IL-2, IL-17, and IFN-γ expression in Jurkat cells and IL-2 and IFN-γ secretion in activated peripheral blood mononuclear cells.

Modestly Elevated Serum Procalcitonin Levels in Patients with Rheumatoid Arthritis Free of Active Infection.

Background and objectives: To investigate the serum procalcitonin (PCT) levels among patients with rheumatoid arthritis (RA) without active infection compared with healthy controls and to understand the relationship of PCT with RA disease activity, and treatment received by patients. Materials and Methods: Patients aged 20 years and above with clinician-confirmed diagnosis of RA and healthy volunteers were included during regular outpatient visits, and those with active infection symptoms and signs were excluded. RA disease activity was measured using the Disease Activity Score-28 for Rheumatoid Arthritis with erythrocyte sedimentation rate (DAS28-ESR). Medications received by the patients were also recorded. Results: A total of 623 patients with RA and 87 healthy subjects were recruited in this study. The mean PCT were significantly higher in patients with RA (6.90 ± 11.81 × 10-3 ng/mL) compared with healthy controls (1.70 ± 6.12 × 10-3 ng/mL) (p < 0.001) and the difference remained statistically significant after adjusting for age and sex. In addition, multiple linear regression analysis showed that a lower rank-transformed PCT serum level was significantly correlated with the use of biologics (p = 0.017) and a high DAS28-ESR score (p = 0.028) in patients with RA. Conclusion: Patients with RA have a significantly higher serum PCT levels compared with healthy controls. The use of biologics and an active RA disease activity were associated with a lower level of PCT in patients with RA. Further investigation is required to determine the optimal cutoff value of PCT among patients with RA and its association with disease activity and biologic usage.

The Essential Role of Peptidylarginine Deiminases 2 for Cytokines Secretion, Apoptosis, and Cell Adhesion in Macrophage.

OBJECTIVE: The study aims to investigate the functional roles of peptidylarginine deiminase 2 (PADI2) in macrophages. METHODS: The clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein-9 nuclease (Cas9) system was used to knockout PADI2 in U937 cells. U937 cells were introduced to differentiate macrophages and were stimulated with lipopolysaccharides (LPS). The protein expression of PADI2, PADI4, and citrullinated proteins were analyzed by Western blotting. The mRNA and protein levels of interleukin 1 beta (IL-1ß), IL-6, and tumor necrosis factor-alpha (TNF-α) were analyzed using RT-PCR and ELISA, respectively. Cell apoptosis was analyzed using flow cytometry. Cell adhesion assay was performed using a commercially available fibrinogen-coated plate. RESULTS: PADI2 knockout could markedly suppress the PADI2 protein expression, but not the PADI4 protein expression. PADI2 knockout decreased the protein levels of citrullinated nuclear factor κB (NF-κB) p65, but not those of citrullinated histone 3, resulting in the decreased mRNA expression levels of IL-1ß and TNF-α in the U937 cells and IL-1ß and IL-6 in the differentiated macrophages and the macrophages stimulated with LPS. The cytokines levels of IL-1ß, IL-6, and TNF-α were all dramatically decreased in the PADI2 knockout group compared with in the controls. PADI2 knockout prevented macrophages apoptosis via the decreased caspase-3, caspase-2, and caspase-9 activation. PADI2 knockout also impaired macrophages adhesion capacity through the decreased protein levels of focal adhesion kinase (FAK), phospho-FAK, paxillin, phospho-paxillin, and p21-activated kinase 1. CONCLUSION: This study showed that PADI2 could promote IL-1ß, IL-6, and TNF-α production in macrophages, promote macrophage apoptosis through caspase-3, caspase-2, and caspase-9 activation and enhance cell adhesion via FAK, paxillin, and PAK1. Therefore, targeting PADI2 could be used as a novel strategy for controlling inflammation caused by macrophages.

Identification of phostensin in association with Eps 15 homology domain-containing protein 1 (EHD1) and EHD4.

Phostensin (PTS) encoded by KIAA1949 is a protein phosphatase 1 (PP1)-binding protein. In order to explore the cellular functions of PTS, we have searched PTS-binding proteins by using co-immunoprecipitation in combination with shotgun proteomics. Here, we report two novel PTS-binding proteins, Eps 15 homology domain-containing protein 1 (EHD1) and EHD4. PTS associated with EHD proteins was also observed in GST pull-down assays. Immunofluorescence microscopy demonstrated that the complex was co-localized at the endocytic vesicles. EHD proteins have been known to play a critical role in regulation of endocytic transport. Overexpression of PTS-ß can attenuate the endocytic trafficking of transferrin.

