IntroductionFollicular helper T (TFH) cells are essential for germinal center reactions and the maintenance of long-lived humoral immunity. Transforming growth factor-β (TGF-β) is a multifunctional cytokine implicated in immune regulation, T-cell differentiation, and the maintenance of cellular stemness. Prior studies have shown that TGF-β promotes stemness across a wide range of cell types and facilitates the differentiation of naïve CD4⁺ T cells into various T helper cell subsets. However, its precise effects on TFH effector function and stem-like properties remain poorly understood.MethodsThe dual regulatory roles of TGF-β1 in modulating TFH effector functions and stem-like properties were investigated using flow cytometry-based phenotyping, co-culture assays with memory B cells, proliferation and apoptosis assays, ELISA for antibody production, and bulk RNA sequencing of naïve-derived and blood-derived TFH cells.ResultsWe found that TGF-β1 treatment in vitro promoted human naïve CD4+ T cells differentiation into CXCR3+ TFH, but significantly attenuated their effector molecule expression and TFH-mediated memory B-cell differentiation and antibody production, whereas it enhanced the expression of stemness-associated molecules in TFH cells both differentiated in vitro from naïve CD4+ T cells and isolated from blood. Notably, TGF-β1 promoted proliferation and reduced apoptosis of naïve-derived TFH cells in vitro, but suppressed proliferation and increased early apoptosis in blood-derived mature TFH cells.DiscussionOur findings indicate that TGF-β1 tunes the balance between TFH effector function and stem-like properties, and show differential regulations of the early phase of TFH differentiation and mature TFH cells, which may have implications for TFH-driven immune pathology and disease.

Coronavirus disease 2019 (COVID-19) is caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1–3 and individuals with COVID-19 have symptoms that can be asymptomatic, mild, moderate or severe4,5. In the early phase of infection, T- and B-cell counts are substantially decreased6,7; however, IgM8–11 and IgG12–14 are detectable within 14 d after symptom onset. In COVID-19-convalescent individuals, spike-specific neutralizing antibodies are variable3,15,16. No specific drug or vaccine is available for COVID-19 at the time of writing; however, patients benefit from treatment with serum from COVID-19-convalescent individuals17,18. Nevertheless, antibody responses and cross-reactivity with other coronaviruses in COVID-19-convalescent individuals are largely unknown. Here, we show that the majority of COVID-19-convalescent individuals maintained SARS-CoV-2 spike S1- and S2-specific antibodies with neutralizing activity against the SARS-CoV-2 pseudotyped virus, and that some of the antibodies cross-neutralized SARS-CoV, Middle East respiratory syndrome coronavirus or both pseudotyped viruses. Convalescent individuals who experienced severe COVID-19 showed higher neutralizing antibody titres, a faster increase in lymphocyte counts and a higher frequency of CXCR3+ T follicular help (TFH) cells compared with COVID-19-convalescent individuals who experienced non-severe disease. Circulating TFH cells were spike specific and functional, and the frequencies of CXCR3+ TFH cells were positively associated with neutralizing antibody titres in COVID-19-convalescent individuals. No individuals had detectable autoantibodies. These findings provide insights into neutralizing antibody responses in COVID-19-convalescent individuals and facilitate the treatment and vaccine development for SARS-CoV-2 infection. COVID-19-convalescent individuals maintain a strong neutralizing antibody response to SARS-CoV-2 that has cross-reactivity to SARS-CoV and MERS-CoV. Neutralizing antibody titres depend on the severity of the disease and are positively correlated with the frequency of CXCR3+ T follicular helper cells and lymphocyte counts.

Summary Seroconversion appeared early after COVID-19 onset, and convalescent sera therapy benefit some critical patients. However, neutralizing antibody (nAb) in convalescents is largely unknown. We found that 97.01% (65/67) of COVID-19 convalescents maintained IgG antibodies with high binding and avidity to SARS-CoV-2 spike subunits S1 and S2, and 95.52% (64/67) had neutralization activity against SARS-CoV-2 pesudovirus, one month after discharge (median ID 50 , 2.75; IQR, 2.34-3.08). Some sera exhibited cross-neutralization against SARS-CoV (76.12%), MERS-CoV (17.91%), or both (10.45%). Interestingly, individuals recovered from severe disease (severe group) had nAbs with binding and neutralization titers higher than non-severe group. Severe group appeared a rapid increase of lymphocytes and a high proportion of circulating CXCR3 + Tfh cells. Interestingly, the later were spike-specific and positively correlated with SARS-CoV-2 nAb titers. All subjects had no autoimmunity. Our findings provide novel insights into nAb responses in COVID-19 convalescents and facilitate treatment and vaccine development for SARS-CoV-2 infection.

