Grape Seed Extract Attenuates Demyelination in Experimental Autoimmune Encephalomyelitis Mice by Inhibiting Inflammatory Response of Immune Cells / 中国结合医学杂志

OBJECTIVE@#To examine the anti-inflammatory effect of grape seed extract (GSE) in animal and cellular models and explore its mechanism of action.@*METHODS@#This study determined the inhibitory effect of GSE on macrophage inflammation and Th1 and Th17 polarization in vitro. Based on the in vitro results, the effects and mechanisms of GSE on multiple sclerosis (MS)-experimental autoimmune encephalomyelitis (EAE) mice model were further explored. The C57BL/6 mice were intragastrically administered with 50 mg/kg of GSE once a day from the 3rd day to the 27th day after immunization. The activation of microglia, the polarization of Th1 and Th17 and the inflammatory factors such as tumor necrosis factor- α (TNF- α), interleukin-1 β (IL-1 β), IL-6, IL-12, IL-17 and interferon-γ (IFN-γ) secreted by them were detected in vitro and in vivo by flow cytometry, enzyme linked immunosorbent assay (ELISA), immunofluorescence staining and Western blot, respectively.@*RESULTS@#GSE reduced the secretion of TNF-α, IL-1 β and IL-6 in bone marrow-derived macrophages stimulated by lipopolysaccharide (P<0.01), inhibited the secretion of TNF-α, IL-1 β, IL-6, IL-12, IL-17 and IFN-γ in spleen cells of EAE mice immunized for 9 days (P<0.05 or P<0.01), and reduced the differentiation of Th1 and Th17 mediated by CD3 and CD28 factors (P<0.01). GSE significantly improved the clinical symptoms of EAE mice, and inhibited spinal cord demyelination and inflammatory cell infiltration. Peripherally, GSE downregulated the expression of toll-like-receptor 4 (TLR4) and Rho-associated kinase (ROCKII, P<0.05 or P<0.01), and inhibited the secretion of inflammatory factors (P<0.01 or P<0.05). In the central nervous system, GSE inhibited the infiltration of CD45+CD11b+ and CD45+CD4+ cells, and weakened the differentiation of Th1 and Th17 (P<0.05). Moreover, it reduced the secretion of inflammatory factors (P<0.01), and prevented the activation of microglia (P<0.05).@*CONCLUSION@#GSE had a beneficial effect on the pathogenesis and progression of EAE by inhibiting inflammatory response as a potential drug and strategy for the treatment of MS.

Neuroprotective Mechanism of Buyang Huanwu Tang on Experimental Autoimmune Encephalomyelitis Mice / 中国实验方剂学杂志

Objective:To explore the neuroprotective effect and mechanism of Buyang Huanwu Tang (BYHWT) on experimental autoimmune encephalomyelitis (EAE) at different stages. Method:The 36 female C57BL/6 mice were immunized subcutaneously with myelin oligodendrocyte glycoprotein peptides (MOG35-55),then randomly divided into 9, 17, 28 d EAE control group. Each BYHWT group was orally given drugs on the 3rd day after immunization (50 g·kg-1·d-1), and EAE control group was given the same volume of normal saline in the same way once a day for 9, 17 and 28 d after immunization. The effect of BYHWT on EAE mice was observed with internationally accepted clinical score. Brain and spinal cord specimens were collected at 9, 17 and 28 d after immunization. The neuroprotective effect of BYHWT was observed by hematoxylin-eosin(HE)staining and solid blue staining (LFB). The expressions of BDNF and GAP-43 in spinal cord and brain were detected by Western blot. Result:After treatment, BYHWT can significantly inhibit myelitis cell infiltration and alleviate myelin loss. Compared with EAE group, the expression of Nogo-A in the spinal cord of each BYHWT group was significantly down-regulated (PPPPConclusion:BYHWT can improve the local nerve growth microenvironment and promote the expression of NTFs, reduce the expressions of neuroinhibitory factors, and play a role in neuroprotection.

The effects of catechin on inflammatory response induced by lipopolysaccharide in BV-2 cells / 药学学报

The effects of catechin on inflammatory response of BV-2 cells were investigated using the lipopolysaccharide (LPS) model. BV-2 cells were incubated with LPS (1 mg·L-1) for 12 h in the microglia inflammatory model in vitro. After catechin and LPS co-incubation for 12 h, MTT, ELISA and Western blot were used to detect cell viability, cytokines, cell migration and protein expression. In addition, transwell assay was conducted to investigate the effect of catechin on cell chemokaxis. Catechin did not show any cytotoxicity effect on BV-2 cells, but reversed the change in cell morphology and inhibited the release of TNF-α and IL-1β, cell chemotaxis and phosphorylation of NF-κB/p65. In conclusion, Catechin could inhibit the LPS-induced inflammatory response in BV-2 cells.