Increased peptidylarginine deiminases expression during the macrophage differentiation and participated inflammatory responses.

OBJECTIVE: To investigate the expression of peptidylarginine deiminases (PADIs) during macrophage differentiation and its role in inflammatory responses. METHODS: The protein expression of PADI2, PADI4, and citrullinated histone 3 in U937 cells, differentiated macrophages, and macrophages stimulated with lipopolysaccharides (LPS) were analyzed by Western blotting. Three PADI inhibitors were used for assessing their effects on the secretion of proinflammatory cytokines in macrophages. The differential expressed citrullinated proteins during macrophage differentiation were probed by self-prepared anti-citrullinated protein antibodies, and the reactive bands were sent for proteomic analyses. Transfection studies were conducted to search for the functions of specific proteins. A specific protein was cloned and citrullinated for its protein binding study. RESULTS: The expression of PADI2 and PADI4 markedly increased during macrophage differentiation, whereas the formation of citrullinated histone 3 increased after stimulated with lipopolysaccharides. Three PADI inhibitors suppressed the LPS mediated proinflammatory cytokines secretion, but did not affect the expression of PADI2 and PADI4. Plasminogen activator inhibitor-2 (PAI-2) was citrullinated during macrophage differentiation. The expression of PAI-2 increased during macrophage differentiation and further increased after stimulated with LPS. Suppressed PAI-2 expression decreased the expression and secretion of proinflammatory cytokines. Decreased PADI2 expression also suppressed the expression of PAI-2 and protein levels of citrullinated PAI-2. The citrullination of PAI-2 inhibited its binding ability to proteasome subunit beta type-1 (PSMB1). CONCLUSION: PADI2 and PADI4 protein levels increased during the macrophage differentiation resulting in protein citrullination, including PAI-2. The increased expression of PAI-2 promoted inflammatory response, and the citrullination of PAI-2 impaired its binding to PSMB1. Therefore, protein citrullination could play a critical role in macrophage differentiation and function.

Dispersion of Poly(urea-formaldehyde)-Based Microcapsules for Self-Healing and Anticorrosion Applications.

An effective dispersant, oleyl phosphate (OP), for the dispersion of poly(urea-formaldehyde)-based microcapsules in a typical epoxy coating material is proposed. Based on electron microscopy observations and rheological and mechanical characterizations, it is observed that the addition of merely 0.5 wt % of OP is sufficient to obtain good dispersion of the microcapsules in the epoxy. In the self-healing and anticorrosion experiments, a microcapsule content of at least 15 wt % is required to efficiently restore the epoxy matrix and provide corrosion protection to underlying low-carbon steel when the particles are not dispersed; however, the amount of microcapsules required to obtain good self-healing and anticorrosion efficiencies can be greatly reduced to only 5 wt % when the microcapsules are dispersed by OP.

The roles of anti-citrullinated protein antibodies in the immunopathogenesis of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a common systemic autoimmune disease. Its major manifestation is persistent joint inflammation, which can lead to bone destruction and severe disability. The immunopathogenesis of RA is very complex, involving both innate and adaptive immune systems. Recently, the discovery of anti-citrullinated protein antibodies (ACPAs) has revolutionized the diagnosis and our understanding of the immunopathogenesis of RA. The presence of ACPAs is also closely linked to the disease activity of RA. Therefore, it is reasonable to believe that ACPAs and protein citrullination are key issues for the development of RA. We have summarized the recent study results in this review. The first theory concerning the pathogenesis of RA proposed that ACPAs link the well-known genetic and environmental risk factors for developing RA. However, due to the close association between joint inflammation and ACPAs, a more direct role of ACPAs in the immunopathogenesis of RA is anticipated. Within the past 10 years, many studies, including some of our own, have shown that ACPAs can promote an inflammatory response through complement activation, formation of neutrophil extracellular traps, and direct binding to key players, including monocytes, osteoclasts, and osteoblasts, in the mediation of bone destruction in the joints of RA patients. We also present some new perspectives and issues that need to be further investigated.

Aberrant expression of interleukin-23-regulated miRNAs in T cells from patients with ankylosing spondylitis.

