IL‐27p28 is essential for parent‐to‐F1 acute graft‐versus‐host disease

Acute graft versus host disease (aGVHD) remains a life‐threatening complication of bone marrow transplantation. Here we show that IL‐27, a member of the IL‐12 cytokine family, plays an essential role in a parent‐to‐F1 murine aGVHD model, using B6 mice as parents and B6D2 mice as F1 recipients. IL‐27 is transiently detectable in the serum of B6D2 recipients of B6 spleen cells, with a peak at day 10. Treatment with anti‐IL‐27p28 mAb MM27.7B1 (αp28Ab), at the time of and six days after B6 cell transfer, blocked GVHD. Protection was associated with host cell survival and undiminished engraftment of donor cells, lack of host B‐cell depletion, increased Th2‐type immunoglobulin production, a decrease in serum IFN‐γ, a drop in anti‐H‐2D^d cytotoxic T lymphocyte activity and an increase in Foxp3⁺ T cells. We therefore conclude that IL‐27 plays a critical role in the parent‐to‐F1 model of aGVHD and that blocking IL‐27 could have therapeutic relevance.     

The influence of glucocorticoid receptor single nucleotide polymorphisms on outcome after haematopoietic stem cell transplantation

Haematopoietic stem cell transplantation (HSCT) remains the only cure for most haematological malignancies, however, the mortality rate remains high. Complications after HSCT include relapse, graft versus host disease (GvHD), graft rejection and infection. Over the last few years several groups, have demonstrated that non‐HLA gene polymorphisms can be predictive of outcome after HSCT. Since the glucocorticoid cortisol is pivotal in the regulation of the immune system, we decided to examine single nucleotide polymorphisms (SNPs; rs6198, rs33388 and rs33389) within the glucocorticoid receptor (GR) and correlate with HSCT outcome. The training set consisted of patients (n = 458) who underwent HSCT for acute leukaemia between 1983 and 2005. In the recipients, the absence of the ACT haplotype and absence of the T allele of rs33388 were associated with decreased OS and the absence of the ACT haplotype, the absence of the T allele of rs33388 and the presence of the ATA haplotype were associated with increased risk of relapse. In addition, the presence of the ACT haplotype in the recipient showed a trend to be associated with increased risk of chronic graft versus host disease (cGvHD). The patients in this cohort received mainly myeloablative conditioning (n = 327). The SNPs in the glucocorticoid receptor were then investigated in a validation set (n = 251) of HSCT patients transplanted for acute leukaemia from 2006. This cohort contained significantly more patients that had received reduced intensity conditioning (RIC). Some of the results could be validated in these patients. However, contrary to the training set, the absence of the haplotype ACT in the donor in this cohort was associated with increased risk of cGvHD. Differences in the conditioning were shown to influence the results. These results are the first to associate GR SNPs with HSCT outcome and demonstrate the inherent problems of replicating SNP association studies in HSCT, due to different pre‐transplant regimens.     

Serum levels of cytokines after bone marrow transplantation: increased IL-8 levels during severe veno-occlusive disease of the liver

Abstract: We investigated the cytokine profile and peak levels of interleukin (IL) -6, IL-8, IL-10 and tumour necrosis factor (TNF) -α levels in 42 patients after allogeneic bone marrow transplantation (BMT). Eleven of them developed veno-occlusive disease (VOD) of the liver. Fourteen patients had moderate-to-severe acute graft-versus-host disease (aGvHD), 10 isolated bacteraemia and 7 had no major complication. Those who developed severe VOD (n = 6) showed a short, very high IL-8 peak (median: 6632 pg/ml, range: 5546 – 10,000 vs. 280 pg/ml, 0 – 2042 in controls, p < 0.01) 1 – 4 d after diagnosis of the liver disease. Five of these patients had high peak levels of IL-6. Five patients with mild VOD showed a lower increase in the cytokines tested. Bilirubin levels, at day of IL-8 peak, did not differ statistically between mild and severe VOD. The highest levels of IL-10 were found in those with aGvHD. IL-8 levels were also increased, but not to the same extent as in patients with severe VOD (p = 0.01 vs. VOD). In patients with bacteraemia, very high levels of IL-6 were seen. In patients without major complications, the levels of cytokines were low. In conclusion, high levels of IL-8 occurred in severe VOD of the liver, which may be of value to determine prognosis.     

