Peripheral CD4^(+)CD8^(+) double positive T cells:A potential marker to evaluate renal impairment susceptibility during systemic lupus erythematosus

Lupus nephritis(LN) has a high incidence in systemic lupus erythematosus(SLE) patients, but there is a lack of sensitive predictive markers. The purpose of the study was to investigate the association between the CD4^(+)CD8^(+)double positive T(DPT) lymphocytes and LN. The study included patients with SLE without renal impairment(SLE-NRI), LN, nephritic syndrome(NS), or nephritis. Peripheral blood lymphocyte subsets were analyzed by flow cytometry. Biochemical measurements were performed with peripheral blood in accordance with the recommendations proposed by the National Center for Clinical Laboratories. The proportions of DPT cells in the LN group were significantly higher than that in the SLE-NRI group(t=4.012, P<0.001), NS group(t=3.240,P=0.001), and nephritis group(t=2.57, P=0.011). In the LN group, the risk of renal impairment increased significantly in a DPT cells proportion-dependent manner. The risk of LN was 5.136 times(95% confidence interval, 2.115–12.473) higher in cases with a high proportion of DPT cells than those whose proportion of DPT cells within the normal range. These findings indicated that the proportion of DPT cells could be a potential marker to evaluate LN susceptibility, and the interference of NS and nephritis could be effectively excluded when assessing the risk of renal impairment during SLE with DPT cell proportion.

CD4^+ T cell-mediated presentation of non-infectious HIV-1 virion antigens to HIV-specific CD8^+ T cells

Background The mechanism of chronic immune activation and impairment of HIV-specific immune responses during chronic infection is not fully understood. However, it is known that high immune activation leads to more rapid progression to AIDS. We hypothesize that CD4^＋ T cell-mediated viral antigen presentation contributes to this pathologic immune activation in HIV-infected individuals. Methods HIV-specific T cells, responding to noninfectious HIV-1 virions as antigen, were measured by flow cytometric assays. These experimental conditions reflect the in vivo condition where noninfectious HIV-1 represents more than 99% of the antigens. Results CD4^＋ T cells purified from HIV-infected individuals were capable of cross presenting exogenous noninfectious HIV-1 virions to HIV-1-specific CD8^＋ T cells. Cross presentation required the entry of HIV-1 to CD4^＋ T cells and antigen translocation from endoplasmic reticulum to the Golgi complex. Blocking CD4^＋ mediated activation of HIV-specific CD8^＋ T cells and redirecting the viral antigens to antigen presenting cells improved HIV-specific T cell responses. Contusions One possible cause of chronic immune activation and impairment of HIV-1 specific T cell responses is represented by HIV-1 harboring CD4^＋ T cells cross presenting HIV-1 antigen to activate CD8^＋ T cells. This new mechanism provides the first evidence that cross presentation of noninfectious HIV-1 virions play a role in the immunopathogenesis of HIV-1 infection.

Conversion of effector CD4^(+)T cells to a CD8^(+)MHC Ⅱ-recognizing lineage

CD4^(+)and CD8^(+)T cells are dichotomous lineages in adaptive immunity.While conventionally viewed as distinct fates that are fixed after thymic development,accumulating evidence indicates that these two populations can exhibit significant lineage plasticity,particularly upon TCR-mediated activation.We define a novel CD4^(-)CD8αβ^(+)MHC Ⅱ-recognizing population generated by lineage conversion from effector CD4^(+)T cells.CD4-CD8αβ^(+)effector T cells downregulated the expression of T helper cell-associated costimulatory molecules and inaeased the expression of cytotoxic T lymphocyte-associated cytotoxic molecules.This shift in functional potential corresponded with a CD8^(+)-lineage skewed transcriptional profile.TCRβ repertoire sequencing and in vivo genetic lineage tracing in acutely infected wild-type mice demonstrated that CD4^(-)CD8αβ^(+)effector T cells arise from fundamental lineage reprogramming of bona fide effector CD4^(+)T cells.Impairing autophagy via functional deletion of the initiating kinase Vps34 or the downstream enzyme Atg7 enhanced the generation of this cell population.These findings suggest that effector CD4^(+)T cells can exhibit a previously unreported degree of skewing towards the CD8^(+)T cell lineage,which may point towards a novel direction for HIV vaccine design.