BACKGROUND: Interleukin (IL)-23 can facilitate the differentiation of IL-17-producing helper T cells (Th17). The IL-23/IL-17 axis is known to play a key role in the immunopathogenesis of ankylosing spondylitis (AS). We hypothesized that the expression of microRNAs (miRNAs, miRs) would be regulated by IL-23 and that these miRNAs could participate in the immunopathogenesis of AS. METHODS: Expression profiles of human miRNAs in K562 cells, cultured in the presence or absence of IL-23 for 3 days, were analyzed by microarray. Potentially aberrantly expressed miRNAs were validated using T-cell samples from 24 patients with AS and 16 control subjects. Next-generation sequencing (NGS) was conducted to search for gene expression and biological functions regulated by specific miRNAs in the IL-23-mediated signaling pathway. RESULTS: Initial analysis revealed that the expression levels of 12 miRNAs were significantly higher, whereas those of 4 miRNAs were significantly lower, in K562 cells after coculture with IL-23 for 3 days. Among these IL-23-regulated miRNAs, the expression levels of miR-29b-1-5p, miR-4449, miR-211-3p, miR-1914-3p, and miR-7114-5p were found to be higher in AS T cells. The transfection of miR-29b-1-5p mimic suppressed IL-23-mediated signal transducer and activator of transcription 3 (STAT3) phosphorylation in K562 cells. After NGS analysis and validation, we found that miR-29b-1-5p upregulated the expression of angiogenin, which was also upregulated in K562 cells after coculture with IL-23. Increased expression of miR-29b-1-5p or miR-211-3p could enhance interferon-γ expression. CONCLUSIONS: Among the miRNAs regulated by IL-23, expression levels of five miRNAs were increased in T cells from patients with AS. The transfection of miR-29b-1-5p mimic could inhibit the IL-23-mediated STAT3 phosphorylation and might play a role in negative feedback control in the immunopathogenesis of AS.

The role of aberrant expression of T cell miRNAs affected by TNF-α in the immunopathogenesis of rheumatoid arthritis.

BACKGROUND: Tumor necrosis factor-alpha (TNF-α) can cause diverse T cell dysfunctions in patients with rheumatoid arthritis (RA). It is involved in the regulation of microRNAs (miRNAs) expression in different cell types. We hypothesized that the expression of T cell miRNAs would be affected by TNF-α, and these miRNAs could participate in the immunopathogenesis of RA. METHODS: Expression profiles of 270 human miRNAs in Jurkat cells, cultured in the presence or absence of TNF-α for 7 days were analyzed by real-time polymerase chain reaction. Potentially aberrantly expressed miRNAs were validated using T cell samples from 35 patients with RA and 15 controls. Transfection studies were conducted to search for gene expression and biological functions regulated by specific miRNAs. RESULTS: Initial analysis revealed 12 miRNAs were significantly lower, whereas the expression level of miR-146a was significantly higher in Jurkat cells after being cultured with TNF-α for 7 days. Decreased expression of miR-139-3p, miR-204, miR-760, miR-524-5p, miR-136, miR-548d-3p, miR-214, miR-383, and miR-887 were noted in RA T cells. Expression levels of miR-139-3p, miR-204, miR-214, and miR-760 were correlated with the use of biologic agents. The transfection of miR-214 mimic suppressed TNF-α-mediated apoptosis of Jurkat cells. Increased phosphorylation of extracellular regulating kinase (ERK) and c-Jun N-terminal kinase (JNK) was noted in RA T cells and Jurkat cells after TNF-α exposure. Transfection of Jurkat cells with miR-214 mimic suppressed both the basal and TNF-α-mediated ERK and JNK phosphoryation. CONCLUSIONS: Among T cell miRNAs affected by TNF-α, the expression levels of nine miRNAs were decreased in T cells from patients with RA. The expression levels of miR-139-3p, miR-204, miR-214, and miR-760 increased in RA patients receiving biologic agents. The transfection of miR-214 reversed the TNF-α-mediated cells apoptosis and inhibited the phosphorylation of ERK and JNK in Jurkat cells.

Targeted Delivery of the HLA-B∗27-Binding Peptide into the Endoplasmic Reticulum Suppresses the IL-23/IL-17 Axis of Immune Cells in Spondylarthritis.

