肾移植受者血清多反应性抗体与HLA抗体的相关性及对移植肾长期存活的影响

发布时间：2018-07-28 12:46

【摘要】：抗体介导的移植物排异反应是导致移植物失功的主要原因，也是影响移植肾长期存活的关键性因素。目前广泛认为，器官移植后移植物发生抗体介导排异反应及移植物失功主要与受者DSA的产生密切相关，而NDSA及非HLA抗体对移植物存活的影响尚未得到足够重视。有文献报道，器官移植受者血清中未产生DSA的情况下，仍然存在针对移植物的抗体介导的排异反应[4,5]。另有研究报道，器官移植患者体内诸如血管紧张素受体、抗波形蛋白抗体、抗过氧化脂类抗体等诸多自身抗体与移植物的排异反应密切相关并影响移植物长期存活[7-10]。综上所述，除了供体特异性抗体之外，非供体特异性HLA抗体及非HLA抗体也是影响移植物存活的重要因素。器官移植致敏后产生的抗体主要为特异性抗供者HLA的抗体，然而在研究中发现，肾移植受者血清中亦可检测出NDSA的出现[6,11-13]，在无特异性免疫致敏的前提下，NDSA产生的原因尚不完全明确。目前普遍接受的解释为交叉反应原理[14-17]，即由于部分HLA之间存在共用的抗原决定簇，故肾移植术后患者血清中产生特异性抗供者HLA抗体的同时，也可以与供者表达的HLA相似的抗原发生反应，从而表现为NDSA的出现。然而有研究发现，肾移植患者在没有产生DSA的情况下也可检测出NDSA，肾移植术后NDSA出现的平均中位时间甚至早于DSA[14]。此种现象显然不能用现有的交叉反应理论来解释。自然抗体（Natural antibodies）是一类未经免疫致敏而广泛存在于人类血清中的抗体[18]。自然抗体由于能够与抗原决定簇毫不相关的多种抗原均发生反应，故表现出多反应性的特点，从而在人类先天免疫中发挥着重要的作用。自然抗体的重要特性是能够与凋亡细胞结合[19-22]。长期以来，对多反应性抗体的研究主要集中于人类自身免疫性疾病领域，而针对多反应性抗体在器官移植领域的研究却罕有报道。鉴于多反应性抗体的特性，在本研究中，我们分别从单克隆B细胞、抗体水平以及肾移植患者血清学水平探寻多反应性抗体与HLA抗体产生的相关性。同时研究肾移植受者血清内预存的多反应性抗体与抗体介导移植肾排异反应的相关性及对移植肾长期存活的影响。结果如下： 1、多反应性抗体与HLA抗体产生的相关性。本研究中，我们将B淋巴细胞从肾移植受者外周血及手术切除的移植肾中分选出来并在体外永生化，成功培育出单克隆B淋巴细胞克隆。通过检测发现所有B淋巴细胞克隆均分泌抗体，部分单克隆抗体可以与多种抗原诸如LPS、dsDNA、胰岛素、细胞裂解产物等发生抗原抗体反应，从而揭示此类单克隆抗体具有多反应性的特点，有别于“一把钥匙开一把锁”的特异性抗原抗体反应。此外，我们通过研究发现，，多反应性抗体能够与凋亡细胞结合。值得关注的是，我们发现部分多反应性的单克隆抗体能够与多种HLA抗原发生反应，从而在单克隆抗体水平发现多反应性抗体与HLA抗体产生的相关性。当将研究扩展至患者血清学水平，我们发现，与HLA抗体阴性的患者相比，HLA抗体阳性的患者血清内多反应性抗体活性显著升高（p=0.009，p0.001，p0.001）。应用Spearman相关性分析发现300例肾移植患者血清中多反应性抗体活性与HLA抗体活性之间存在显著相关性（p=0.004，p0.001，p=0.019）。在HLAI类抗体阳性的患者中，与血清多反应性抗体活性低的患者相比，血清多反应性抗体活性高的患者能够与更多的HLAI类抗原发生反应（p=0.030），因而从血清学水平揭示了多反应性抗体与HLA抗体的相关性。 2、肾移植受者术前血清多反应性抗体与移植肾抗体介导排异反应及移植肾长期存活的相关性。我们通过对300例肾移植患者血清标本研究发现，肾移植术前患者血清多反应性抗体IgG的活性较正常人群显著性升高（p=0.011），而这种差异并非由于肾移植患者及正常人群血清中IgG抗体浓度的不同所致。在此项回顾性研究中，300例患者随访时间为81.2±35.3个月。46例患者在观察期内出现移植肾失功，我们通过研究发现移植肾失功的患者术前血清多反应性IgG活性显著高于移植肾功能正常的患者（p0.001）。而且在46例移植肾失功的患者中，因抗体介导排异反应导致移植肾失功的患者血清多反应性IgG活性显著高于因其他原因导致移植肾失功的患者（p=0.033)。Kaplan-Meier生存曲线显示，移植前血清中多反应性抗体活性高的患者移植肾存活率显著降低（p0.001），而多反应性IgG独立于HLA抗体之外对移植肾存活产生影响。进一步行Cox比例风险模型分析显示，血清多反应性抗体IgG活性是移植肾失功的独立危险因素（Hazard ratio=2.271，p0.001）。本研究发现，肾移植患者血清多反应性IgG抗体对凋亡细胞的反应主要由IgG1亚型及IgG3亚型介导。多反应性抗体与凋亡细胞反应后可激活补体途径，致使C4d沉积于靶细胞表面。进一步分析显示，IgG1及IgG3亚型对凋亡细胞的反应性与C4d在靶细胞表面沉积强度呈显著相关性（p0.001，p=0.005）。本研究分别从单克隆抗体水平及肾移植受者血清学水平研究发现多反应性抗体与HLA抗体的产生具有显著相关性，在交叉反应理论之外，为器官移植患者HLA抗体尤其是NDSA的产生原因提供新的依据；并发现多反应性抗体IgG与肾移植术后抗体介导的排异反应显著相关，并成为移植物失功的独立危险因素。本研究为肾移植患者术前风险评估提供了新的依据。在器官移植术前除常规检测HLA抗体之外，对患者血清多反应性抗体的监测有利于提高器官移植后移植物失功风险评估的准确性，并为器官移植术前高风险患者的去抗体治疗提供了新的理论及实验依据。
