冠状病毒疫苗研究进展

进入21世纪以来,严重急性呼吸综合征冠状病毒(severe acute respiratory syndrome coronavirus, SARS-CoV)、中东呼吸综合征冠状病毒(Middle East respiratory syndromecoronavirus, MERS-CoV)及最新出现的严重急性呼吸综合征冠状病毒-2(severe acute respiratory syndrome coronavirus-2, SARS-CoV-2)等高致病性冠状病毒先后在人群中暴发流行,成为影响地区、国家乃至全球的重大公共卫生事件,研发特异性疫苗成为防控病毒流行的当务之急。本文综述了SARS-CoV和MERS-CoV疫苗的研究进展,望对SARS-CoV-2疫苗研制提供参考。     

新型冠状病毒与血管紧张素转换酶2对心血管系统的影响

自2019年12月起,爆发由新型冠状病毒(SARS-CoV-2)感染的新型冠状病毒肺炎(COVID-19),疫情迅速蔓延至全球。COVID-19以呼吸系统症状为主,但部分病例出现心血管系统损害。合并心血管系统基础疾病患者,会导致死亡率增加。SARS-CoV-2属于冠状病毒科β冠状病毒属,SARS-CoV-2与严重急性呼吸综合征冠状病毒具有79.5%的同源性,通过受体血管紧张素转换酶2(ACE2)入侵人体细胞。而表达ACE2的Ⅱ型肺上皮细胞是SARS-CoV-2感染的主要靶细胞。因此,了解SARS-CoV-2所致心血管系统损害及相关的机制,对SARS-CoV-2疫苗和药物的研制及降低病死率具有重要的意义。     

SARS-CoV-2的研究进展

新型冠状病毒(SARS-CoV-2)属冠状病毒家族中的新成员,从感染人体引起第一例不明原因肺炎开始,已导致世界累计确诊人数超2 000万人。此种病原体感染性之强,传播之迅速,远超2003年流行的SARS冠状病毒(SARS-CoV)。目前,各国学者已广泛开展SARS-CoV-2的研究,更为深入解析该病毒的微生物学特性的同时,也在逐步开展疫苗、治疗性抗体和抗病毒药物的研究工作。尤其是在疫苗研发中,我国已取得令人瞩目的 成果。本文基于当前SARS-CoV-2最新研究进展,对该病原体的生物学特性、起源、感染与宿主免疫反应、传播及治疗等做出研究与探讨。     

病毒感染性疾病中血小板减少机制的研究进展

血小板的生物学功能是参与凝血和维持血管系统完整性。血小板减少症见于多种血液系统疾病、放化疗损伤及药物相关性血小板减少,也可作为多种疾病的伴发症状,如肿瘤和感染等。病毒感染性疾病可导致血小板减少,其可能的机制主要有血小板生成减少和消耗增加,且不同病毒导致血小板减少的机制也不尽相同。本文对新型冠状病毒(SARS-CoV-2)、汉滩病毒、登革热病毒及人类免疫缺陷病毒(HIV)等病毒感染性疾病中血小板减少机制研究进展进行综述,特别是新近发现的注射新冠疫苗后发生血栓性血小板减少可能的机制,可为病毒感染性疾病中血小板减少的预防与治疗提供新的思路。     

新型冠状病毒引起肝损伤的研究现状

2019年新型冠状病毒(SARS-CoV-2)引起的病毒性肺炎在中国武汉爆发,对公众健康构成重大威胁。SARS-CoV-2与严重急性呼吸综合征相关冠状病毒(SARS-CoV)和中东呼吸综合征相关冠状病毒(MERS-CoV)高度同源,都会导致严重的呼吸道症状。除呼吸系统症状外,相当比例的SARS和SARS-CoV-2感染者出现不同程度的肝损伤,但其流行病学特征和发生机制尚不明确。在总结SARS-CoV-2流行病学基础上,重点阐述SARS-CoV-2及其所致肝损伤的可能机制和有效治疗方案的研究现状,为SARS-CoV-2感染者肝损伤的防治提供参考和研究思路。     

动物冠状病毒检测方法的研究进展

2019年12月底以来,严重急性呼吸系统综合征冠状病毒2(severe acute respiratory syndrome coronavirus 2,SARS-CoV-2)给全球公共卫生安全带来了严重的威胁.包括SARS-CoV-2在内的多种冠状病毒都有动物源性.因此,加强对动物冠状病毒的监测尤为重要.本文针对检测...     

