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,单击此处编辑母版标题样式,单击此处编辑母版文本样式,第二级,第三级,第四级,第五级,*,*,Macrophages from patients with atopic dermatitis show a reduced CXCL10 expression in response to staphylococcal a-toxin.,1,CONTENTS,Introduction,Results,Materials and Methods,Discussion,2,Introduction,keywords,present study,purpose,background,3,Keywords,atopic dermatitis; (,特应性皮炎,),chemokines;,（趋化因子）,human macrophages;,（人巨噬细胞）,IP-10/CXCL10;,MDC/CCL22;,psoriasis;,（牛皮癣；银屑病）,Staphylococcus aureus;,（金黄色葡萄球菌）,-toxin.,（,-,毒素）,4,Background:,Patients with atopic dermatitis (AD) and,psoriasis,are frequently colonized with Staphylococcus aureus (S. aureus). one-third of them producing -toxin, which is cor-related with the severity of eczema in AD.,Distinct expression of chemokine (C-C motif) ligand (,配体,CCL,) and chemokine (C-X-C motif) ligand (,配体,CXCL,) chemokines has been documented in both diseases.,the effects of a-toxin on human macrophages in patients with AD and psoriasis?,5,Present study,Recent studies demonstrated that infiltration of inflammatory cells into tissues is regulated by chemokines. Two main subfamilies, recently renamed chemokine (C-C motif) ligand (CCL) and chemokine (C-X-C motif) ligand (CXCL) chemo-kines, have been distinguished,.,Several studies have suggested a crucial involvement of,CCL,chemokines in inflammation.,Chemokines and their receptors are implicated in the development of symptoms of AD and psoriasis .,CXCL10,plays a role in pathogenesis of psoriasis .,6,Purpose,The effects of -toxin on human macrophages still remain unclear.,We show here for the first time that -toxin induces the Th1-related chemokine CXCL10 in human macrophages.,we studied the effects of -toxin on Th1- and Th2-related chemokines in macrophages from patients with AD and psoriasis where the intrinsic abnormal and different chemokines production profile is well defined.,7,Materials and Methods,Materials and Methods,Patients,Cell isolation and culture,Statistical analysis,Cytokine assessment by ELISA,Western blot,8,Methods,macrophage-derived chemokine (MDC)/CCL22,Cell surface markers expression and chemotaxis,IFN-c-induced protein of 10-kDa (IP-10)/CXCL10,qRTPCR or ELISA,flow cytometry,Boyden chamber technique,9,Patients,healthy donors and patients with AD or psoriasis,Cell isolation and culture,Peripheral blood mononuclear cells (PBMCs) were isolated by,Lymphoprep density-gradient centrifugation,from healthy donors and from patients with AD and psoriasis,Chemotaxis assay,The chemotactic activity of supernatants from a-toxin stimulated macrophages on lymphocytes (CD4+ T cells) was determined using a,Boyden chamber technique,10,Western blot,macrophages were prestimulated with a-toxin (100 ng/ml), IFN-c (10 ng/ml), TNF-a (10 ng/ml) or LPS,.,Cell extracts were subjected to Western blot analysis,Cytokine assessment by ELISA,Cell-free culture supernatants were harvested and analysed for CXCL10 or CCL22 , using a commercially available enzyme-linked immunosorbent assay (ELISA) system .,11,Results,One,Induction of IP-10/CXCL10 by staphylococcal a-toxin in human macrophages,Three,Low effect of a-toxin on CXCL10 induction (Th1-related chemokine) in macrophages from patients with AD,Two,a-Toxin-stimulated macrophages could induce the migration of human CD4+ T cells via CXCR3,Four,Effect of a-toxin on MDC production (Th2-related chemokine) in macrophages from patients with chronic inammatory skin diseases,12,Induction of IP-10/CXCL10 by staphylococcal a-toxin in human macrophages,Microarray analysis revealed that CXCL10 was the most strongly up-regulated gene in macrophages.,An increased expression was not observed for other chemokines.,The mean ratio for all these chemokines was between 0.2 and 2 when comparing not stimulated with a-toxin-stimulated macrophages .,13,Figure 1 Induction of chemokine (C-X-C motif) ligand (CXCL)10 following a-toxin stimulation in human macrophages,at the mRNA level.,Figure 2 Induction of chemokine (C-X-C motif) ligand (CXCL)10 following a-toxin stimulation in human macrophages,at the protein level,.,14,Figure 3 Punch biopsies (3 mm) from healthy individuals were left either unstimulated (A) or stimulated with a-toxin (100 ng/ ml) (B) or IFN-c (100 ng/ml) (C) for 24 h at 37C. 5-lm parafn sections were stained for CXCL10 along with appropriate isotype as well as CD68.,15,a-Toxin-stimulated macrophages could induce the migration of human CD4+ T cells via CXCR3,Both inhibitors(anti-IP-10 antibody&anti-CXCR3 antibody) abrogated the migration induced by a-toxin-treated macrophage supernatants .,a-Toxin in cell culture medium without macrophages did not directly induce the migration of human CD4+ lymphocytes.,16,Figure 4 Chemotactic activity of a-toxin-stimulated macrophages and inhibition of chemokine (C-X-C motif) ligand (CXCL)10-induced chemotaxis using a neutralizing anti-CXCL10 or anti-CXCR3 Ab.,17,Low effect of a-toxin on CXCL10 induction (Th1-related chemokine) in macrophages from patients with AD,a-toxin induced signicantly lower levels of CXCL10 expression or secretion in patients with AD compared with healthy controls as well as psoriasis at all time points and doses tested.,18,Figure 5 Macrophages from patients with atopic dermatitis (AD) and psoriasis (PSO) as well as from healthy controls (HC) were left either unstimulated (medium control) or stimulated with a-toxin as indicated or with IFN-c (10 ng/ml) or TNF-a (20 ng/ml), respectively, for the indicated periods of time.,19,Effect of a-toxin on MDC production (Th2-related chemokine) in macrophages from patients with chronic infammatory skin diseases,Macrophages from patients with AD as well as patients with psoriasis showed an intrinsically higher CCL22 production compared to healthy controls by trend. On the whole, a-toxin did not regulate CCL22 secretion.,20,Thank you,21,
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