目的:探讨叶氏狼疮方在诱导狼疮肾炎(lupus nephritis,LN)患者外周血T淋巴细胞免疫耐受中有无作用。方法:密度梯度离心法提取LN患者及健康对照外周血单个核细胞(PBMCs),设立4个培养亚组:空白(blank)、植物血凝素(PHA)、狼疮方血清(lupus...目的:探讨叶氏狼疮方在诱导狼疮肾炎(lupus nephritis,LN)患者外周血T淋巴细胞免疫耐受中有无作用。方法:密度梯度离心法提取LN患者及健康对照外周血单个核细胞(PBMCs),设立4个培养亚组:空白(blank)、植物血凝素(PHA)、狼疮方血清(lupus formula serum,LFS)、PHA+LFS组。双色标记流式细胞术检测PBMCs及其条件培养后CD3+T细胞表面分子CD28,CD40L,LFA-1a和CTLA-4的表达水平。多组间比较采用单因素方差分析。结果:活动性LN组外周血T细胞表面CD28、CD40L、LFA-1a和CTLA-4表达水平较健康对照组明显上调(P均<0.01);PHA可诱导健康对照T细胞表面CD28和CD40L表达明显上调(P均<0.01),但对其CTLA-4和LFA-1a的表达无明显影响(P均>0.05);而PHA对活动性LN组外周血T细胞表面上述指标表达均有明显上调作用(P均<0.05);叶氏狼疮方对健康人外周血T细胞表面上述指标的表达无明显影响,但可逆转活动性LN患者外周血T细胞表面的CD40L、LFA-1a过度表达(P均<0.05),对其CD28和CTLA-4过度表达无明显影响(P均>0.05);且叶氏狼疮方对PHA所诱导的活动性LN患者外周血T细胞表面CD28、CD40L和LFA-1a表达增加亦有明显抑制。结论:叶氏狼疮方可能主要通过下调T细胞CD40L和LFA-1a表达水平来诱导LN患者外周T细胞免疫耐受重建。展开更多
Regulatory T cells （Treg） play important roles in immune system homeostasis, and may also be involved in tumor immunotolerance by suppressing Th1 immune response which is involved in anti-tumor immunity. We have pre...Regulatory T cells （Treg） play important roles in immune system homeostasis, and may also be involved in tumor immunotolerance by suppressing Th1 immune response which is involved in anti-tumor immunity. We have previously reported that immunization with attenuated activated autologous T cells leads to enhanced anti-tumor immunity and upregulated Thl responses in vivo. However, the underlying molecular mechanisms are not well understood. Here we show that Treg function was significantly downregulated in mice that received immunization of attenuated activated autologous T cells. We found that Foxp3 expression decreased in CD4＋CD25＋ T cells from the immunized mice. Moreover, CD4＋CD25＋Foxp3＋ Treg obtained from immunized mice exhibited diminished immunosuppression ability compared to those from naive mice. Further analysis showed that the serum of immunized mice contains a high level ofanti-CD25 antibody （about 30 ng/ml, p〈0.01 vs controls）. Consistent with a role ofanti-CD25 response in the downregulation of Treg, adoptive transfer of serum from immunized mice to naive mice led to a significant decrease in Treg population and function in recipient mice. The triggering of anti-CD25 response in immunized mice can be explained by the fact that CD25 was induced to a high level in the ConA activated autologous T cells used for immunization. Our results demonstrate for the first time that immunization with attenuated activated autologous T cells evokes anti-CD25 antibody production, which leads to impeded CD4＋CD25＋Foxp3＋ Treg expansion and function in vivo. We suggest that dampened Treg function likely contributes to enhanced Thl response in immunized mice and is at least part of the mechanism underlying the boosted anti-tumor immunity.展开更多
Objective： The use of donor-derived immature dendritic cells （imDC） has become a promising approach to induce immune tolerance or immune hyporesponsiveness. However, donor-derived imDC needs to be harvested for a f...Objective： The use of donor-derived immature dendritic cells （imDC） has become a promising approach to induce immune tolerance or immune hyporesponsiveness. However, donor-derived imDC needs to be harvested for a few days and transfused into the recipient in 5-10 days before transplantation, which is practically impossible in a clinical setting where donor organs are mainly harvested from cadavers. Moreover, donor-derived imDC might be cleared by allogeneic reaction offsetting induced immune tolerance or immune hyporesponsiveness. In our study, we further explored the underlying mechanism of immune hyporesponsiveness induced by donor-antigen-unloaded recipient-derived imDC by transfusing these imDC into rats in 1 day before liver transplantation. This paper is to study the mechanism of immune hyporesponsiveness induced