Tfh-cell-derived interleukin 21 sustains effector CD8+ T cell responses during chronic viral infection

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一例X-连锁高IgM综合征合并进行性多灶性脑白质病的基因变异分析

目的对1例临床拟诊为X-连锁高IgM综合征(X-linked hyper-IgM syndrome,XHIM)并发进行性多灶性脑白质病变(progressive multifocal leukoencephalopathy,PML)的患儿CD40L基因及人类嗜神经多瘤病毒(Jamestown Canyon virus,JCV)进行检测,明确致病原因,为临床诊断提供依据。方法采集患儿及父母外周血提取基因组NDA,设计扩增CD40L基因5个外显子及外显子-内含子连接区的特异性引物并进行PCR扩增,产物测序结果与GenBank中CD40L基因序列分析对比确定有无变异。用两对JCV特异性引物对患儿外周血DNA行巢式PCR扩增,目的条带PCR产物测序结果与GenBank中JCV序列对比确定患儿是否存在JCV感染。结果测序结果显示患儿为CD40L c.506 A>C(p.Tyr169Ser)错义变异半合子,该变异位于CD40L肿瘤坏死因子同源区结构域,引起CD40L蛋白疏水作用及结构稳定性丧失,经PolyPhen2及SIFT蛋白功能预测软件预测分别为很可能有害变异(probably damaging,score=1.00)及有害变异(deleterious,score=-8.868),该变异尚未见文献报道。患儿母亲携带CD40L c.506 A>C(p.Tyr169Ser)半合子变异,父亲未检测到该变异。患儿外周血DNA巢式PCR产物经凝胶电泳后发现JCV目的条带,测序结果与GenBank中JCV基因序列比对同源性达到99%,表明患儿外周血DNA中含有JCV,有JCV感染。结论CD40L基因分析及JCV检测结果确诊患儿为XHIGM,其并发的PML与JCV感染有关。     

Cryo-thermal therapy inducing MI macrophage polarization created CXCL10 and IL-6-rich pro-inflammatory environment for CD4+ T cell-mediated anti-tumor immunity

Purpose: We previously developed a novel cryo-thermal therapy to treat malignant mammary carcinoma and melanoma in a mouse model; long-term survival and CD4⁺ T cell orchestrating anti-tumor immune memory response were achieved. Moreover, cryo-thermal-induced CD4⁺ T cell differentiation into Th1 and CD4⁺CTL sub-lineages, in which M1 macrophage polarization played a direct, important role. In particular, cryo-thermal therapy triggered M1 macrophage polarization with up-regulated expression of C–X–C motif ligand 10 (CXCL10) and Interleukin 6 (IL-6). But whether CXCL10 and IL-6 contribute to CD4⁺ T cell-mediated anti-tumor immunity remains unclear. In this study, the role of cryo-thermal-induced CXCL10 and IL-6 in anti-tumor immunity was determined.

Methods: The level of CXCL10 and IL-6 in spleen and serum was determined by RT-PCR and ELISA on day 14 after cryo-thermal therapy. Splenic dendritic cells (DCs) and macrophages were isolated from cryo-thermal-treated mice on day 5 and 14, and the level of CXCL10 and IL-6 in macrophages and DCs was determined by ELISA. The transwell migration assay was performed to study immune cell migration. In vivo neutralization of CXCL10 or IL-6 was performed to investigate the phenotypic changes of immune cells.

Results: Cryo-thermal therapy induced M1 macrophage polarization with up-regulation of CXCL10 and IL-6 expression in spleen. CXCL10 and IL-6 promoted DCs migration and maturation, and subsequently promoted CD4⁺ T cell migration and differentiation into Th1 and CD4⁺ CTL, moreover, reduced myeloid-derived suppressor cells (MDSCs) accumulation.

Conclusions: Cryo-thermal-induced CXCL10 and IL-6 created acute inflammatory environment to initiate a systemically cascading innate and adaptive anti-tumor immunity, which was more permissive for tumor eradication.     