新型冠状病毒相关自发性脾破裂的诊治

新型冠状病毒肺炎(COVID-19)是由新型冠状病毒(SARS-CoV-2)感染引起的主要经呼吸道传播的急性病毒性疾病，其不仅侵袭人体呼吸系统，还损伤人体多个器官系统。现已有证据显示SARS-CoV-2与自发性脾破裂之间可能存在因果联系，本文认识到SARS-CoV-2相关性自发性脾破裂发生的可能性，并探讨其发生机制和诊治方案，避免临床上对其漏诊误诊。     

慢性HBV感染者接种新型冠状病毒疫苗的安全性及有效性研究

目的?分析慢性HBV感染者全程接种3剂新型冠状病毒（severe acute respiratory syndrome coronavirus 2, SARS-CoV-2）疫苗的安全性及有效性。方法?纳入2021年10月—2022年8月在解放军总医院第五医学中心就诊并自愿接种SARS-CoV-2疫苗的慢性HBV感染者107例，在第0（基线）、1、2、4、7、8月时进行访视并定期检测血常规、血生化及凝血四项等实验室指标。通过记录慢性HBV感染者接种每剂SARS-CoV-2疫苗后28 d内的不良反应及实验室指标的动态变化特点以评估其安全性。在随访时留取血浆样本检测中和抗体滴度以评估其有效性，同时分析抗体滴度可能的影响因素。结果?慢性HBV感染者接种3剂SARS-CoV-2疫苗后不良反应发生率依次为22.43%（24/107）、19.79%（19/96）和16.67%（12/72），最常见的局部不良反应为接种部位疼痛，最常见的全身性的不良反应为疲劳、乏力；接种疫苗后生化学及病毒学指标未出现恶化；接种每剂SARS-CoV-2疫苗后中和抗体滴度均显著上升（P均＜0.05），而后随着时间逐渐下降；抗体阳性组受试者年龄低于阴性组受试者[（42.27±9.40）岁 vs. 48.00（43.00，53.50）岁，P = 0.040]。结论?慢性HBV感染者接种SARS-CoV-2疫苗后具有良好的安全性及有效性，不良反应发生率低且未发生疫苗相关严重不良事件；接种第3剂加强针后产生的中和抗体反应最为强烈；而年龄则会影响慢性HBV感染者接种疫苗后的抗体滴度。     

新型冠状病毒(SARS-CoV-2)变异的研究进展

2019年12月底,湖北省武汉市暴发了由新型冠状病毒(SARS-CoV-2)引起的肺炎疫情.迄今为止,该病毒引起的疫情仍在全球流行,累计感染人数超1.8亿.随着SARS-CoV-2在人群间的不断传播,其基因组不断发生变异,从SARS-CoV-2首次出现S蛋白D614G突变到被世界卫生组织列为关切的Alpha、Beta、...     

《2021年欧洲肝病学会专家立场声明:慢性肝病、肝胆恶性肿瘤及肝移植受者COVID-19疫苗的接种》摘译

正2019年以来,严重急性呼吸综合征冠状病毒2(severe acute respiratory syndrome coronavirus 2,SARS-CoV-2)引起的冠状病毒病-2019(coronavirus disease 2019,COVID-19;国内习惯称为新型冠状病毒肺炎)疫情肆虐全球。据世界卫生组织(WHO)评估,截至2021年1月,全球已有近1亿人感染SARSCoV-2。有慢性肝病(chronic liver diseases,CLD)的人群,     

Laboratory testing for suspected COVID-19 vaccine–induced (immune) thrombotic thrombocytopenia

COVID-19 (coronavirus disease 2019) represents a pandemic, and several vaccines have been produced to prevent infection and/or severe sequelae associated with SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection. There have been several reports of infrequent post vaccine associated thrombotic events, in particular for adenovirus-based vaccines. These have variously been termed VIPIT (vaccine-induced prothrombotic immune thrombocytopenia), VITT (vaccine-induced [immune] thrombotic thrombocytopenia), VATT (vaccine-associated [immune] thrombotic thrombocytopenia), and TTS (thrombosis with thrombocytopenia syndrome). In this report, the laboratory test processes, as utilised to assess suspected VITT, are reviewed. In published reports to date, there are notable similarities and divergences in testing approaches, potentially leading to identification of slightly disparate patient cohorts. The key to appropriate identification/exclusion of VITT, and potential differentiation from heparin-induced thrombocytopenia with thrombosis (HITT), is identification of potentially differential test patterns. In summary, testing typically comprises platelet counts, D-dimer, fibrinogen, and various immunological and functional assays for platelet factor 4 (PF4) antibodies. In suspected VITT, there is a generally highly elevated level of D-dimer, thrombocytopenia, and PF4 antibodies can be identified by ELISA-based assays, but not by other immunological assays typically positive in HITT. In addition, in some functional platelet activation assays, standard doses of heparin have been identified to inhibit activation in suspected VITT, but they tend to augment activation in HITT. Conversely, it is also important to not over-diagnose VITT, given that not all cases of thrombosis post vaccination will have an immune basis and not all PF4-ELISA positive patients will be VITT.     

