Hepatotoxicity of immune checkpoint inhibitors: An evolving picture of risk associated with a vital class of immunotherapy agents

Immune checkpoint inhibitors (ICIs) block CTLA‐4, PD‐1 and PD‐L1, or other molecules that control antitumour activities of lymphocytes. These products are associated with a broad array of immune‐related toxicities affecting a variety of organs, including the liver. ICI‐associated immune‐mediated hepatitis (IMH) ranges in severity between mild and life‐threatening and is marked by findings that bear both similarities as well as differences with idiopathic autoimmune hepatitis. Hepatotoxic events are often detected in clinical trials of ICIs that are powered for efficacy. Risk levels for ICI‐induced liver injury may be impacted by the specific checkpoint molecule targeted for treatment, the ICI dose levels, and the presence of a pre‐existing autoimmune diathesis, chronic infection or tumour cells which infiltrate the liver parenchyma. When patients develop liver injury during ICI treatment, a prompt assessment of the cause of injury, in conjunction with the application of measures to optimally manage the adverse event, should be made. Strategies to manage the risk of IMH include the performance of pretreatment liver tests with regular monitoring during and after ICI treatment and patient education. Using Common Terminology Criteria for Adverse Events developed at the National Cancer Institute to measure the severity level of liver injury, recommended actions may include continued ICI treatment with close patient monitoring, ICI treatment suspension or discontinuation and/or administration of corticosteroids or, when necessary, a non‐steroidal immunosuppressive agent. The elucidation of reliable predictors of tumour‐specific ICI treatment responses, as well as an increased susceptibility for clinically serious immune‐related adverse events, would help optimize treatment decisions for individual patients.     

Influence of Echinococcus multilocularis infection on reproduction and cellular immune response of mice

The influence of secondary Echinococcus multilocularis infection on reproduction and cellular immune response of mice was studied in BALB/c mice infected with 2000 E. multilocularis protoscoleces. Of the total infected mothers, 11·7% did not give birth and 10% of uninfected ones did not deliver. Both, healthy and infected mothers, produced on average 6–7 offspring per litter. The changes in production of seral IFN‐γ, TNF and IL‐10 did not significantly influence the course of gravidity. On the other hand, more intensive metacestode growth was observed after the delivery. This study confirmed the ability of host organism to adapt to severe damage caused by E. multilocularis, not only in normal conditions, but also during pregnancy.     

免疫检查点抑制剂相关致命性不良反应及管理策略

免疫检查点抑制剂(immune checkpointinhibitors, ICIs)是针对程序性死亡受体1(programmed cell death protein 1,PD-1)/程序性死亡受体配体1(programmed cell death protein ligand1,PD-L1)和细胞毒性T淋巴细胞相关抗原4(cytotoxic T lymphocyte associated antigen-4,CTLA-4)的单克隆抗体,主要通过阻断上述免疫检查点通路的抑制性免疫调节作用从而增强人体的抗肿瘤免疫反应。     

多种药物对体外培养泡球蚴作用的观察

目的观察6种药物单独或联合用药对体外培养泡球蚴的作用。方法泡球蚴体外培养5周后,收集囊泡,随机分为17组,每组约120~140个囊泡,分别加入不同药物进行培养,①单独用药组:阿苯达唑组(1μg/ml、10μg/ml)、伊曲康唑组(0.7 mg/ml)、伊维菌素组(1.75 mg/ml)、米替福新组(0.5μg/ml、2.5μg/ml和7.5μg/ml)、硝唑尼特组(0.1μg/ml、1μg/ml和10μg/ml)、利福平组(10μg/ml);②联合用药组:硝唑尼特联合阿苯达唑10μg/ml+10μg/ml组、10μg/ml+1μg/ml组、1μg/ml+10μg/ml组和1μg/ml+1μg/ml组;③对照组:二甲基亚砜组(2μl/ml)和空白对照组。培养6周,观察囊泡塌陷情况,对囊泡进行计数,并绘制囊泡曲线。6周后停药,联合用药组连续观察3周、3个月和6个月,观察泡球蚴囊泡生长情况。将各药物组体外培养的泡球蚴接种于雌性BALB/c小鼠,饲养8周后处死并剖检观察小鼠腹腔内有无泡球蚴生长并称重,测试泡球蚴活力。结果伊维菌素、米替福新和利福平对泡球蚴无抑制或杀灭作用。阿苯达唑、伊曲康唑、硝唑尼特,及硝唑尼特与阿苯...     