A Novel Missense Mutation in Peripheral Myelin Protein-22 Causes Charcot-Marie-Tooth Disease / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Charcot-Marie-Tooth disease (CMT) is the most common inherited peripheral neuropathy. A great number of causative genes have been described in CMT, and among them, the heterozygous duplication of peripheral myelin protein-22 (PMP22) is the major cause. Although the missense mutation in PMP22 is rarely reported, it has been demonstrated to be associated with CMT. This study described a novel missense mutation of PMP22 in a Chinese family with CMT phenotype.</p><p><b>METHODS</b>Targeted next-generation sequencing (NGS) was used to screen the causative genes in a family featured with an autosomal dominant demyelinating form of CMT. The potential variants identified by targeted NGS were verified by Sanger sequencing and classified according to the American College of Medical Genetics and Genomics standards and guidelines. Further cell transfection studies were performed to characterize the function of the novel variant.</p><p><b>RESULTS</b>Using targeted NGS, a novel heterozygous missense variant in PMP22 (c.320G>A, p.G107D) was identified. In vitro cell functional studies revealed that mutant PMP22 protein carrying p.G107D mutation lost the ability to reach the plasma membrane, was mainly retained in the endoplasmic reticulum, and induced cell apoptosis.</p><p><b>CONCLUSIONS</b>This study supported the notion that missense mutations in PMP22 give rise to a CMT phenotype, possibly through a toxic gain-of-function mechanism.</p>

Influence of gamma irradiation on phenotype and function of human dendritic cells in vitro / 中国实验血液学杂志

To determine whether gamma irradiation influences phenotype and function of human dendritic cells (DC) in vitro, dendritic cells were induced from the peripheral blood mononuclear cells of multiple sclerosis patients with RPMI 1640 medium containing recombinant human GM-CSF (rhGM-CSF, 800 U/ml) and recombinant human IL-4 (rhIL-4, 500 U/ml). Phenotypic changes were monitored by light microscopy. Lipopolysaccharide at a concentration of 5 micro g/ml was added into the cultures after 6 days of growth for DC complete maturation, and the cells were cultured for another 24 hours. The harvested DC on day 7 were divided equally into several parts. One part was used as non-irradiated DC (naive DC) while the other parts were irradiated by gamma ray at a dose of 25 Gy and 30 Gy respectively. Cell surface molecules were analyzed by flow cytometry. The capability of DC to stimulated autologous T cell proliferation were determined. The results showed that gamma irradiation reduced expression of CD86, CD80 and HLA-DR molecules on dendritic cells, especially CD86 molecules. Dendritic cells effectively stimulated autologous T cells proliferation while irradiated DC in all groups showed profound decrease of capability to promote T cells proliferation. It is concluded that gamma irradiation of dendritic cells not only influenced phenotype of DC but also altered their function as stimulator cells in mixed lymphocyte reaction.

Modulation effects of human immature and mature dendritic cells on glatiramer acetate specific T cell lines in vitro / 中国实验血液学杂志

A large body of evidence demonstrates that dendritic cells (DC) play a pivotal role in the control of immunity by priming and tolerizing T cells. In multiple sclerosis (MS), autoreactive T cells are proposed to play a pathogenic role by secreting pro-inflammatory cytokines, but comparison studies on the effects of immature and mature dendritic cells on the cytokines profile of antigen-specific T cell lines are lacking. To evaluate the actions of dendritic cell maturation on T cell polarization, the effects of immature and mature dendritic cells derived from MS patients on in vitro proliferative responses, and cytokine production by glatiramer acetate (GA)- specific T cell lines (TCL) derived from MS patients were analyzed. The results demonstrated that it is easy to derive GA-specific TCL from MS patients with high specificity; lipopolysaccharide can efficiently induce DC maturation within 24 hours at a concentration of 5 micro g/ml; mature DC showed higher co-stimulatory capacity of GA-specific TCLs than immature DC. GA-specific TCLs produce dominantly IL-2, IL-4, IFN-gamma and IL-10, but low levels of IL-6. In contrast to immature DC, mature DC enhanced capacity to induce IL-6 and IL-10 secretion, but down-regulate IL-2, IL-4 and IFN-gamma production by GA- specific TCLs. It is concluded that DC maturation status modulating proliferation of TCL and production of cytokines may represent another focus for the study on both immuno-pathogenesis and immunotherapeutic interventions in MS.