敲除BRCC3加重异基因造血干细胞移植受体小鼠急性移植物抗宿主病

目的研究BRCC3的缺失对急性移植物抗宿主病(aGVHD)的影响, 并初步探讨其机制。方法 12只C57BL/6J小鼠作为受体, 分为野生型(WT组)和BRCC3-/-型(KO组), 每组6只。采用两次4.5 Gy60Co全身照射(TBI), 两次照射间隔30 min, 6 h后通过尾静脉输注BABL/c小鼠1×107骨髓细胞和8×106脾脏细胞建立aGVHD小鼠模型。在aGVHD小鼠模型中特异性敲除BRCC3, 通过组织病理学检测靶器官损伤;酶联免疫吸附试验(ELISA)和细胞因子微球检测技术(CBA)检测小鼠血清中细胞因子的水平;移植后第9天分离脾脏、肝脏和小肠淋巴细胞, 运用流式细胞术检测靶器官中T细胞的浸润和活化。结果受体小鼠BRCC3的缺失可导致aGVHD小鼠生存期明显缩短(P<0.05);靶器官肝脏损伤明显加重;脾脏中CD8+ T细胞和CD8+CD25+ T细胞的比例明显升高(t=6.53、5.52, P<0.05);肝脏中CD8+ T细胞和CD8+CD25+ T细胞的比例明显升高(t=3.74、3.19, P<0.05)以及小肠中CD8+ T细胞、CD8...     

苦参碱联合环孢素A减轻小鼠急性移植物抗宿主病

背景：苦参碱具有降低白细胞介素2浓度的作用,作为化疗辅助用药已用于临床。　 目的：探讨苦参碱联合环孢素A对小鼠异基因骨髓移植后急性移植物抗宿主病发生发展的影响,及苦参碱可能的作用机制。 方法：C57BL/6小鼠作为供鼠,BABL/C小鼠为受鼠,建立小鼠同种异基因骨髓移植模型。BALB/C受鼠随机分为7组：空白对照组、单纯照射组、骨髓移植组及足量环孢素A、半量环孢素A、足量苦参碱组、足量苦参碱联合半量环孢素A组。 结果与结论：足量苦参碱联合半量环孢素A组小鼠生存时间明显长于其他组。进行骨髓移植的小鼠均出现不同程度病理改变,越早程度越重。移植后7 d,与骨髓移植组比较,其他移植组小鼠血清γ-干扰素质量浓度下降,白细胞介素4质量浓度差异无显著性意义。提示,苦参碱能够减轻小鼠异基因骨髓移植后致死性急性移植物抗宿主病的发生,生存时间延长。苦参碱与环孢素A作用相似,二者联用,有协同作用。     

急性移植物抗宿主病细胞因子基因多态性与环孢素A血药浓度的相关性研究

目的：探讨异基因造血干细胞移植（allo-HSCT）中急性移植物抗宿主病细胞因子基因多态性与环孢素A的关系。方法：选取2014年1月至2015年12月行allo-HSCT受者32例作为研究对象,采用聚合酶链式反应（PCR）联合基因测序法检测目的基因特殊SNP位点基因分型,测定术后早期出现aGVHD的重要时期CsA的血药浓度,观察受者术后出现aGVHD的情况,比较TGF-β1+869（C/T）不同基因分型CsA血药浓度的差异,分析TGF-β1+869（C/T）基因多态性与CsA的关系及对aGVHD的影响。结果：本组32例患者的TGF-β+869基因多态性研究结果为16例C/C基因型占50.00%,9例T/T基因型占28.12%,7例C/T基因型占21.88%;C/T型患者中的4例出现重度aGVHD占57.14%,发生率与C/C型、T/T型患者比较有显著性差异（P<0.01）。3种基因型患者的CsA血药谷浓度均能达到200~300 μg·L^-1的目标值,比较3种基因型患者的CsA血谷浓度,C/T型及T/T型患者均高于C/C型患者（P<0.01）。结论：本组患者发生重度aGVHD与CsA血药浓度无明显相关性,TGF-β1+869基因型为C/T杂合子的allo-HSCT受者预防GVHD应调整免疫抑制方案,加用霉酚酸酯或使用其它免疫抑制剂以减少重度aGVHD发生。     

蛋白质芯片技术检测异基因造血干细胞移植患者血浆IL-1α、IL-2和VEGF含量的变化与aGVHD关系的初步探讨

目的通过动态观察异基因造血干细胞移植（allo-HSCT）患者血浆中IL-1α、IL-2及血管内皮细胞生长因子（vascular endothelial growth factor，VEGF）含量的变化．探讨其变化与临床急性移植物抗宿主病（aGVHD）发生的关系。方法32例以异基因外周血造血干细胞移植（allo-PBSCT）治疗的患者．其中男性24例．女性8例．中位年龄36（22-47）岁。根据移植后aGVHD发生情况分成2组，分别为aGVHD组（24例）和无aGVHD组（8例）。对2组患者采用蛋白质芯片方法动态检测外周血细胞因子IL-1α、IL-2和VEGF的含量．来探讨细胞因子IL-1α、IL-2和VEGF的含量变化与aGVHD发生的关系。结果aGVHD组的IL-2含量在移植后第2周升高并达到峰值．显著高于移植前含量及相同时间无aGVHD组（P〈0．01），IL-2的含量变化要早于aGVHD症状出现．而且随着aGVHD的症状控制．IL-2的含量下降。无aGVHD组IL-1α含量在移植后的第3、5周显著高于aGVHD组（P〈0．01）；血浆VEGF的含量在移植后的第5周无aGVHD组患者达到最高浓度．而在aGVHD组，VEGF的最高浓度值出现在移植后的第7周。结论IL-2在allo-PBSCT后含量突然升高提示即将发生急性aGVHD；无aGVHD组VEGF的峰值要比aGVHD组提前1周出现．可能预示着无aGVHD的患者比aGVHD患者造血重建更早；2组IL-1α含量在整个观察期内波动较大。通过蛋白质芯片技术检测对IL-2含量并动态观察以预测aGVHD．可能是一个颇具前景的研究方向。     