[Abstract]:Antibody mediated graft rejection is the main cause of graft dysfunction and is also a key factor affecting the long-term survival of the transplanted kidney. It is widely believed that antibody mediated rejection and graft dysfunction after organ transplantation are closely related to the production of DSA, while NDSA and non HLA antibodies are responsible for graft survival. It has not been paid much attention to. It is reported that there is still another study on antibody mediated rejection mediated by antibodies against graft in the absence of DSA in the serum of organ transplant recipients. [4,5]., such as angiotensin receptor, anti wave protein antibody, anti oxidative lipid antibody, and so on in the body transplant patients. Antibodies are closely related to graft rejection and affect the long-term survival of the grafts in [7-10].. In addition to donor specific antibodies, non donor specific HLA antibodies and non HLA antibodies are also important factors affecting graft survival.
The antibody produced by organ transplant sensitization is mainly a specific anti donor HLA antibody. However, in the study, it is found that the serum of NDSA can also detect the emergence of [6,11-13] in the serum of the renal transplant recipients. The reasons for the production of NDSA are not completely clear on the premise of no specific immuno sensitization. The commonly accepted interpretation is the principle of cross reaction [14-17], That is, due to the existence of a common antigenic determinant among some of the HLA, the specific donor HLA antibody in the serum of the patients after renal transplantation can also be reacted with the HLA similar antigen expressed by the donor, thus showing the appearance of NDSA. However, some studies have found that the renal transplant patients can also be detected without DSA. Out of NDSA, the average median time of NDSA after renal transplantation was even earlier than that of DSA[14].. This phenomenon obviously can not be explained by the existing cross reaction theory.
Natural antibody (Natural antibodies) is a kind of unimmunized and widely existed antibody [18]. natural antibody response to various antigens that are not related to antigenic determinants. Therefore, it is characterized by multi reactivity and plays an important role in human innate immunity. The feature is to be able to combine [19-22]. with apoptotic cells
For a long time, the study of multi reactive antibodies is mainly concentrated in the field of human autoimmune diseases. However, the study of multi reactive antibody in the field of organ transplantation is rare. In view of the characteristics of the multi reactive antibody, we have explored the level of monoclonal B, the level of antibody and the serological level of renal transplant patients. The correlation between multiple reactive antibodies and the production of HLA antibodies. The correlation between the prestored multiple reactive antibodies in the renal transplant recipients and the antibody mediated rejection of the renal allograft and the long-term survival of the renal allograft were investigated.
