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.

Metabolic regulation of immune cells in proinflammatory microenvironments and diseases during ageing.

The process of ageing includes molecular changes within cells and interactions between cells, eventually resulting in age-related diseases. Although various cells (immune cells, parenchymal cells, fibroblasts and endothelial cells) in tissues secrete proinflammatory signals in age-related diseases, immune cells are the major contributors to inflammation. Many studies have emphasized the role of metabolic dysregulation in parenchymal cells in age-related inflammatory diseases. However, few studies have discussed metabolic modifications in immune cells during ageing. In this review, we introduce the metabolic dysregulation of major nutrients (glucose, lipids, and amino acids) within immune cells during ageing, which leads to dysfunctional NAD + metabolism that increases immune cell senescence and leads to the acquisition of the corresponding senescence-associated secretory phenotype (SASP). We then focus on senescent immune cell interactions with parenchymal cells and the extracellular matrix and their involvement in angiogenesis, which lead to proinflammatory microenvironments in tissues and inflammatory diseases at the systemic level. Elucidating the roles of metabolic modifications in immune cells during ageing will provide new insights into the mechanisms of ageing and therapeutic directions for age-related inflammatory diseases.

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.

Lupus Myocarditis: A Case-Control Study from China.

BACKGROUND: Myocarditis is an uncommon but serious manifestation of systemic lupus erythematosus (SLE). This study aimed to investigate clinical characteristics and outcomes of lupus myocarditis (LM) and to determine risk factors of LM in hospitalized Chinese patients with SLE. METHODS: We conducted a retrospective case-control study. A total of 25 patients with LM from 2001 to 2012 were enrolled as the study group, and 100 patients with SLE but without LM were randomly pooled as the control group. Univariable analysis was performed using Chi-square tests for categorical variables, and the Student's t-test or Mann-Whitney U-test was performed for continuous variables according to the normality. RESULTS: LM presented as the initial manifestation of SLE in 7 patients (28%) and occurred mostly at earlier stages compared to the controls (20.88 ± 35.73 vs. 44.08 ± 61.56 months, P = 0.008). Twenty-one patients (84%) experienced episodes of symptomatic heart failure. Echocardiography showed that 23 patients (92%) had decreased left ventricular ejection fraction (<50%) and all patients had wall motion abnormalities. A high SLE Disease Activity Index was the independent risk factor in the development of LM (odds ratio = 1.322, P < 0.001). With aggressive immunosuppressive therapies, most patients achieved satisfactory outcome. The in-hospital mortality was not significantly higher in the LM group than in the controls (4% vs. 2%,P = 0.491). CONCLUSIONS: LM could result in cardiac dysfunction and even sudden death. High SLE disease activity might potentially predict the occurrence of LM at the early stage of SLE. Characteristic echocardiographic findings could confirm the diagnosis of LM. Early aggressive immunosuppressive therapy could improve the cardiac outcome of LM.

Regulation of Cell Cycle Regulators by SIRT1 Contributes to Resveratrol-Mediated Prevention of Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is a major cause of morbidity and mortality in rheumatic diseases. Vascular remodeling due to the proliferation of pulmonary arterial smooth muscle cells (PASMCs) is central to the development of PAH. To date, it is still unclear if Silence Information Regulator 1 (SIRT1) regulates cell cycle regulators in the proliferation of PASMCs and contributes to prevention of PAH by resveratrol. In this study, we found that a significant decrease of SIRT1 expression levels in platelet-derived growth factor BB (PDGF-BB) treated human PASMCs (HPASMCs) and in monocrotaline (MCT) induced PAH rat. Overexpression of SIRT1 induced G1 phase arrest and increased p21 expression but decreased cyclin D1 expression in PDGF-BB treated HPASMCs. Moreover, resveratrol attenuated pulmonary arterial remodeling, decreased pulmonary arterial pressure, and upregulated SIRT1 and p21 expression but downregulated cyclin D1 expression in MCT induced PAH rat. Notably, knockdown of SIRT1 eliminated the regulation of resveratrol on p21 and cyclin D1 expression in PDGF-BB treated HPASMCs. These results demonstrated that SIRT1 mediated the regulation of resveratrol on the expression of cell cycle regulatory molecules. It suggests that SIRT1 exerts a protective role in PAH associated with rheumatic diseases and can be a potential treatment target.