Ankylosing spondylitis (AS) is highly associated with the expression of human leukocyte antigen-B27 (HLA-B∗27). HLA-B∗27 heavy chain (B27-HC) has an intrinsic propensity to fold slowly, leading to the accumulation of the misfolded B27-HC in the endoplasmic reticulum (ER) and formation of the HLA-B∗27 HC homodimer, (B27-HC)2, by a disulfide linkage at Cys-67. (B27-HC)2 displayed on the cell surface can act as a ligand of the killer-cell Ig-like receptor (KIR3DL2). (B27-HC)2 binds to KIR3DL2 of NK and Th17 cells and activates both cells, resulting in the activation of the IL-23/IL-17 axis to launch the inflammatory reaction in AS patients. However, activation of the IL-23/IL-17 axis originally derived from the HLA-B∗27 misfolding in the ER needs to be characterized. In this study, we delivered two HLA-B∗27-binding peptides, KRGILTLKY and SRYWAIRTR, into the ER by using a tat-derived peptide (GRKKRRQRRR)-His6-ubiquitin (THU) vehicle. Both peptides are derived from the human actin and nucleoprotein of influenza virus, respectively. Our results demonstrated that targeted delivery of both HLA-B∗27-binding peptides into the ER can promote the HLA-B∗27 folding, decrease the levels of (B27-HC)2, and suppress the activation of the IL-23/IL-17 axis in response to lipopolysaccharide. Our findings can provide a new therapeutic strategy in AS.

Increased Serum Levels of Anti-Carbamylated 78-kDa Glucose-Regulated Protein Antibody in Patients with Rheumatoid Arthritis.

The objective of this study was to investigate the presence and titer of anti-carbamylated 78-kDa glucose-regulated protein (anti-CarGRP78) antibody in serum from controls, and patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and primary Sjögren syndrome (pSS). Thirty-three RA patients, 20 SLE patients, 20 pSS patients, and 20 controls were enrolled from our outpatient clinic. GRP78 was cloned and carbamylated. Serum titers of anti- cyclic citrullinated peptides (anti-CCP), anti-GRP78, and anti-CarGRP78 were measured with an enzyme-linked immunosorbent assay. No differences in serum titers of anti-GRP78 antibody in patients with RA, SLE, or pSS compared with the controls were observed. Serum levels of anti-carGRP78 antibody in patients with RA, but not SLE or pSS, were significantly higher compared with the controls (OD405 0.15 ± 0.08 versus 0.11 ± 0.03, p = 0.033). There was a positive correlation between the serum levels of anti-GRP78 antibody, but not anti-CarGRP78 antibody, with the levels of anti-CCP antibody in patients with RA. Both anti-GRP78 and anti-carGRP78 antibodies failed to correlate with C-reactive protein levels in patients with RA. In conclusion, we demonstrated the presence of anti-CarGRP78 antibody in patients with RA. In addition, the serum titer of anti-CarGRP78 antibody was significantly elevated in patients with RA compared with the controls. Anti-CarGRP78 antibody could also be detected in patients with SLE or pSS.

Increased expression of long noncoding RNAs LOC100652951 and LOC100506036 in T cells from patients with rheumatoid arthritis facilitates the inflammatory responses.

The aim of this study was to evaluate whether the presence of aberrantly expressed lncRNAs could promote T cell inflammatory responses in patients with RA. The expression levels of 10 potential aberrantly expressed lncRNAs were evaluated in T cells from 39 patients with RA and 17 controls using real-time reverse transcription polymerase chain reaction. The aberrantly expressed lncRNAs were measured in Jurkat cells co-cultured with or without ionomycin and phorbol 12-myristate 13-acetate. Transfection studies using small interfering RNA (siRNA) were conducted for biological functions, and microarray analysis was performed to search for target genes of specific lncRNAs. We confirmed that the expression levels of LOC100652951 and LOC100506036 were higher in RA T cells compared with controls. RA patients treated with biologic agents had lower expression levels of LOC100652951, and female RA patients had lower LOC100506036 expression levels after multivariate analysis. After activation, the expression levels of LOC100506036, but not LOC100652951, increased in Jurkat cells. Transfection of siRNA targeting LOC100506036 inhibited interferon gamma production and the expression of nuclear factor of activated T cells in activated Jurkat cells. After the microarray analysis with validation, inhibition of LOC100506036 expression by siRNA leaded to the decreased expression of sphingomyelin phosphodiesterase 1 (SMPD1). In conclusion, the expression levels of LOC100652951 and LOC100506036 were increased in RA T cells. Treatment with biologic agents could lower the expression of LOC100652951 in RA T cells. LOC100506036 could regulate the expression of SMPD1 and NFAT1 and could contribute to the inflammatory responses in RA.