白黎芦醇体外对多房棘球蚴原头节和微囊的作用研究

目的 研究白藜芦醇(Resveratrol,RES)体外对多房棘球蚴原头节(Protoscoleces,PSCs)和微囊的作用效果,为多房棘球蚴病的临床用药提供新的依据.方法 PSCs体外经不同浓度(5、10、20、40、80、100、200和400 μmol/L)的RES作用7 d,采用台盼蓝染色观察其活力与形态变化...     

多房棘球蚴亮氨酸氨基肽酶优势抗原表位的鉴定

目的 根据预测多房棘球蚴亮氨酸氨基肽酶(Leucine aminopeptidase,LAP)抗原表位的结果,进一步鉴定其优势的T细胞和B细胞抗原表位.方法 构建重组质粒pCzn1-LAP后进行蛋白原核表达与纯化,将纯化的LAP蛋白进行动物免疫后,利用脾脏淋巴细胞增殖、流式细胞术、ELISA pot、ELISA等方法鉴...     

多房棘球绦虫原头蚴对体外培养宿主肝细胞MAPK信号通路影响的初步研究

目的探讨多房棘球绦虫原头蚴对体外培养宿主肝细胞丝裂素活化蛋白激酶(mitogen activated protein kinase,MAPK)信号通路的影响。方法采用宿主大鼠肝细胞BRL3A和人源肝癌细胞系HepG2,分别与多房棘球绦虫原头蚴体外无血清共培养。实验组(与多房棘球绦虫原头蚴共培养)每瓶细胞接种约3×103个原头蚴;处理组(U0126)每瓶细胞先用20μmol/L U0126处理3 h,再接种约3×103个原头蚴,继续共培养,分别在12、24、48、72、96和120 h收集细胞,同时收集对照组(无血清培养基)细胞。利用免疫印迹技术(Western blot)检测p-ERK1/2、p-p38和p-JNK的变化。结果在48 h,肝细胞BRL3A共培养组p-ERK1/2表达活性与对照组相比差异有统计学意义(P<0.05),在12、24和48 h,p-JNK和p-p38表达活性分别有微弱升高;在72和96 h肝细胞HepG2共培养组p-ERK1/2表达活性与对照组相比差异有统计学意义(P<0.05),p-JNK和p-p38表达活性无明显变化;U0126处理组p-ERK1/2活性受抑制。结论多房棘球绦虫原...     

多房棘球蚴线粒体NADH脱氢酶1基因分析及PCR检测方法

目的 扩增多房棘球蚴线粒体NADH脱氢酶亚基1(NADH dehydrogenase subunit1,ND1)基因全序列,测序并进行同源性比较.建立检测多房棘球蚴感染的PCR方法,应用于人和动物感染多房棘球蚴的快速检测和鉴定.方法 根据多房棘球绦虫线粒体DNA全序列,设计引物扩增ND1基因并进行测序,获得多房棘球蚴ND1基因全序列.对该序列与其它常见棘球绦虫的ND1基因序列进行同源性分析.结果 多房棘球蚴线粒体ND1基因序列与国外报告的多房棘球绦虫的同源性达到98.8％,与细粒棘球绦虫的同源性为86.2％,与少节棘球绦虫的同源性为84.6％,与伏氏棘球绦虫的同源性为87.0％.结论 多房棘球蚴线粒体ND1基因与其他棘球绦虫相应基因存在显著差异.PCR方法可以用于多房棘球蚴感染的快速检测和鉴定.     