Analysis of the role of palmitoleic acid in acute anterior uveitis

PurposeTo study the role of palmitoleic acid (PA) in the pathogenesis of acute anterior uveitis (AAU).MethodsPA levels in feces from AAU patients were measured by gas chromatography coupled with a mass spectrometer (GC-MS) and compared with samples obtained from healthy individuals. Enzyme linked immunosorbent assay (ELISA) and flow cytometry (FCM) were used to assess the effect of PA on dendritic cells (DCs) and CD4⁺T cells obtained from mice, AAU patients and healthy individuals. C57BL/6 mice were fed with PA or vehicle and experimental autoimmune uveitis (EAU) was induced with a human retinal IRBP651-670 peptide. Disease severity of EAU was evaluated by clinical manifestation and histology. Differentiation of splenic Type 1 helper T cells (Th1) and Th17 cells was evaluated by FCM. Tandem mass tag (TMT)-based proteomics analysis was used to identify differentially expressed proteins following incubation of DCs with PA.ResultsThe fecal concentration of PA was increased in AAU patients as compared with healthy individuals. In vitro, PA promoted apoptosis of DCs and inhibited the secretion of TNF-α from mouse bone-marrow-derived dendritic cells (BMDCs) as well as in DCs from AAU patients and healthy individuals. It only decreased DCs surface marker expression and IL-12p70 secretion in BMDCs and healthy individuals DCs but not in AAU patient DCs. PA-treated BMDCs inhibited Th cell differentiation from mouse naïve CD4⁺T cells and IL-17 and IFN-γ secretion in co-culture supernatants. PA also inhibited the differentiation of Th cells and secretion of IFN-γ and IL-17 in CD4⁺T cells from mice, AAU patients and healthy individuals. In vivo, PA-treated EAU mice showed milder clinical and histopathological intraocular manifestations as compared with the control group. PA feeding inhibited differentiation of splenic Th17 cells, whereas Th1 cells were not affected. Up to 30 upregulated and 77 downregulated proteins were identified when comparing PA-treated DCs with controls.ConclusionAn increased expression of fecal PA was observed in AAU patients. PA was shown to have immunoregulatory effects on DCs and CD4⁺T cells and attenuated disease severity in EAU mice.     

DARTS,Bites and ADCs for the Treatment of AML and for Conditioning for Allogeneic Stem Cell Transplantation

There are over 50 bispecific antibodies that are currently being tested for the treatment of patients with acute leukemias and other hematologic malignancies. In addition to bispecific therapies both chimeric antigen receptor T (CART) cells and antibody drug conjugates (ADC) are being tested for both the treatment of acute leukemias and for conditioning for allogeneic stem cell transplantation. These are being developed to replace chemotherapy and radiation therapy in conditioning regimens to     

circZBTB44通过调控AKT通路促进肾透明细胞癌增殖和迁移的研究

目的 探究circZBTB44在肾透明细胞癌中的表达情况及探讨circZBTB44在肾透明细胞癌中的生物学功能及促进肾透明细胞癌细胞进展的可能机制。方法 通过qRT-PCR法检测circZBTB44在我院21对肾透明细胞癌组织及细胞系786-O、ACHN中的表达水平;通过放线菌素D实验、RNase R实验、核浆分离实验鉴定circZBTB44的特征;通过siRNA下调circZBTB44的表达后,通过MTS细胞增殖实验、克隆形成实验及Transwell迁移实验来探究circZBTB44对肾透明细胞癌786-O、ACHN的生物学功能影响;通过蛋白免疫印迹实验来探究circZBTB44调控的下游通路。结果 circZBTB44在肾透明细胞癌组织和细胞株786-O和ACHN中明显高表达。下调circZBTB44的表达可明显抑制786-O和ACHN的增殖和迁移能力。下调circZBTB44可抑制肾透明细胞癌786-O和ACHN细胞中AKT的磷酸化。结论 circZBTB44在肾透明细胞癌中高表达。circZBTB44可能通过调控AKT信号通路促进肾透明细胞癌的增殖和迁移。