Longitudinal Aspects of VITT

In hundreds of patients worldwide, vaccination against COVID-19 with adenovirus vector vaccines (ChAdOx1 nCoV-19; Ad26.COV2.S) triggered platelet-activating anti-platelet factor 4 (PF4) antibodies inducing vaccine-induced immune thrombotic thrombocytopenia (VITT). In most VITT patients, platelet-activating anti-PF4-antibodies are transient and the disorder is discrete and non-recurring. However, in some patients platelet-activating antibodies persist, associated with recurrent thrombocytopenia and sometimes with relapse of thrombosis despite therapeutic-dose anticoagulation. Anti-PF4 IgG antibodies measured by enzyme-immunoassay (EIA) are usually detectable for longer than platelet-activating antibodies in functional assays, but duration of detectability is highly assay-dependent. As more than 1 vaccination dose against COVID-19 is required to achieve sufficient protection, at least 69 VITT patients have undergone subsequent vaccination with an mRNA vaccine, with no relevant subsequent increase in anti-PF4 antibody titers, thrombocytopenia, or thrombotic complications. Also, re-exposure to adenoviral vector-based vaccines in 5 VITT patients was not associated with adverse reactions. Although data are limited, vaccination against influenza also appears to be safe. SARS-CoV-2 infection reported in 1 patient with preceding VITT did not influence anti-PF4 antibody levels. We discuss how these temporal characteristics of VITT provide insights into pathogenesis.     

Thrombosis and thrombocytopenia in COVID-19 and after COVID-19 vaccination

Thrombosis that occurs in coronavirus disease 19 (COVID-19) is a serious complication and a critical aspect of pathogenesis in the disease progression. Although thrombocytopenia is uncommon in the initial presentation, it may also reflect disease severity due to the ability of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to activate platelets. This occurs directly through the spike protein-angiotensin converting enzyme 2 (ACE2) interaction and indirectly by coagulation and inflammation activation. Dysregulation in both innate and adaptive immune systems is another critical factor that causes thrombosis and thrombocytopenia in COVID-19.Vaccination is the most potent and effective tool to mitigate COVID-19; however, rare side effects, namely vaccine-induced immune thrombotic thrombocytopenia (VITT)/thrombosis with thrombocytopenia syndrome (TTS) can occur following adenovirus-vectored vaccine administration. VITT/TTS is rare, and thrombocytopenia can be the clue to detect this serious complication. It is important to consider that thrombocytopenia and/or thromboembolism are not events limited to post-vaccination with vectored vaccine, but are also seen rarely after vaccination with other vaccines.Various conditions mimic VITT/TTS, and it is vital to achieving the correct diagnosis at an earlier stage. Antiplatelet factor 4 (PF4) antibody detection by the enzyme-linked immunosorbent assay (ELISA) is used for diagnosing VITT/TTS. However, false-positive rates also occur in vaccinated people, who do not show any thrombosis or thrombocytopenia. Vaccinated people with messenger RNA vaccine can show positive but low density and non-functional in terms of platelet aggregation, it is vital to check the optical density. If anti-PF4 ELISA is not available, discriminating other conditions such as antiphospholipid syndrome, thrombotic thrombocytopenic purpura, immune thrombocytopenic purpura, systemic lupus erythematosus, and hemophagocytic syndrome/hemophagocytic lymphohistiocytosis is critical when the patients show thrombosis with thrombocytopenia after COVID-19 vaccination.     

Treatment of vaccine-induced immune thrombotic thrombocytopenia (VITT)

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a novel prothrombotic disorder characterized by thrombosis, thrombocytopenia, and disseminated intravascular coagulation identified in hundreds of recipients of ChAdOx1 nCoV-19 (Oxford/AstraZeneca), an adenovirus vector coronavirus disease 2019 (COVID-19) vaccine. VITT resembles heparin-induced thrombocytopenia (HIT) in that patients have platelet-activating anti-platelet factor 4 antibodies; however, whereas heparin typically enhances platelet activation by HIT antibodies, VITT antibody-induced platelet activation is often inhibited in vitro by pharmacological concentrations of heparin. Further, the thrombotic complications in VITT feature much higher frequencies of atypical thrombosis, most notably cerebral vein thrombosis and splanchnic vein thrombosis, compared with HIT. In this review, we outline the treatments that have been used to manage this novel condition since its recognition in March 2021, including anticoagulation, high-dose intravenous immune globulin, therapeutic plasma exchange, corticosteroids, rituximab, and eculizumab. We discuss the controversial issue of whether heparin, which often inhibits VITT antibody-induced platelet activation, is harmful in the treatment of VITT. We also describe a case of “long VITT,” describing the treatment challenges resulting from platelet-activating anti-PF4 antibodies that persisted for more than 9 months.     