多房棘球绦虫Em14-3-3抗原编码基因研究进展

本文介绍了多房棘球绦虫Em14-3-3蛋白，并对Em14-3-3抗原编码基因的研究进展进行了综述。     

甘肃省合作牦牛自然感染多房棘球蚴的调查

本文报告甘肃合作牦牛自然感染多房棘球蚴，感染率１．６％。病理学观察，多房棘球蚴发育不良，无生发层与原头节存在，与人体多房棘球蚴病理观察相似。     

青海省多房棘球绦虫分子种系发生研究

目的　对青海省多房棘球绦虫分离株Cox1、Nad1基因进行扩增和测序，并构建系统进化树及分子钟，为推测其进化关系及起源提供参考依据。方法　收集2017年青海省人民医院收治的22份肝多房棘球蚴病患者术后标本，对多房棘球蚴线粒体DNA Cox1、Nad1基因进行PCR扩增和测序，并与GenBank数据库中的棘球属绦虫Cox1基因和Nad1基因序列进行比对，观察种内变异情况并构建进化树和分子钟。结果　所有多房棘球绦虫标本与GenBank数据库中检索的多房棘球绦虫亚洲株Cox1、Nad1基因序列同源性均达99%以上，共获得6种不同基因型，其中有2个分离株在GenBank数据库中未找到同源性100%的基因序列。系统进化树显示，收集的多房棘球绦虫标本与多房棘球绦虫亚洲株聚类效果明显；分子钟推测青海地区多房棘球绦虫起源于9.4万年前。结论　本研究发现青海省多房棘球绦虫存在6种不同基因型，其中首次发现2种基因型。青海地区多房棘球绦虫优势株为亚洲株，起源时间约在9.4万年前。     

青海省多房棘球绦虫分子种系发生研究

目的　对青海省多房棘球绦虫分离株Cox1、Nad1基因进行扩增和测序,并构建系统进化树及分子钟,为推测其进化关系及起源提供参考依据。方法　收集2017年青海省人民医院收治的22份肝多房棘球蚴病患者术后标本,对多房棘球蚴线粒体DNA Cox1、Nad1基因进行PCR扩增和测序,并与GenBank数据库中的棘球属绦虫Cox1基因和Nad1基因序列进行比对,观察种内变异情况并构建进化树和分子钟。结果　所有多房棘球绦虫标本与GenBank数据库中检索的多房棘球绦虫亚洲株Cox1、Nad1基因序列同源性均达99%以上,共获得6种不同基因型,其中有2个分离株在GenBank数据库中未找到同源性100%的基因序列。系统进化树显示,收集的多房棘球绦虫标本与多房棘球绦虫亚洲株聚类效果明显;分子钟推测青海地区多房棘球绦虫起源于9.4万年前。结论　本研究发现青海省多房棘球绦虫存在6种不同基因型,其中首次发现2种基因型。青海地区多房棘球绦虫优势株为亚洲株,起源时间约在9.4万年前。     

MEK1/2抑制剂曲美替尼对体外多房棘球蚴原头节的作用研究北大核心CSCD

目的研究MEK1/2抑制剂曲美替尼对体外多房棘球绦虫(Echinococcus multilocularis,Em)活性的影响,探讨MEK1/2作为治疗泡型棘球蚴病靶点的可能性。方法建立Em空白对照组、DMSO及1μmol/L、50μmol/L、100μmol/L曲美替尼组,利用活力染色与扫描电镜(scanning electron microscope,SEM)分别观察虫体活性及体表结构变化;以Swiss Model同源建模合理构建Em MEK1/2分子的三级结构进行分析。结果1μmol/L曲美替尼干预3 d、6 d、9 d的原头节活性分别为(84.72±0.27)%、(87.32±2.01)%、(85.12±3.64)%,低于相同时间DMSO组,且差异有统计学意义(t>2.78,P<0.05);当1μmol/L曲美替尼干预时间延长至12 d,原头节活性降至(37.82±2.12)%,与DMSO组比较差异具有统计学意义(t=32.90,P<0.01)。SEM和活力染色显示曲美替尼组虫体活性与干预浓度成正比,同源建模结果显示Em与人MEK1/2分子具有较高的同源性。结论综合分析活力染色、SEM与同源建模结果,证实体外1μmol/L的MEK1/2抑制剂曲美替尼对虫体活性具有较好的抑制作用,为临床治疗提供了理论依据。     

Lower expression of PD-1 and PD-L1 in peripheral blood from patients with chronic ITP

Background: T-cell dysregulation is a major event involved in immune thrombocytopenic purpura (ITP). Increasing data have indicated that abnormal expression of T-cell immunosuppressive receptors, such as programmed death (PD) 1 and cytotoxic T lymphocyte antigen-4 (CTLA-4), may be related to autoimmune disease pathogenesis.

