MsNRAMP2 Enhances Tolerance to Iron Excess Stress in Nicotiana tabacum and MsMYB Binds to Its Promoter.

Iron(Fe) is a trace metal element necessary for plant growth, but excess iron is harmful to plants. Natural resistance-associated macrophage proteins (NRAMPs) are important for divalent metal transport in plants. In this study, we isolated the MsNRAMP2 (MN_547960) gene from alfalfa, the perennial legume forage. The expression of MsNRAMP2 is specifically induced by iron excess. Overexpression of MsNRAMP2 conferred transgenic tobacco tolerance to iron excess, while it conferred yeast sensitivity to excess iron. Together with the MsNRAMP2 gene, MsMYB (MN_547959) expression is induced by excess iron. Y1H indicated that the MsMYB protein could bind to the "CTGTTG" cis element of the MsNRAMP2 promoter. The results indicated that MsNRAMP2 has a function in iron transport and its expression might be regulated by MsMYB. The excess iron tolerance ability enhancement of MsNRAMP2 may be involved in iron transport, sequestration, or redistribution.

Methylprednisolone pulse therapy promotes the differentiation of regulatory T cells by inducing the apoptosis of CD4+ T cells in patients with systemic lupus erythematosus.

OBJECTIVES: To investigate the differentiation of regulatory T cells (Tregs) induced by methylprednisolone (MP) pulse therapy in patients with Systemic Lupus Erythematosus (SLE). METHODS: We enrolled 30 patients with SLE and analyzed peripheral blood mononuclear cells (PBMCs) before and after MP pulse therapy. Peripheral Tregs, apoptosis of PBMCs subsets, and TGFß production by monocytes was quantified by flow cytometry. Proliferation and IFN-γ production of CD4+ T cells were measured. Furthermore, TGFß1 production by human monocyte-derived macrophages (HMDM) stimulated with MP-treated CD4+ T cells were quantified by ELISA. RESULTS: Peripheral Tregs was significantly increased after MP pulse therapy (6.76 ± 1.46% vs. 3.82 ± 1.02%, p < 0.01), with an expansion of Nrp1- induced Tregs (4.54 ± 0.46% vs. 1.75 ± 0.38%, p < 0.01). Proliferation and IFN-γ production of CD4+ T cells were significantly decreased after MP pulse therapy. MP pulse therapy induced CD4+ T cell apoptosis (early apoptosis, 26.34 ± 3.54% vs. 14.81 ± 2.89%, p < 0.01) and TGFß expression on monocytes (6.02% vs. 2.45%, p < 0.01). Furthermore, MP induced CD4+ T cell apoptosis in vitro, which stimulated HMDM to produce TGFß. Moreover, elevated TGFß level in supernatant from HMDM stimulated with MP-treated CD4+ T cells promoted Tregs differentiation. CONCLUSIONS: MP pulse therapy induces CD4+ T cell apoptosis, which promotes monocytes to produce TGFß and further facilitates Tregs differentiation. Newly-differentiated Tregs suppress proliferation and IFN-γ production of CD4+ T cells and contribute to immunoregulatory milieu after MP pulse therapy.

Metagenomic profiling of the pro-inflammatory gut microbiota in ankylosing spondylitis.

OBJECTIVE: Gut dysbiosis has been reported implicated in ankylosing spondylitis (AS), a common chronic inflammatory disease mainly affects sacroiliac joints and spine. Utilizing deep sequencing on the feces of untreated AS patients, our study aimed at providing an in-depth understanding of AS gut microbiota. METHODS: We analyzed the fecal metagenome of 85 untreated AS patients and 62 healthy controls by metagenomic shotgun sequencing, and 23 post-treatment feces of those AS patients were collected for comparison. Comparative analyses among different cohorts including AS, rheumatoid arthritis and Behcet's disease were performed to uncover some common signatures related to inflammatory arthritis. Molecular mimicry of a microbial peptide was also demonstrated by ELISpot assay. RESULTS: We identified AS-enriched species including Bacteroides coprophilus, Parabacteroides distasonis, Eubacterium siraeum, Acidaminococcus fermentans and Prevotella copri. Pathway analysis revealed increased oxidative phosphorylation, lipopolysaccharide biosynthesis and glycosaminoglycan degradation in AS gut microbiota. Microbial signatures of AS gut selected by random forest model showed high distinguishing accuracy. Some common signatures related to autoimmunity, such as Bacteroides fragilis and type III secretion system (T3SS), were also found. Finally, in vitro experiments demonstrated an increased amount of IFN-γ producing cells triggered by a bacterial peptide of AS-enriched species, mimicking type II collagen. CONCLUSIONS: These findings collectively indicate that gut microbiota was perturbed in untreated AS patients with diagnostic potential, and some AS-enriched species might be triggers of autoimmunity by molecular mimicry. Additionally, different inflammatory arthritis shared some common microbial signatures.