4-1BB signaling activates glucose and fatty acid metabolism to enhance CD8^(+) T cell proliferation

4-1BB(CD137)is a strong enhancer of the proliferation of CD8^(+)T cells.Since these cells require increased production of energy and biomass to support their proliferation,we hypothesized that 4-1BB signaling activated glucose and fatty acid metabolism.We found that treatment with agonistic anti-4-1BB mAb promoted the proliferation of CD8^(+)T cells in vitro,increasing their size and granularity.Studies with a glycolysis inhibitor and a fatty acid oxidation inhibitor revealed that CD8^(+)T cell proliferation required both glucose and fatty acid metabolism.Anti-4-1BB treatment increased glucose transporter 1 expression and activated the liver kinase B1(LKB1)-AMP-activated protein kinase(AMPK)-acetyl-CoA carboxylase(ACC)signaling pathway,which may be responsible for activating the metabolism of glucose and fatty acids.We also examined whether blocking glucose or fatty acid metabolism affected cell cycle progression and the anti-apoptotic effect of 4-1BB signaling.The increase of anti-apoptotic factors and cyclins in response to anti-4-1BB treatment was completely prevented by treating CD8^(+)T cells with the fatty acid oxidation inhibitor,etomoxir,but not with the glycolysis inhibitor,2-deoxy-D-glucose.We conclude that anti-4-1BB treatment activates glucose and fatty acid metabolism thus supporting the increased demand for energy and biomass,and that fatty acid metabolism plays a crucial role in enhancing the cell cycle progression of anti-CD3-activated CD8^(+)T cells in vitro and the anti-apoptotic effects of 4-1BB signaling on these cells.

The OX40-TRAF6 axis promotes CTLA-4 degradation to augment antitumor CD8^(+)T-cell immunity

Immune checkpoint blockade(ICB),including anti-cytotoxic T-lymphocyte associated protein 4(CTLA-4),benefits only a limited number of patients with cancer.Understanding the in-depth regulatory mechanism of CTLA-4 protein stability and its functional significance may help identify ICB resistance mechanisms and assist in the development of novel immunotherapeutic modalities to improve ICB efficacy.Here,we identified that TNF receptor-associated factor 6(TRAF6)mediates Lys63-linked ubiquitination and subsequent lysosomal degradation of CTLA-4.Moreover,by using TRAF6-deficient mice and retroviral overexpression experiments,we demonstrated that TRAF6 promotes CTLA-4 degradation in a T-cell-intrinsic manner,which is dependent on the RING domain of TRAF6.This intrinsic regulatory mechanism contributes to CD8+T-cell-mediated antitumor immunity in vivo.Additionally,by using an OX40 agonist,we demonstrated that the OX40-TRAF6 axis is responsible for CTLA-4 degradation,thereby controlling antitumor immunity in both tumor-bearing mice and patients with cancer.Overall,our findings demonstrate that the OX40-TRAF6 axis promotes CTLA-4 degradation and is a potential therapeutic target for the improvement of T-cell-based immunotherapies.

Tcf1 at the crossroads of CD4^＋ and CD8^＋ T cell identity

Transcription factors and DNA/histone modification enzymes work in concert to establish and maintain cell identity. CD4^＋ and CD8^＋ T cells are key players in cellular immunity with distinct functions. Recent studies offer novel insights into how their identities are established in the thymus and maintained in the periphery during immune responses. During thymic maturation, Thpok, HDAC1 and HDAC2 guard CD4^＋ T cells from activation of CD8^＋ cytotoxic genes, and Tcfl and Left utilize their intrinsic HDAC activity to shut down CD4^＋ lineage-associated genes in CD8^＋ T cells. In activated CD4＋ T cells, Tcfl and Left act upstream of the Bc16-Blimpl axis to direct differentiation of follicular helper T （Tfh） cells, and prevent diversion of Tfh to IL-17-producing cells. In parallel, T-bet, together with Eomes or Blimpl, ensures proper induction of the cytotoxic program in CD8^＋ effectors elicited by acute infection, and prevents generation of pathogenic, IL-17-producing CD8^＋ effector T cells. Antigen persistence due to chronic viral infection leads to CD8^＋ T cell exhaustion. A portion of exhausted CD8^＋ T cells has the capacity to activate the Tfh program in a Tcfl-dependent manner. Those Tfh-like CD8^＋ T cells exhibit enhanced proliferative capacity in response to PD-1 blockage therapy and are more effective in curtailing viral replication. Thus, dissecting the molecular aspects of T cell identity, during development and immune responses, may lead to new therapies for treating autoimmunity, tumors, and persistent infections.

Research progress on the role of immune cells in Brucella infection

Brucellosis is one of the most prevalent zoonoses in the world. Incidence of the disease has increased significantly in recent years and has seriously affected the health of human beings and the development of animal husbandry. The pathogenesis of brucellosis remains unclear. Current studies suggest that this disease may be related to changes in natural killer cells, dendritic cells, and macrophages in immune cell subsets. Brucellosis may be also related to T helper(Th) 1 cell/Th2 cell imbalance in the CD4^+ T cell subset, immunoregulation of regulatory T cells and Th17 cells, and the mechanism of action of CD8^+ T cell. This paper aims to review the research progress on these inherent immune cells, the CD4^+ T cell subset, and CD8^+ T cells in Brucella infection.