Study on like-stem characteristics of tumor sphere cells in human gastric cancer line HGC-27.

Stem-like cancer cells are called cancer stem cells (CSCs) or tumor stem cells (TSCs). Methods for sorting CSCs are mainly based on the marker (CD133+/CD44+) or side population cells. However, CD133+/CD44+ cells or side population cells are very rare or even undetectable. In the present study, the tumor sphere of human gastric cancer (HGC) cell line HGC-27 was used for CSCs enrichment, and stem-like characteristics were verified by Hoechst 33342 staining technology, cell growth rate assays, sphere differentiation assay, clone formation, chemotherapy resistance study and tumor formation in an animal model. Our results demonstrated that the tumor sphere cells of HGC-27 cell line could be used to enrich CSCs, which may contribute to human gastric cancer stem cell biology research.

Repression of P66Shc expression by SIRT1 contributes to the prevention of hyperglycemia-induced endothelial dysfunction.

RATIONALE: Inactivation of the p66Shc adaptor protein confers resistance to oxidative stress and protects mice from aging-associated vascular diseases. However, there is limited information about the negative regulating mechanisms of p66Shc expression in the vascular system. OBJECTIVE: In this study, we investigated the role of SIRT1, a class III histone deacetylase, in the regulation of p66Shc expression and hyperglycemia-induced endothelial dysfunction. METHODS AND RESULTS: Expressions of p66Shc gene transcript and protein were significantly increased by different kinds of class III histone deacetylase (sirtuin) inhibitors in human umbilical vein endothelial cells and 293A cells. Adenoviral overexpression of SIRT1 inhibited high-glucose-induced p66Shc upregulation in human umbilical vein endothelial cells. Knockdown of SIRT1 increased p66Shc expression and also increased the expression levels of plasminogen activator inhibitor-1 expression, but decreased manganese superoxide dismutase expression in high-glucose conditions. However, knockdown of p66Shc significantly reversed the effects of SIRT1 knockdown. In addition, p66Shc overexpression significantly decreased manganese superoxide dismutase expression and increased plasminogen activator inhibitor-1 expression in high-glucose conditions, which were recovered by SIRT1 overexpression. Moreover, compared to streptozotocin-induced wild-type diabetic mice, endothelium-specific SIRT1 transgenic diabetic mice had decreased p66Shc expression at both the mRNA and the protein levels, improved endothelial function, and reduced accumulation of nitrotyrosine and 8-OHdG (markers of oxidative stress). We further found that SIRT1 was able to bind to the p66Shc promoter (-508 bp to -250 bp), resulting in a decrease in the acetylation of histone H3 bound to the p66Shc promoter region. CONCLUSION: Our findings indicate that repression of p66Shc expression by SIRT1 contributes to the protection of hyperglycemia-induced endothelial dysfunction.

SIRT1 suppresses activator protein-1 transcriptional activity and cyclooxygenase-2 expression in macrophages.

SIRT1 (Sirtuin type 1), a mammalian orthologue of yeast SIR2 (silent information regulator 2), has been shown to mediate a variety of calorie restriction (CR)-induced physiological events, such as cell fate regulation via deacetylation of the substrate proteins. However, whether SIRT1 deacetylates activator protein-1 (AP-1) to influence its transcriptional activity and target gene expression is still unknown. Here we demonstrate that SIRT1 directly interacts with the basic leucine zipper domains of c-Fos and c-Jun, the major components of AP-1, by which SIRT1 suppressed the transcriptional activity of AP-1. This process requires the deacetylase activity of SIRT1. Notably, SIRT1 reduced the expression of COX-2, a typical AP-1 target gene, and decreased prostaglandin E(2) (PGE(2)) production of peritoneal macrophages (pMPhis). pMPhis with SIRT1 overexpression displayed improved phagocytosis and tumoricidal functions, which are associated with depressed PGE(2). Furthermore, SIRT1 protein level was up-regulated in CR mouse pMPhis, whereas elevated SIRT1 decreased COX-2 expression and improved PGE(2)-related macrophage functions that were reversed following inhibition of SIRT1 deacetylase activity. Thus, our results indicate that SIRT1 may be a mediator of CR-induced macrophage regulation, and its deacetylase activity contributes to the inhibition of AP-1 transcriptional activity and COX-2 expression leading to amelioration of macrophage function.