苦参碱对异基因造血干细胞移植后小鼠细胞因子影响的实验研究

目的：探讨苦参碱对畀基因造血干细胞移植后小鼠细胞因子的影响。方法：用雄性C57BL／6小鼠作为供鼠，雄性BABUC小鼠为受鼠，输入供鼠T淋巴细胞诱导出急性移植物抗宿主反应。BALB／C受鼠随机分为7组：A空白对照组、B单纯照射组、C骨髓移植组、D足量CsA组、E半量CsA组、F苦参碱组、G苦参碱联合半量CsA组。利用酶联免疫吸附试验测定各组小鼠血清INF-γ、IL-4等细胞因子浓度变化，并进行组间比较。结果：移植后7天，4组实验组小鼠（足量CsA、半量CsA、苦参碱、苦参碱联合半量CsA组）血清INF-γ浓度较骨髓移植组降低，足量CsA组与苦参碱联合半量CsA组及苦参碱组的差异，均具有显著统计学意义（P〈0．01）；4组实验组小鼠血清IL-4浓度较骨髓移植组无明显变化，差异无显著统计学意义（P〉0．01）。结论：苦参碱可降低小鼠血清INF-γ浓度，但对IL-4浓度的影响不明显。     

The Tim-3/galectin-9 pathway involves in the homeostasis of hepatic Tregs in a mouse model of concanavalin A-induced hepatitis

T cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) is a negative regulator of interferon (IFN)-γ-secreting CD4⁺ Th1 cells and plays a key role in autoimmune diseases. Here, we report that galectin-9 expression was increased in hepatic CD4⁺CD25⁺ T cells in a mouse model of concanavalin A (Con A)-induced hepatitis. Moreover, Tim-3 showed increased levels in CD4⁺CD25⁺ Foxp3⁺ regulatory T cells (Tregs). Further analyses showed that blocking the Tim-3/galectin-9 pathway resulted in the suppression of Tregs in vitro, thereby significantly increasing interferon (IFN)-γ production from hepatic Teffs. Moreover, blockade of Tim-3 in vivo with an anti-Tim-3 antibody exacerbated the acute hepatitis, possibly by increased IFN-γ production. Furthermore, we found that in vitro activation of CD4⁺CD25⁻ T cells with the T cell receptor (TCR) plus interleukin 2 (IL-2) up-regulated Tim-3 expression. And the induced Tim-3 interacted with galectin-9 to induce CD4⁺ T cell apoptosis which could be partly reversed by blocking Tim-3 signaling. Our results suggested that the Tim-3/galectin-9 pathway plays a critical role in the homeostasis of hepatic Tregs through the elimination induction in Teffs and the inhibition of IFN-γ release, which contributes to the pathogenesis of liver damage and constitutes at least part of the mechanism underlying the induction of hepatitis by Con A.     

高压氧对异基因骨髓移植后aGVHD影响的实验研究

本研究探讨高压氧（HBO）在抗异基因骨髓移植后aGVHD和提高生存率中的作用及机制。给受致死量照射的C57BL／6受鼠注入供鼠BALB／c骨髓与脾淋巴细胞的混合液，分别给与高压氧、环孢素A、氨甲蝶呤处理。用流式细胞仪检测脾T细胞及其亚群和黏附分子CD3^＋、CD4^＋、CD8^＋、CD4^＋CD11a^＋、CD4^＋CD18^＋、CD8^＋CD11a^＋、CD8^＋CD18^＋的表达，用ELISA和RT-PCR法检测细胞因子IL-2、IL-4、IL-10和TNFα的表达。结果表明：HBO组小鼠11天的活存率明显高于异基因移植组及CsA＋MTX组；HBO和CsA＋MTX可明显降低脾组织中CD3^＋、CD4^＋、CD8^＋、CD4^＋CD11a^＋、CD4^＋CD18^＋、CD8^＋CD11a^＋、CD8^＋CD18^＋的表达（P〈0.05）；HBO组小鼠血清IL-2和TNFα的浓度及脾组织中IL-2和TNFα mRNA的含量都低于异基因骨髓移植组（P〈0.05），但高于CsA＋MTX组；HBO组小鼠脾组织中IL-4和IL-10 mRNA的含量显著高于异基因骨髓移植组和CsA＋MTX组（P〈0、05）。结论：高压氧在对抗aGVHD和提高活存率方面优于环孢素A和氨甲蝶呤，其抗aGVHD的作用机制与调节促炎／抑炎细胞因子的分泌和黏附分子的表达及抑制T淋巴细胞作用有关。