Hepatitis C virus (HCV) and hepatitis B virus (HBV) coinfection reciprocally influences viral replication and host defence responses. This study aimed to investigate the impact of HBV coinfection on circulating T follicular helper cell (cTfh) distribution and the HCV neutralizing antibody (nAb) response. HCV neutralizing antibody responses were measured in individuals with HCV monoinfection (n = 83) and HBV/HCV coinfection (n = 78) using the HCV pseudoparticle neutralization assay. The frequencies of cTfh cells and their subsets in HCV monoinfection (n = 34) and HBV/HCV coinfection (n = 30) were analysed by flow cytometry. The correlations of clinical parameters, cTfh cells and neutralizing antibody responses were analysed. Compared with HCV monoinfection, the HBV coinfection group showed significantly lower HCV neutralizing antibody responses (P < 0.001) and a decreased frequency of circulating Th1‐like Tfh cells (Tfh1) (P = 0.004). In HCV monoinfection, the frequency of the Tfh1 subset was positively correlated with HCV neutralizing antibody responses (R = 0.378, P = 0.03), but this correlation was lost under HBV/HCV coinfection (R = 0.115, P = 0.551). In contrast, the frequency of circulating Th2‐like Tfh cells (Tfh2) was negatively correlated with the HCV neutralizing antibody responses (R = 0.404, P = 0.003). Further analysis showed that HBV coinfection enhanced the Tfh2 subset composition within cTfh cells (P < 0.001), which was associated with serum HBsAg in HBV/HCV coinfection (R = 0.521, P = 0.003). As expected, HBsAg also exhibited an inverse association with HCV neutralizing antibody responses in HBV/HCV coinfection (R = 0.59, P < 0.001). In contrast to HCV monoinfection, HBV/HCV coinfection leads to altered cTfh cell distribution and impaired HCV neutralizing antibody responses, which are associated with HBsAg. These findings will be helpful for better understanding the immunopathogenesis of HBV/HCV coinfection.

Circulating T follicular helper (cTfh) cells have been identified as counterparts of germinal center Tfh (GC Tfh) cells in humans and can support T-dependent B cell maturation and antibody production in vitro . However, the role of cTfh cells in neutralizing antibody (nAb) responses in HCV infection remains unclear. Here, we characterized the phenotype and function of cTfh cells and demonstrated the associations of cTfh cells and their subsets with nAb responses in HCV infection. A total of 38 HCV-infected individuals and 28 healthy controls were enrolled from a pool of injection drug users. The frequency and function of blood Tfh cells were analyzed by flow cytometry. The titers and breadths of serum nAbs were measured using HCV pseudo-particle neutralization assays. Herein, we report several key observations. First, HCV infection skewed cTfh toward CXCR3 + cTfh cell differentiation. Second, the frequency of CXCR3 + cTfh cells positively correlated with HCV nAb titers and breadths. Third, CXCR3 + cTfh cells showed higher expression of Tfh-associated molecules (PD-1, ICOS, IL-21, Bcl-6) compared with CXCR3 − cTfh cells from individuals with HCV infection. Coculture of cTfh cells and autologous memory B cells in vitro indicated that CXCR3 + cTfh cells show a superior ability to support HCV E2-specific B cell expansion compared with CXCR3 − cTfh cells from individuals with HCV infection. HCV infection skews cTfh cells toward CXCR3-biased Tfh cell differentiation, which positively correlates with the magnitude and breadth of the HCV nAb response. It is our hope that these findings will provide insights for the rational design of a nAb-based HCV vaccine.

目的:糖皮质激素临床应用广泛,其免疫作用机制尚未完全阐明,本研究旨在分析糖皮质激素体外短期处理对CD8+T细胞活化及其功能的影响.方法:收集32例健康志愿者外周血单个核细胞,同一样本分为地塞米松处理组及对照组,体外处理3天后流式细胞仪检测:CD8+T细胞总数及其亚群的分布、活化、凋亡;CD8+T细胞功能及其关键转录因子的表达情况.结果:地塞米松处理后与对照组相比:① 地塞米松短期处理后CD8+T细胞比例减少且呈剂量依赖性,主要表现为效应性T细胞减少(P<0.001);② 地塞米松抑制CD8+T细胞活化(P=0.044),促进亚群效应性T细胞晚期凋亡(P=0.032);③ 地塞米松上调抑制性受体:程序性死亡受体-1(PD-1)、T细胞免疫球蛋白粘蛋白分子-3(Tim-3)的表达(P=0.009;P=0.023);④ 地塞米松抑制CD8+T细胞穿孔素(Perforin)、颗粒酶B(Granzyme B)等功能性分子及胞内细胞因子γ干扰素(IFN-γ)、肿瘤坏死因子α(TNF-α)的分泌(P=0.008;P=0.001;P=0.05;P=0.022).⑤ 地塞米松处理后,CD8+T细胞转录因子脱中胚蛋白(Eomes)表达下降,且Eomes与T盒转录因子(T-bet)比值降低(P<0.001;P=0.005).结论:地塞米松体外短期处理外周血单个核细胞可显著抑制效应性CD8+T细胞活化与分化并促进其凋亡.功能上通过抑制转录因子Eomes表达,影响CD8+T细胞功能并促进其耗竭.