Early biochemical response to ursodeoxycholic acid and long-term prognosis of primary biliary cirrhosis: results of a 14-year cohort study.

UNLABELLED: The biochemical response to ursodeoxycholic acid (UDCA) in primary biliary cirrhosis is a strong predictor of long-term outcome and thus facilitates the rapid identification of patients needing new therapeutic approaches. Numerous criteria for predicting outcome of treatment have been studied based on biochemical response to UDCA at 1 year. We sought to determine whether an earlier biochemical response at 3 or 6 months could as efficiently identify patients at risk of poor outcome, as defined by liver-related death, liver transplantation, and complications of cirrhosis. We analyzed the prospectively collected data of 187 patients with a median follow-up of 5.8 years (range, 1.3-14 years). The survival rates without adverse outcome at 5 years and 10 years were 86% and 63%. Under UDCA therapy, laboratory liver parameters experienced the most prominent improvement in the first 3 months (P < 0.0001) and then stayed relatively stable for the following months. The Paris, Barcelona, Toronto, and Ehime definitions, but not the Rotterdam definition, applied at 3, 6, and 12 months significantly discriminated the patients in terms of long-term outcome. Compared with biochemical responses evaluated after 1 year of UDCA therapy, biochemical responses at the third month demonstrated higher positive predictive value (PPV) but lower negative predictive value (NPV) and increased negative likelihood ratio (NLR) by all definitions; biochemical responses at the sixth month showed higher or the same PPV and NPV and lower NLR by all definitions. CONCLUSION: For the previously published criteria, biochemical responses at the sixth month can be used in place of those evaluated after 1 year of UDCA therapy. Our findings justify a more rapid identification of patients who need new therapeutic approaches.

Clinical characteristics of multicentric reticulohistiocytosis and distinguished features from rheumatoid arthritis: a single-center experience in China.

OBJECTIVE: To investigate the clinical features of multicentric reticulohistiocytosis (MRH). METHODS: The clinical manifestations, laboratory examination results and histologic characteristics of eleven patients with MRH were collected and compared with those of 33 patients with rheumatoid arthritis. RESULTS: In total, 72.7% of the MRH patients were women. The median age was 46 years (range 33-84 years). Diagnosed by specific pathologic features, all MRH patients exhibited cutaneous involvement. The dorsa of the hands, arms, face and auricle were the most commonly affected areas. Nodules were also located on the legs, scalp, trunk, neck, and even the hypoglossis and buccal mucosa. Ten MRH patients (90.9%) had symmetric polyarthritis. Compared with rheumatoid arthritis (RA) patients, MRH patients were more likely to have distal interphalangeal joint (DIP) involvement (63.6% vs 24.2%, P = 0.017) and less likely to have elbow (36.4% vs 72.7%, P = 0.003), ankle (45.5% vs 93.9%, P < 0.001) and metacarpophalangeal joint (MCP) (36.4% vs 78.8%, P = 0.009) involvement. Positivity for rheumatoid factor (RF) (36.4% vs 84.6%, P = 0.001) and anti-CCP antibody (9.1% vs 81.8%, P = 0.000), as well as the median RF titer [43.8 (31.7-61.0) vs 175.4 (21.3-940.3), P = 0.021], in MRH patients was lower than in RA patients. Elevation of the erythrocyte sedimentation rate (ESR) was also less common in MRH patients than in RA patients (36.4% vs 72.7%, P = 0.030). After treatment with median- to large-dose corticosteroids and disease-modifying antirheumatic drugs, 8 patients achieved complete remission and 2 patients partial remission (skin lesions ameliorated, joint lesions not ameliorated). CONCLUSION: Always pathologically diagnosed, MRH is a systemic disease involving RA-like erosive polyarthritis and a specific distribution of skin nodules characterized by "coral beads". More DIP involvement and less elbow, ankle and MCP involvement are seen in MRH than in RA. In addition, less positivity and lower-titer RF, uncommon presence of anti-CCP antibodies and ESR elevation may be helpful to distinguish MRH from RA.