by donor-antigen-unloaded recipient-derived imDC and its protection of liver grafts in rats. Methods： 40 SD rats （donor） and 40 male Wistar rats （recipient） were randomly divided into 4 groups： control, cyclosporine A （CsA）, mature DC （mDC）, and imDC; with 10 SD rats and 10 Wistar rats for each group. Animal models of acute graft rejection were established with these rats. Corresponding treatments were given before or after transplantation. In the control group, Wistar rats received no treatment other than liver transplantation. In the CsA group, Wistar rats underwent liver transplantation plus CsA treatment （10 mg/kg·d） in the starting day 2 after transplantation. For the mDC group, recipient-derived mDC （1 × 10^6/rat） were infused intravenously via the dorsal vein of the penis to recipient rats. For the imDC group, imDC （1× 10^6/rat） were injected into recipient rats via the dorsal vein of the penis. In each group, 5 recipients were executed at 10 days after transplantation; the remaining five recipients were kept for the observation of survival time. Blood samples were collected for the measurement of ALT and TBIL; IL-2, IFN-γ, IL-4 and IL-10 and levels were measured with double-antibody sandwich ELISA. Liver tissue was harvested for HE staining and the observation of histological features. Acute rejection was evaluated with Banff classification. Expression levels of Fas-L/Fas in the grafts were detected by iminunohistochemieal staining; and western blot was used to detect the expression level of Scurfin. Results： The median survival times （MST） of the liver allografls in the CsA and imDC group were significantly longer than those in the control or mDC group （P〈0.05）. The serum levels of ALT and TBIL in the control and mDC groups were significantly higher than those of the CsA or imDC group （P〈0.05）. Compared with the CsA anti imDC group, the levels of IL-2 and IFN-γ were higher but the levels of IL-4 and IL-10 were lower than those of the control and mDC groups （P〈0.01）. Slight or no rejection reaction was found in the CsA anti imDC groups （P〈0.05）. The expression level of Scurfin protein in CD4^＋ CD25^＋ T cells of the imDC group was significantly higher than that of three other groups （P〈 0.05）. Conclusion： Donor-antigen-unloaded recipient- derived imDC is an effective treatment in inducing immune hyporesponsiveness by blocking indirect recognition in rat liver transplantation model. Survival span was significantly prolonged by its protective effect. The mechanism of immune hyporesponsiveness induced hy imDC transfusion may involve the preprocesses of T cell apoptosis induction, immune tolerance or hyporesponsiveness in T cells, induction of the shift in TH1/TH2 balance, selection activation of Th2 subset, or induction of regulatory T cell.展开更多
文摘目的:探讨叶氏狼疮方在诱导狼疮肾炎(lupus nephritis,LN)患者外周血T淋巴细胞免疫耐受中有无作用。方法:密度梯度离心法提取LN患者及健康对照外周血单个核细胞(PBMCs),设立4个培养亚组:空白(blank)、植物血凝素(PHA)、狼疮方血清(lupus formula serum,LFS)、PHA+LFS组。双色标记流式细胞术检测PBMCs及其条件培养后CD3+T细胞表面分子CD28,CD40L,LFA-1a和CTLA-4的表达水平。多组间比较采用单因素方差分析。结果:活动性LN组外周血T细胞表面CD28、CD40L、LFA-1a和CTLA-4表达水平较健康对照组明显上调(P均<0.01);PHA可诱导健康对照T细胞表面CD28和CD40L表达明显上调(P均<0.01),但对其CTLA-4和LFA-1a的表达无明显影响(P均>0.05);而PHA对活动性LN组外周血T细胞表面上述指标表达均有明显上调作用(P均<0.05);叶氏狼疮方对健康人外周血T细胞表面上述指标的表达无明显影响,但可逆转活动性LN患者外周血T细胞表面的CD40L、LFA-1a过度表达(P均<0.05),对其CD28和CTLA-4过度表达无明显影响(P均>0.05);且叶氏狼疮方对PHA所诱导的活动性LN患者外周血T细胞表面CD28、CD40L和LFA-1a表达增加亦有明显抑制。结论:叶氏狼疮方可能主要通过下调T细胞CD40L和LFA-1a表达水平来诱导LN患者外周T细胞免疫耐受重建。
基金This work was supported by National Natural Science Foundation of China(No.30671945)Science and Technology Commission of Shanghai Municipality(Nos.06JC14044,05ZR14055,054319928,04DZ14902)+2 种基金Shanghai Municipal Education(No.05BZ26)Shanghai Leading Academic Discipline Project(T0206)Science Foundation of Shanghai Institute of Immunology(No.07-A04,to Ningli Li).