Anti-citrullinated protein antibodies promote apoptosis of mature human Saos-2 osteoblasts via cell-surface binding to citrullinated heat shock protein 60.

We hypothesized that anti-citrullinated protein antibodies (ACPAs) react with osteoblast surface citrullinated proteins and affect cell function, leading to joint damage in patients with rheumatoid arthritis (RA). First, we purified ACPAs by cyclic citrullinated peptide (CCP)-conjugated affinity column chromatography. The cognate antigens of ACPAs on Saos-2 cells, a sarcoma osteogenic cell line generated from human osteoblasts, were probed by ACPAs, and the reactive bands were analyzed using proteomic analyses. We found that ACPAs bind to Saos-2 cell membrane, and several protein candidates, including HSP60, were identified. We then cloned and purified recombinant heat shock protein 60 (HSP60) and citrullinated HSP60 (citHSP60) and investigated the effect of ACPAs on Saos-2 cell. We confirmed that HSP60 obtained from Saos-2 cell membrane were citrullinated and reacted with ACPAs, which induces Saos-2 cells apoptosis via binding to surface-expressed citHSP60 through Toll-like receptor 4 signaling. ACPAs promoted interleukin (IL)-6 and IL-8 expression in Saos-2 cells. Finally, sera from patients with RA and healthy controls were examined for their titers of anti-HSP60 and anti-citHSP60 antibodies using an enzyme-linked immunosorbent assay. The radiographic change in patients with RA was evaluated using the Genant-modified Sharp scoring system. Patients with RA showed higher sera titers of anti-citHSP60, but not anti-HSP60, antibodies when compared with controls. In addition, the anti-citHSP60 level was positively associated with increased joint damage in patients with RA. In conclusion, Saos-2 cell apoptosis was mediated by ACPAs via binding to cell surface-expressed citHSP60 and the titer of anti-citHSP60 in patients with RA positively associated with joint damage.

Anti-citrullinated protein antibodies suppress let-7a expression in monocytes from patients with rheumatoid arthritis and facilitate the inflammatory responses in rheumatoid arthritis.

We hypothesized that anti-citrullinated protein antibodies (ACPAs) could affect the expression of miRNAs in monocytes and contribute to the inflammatory responses in rheumatoid arthritis (RA). The expression profiles of 270 human miRNAs, co-cultured with ACPAs or human immunoglobulin G (IgG), were analyzed using real-time polymerase chain reaction. Ten miRNAs exhibited differential expression in U937 cells after co-cultured with ACPAs compared with human IgG. The expression levels of these miRNAs were investigated in monocytes from 21 ACPA-positive RA patients and 13 controls. Among these miRNAs, the expression levels of let-7a was decreased in monocytes from ACPA-positive RA patients. The expression levels of let-7a showed a negative correlation with positivity of rheumatoid factor in patients sampled. We found that transfection of U937 cells with let-7a mimic suppressed K-Ras protein expression. In the ACPA-mediated signaling pathway, transfection of U937 cells with let-7a mimic suppressed the ACPA-enhanced phosphorylation of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase 1/2 (ERK1/2), and the expression and secretion of interleukin (IL)-1ß. In conclusion, ACPA-mediated decreased let-7a expression in monocytes from ACPA-positive RA patients. Decreased let-7a expression was associated with the positivity of RF in ACPA-positive RA patients. The decreased expression of let-7a could facilitate the inflammatory pathway via enhanced ACPA-mediated phosphorylation of ERK1/2 and JNK and increased expression of IL-1ß through an increase in the expression of Ras proteins.

An extensive DeBakey type IIIb aortic dissection with massive right pleural effusion presenting as abdominal pain and acute anemia: particular case report.

We describe the case of a 79-year-old male presented with sudden onset of abdominal pain and mild breathlessness, and complicated acute progressive anemia with haemoglobin which declined from 120 g/L to 70 g/L within five days. An urgent computed tomography angiography showed acute thoracic aortic dissection, DeBakey type IIIb, a dissecting aneurysm in the proximal descending thoracic aorta starting immediately after the origin of the left subclavian artery and extending distally below the renal arteries with evidence of rupture into the right pleural cavity for massive pleural effusion. Plasma D-dimer, brain natriuretic peptide and C reactive protein level were elevated. Our case showed that D-dimer can be used as a 'rule-out' test in patients with suspected aortic dissection. A raised BNP may exert a protective role through anti-inflammatory endothelial actions in the systemic circulation.