Contrast-enhanced ultrasound for evaluating arteritis activity in Takayasu arteritis patients.

INTRODUCTION: Contrast-enhanced ultrasound (CEUS) was recently used to evaluate vascularization within the carotid artery wall, and this process of vascularization was correlated with arteritis activity. We aimed to use CEUS to evaluate disease activity in Takayasu arteritis (TAK) patients. METHOD: We used CEUS to analyze 28 consecutive TAK patients. Disease activity was assessed according to the NIH criteria. We measured CEUS grades and assessed the correlation between contrast features and disease activity. RESULTS: Higher erythrocyte sedimentation rates (ESRs) were found (35 ± 28.7 vs. 13 ± 7.4 mm/h, p < 0.01), and CEUS carotid wall enhancement was more frequently (100% vs. 36.6%, p < 0.01) seen in TAK patients in the active phase than in those in the inactive phase. With increasing CEUS grades, both the artery wall thickness and ESR increased, and patients were more likely to be in the active phase (0 in grade 0, 42.9% in grade 1, and 75% in grade 2). Receiver operating characteristic (ROC) curve analysis showed that CEUS had an area under the ROC curve (AUC) of 0.872 (95% CI 0.785-0.959, p < 0.01), demonstrating good diagnostic accuracy. CONCLUSIONS: Higher CEUS grades reliably identify patients with active TAK.Key Points• No CEUS vascularization is obviously relative with the inactive disease of TAK patients.• Obvious CEUS vascularization is obviously relative with the active disease of TAK patients.

Exploring pathogenesis of primary biliary cholangitis by proteomics: A pilot study.

AIM: To explore the pathogenesis of primary biliary cholangitis (PBC) by identifying candidate autoantibodies in serum samples by proteomics and bioinformatics. METHODS: Nine antimitochondrial antibody (AMA)-positive PBC patients and nine age- and sex-matched AMA-negative PBC patients were recruited. Antigen enrichment technology was applied to capture autoantigens of human intrahepatic biliary epithelial cells (HiBECs) that are recognized by autoantibodies from the sera of PBC patients. Candidate autoantigens were identified by label-free mass spectrometry. Bioinformatics analysis with MaxQuant software (version 1.5.2.8), DAVID platform, and Cytoscape v.3.0 allowed illustration of pathways potentially involved in the pathogenesis of PBC. RESULTS: In total, 1081 candidate autoantigen proteins were identified from the PBC patient pool. Among them, 371 were determined to be significantly differentially expressed between AMA-positive and -negative PBC patients (P < 0.05). Fisher's exact test was performed for enrichment analysis of Gene Ontology protein annotations (biological processes, cellular components, and molecular functions) and the Kyoto Encyclopedia of Genes and Genomes pathways. Significantly different protein categories were revealed between AMA-positive and -negative PBC patients. As expected, autoantigens related to mitochondria were highly enriched in AMA-positive PBC patients. However, lower levels of AMA were also detected in AMA-negative PBC patients. In addition, autoantigens of AMA-negative PBC patients were mainly involved in B-cell activation, recognition of phagocytosis, and complement activation. CONCLUSION: AMA-negative PBC individuals may not exist, but rather, those patients exhibit pathogenesis pathways different from those of AMA-positive PBC. Comprehensive research is needed to confirm these observations.

MicroRNA profiling in Chinese patients with primary Sjögren syndrome reveals elevated miRNA-181a in peripheral blood mononuclear cells.