Antibody-mediated procoagulant platelets in SARS-CoV-2-vaccination associated immune thrombotic thrombocytopenia

The COVID-19 pandemic has resulted in significant morbidity and mortality worldwide. In order to prevent severe infection, mass COVID-19 vaccination campaigns with several vaccine types are currently underway. We report pathological and immunological findings in eight patients who developed vaccine-induced immune thrombotic thrombocytopenia (VITT) after administration of SARS-CoV-2 vaccine ChAdOx1 nCoV-19. We analyzed patient material using enzyme immune assays, flow cytometry and heparin-induced platelet aggregation assay and performed autopsies on two fatal cases. Eight patients (five females, three males) with a median age of 41.5 years (range, 24-53) were referred to us with suspected thrombotic complications 6 to 20 days after ChAdOx1 nCoV-19 vaccination. All patients had thrombocytopenia at admission. Patients had a median platelet count of 46.5x109/L (range, 8-92). Three had a fatal outcome and five were successfully treated. Autopsies showed arterial and venous thromboses in various organs and the occlusion of glomerular capillaries by hyaline thrombi. Sera from VITT patients contain high-titer antibodies against platelet factor 4 (PF4) (optical density [OD] 2.59±0.64). PF4 antibodies in VITT patients induced significant increase in procoagulant markers (Pselectin and phosphatidylserine externalization) compared to healthy volunteers and healthy vaccinated volunteers. The generation of procoagulant platelets was PF4 and heparin dependent. We demonstrate the contribution of antibody-mediated platelet activation in the pathogenesis of VITT.     

Vaccine-induced immune thrombotic thrombocytopenia

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is primarily a complication of adenoviral vector-based covid-19 vaccination. In VITT, thrombocytopenia and thrombosis mediated by anti-platelet factor 4 (PF4) antibodies can be severe, often characterized by thrombosis at unusual sites such as the cerebral venous sinus and splanchnic circulation. Like in heparin-induced thrombocytopenia (HIT) and spontaneous HIT, VITT antibodies recognize PF4-polyanion complexes and activate PF4-treated platelets but additionally bind to un-complexed PF4, a critical finding that could be leveraged for more specific detection of VITT. Intravenous immunoglobulin and non-heparin-based anticoagulation remain the mainstay of treatment. Second dose/boosters of mRNA covid-19 vaccines appear safe in patients with adenoviral vector-associated VITT. Emerging data is consistent with the possibility that ultra-rare cases of VITT may be seen in the setting of mRNA and virus-like particle (VLP) technology-based vaccinations and until more data is available, it is prudent to consider VITT in the differential diagnosis of all post-vaccine thrombosis and thrombocytopenia reactions.     

Epidemiology of VITT

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a life-threatening syndrome of aggressive thrombosis, often profound thrombocytopenia, and frequently overt disseminated intravascular coagulation. It has been associated with 2 adenovirus vector COVID-19 vaccines: ChAdOx1 nCoV-19 (AstraZeneca) and Ad26.COV2.S (Janssen). Unlike the myriad of other conditions that cause thrombosis and thrombocytopenia, VITT has an important distinguishing feature: affected individuals have platelet activating anti-PF4 antibodies that appear in a predictable time frame following vaccination. The reported incidence of VITT differs between jurisdictions; it is dependent on accurate ascertainment of cases and accurate estimates of the size of the vaccinated population. The incidence ranges from 1 case per 26,500 to 127,3000 first doses of ChAdOx1 nCoV-19 administered. It is estimated at 1 case per 518,181 second doses of ChAdOx1 nCoV-19 administered, and 1 case per 263,000 Ad26.COV2.S doses administered. There are no clear risk factors for VITT, including sex, age, or comorbidities. VITT is a rare event, but its considerable morbidity and mortality merit ongoing pharmacovigilance, and accurate case ascertainment.     