小鼠乳腺癌4T1细胞中肿瘤干细胞样细胞的富集和鉴定

目的：无血清培养基（serum-free medium,SFM）悬浮培养小鼠乳腺癌细胞株4T1,富集并鉴定4T1细胞株中肿瘤干细胞样细胞。方法：通过含EGF、bFGF和B27等细胞因子的SFM培养富集4T1细胞中肿瘤干细胞样细胞,将其接种于含血清培养基（serum-supplemented medium,SSM）,观察4T1肿瘤干细胞样细胞分化情况。应用细胞表面标志CD44^＋CD24^-/low和Hoechst33342染色法检测4T1细胞中肿瘤干细胞样细胞的比例,小鼠致瘤实验验证不同培养条件下4T1肿瘤干细胞样细胞的致瘤能力。结果：4T1细胞在SFM中能够存活、增殖,并形成细胞球,细胞球可连续传代,若重新接种于SSM中可贴壁分化。4T1细胞球中CD44^＋CD24^-/low细胞比例为6.4%68.9%,侧群（side population,SP）细胞比例为7.3%61.2%,均显著高于SSM中培养的4T1细胞（P〈0.05）;随着SFM中细胞球传代次数增加,CD44^＋CD24^-/low细胞和SP细胞的比例逐渐升高。小鼠致瘤实验结果显示,富集了肿瘤干细胞的细胞球比常规培养4T1细胞的致瘤性更强。结论：乳腺癌4T1细胞可在含多种生长因子的SFM中悬浮生长并形成细胞球,4T1细胞中含有的乳腺癌干细胞样细胞可通过SFM培养法富集。

Efficient control of chronic LCMV infection by a CD4 T cell epitope-based heterologous prime-boost vaccination in a murine model

CD4^(+)T cells are essential for sustaining CD8^(+)T cell responses during a chronic infection.The adoptive transfer of virus-specific CD4^(+)T cells has been shown to efficiently rescue exhausted CD8^(+)T cells.However,the question of whether endogenous virus-specific CD4^(+)T cell responses can be enhanced by certain vaccination strategies and subsequently reinvigorate exhausted CD8^(+)T cells remains unexplored.In this study,we developed a CD4^(+)T cell epitope-based heterologous prime-boost immunization strategy and examined the efficacy of this strategy using a mouse model of chronic lymphocytic choriomeningitis virus(LCMV)infection.We primed chronically LCMV-infected mice with a Listeria monocytogenes vector that expressed the LCMV glycoprotein-specific I-Ab-restricted CD4^(+)T cell epitope GP61–80(LM-GP61)and subsequently boosted the primed mice with an influenza virus A(PR8 strain)vector that expressed the same CD4^(+)T cell epitope(IAV-GP61).This heterologous prime-boost vaccination strategy elicited strong anti-viral CD4^(+)T cell responses,which further improved both the quantity and quality of the virusspecific CD8^(+)T cells and led to better control of the viral loads.The combination of this strategy and the blockade of the programmed cell death-1(PD-1)inhibitory pathway further enhanced the anti-viral CD8^(+)T cell responses and viral clearance.Thus,a heterologous prime-boost immunization that selectively induces virus-specific CD4^(+)T cell responses in conjunction with blockade of the inhibitory pathway may represent a promising therapeutic approach to treating patients with chronic viral infections.

Enhancement of antitumor immunity by low-dose total body irradiation is associated with selectively decreasing the proportion and number of T regulatory cells

Low-dose total body irradiation （LTBI） is used in the treatment of some cancers mainly for immune enhancement rather than cell killing. However, the mechanism underlying LTBI remains unknown. In this study, by analyzing the immune patterns of lymphocytes, we found that the percentage and absolute number of CD4^＋CD25^＋Foxp3^＋ regulatory T cells are markedly decreased in naive mice following treatment with LTBI. On the contrary, the CD4^＋CD44^＋/CD8^＋CD44^＋ effector-memory T cells are greatly increased. Importantly, naive mice treated with dendritic cell-gp100 tumor vaccines under LTBI induced an enhancement of antigen-specific proliferation and cytotoxicity as well as interferon-γ, （IFN-γ） secretion against FIO melanoma tumor challenge, compared to treatment with either the tumor vaccine or LTBI alone. Consequently, the treatment resulted in a reduced tumor burden and prolonged mouse survival. Our data demonstrate that LTBI＇s enhancement of antitumor immunity was mainly associated with selectively decreasing the proportion and number of T regulatory cells, implying the potential application of the combination of LTBI and a tumor vaccine in antitumor therapy.