Methods: We analyzed the expression levels of PD-1, its ligand PD-L1, and CTLA-4 in peripheral blood mononuclear cells from 18 patients with chronic ITP by real-time polymerase chain reaction, and samples from 20 healthy individuals served as control.

Results: The results demonstrated significantly lower expression of PD-1 (median: 0.0015) and PD-L1 (median: 0.0572) in chronic ITP patients compared with healthy individuals (PD-1: median: 0.0117, P?<?0.0001; PD-L1: median: 0.5428, P?<?0.0001), while there was no significant difference in the CTLA-4 expression level between the chronic ITP patients (median: 0.0818) and healthy individuals (median: 0.1667) (P?=?0.219). Moreover, a positive correlation between the expression levels of PD-1 and PD-L1 (r_s?=?0.486, P?=?0.041) and CTLA-4 and PD-1 (r_s?=?0.643, P?=?0.004) in the chronic ITP patients was found.

Conclusion: Consistently lower expression of T-cell immunosuppressive receptors is a common characteristic of chronic ITP, which may be associated with its pathogenesis.     

多房棘球绦虫elp基因核酸疫苗诱生免疫应答的实验研究

目的观察多房棘球绦虫elp基因核酸疫苗免疫BALB/c小鼠引起的体液和细胞免疫应答。方法30只BALB/c小鼠随机分成3组:Ⅰ组,肌注空质粒(pcDNA3.1)100μg/鼠;Ⅱ组,肌注多房棘球绦虫elp基因真核表达重组质粒(pcD-ELP)100μg/鼠;NS组,肌注等体积生理盐水。每2周免疫1次,共3次。ELISA方法检测免疫后不同时间产生的特异性IgG1,IgG2a和IgG 2 b水平。双抗体夹心ELISA法检测免疫鼠脾脏单个核细胞(PMNC)在受到特异抗原刺激后,产生IL-4I、L-12和IFN-γ的能力,3H-TdR掺入法检测脾淋巴细胞的增值能力。结果首次免疫后4周,Ⅱ组小鼠血清可检测到特异性IgG1I、gG2a和IgG2b抗体,随免疫时间和免疫次数的增加3种特异性抗体的OD值逐渐增高,实验结束时(首次免疫后7周)3种特异性抗体OD值均最高。Ⅰ组和Ⅱ组小鼠脾PMNC培养上清中IFN-γ浓度分别为50 pg/ml和55 pg/ml,受特异抗原刺激后分别为44 pg/ml和101.5 pg/ml。NS组IFN-γ及各组IL-4和IL-12均未检出。Ⅰ组和Ⅱ组小鼠脾淋巴细胞增殖能力增高,其淋巴细胞CPM值分别为NS组的85倍和123倍,受到特异抗原或刀豆素刺激其淋巴细胞增殖更明显。结论多房棘球绦虫elp基因核酸疫苗可刺激BALB/c小鼠同时产生很强的细胞免疫和体液免疫应答。     

Arginine-dependent generation of reactive nitrogen intermediates is instrumental in the in vitro killing of protoscoleces of Echinococcus multilocularis by activated macrophages

The interaction between protoscoleces of Echinococcus multilocularis and activated murine macrophages was examined in this study. Marked protoscolicidal activity was displayed by peritoneal macrophages (PM) activated with interferon-γ (IFN-γ), or IFN-y plus lipopolysaccharide. Pretreatment of the parasites with heat-inactivated specific murine infection serum, but not with normal serum rendered them more susceptible to PM killing. N^G-monomethyl-Larginine, a competitive inhibitor of L-arginine completely inhibited the killing activity of activated PM. while reconstitution of arginine-free medium with L-arginine restored the killing properties of the activated PM. The results show that activated PM have the ability to kill E. multilocularis protoscoleces in vitro and suggest that reactive nitrogen intermediates (RNl) play an important role in the mechanism. An oxygen-mediated mechanism did not appear to play a role because scavengers of reactive oxygen species did not reduce the killing activity. The arginine-dependent killing mechanism was enhanced by superoxide dismutase (SOD), probably because SOD might prolong the effect of nitric oxide. Secretion of RNI by activated macrophages may be capable of a significant role in preventing of the dissemination of E. multilocularis infection in vivo.     