文摘Regulatory T cells （Treg） play important roles in immune system homeostasis, and may also be involved in tumor immunotolerance by suppressing Th1 immune response which is involved in anti-tumor immunity. We have previously reported that immunization with attenuated activated autologous T cells leads to enhanced anti-tumor immunity and upregulated Thl responses in vivo. However, the underlying molecular mechanisms are not well understood. Here we show that Treg function was significantly downregulated in mice that received immunization of attenuated activated autologous T cells. We found that Foxp3 expression decreased in CD4＋CD25＋ T cells from the immunized mice. Moreover, CD4＋CD25＋Foxp3＋ Treg obtained from immunized mice exhibited diminished immunosuppression ability compared to those from naive mice. Further analysis showed that the serum of immunized mice contains a high level ofanti-CD25 antibody （about 30 ng/ml, p〈0.01 vs controls）. Consistent with a role ofanti-CD25 response in the downregulation of Treg, adoptive transfer of serum from immunized mice to naive mice led to a significant decrease in Treg population and function in recipient mice. The triggering of anti-CD25 response in immunized mice can be explained by the fact that CD25 was induced to a high level in the ConA activated autologous T cells used for immunization. Our results demonstrate for the first time that immunization with attenuated activated autologous T cells evokes anti-CD25 antibody production, which leads to impeded CD4＋CD25＋Foxp3＋ Treg expansion and function in vivo. We suggest that dampened Treg function likely contributes to enhanced Thl response in immunized mice and is at least part of the mechanism underlying the boosted anti-tumor immunity.
文摘Objective： The use of donor-derived immature dendritic cells （imDC） has become a promising approach to induce immune tolerance or immune hyporesponsiveness. However, donor-derived imDC needs to be harvested for a few days and transfused into the recipient in 5-10 days before transplantation, which is practically impossible in a clinical setting where donor organs are mainly harvested from cadavers. Moreover, donor-derived imDC might be cleared by allogeneic reaction offsetting induced immune tolerance or immune hyporesponsiveness. In our study, we further explored the underlying mechanism of immune hyporesponsiveness induced by donor-antigen-unloaded recipient-derived imDC by transfusing these imDC into rats in 1 day before liver transplantation. This paper is to study the mechanism of immune hyporesponsiveness induced by donor-antigen-unloaded recipient-derived imDC and its protection of liver grafts in rats. Methods： 40 SD rats （donor） and 40 male Wistar rats （recipient） were randomly divided into 4 groups： control, cyclosporine A （CsA）, mature DC （mDC）, and imDC; with 10 SD rats and 10 Wistar rats for each group. Animal models of acute graft rejection were established with these rats. Corresponding treatments were given before or after transplantation. In the control group, Wistar rats received no treatment other than liver transplantation. In the CsA group, Wistar rats underwent liver transplantation plus CsA treatment （10 mg/kg·d） in the starting day 2 after transplantation. For the mDC group, recipient-derived mDC （1 × 10^6/rat） were infused intravenously via the dorsal vein of the penis to recipient rats. For the imDC group, imDC （1× 10^6/rat） were injected into recipient rats via the dorsal vein of the penis. In each group, 5 recipients were executed at 10 days after transplantation; the remaining five recipients were kept for the observation of survival time. Blood samples were collected for the measurement of ALT and TBIL; IL-2, IFN-γ, IL-4 and IL-10 and levels were measured with double-antibody sandwich ELISA. Liver tissue was harvested for HE staining and the observation of histological features. Acute rejection was evaluated with Banff classification. Expression levels of Fas-L/Fas in the grafts were detected by iminunohistochemieal staining; and western blot was used to detect the expression level of Scurfin. Results： The median survival times （MST） of the liver allografls in the CsA and imDC group were significantly longer than those in the control or mDC group （P〈0.05）. The serum levels of ALT and TBIL in the control and mDC groups were significantly higher than those of the CsA or imDC group （P〈0.05）. Compared with the CsA anti imDC group, the levels of IL-2 and IFN-γ were higher but the levels of IL-4 and IL-10 were lower than those of the control and mDC groups （P〈0.01）. Slight or no rejection reaction was found in the CsA anti imDC groups （P〈0.05）. The expression level of Scurfin protein in CD4^＋ CD25^＋ T cells of the imDC group was significantly higher than that of three other groups （P〈 0.05）. Conclusion： Donor-antigen-unloaded recipient- derived imDC is an effective treatment in inducing immune hyporesponsiveness by blocking indirect recognition in rat liver transplantation model. Survival span was significantly prolonged by its protective effect. The mechanism of immune hyporesponsiveness induced hy imDC transfusion may involve the preprocesses of T cell apoptosis induction, immune tolerance or hyporesponsiveness in T cells, induction of the shift in TH1/TH2 balance, selection activation of Th2 subset, or induction of regulatory T cell.