Laboratory testing for VITT antibodies

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a highly prothrombotic disorder that like heparin-induced thrombocytopenia (HIT) is caused by platelet-activating antibodies that recognize platelet factor 4 (PF4). However, unlike HIT—where heparin at low concentrations (0.1-0.5 U/mL) typically enhances antibody-induced platelet activation, platelet activation by VITT sera is usually inhibited by heparin. Further, conventional platelet activation assays for HIT, such as the serotonin-release assay (SRA) and heparin-induced platelet activation (HIPA) test, often yield negative or atypical results when testing VITT sera. Nevertheless, VITT (like HIT) is a “clinical-pathological” disorder whereby laboratory detectability of platelet-activating anti-PF4 antibodies is crucial for diagnosis. VITT antibodies follow 2 fundamental principles of HIT laboratory testing: (1) high probability of a positive PF4-dependent enzyme-immunoassay (EIA), and (2) high probability of a positive platelet activation assay. However, optimal detection of VITT in platelet activation assays requires the addition of PF4, for example, PF4-enhanced SRA (PF4-SRA) and PF4-enhanced HIPA (PIPA). A novel whole blood assay, called the PF4-induced flow cytometry-based platelet activation (PIFPA) assay, exhibits high sensitivity and specificity for VITT. HIT and VITT sera/plasmas differ in their reactivity in rapid HIT immunoassays (90-97% sensitivity for HIT, <25% sensitivity for VITT), consistent with distinct antigen sites on PF4 recognized by HIT and VITT antibodies.     

Platelet factor 4 triggers thrombo-inflammation by bridging innate and adaptive immunity

Platelet factor 4 (PF4, synonym: CXCL4) is an evolutionary old chemokine with proposed roles in hemostasis and antimicrobial defense. In addition, PF4 has attracted considerable attention as a crucial mediator of one of the most prothrombotic adverse drug effects affecting blood cells, heparin-induced thrombocytopenia (HIT). Interest in PF4 substantially increased in 2021 when it was identified as the target antigen in the life-threatening adverse effect, vaccine-induced immune thrombotic thrombocytopenia (VITT). We address the concept that a major biological function of PF4—a strongly cationic chemokine—is to bind to negatively-charged prokaryotic microorganisms, resulting in structural changes in PF4 that trigger a danger signal recognized by the adaptive immune system. Application of biophysical tools has provided substantial insights into the molecular mechanisms by which PF4 becomes immunogenic, providing insights into a new mechanism of autoimmunity. Binding of autoantibodies with high affinity induces conformational change(s) in the endogenous protein, which are then recognized as foreign antigen, as exemplified by the prothrombotic disorders, autoimmune HIT and VITT. The final part of our review summarizes current assays for HIT and VITT, explaining how structural aspects of anti-PF4 pathobiology relate to assay design and performance characteristics. Currently, functional (platelet activation) assays using washed platelets detect HIT antibodies when heparin is added, and VITT antibodies when PF4 is added. Solid-phase PF4-dependent immunoassays using microtiter plates are sensitive for both HIT and VITT antibodies, while rapid immunoassays, in which the PF4/heparin antigen is coated on beads, are sensitive and specific for HIT, but not for VITT antibodies.     

Pathogenesis of vaccine-induced immune thrombotic thrombocytopenia (VITT)

Vaccine-induced immune thrombotic thrombocytopenia (VITT; synonym, thrombosis with thrombocytopenia syndrome, is associated with high-titer immunoglobulin G antibodies directed against platelet factor 4 (PF4). These antibodies activate platelets via platelet FcγIIa receptors, with platelet activation greatly enhanced by PF4. Here we summarize the current concepts in the pathogenesis of VITT. We first address parallels between heparin-induced thrombocytopenia and VITT, and provide recent findings on binding of PF4 to adenovirus particles and non-assembled adenovirus proteins in the 2 adenovirus vector-based COVID-19 vaccines, ChAdOx1 nCoV-19 and Ad26.COV2.S. Further, we discuss the potential role of vaccine constituents such as glycosaminoglycans, EDTA, polysorbate 80, human cell-line proteins and nucleotides as potential binding partners of PF4. The immune response towards PF4 in VITT is likely triggered by a proinflammatory milieu. Human cell-line proteins, non-assembled virus proteins, and potentially EDTA may contribute to the proinflammatory state. The transient nature of the immune response towards PF4 in VITT makes it likely that—as in heparin-induced thrombocytopenia —marginal zone B cells are key for antibody production. Once high-titer anti-PF4 antibodies have been formed 5 to 20 days after vaccination, they activate platelets and granulocytes. Activated granulocytes undergo NETosis and the released DNA also forms complexes with PF4, which fuels the Fcγ receptor-dependent cell activation process, ultimately leading to massive thrombin generation. Finally, we summarize our initial observations indicating that VITT-like antibodies might also be present in rare patients with recurrent venous and arterial thrombotic complications, independent of vaccination.