苦参碱和阿苯达唑联合治疗小鼠泡球蚴病的机制探讨

目的 探讨泡球蚴病小鼠经苦参碱（matrine, Mat）阿苯达唑（albendazole, ABZ）及两药 联用治疗后机体的免疫应答状态和小鼠肝酶代谢的变化。 方法 泡球蚴病小鼠（AE小鼠）经Mat、ABZ及其联合用药治疗60 d后，检测小鼠血清中白细胞介素-2（IL-2）、IL-4、IL-6、肿瘤坏死因子-α（TNF-α）及肝组织匀浆中NO、一氧化氮合酶（NOS）、诱导型一氧化氮合酶（iNOS）、乳酸（LD）、乳酸脱氢酶（LDH）、钠钾ATP酶和钙镁ATP酶的含量及活性。 结果 药物治疗组血清中IL?鄄4、IL-6和TNF-α的含量较感染对照组降低（P<0.05），IL?鄄2的含量高于感染对照组（P<0.05），其中联合用药组IL-4和TNF-α下降较感染对照组明显（P<0.05）；各用药组肝组织LDH、NO、NOS和iNOS均较感染对照组明显下降（P<0.05），iNOS在Mat组及联合用药组下降幅度明显大于ABZ组（P<0.05），LD仅在联合用药组下降（P<0.05）；各用药组钠钾ATP酶和钙镁ATP酶较之对照组升高（P<0.05），且Mat组及联合用药组钙镁ATP酶与ABZ组差异有统计学意义（P<0.05）。 结论 AE小鼠经治疗后，Th1型细胞因子分泌增加，Th2型细胞因子分泌减少，说明Mat能提高小鼠机体的免疫力，并能显著改善小鼠肝功能，这可能与Mat作用钙离子通道和逆转耐药性有关。     

泡球蚴囊液对大鼠肝星状细胞MAPK信号通路5个相关基因的影响

目的探讨泡球蚴囊液对大鼠肝星状细胞丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路5个相关基因的影响。方法采用大鼠肝星状细胞株HSC-T6与不同蛋白浓度的泡球蚴囊液(0.01、0.025、0.05、0.1、0.2、0.4、0.9、1.7、3.4、6.8和13.5 mg/ml)共培养24 h后收集细胞,同时收集对照组(未加泡球蚴囊液干预)细胞,显微镜下观察HSC-T6细胞的形态变化。利用荧光实时定量PCR检测大鼠肝星状细胞的细胞外调节蛋白激酶(ERK)、c-Jun氨基末端激酶(JNK)和丝裂原活化蛋白激酶(p38)的表达情况。结果蛋白浓度为13.5 mg/ml泡球蚴囊液与HSC-T6细胞共培养24 h后,大部分细胞发生固缩,呈脱落前兆。6.8 mg/ml囊液组部分细胞固缩,呈脱落前兆;而部分HSC-T6细胞收缩,呈长梭形,细胞生长出许多细长的伪足,是正常状态的2倍以上。3.4 mg/ml囊液组部分细胞呈收缩的长梭形,生长出许多细长的伪足,但大部分细胞仍呈现为贴壁的胞体较大的不规则多边形,且伸出较短的伪足与其他细胞连接成片状。1.7 mg/ml囊液组与对照组细胞形态上无差异。实时荧光定量PCR检测显示,泡球蚴囊液浓度0.4 mg/ml时,ERK1/2、JNK1/2和p38 m RNA水平显著增加;囊液浓度为6.8 mg/ml时,ERK1/2、JNK1/2和p38 m RNA较对照组高表达(P0.05)。结论浓度为6.8 mg/ml的泡球蚴囊液可显著影响大鼠肝星状细胞ERK1/2、JNK1/2和p38 m RNA水平。