Cellular metabolism regulates the differentiation and function of T-cell subsets

Tcells are an important component of adaptive immunity and protect the host from infectious diseases and cancers.However,uncontrolled T cell immunity may cause autoimmune disorders.In both situations,antigen-specific T cells undergo clonal expansion upon the engagement and activation of antigens.Cellular metabolism is reprogrammed to meet the increase in bioenergetic and biosynthetic demands associated with effector T cell expansion.Metabolites not only serve as building blocks or energy sources to fuel cell growth and expansion but also regulate a broad spectrum of cellular signals that instruct the differentiation of multiple T cell subsets.The realm of immunometabolism research is undergoing swift advancements.Encapsulating all the recent progress within this concise review in not possible.Instead,our objective is to provide a succinct introduction to this swiftly progressing research,concentrating on the metabolic intricacies of three pivotal nutrient classes—lipids,glucose,and amino acids—in T cells.We shed light on recent investigations elucidating the roles of these three groups of metabolites in mediating the metabolic and immune functions of T cells.Moreover,we delve into the prospect of“editing”metabolic pathways within T cells using pharmacological or genetic approaches,with the aim of synergizing this approach with existing immunotherapies and enhancing the efficacy of antitumor and antiinfection immune responses.

Double negative T cells,a potential biomarker for systemic lupus erythematosus

Systemic lupus erythematosus(SLE)is an autoimmune disease that is a challenge to diagnose and treat.There is an urgent need for biomarkers to help define organ involvement,and more effective therapies.A unique population of T cells,the CD3^(+)CD4^(−)CD8^(−)(DNeg)cells,is significantly increased in lupus patients.Twentyseven cases(53%)of pediatric SLE patients had elevated DNeg cells in their peripheral blood,which correlated with kidney function(R^(2)=0.54).Significant infiltration of DNeg cells was observed in both adult and pediatric lupus kidneys by immunofluorescence.For the first time,this study provides direct evidence that DNeg cells facilitate kidney injury in preclinical 8-week-old MRL/lpr lupus mice.In lupus mice,the increase in DNeg cells tracked with worsening disease and correlated with kidney function(R^(2)=0.85).Our results show that DNeg cells per se can cause kidney dysfunction,increase in number with increase in disease pathology,and could serve as a potential biomarker.

Key immunity characteristics of diverse stages of brucellosis in rural population from Inner Mongolia, China

Background: Brucellosis poses a serious threat to human and animal health,particularly in developing countries such as China.The Inner Mongolia Autonomous Region is one of the most severely brucellosis-endemic provinces in China.Currently,the host immune responses functioning to control Brucella infection and development remain poorly understood.The aim of this study is to further clarify the key immunity characteristics of diverse stages of brucellosis in Inner Mongolia.Methods: We collected a total of 733 blood samples from acute(n=137),chronic(n=316),inapparent(n=35),recovery(n=99),and healthy(n=146)groups from the rural community of Inner Mongolia between 2014 and 2015.The proportions of CD4^(+),CD8^(+),Th1,Th2,and Th17 T cells in peripheral blood and the expression of TLR2 and TLR4 in lymphocytes,monocytes and granulocytes were examined using flow cytometry analysis.The differences among the five groups were compared using one-way ANOVA and the Kruskal–Wallis method,respectively.Results: Our results revealed that the proportions of CD4^(+) and CD8^(+) T cells were significantly different among the acute,chronic,recovery,and healthy control groups(P<0.05),with lower proportions of CD4^(+) T cells and a higher proportion of CD8^(+) T cells in the acute,chronic,and recovery groups.The proportion of Th1 cells in the acute,chronic,and inapparent groups was higher than that in the healthy and recovery groups;however,there was no significant difference between patients and healthy individuals(P>0.05).The proportion of Th2 lymphocytes was significantly higher in the acute and healthy groups than in the inapparent group(P<0.05).The proportion of Th17 cells in the acute group was significantly higher than that in the healthy control,chronic,and inapparent groups(P<0.05).Finally,the highest expression of TLR4 in lymphocytes,monocytes and granulocytes was observed in the recovery group,and this was followed by the acute,chronic,healthy control,and inapparent groups.There was a significant difference between the recovery group and the other groups,except for the acute group(P<0.05).Moreover,a correlation in TLR4 expression was observed in lymphocytes,monocytes and granulocytes among the five groups(r>0.5),except for the inapparent group between lymphocytes and granulocytes(r=0.34).Conclusions: Two key factors(CD8^(+)T cells and TLR4)in human immune profiles may closely correlate with the progression of brucellosis.The detailed function of TLR4 in the context of a greater number of cell types or tissues in human or animal brucellosis and in larger samples should be further explored in the future.