Human paraoxonase gene cluster transgenic overexpression represses atherogenesis and promotes atherosclerotic plaque stability in ApoE-null mice.

The paraoxonase (PON) gene cluster consists of the PON1, PON2, and PON3 genes, each of which can individually inhibit atherogenesis. To analyze the functions of the PON gene cluster (PC) in atherogenesis and plaque stability, human PC transgenic (Tg) mice were generated using bacterial artificial chromosome. The high-density lipoprotein from Tg mice exhibited increased paraoxonase activity. When crossed to the ApoE-null background and challenged by high-fat diet, PC Tg/ApoE-null mice formed significantly fewer atherosclerotic lesions. However overexpression of the PC transgene had no additive effect on atherosclerosis compared to the overexpression of the single PON1 or PON3 transgene. Plaques from PC Tg/ApoE-null mice exhibited increased levels of collagen and smooth muscle cells, and reduced levels of macrophages and lipid, compared with those from ApoE-null mice, indicating lesions of PC Tg/ApoE-null mice had characteristics of more stable plaques than those of ApoE-null mice. PC transgene enhanced high-density lipoprotein ability to protect low-density lipoprotein against oxidation in vitro. Serum intercellular adhesion molecule-1 and monocyte chemoattractant protein-1 were also repressed by PC transgene. Proatherogenic reactions of Tg mouse peritoneal macrophages induced by oxidized low-density lipoprotein were inhibited by PC transgene, as indicated by reduced reactive oxygen species generation, inflammation, matrix metalloproteinase-9 expression, and foam cell formation. Our results demonstrate that the PC transgene not only represses atherogenesis but also promotes atherosclerotic plaque stability in vivo. PC may therefore be a useful target for atherosclerosis treatment.

Screening and characterization of a bioflocculant produced by Aeromonas sp.

OBJECTIVE: To isolate the bioflocculant-producing bacteria from activated sludge and investigate the flocculating characteristics of the newly isolated bioflocculant. METHODS: Bacteria were screened from activated sludge samples to isolate bioflocculant-producing bacteria. Flocculating activity was used as a measure of the flocculating capability of the bioflocculant. RESULTS: A novel bioflocculant-producing bacterium was isolated, which was identified to belong to genus Aeromonas and named as Aeromonas sp. N11. Flocculating activity increased in the presence of K+, Na+, or Ca2+. The highest flocculating activities for kaolin suspension were obtained in acidic pH ranges, and optimum pHs for it were 3.0, 4.0, and 5.0 with 1 mmol/L K+, Ca+, and Na+ present, respectively. The highest flocculating activities for soil suspension were observed at pH 8.0. The bioflocculant had a good flocculating activity and could achieve a flocculating activity of 92.4% for kaolin suspension at a dosage of only 1 mgxL(-1), and its activity in kaolin suspension was decreased by only 9.2% after heating at 100 degrees C for 60 min. CONCLUSION: The bioflocculant produced by Aeromonas sp. N11 has strong flocculating activity and high stability, which affords high possibility of its practical use.

[Killing cell by granzyme].

Granzyme is an effector molecule of activated cytotoxic T cells and natural killer cells. It mainly mediates cell apoptosis. Its function could be explained by its molecular characteristics to some extent. Its cytotoxic effect is related to some other factors contributing to apoptosis induction. It deserves studying if perforin mediates entrance of granzyme into cells. As potential substrates of granzyme caspases and their substrates have been paid much attention to.