OBJECTIVE: Characterized by chronic inflammation, dysfunction of exocrine glands, and systemic autoimmunity, primary Sjögren syndrome (pSS) is a common autoimmune disease in elderly women. Our study was performed to explore the potential involvement of microRNA (miRNA) in Chinese patients with pSS. METHODS: Using microarrays, miRNA expression in peripheral blood mononuclear cells (PBMC) was profiled in 4 female patients with pSS and 3 healthy participants, followed by a large-scale study of 33 patients and 10 healthy individuals. Compared to the healthy participants, 202 miRNA were upregulated and 180 were downregulated in the patients with pSS. To confirm this finding, a set of regulated miRNA was further examined in a large patient group, using quantitative reverse transcriptase-PCR assays. RESULTS: MiR-181a was the miRNA that most profoundly differed between patients with pSS and healthy individuals; however, similar miRNA-181a expression profiles were found in groups with different disease phenotypes. Together, these observations suggested that an elevated miRNA-181a level is a general phenomenon in Chinese patients with pSS. CONCLUSION: In addition to the elevated miR-181a levels, our study led to the speculation that elevated miR-181a levels in the PBMC of these patients compromise the maturation of B cells, enabling them to recognize and attack autoantigens and resulting in disease phenotypes. In addition to the regulation of human miRNA, many virus-derived miRNA were unexpectedly upregulated in the patients with pSS, suggesting that viral infection of PBMC plays a role in this disease.

Allogeneic bone marrow mesenchymal stem cell transplantation in patients with UDCA-resistant primary biliary cirrhosis.

The objective of this study was to evaluate the safety and efficacy of allogeneic bone marrow mesenchymal stromal/stem cell transplantation (BM-MSCT) for patients with ursodeoxycholic acid (UDCA)-resistant primary biliary cirrhosis (PBC). Ten patients were enrolled in this trial of BM-MSCT. All patients were permitted to concurrently continue their previous UDCA treatment. The efficacy of BM-MSCT in UDCA-resistant PBC was assessed at various time points throughout the 12-month follow up. No transplantation-related side effects were observed. The life quality of the patients was improved after BM-MSCT as demonstrated by responses to the PBC-40 questionnaire. Serum levels of ALT, AST, γ-GT, and IgM significantly decreased from baseline after BM-MSCT. In addition, the percentage of CD8+ T cells was reduced, while that of CD4+CD25+Foxp3+ T cells was increased in peripheral lymphocytic subsets. Serum levels of IL-10 were also elevated. Notably, the optimal therapeutic outcome was acquired in 3 to 6 months and could be maintained for 12 months after BM-MSCT. In conclusion, allogeneic BM-MSCT in UDCA-resistant PBC is safe and appears to be effective.

Connective tissue diseases in primary biliary cirrhosis: a population-based cohort study.

AIM: To establish the frequency and clinical features of connective tissue diseases (CTDs) in a cohort of Chinese patients with primary biliary cirrhosis (PBC). METHODS: Three-hundred and twenty-two Chinese PBC patients were screened for the presence of CTD, and the systemic involvement was assessed. The differences in clinical features and laboratory findings between PBC patients with and without CTD were documented. The diversity of incidence of CTDs in PBC of different countries and areas was discussed. For the comparison of normally distributed data, Student's t test was used, while non-parametric test (Wilcoxon test) for the non-normally distributed data and 2 × 2 χ(2) or Fisher's exact tests for the ratio. RESULTS: One-hundred and fifty (46.6%) PBC patients had one or more CTDs. The most common CTD was Sjögren's syndrome (SS, 121 cases, 36.2%). There were nine cases of systemic sclerosis (SSc, 2.8%), 12 of systemic lupus erythematosus (SLE, 3.7%), nine of rheumatoid arthritis (RA, 2.8%), and 10 of polymyositis (PM, 3.1%) in this cohort. Compared to patients with PBC only, the PBC + SS patients were more likely to have fever and elevated erythrocyte sedimentation rate (ESR), higher serum immunoglobulin G (IgG) levels and more frequent rheumatoid factor (RF) and interstitial lung disease (ILD) incidences; PBC + SSc patients had higher frequency of ILD; PBC + SLE patients had lower white blood cell (WBC) count, hemoglobin (Hb), platelet count, γ-glutamyl transpeptidase and immunoglobulin M levels, but higher frequency of renal involvement; PBC + RA patients had lower Hb, higher serum IgG, alkaline phosphatase, faster ESR and a higher ratio of RF positivity; PBC + PM patients had higher WBC count and a tendency towards myocardial involvement. CONCLUSION: Besides the common liver manifestation of PBC, systemic involvement and overlaps with other CTDs are not infrequent in Chinese patients. When overlapping with other CTDs, PBC patients manifested some special clinical and laboratory features which may have effect on the prognosis.