1. Correlation between polyreactive antibody and HLA antibody production.
In this study, we selected and immortalized B lymphocytes from peripheral blood and surgical excised kidneys of renal transplant recipients and successfully produced monoclonal B lymphocyte clones. Through detection, we found that all B lymphocyte clones secreted antibodies, and some monoclonal anti bodies could be associated with a variety of antigens such as LPS, dsDNA, insulin, The antigen antibody reaction of the cell lysis product, which reveals the multi reactivity characteristic of the monoclonal antibody, is different from the specific antigen antibody reaction of "a key opening a lock". In addition, we have found that the multi reactive antibody can combine with the apoptotic cells. The specific monoclonal antibodies can react with a variety of HLA antigens, thus finding the correlation between the multi reactive antibody and the HLA antibody at the monoclonal antibody level. When the study was extended to the serological level of the patient, we found that the activity of the serum multi reactive antibody in the patients with HLA antibody positive was significantly higher than that of the patients with the negative HLA antibody. Increase (p=0.009, p0.001, p0.001). Application of Spearman correlation analysis found that there was a significant correlation between the activity of multi reactive antibody and the activity of HLA antibody in 300 patients with renal transplantation (p=0.004, p0.001, p=0.019). In patients with positive HLAI antibody, serum multi reactivity resistance was compared with those with low serum reactivity antibody. Patients with high body activity can react with more HLAI class antigens (p=0.030), thus revealing the correlation between reactive antibodies and HLA antibodies from serological level.
2, the correlation between preoperative serum reactivity antibody and renal transplant antibody rejection and long term survival of renal transplant recipients.
In 300 cases of renal transplant patients, we found that the activity of serum multi reactive antibody IgG in patients with renal transplantation was significantly higher than that in the normal population (p=0.011), and this difference was not due to the difference in the serum level of IgG in the renal transplant patients and the normal population. In this retrospective study, 300 patients followed. The time of visits was 81.2 + 35.3 months after the observation period of.46 patients with renal allograft dysfunction. We found that the serum reactivity IgG activity in patients with renal allograft dysfunction was significantly higher than that of normal renal transplantation (p0.001). And in 46 patients with renal allograft dysfunction, antibody mediated rejection resulted in renal allograft dysfunction. The patients' serum reactivity IgG activity was significantly higher than that of patients with other causes of graft dysfunction (p=0.033).Kaplan-Meier survival curve. The survival rate of renal allograft in patients with high reactive antibody activity before transplantation was significantly lower (p0.001), while multi reactive IgG was independent of HLA antibody to the survival of transplanted kidney. Further analysis of Cox proportional hazards model showed that IgG activity of serum reactive reactivity was an independent risk factor for renal allograft dysfunction (Hazard ratio=2.271, p0.001).
This study found that the reaction of serum multi reactive IgG antibody to apoptotic cells in renal transplantation patients is mainly mediated by IgG1 subtype and IgG3 subtype. The reaction of multi reactive antibody and apoptotic cells can activate complement pathway and cause C4d to be deposited on the surface of target cells. Further analysis shows that the responsiveness of IgG1 and IgG3 subtypes to the apoptotic cells and C4d are in the target. There was a significant correlation between cell surface deposition intensity (p0.001, p=0.005).
In this study, we found a significant correlation between the production of multiple reactive antibodies and the production of HLA antibodies from the level of monoclonal antibodies and the serological level of renal transplant recipients. In addition to the theory of cross reaction, a new basis for the cause of HLA antibody, especially NDSA, in organ transplant patients was provided, and the multiple reactive antibody IgG and renal transplantation were found. This study provides a new basis for the preoperative risk assessment of renal transplant recipients. In addition to the routine detection of HLA antibodies before organ transplantation, the monitoring of serum multi reactive antibodies is beneficial to the improvement of graft dysfunction after organ transplantation. The accuracy of the risk assessment provides a new theoretical and experimental basis for anti antibody therapy in high-risk patients before organ transplantation.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R699.2

【共引文献】