小鼠皮肤移植耐受诱导及微嵌合体检测

目的　探求简便易行、安全有效、有临床应用前景的移植耐受诱导方法，研究微嵌合体的形成情况及其与移植耐受的关系。方法　雄性ＢＡＬＢ／ｃ 和雌性Ｃ５７ＢＬ／６ 小鼠作为皮肤移植的供体和受体，移植前对受体小鼠单独或联合应用５Ｇｙ（５００ｒａｄ ，１Ｇｙ ＝ １００ｒａｄ） 全身照射，０ ．１ ｍｌ 抗淋巴细胞血清注射，３ ×１０７ 供体骨髓细胞静脉输注，１５０ｍｇ／ｋｇ 体重环磷酰胺腹腔注射。应用多聚酶链式反应（ＰＣＲ） 检测微嵌合体的形成。结果　单纯照射不能延长移植物的存活，也没有微嵌合体形成，而其它各实验组的皮肤移植物存活时间均较对照组明显延长，混合淋巴细胞反应和迟发型超敏反应也表现为供者特异性的降低，多种组织的微嵌合体检查均呈阳性。结论　采用比较温和的非照射的预处理方法，也可获得一定程度的免疫耐受，达到延长移植物存活的目的；移植前输注供体骨髓细胞能够促进微嵌合体的形成及移植物的存活。     

023 供体骨髓输注在肾脏移植耐受中的疗效和机制

同种移植免疫耐受的诱导和维持是控制移植后排斥反应,提高移植物存活的关键.动物实验及临床资料表明骨髓输注具有诱导免疫耐受的作用,其机制包括诱导T细胞无能,克隆清除,提供可溶性HLA抗原、抑制细胞及否决细胞,调节细胞表面分子及细胞因子,促进嵌合体的形成.因此,供体骨髓输注在诱导移植免疫耐受中具有相当的应用前景.     

肾移植受者游离血浆中微嵌合体与移植肾免疫耐受的相关性研究

目的:研究在移植受者血液标本中供者和受者源性DNA嵌合体存在是否可以作为预测移植器官免疫状态的一个标准.方法:126名获得男性肾脏的女性移植受者被纳入该项研究,通过RT-PCR方法检测这些女性受者血浆中Y染色体的特异基因序列SRY1,DYZ11st,DYZ12nd.结果:126名获男性肾脏的女性肾移植受者中,97名(77%)血浆中可检测到SRY1,DYZ11st,DYZ12nd序列.移植肾功能维持正常的平均持续时间,在微嵌合体阳性组及微嵌合体阴性组分别为8.7年和5.4年;肾移植术后急性排斥反应发生率在微嵌合体阳性组和阴性组分别为10%和28%;微嵌合体阳性病人血清肌酐水平明显低于微嵌合体阴性病人.结论:肾移植术后某些受者血浆中存在DNA微嵌合体,通过使用定量RT-PCR检测血浆中DNA微嵌合体可能成为衡量移植肾耐受状态的一个预测指标.     

骨髓干细胞输注联合低剂量他克莫司在肾移植中诱导嵌合体的形成

背景:骨髓细胞嵌合体通过有效的免疫耐受机制,使其成为临床用于移植免疫耐受的主要候选方法。目的:探讨大鼠肾移植及活体肾移植中骨髓干细胞移植对免疫耐受的诱导作用及机制。方法:①大鼠肾移植:40只Wistar大鼠(雌雄各半)随机分为实验组和对照组,进行肾移植手术,术前经门静脉分别注射骨髓干细胞及生理盐水,30d后观察肉眼和光镜下排斥反应及检测嵌合体的形成。②活体肾移植:实验组为夫妻或亲属共5对,供者均为男性,受者为女性。术前应用他克莫司口服,供者皮下注射生白细胞药物,采集白细胞悬浊液,以流式细胞仪测定CD34细胞比例,0.7%以上为骨髓干细胞含量合格,并通过外周静脉输注至受者体内,10d后同法再输注1次,1周内行移植手术,术后复查肾功,于半年后采用荧光原位杂交的方法检测Y基因片段。对照组为正常活体肾移植手术,未行骨髓干细胞移植。结果与结论:动物实验组大体和光镜下排斥反应均较轻微,嵌合体检测均为阳性,活体实验组嵌合体检测4例为阳性,而对照组仅1例为阳性。提示应用他克莫司并输注供者骨髓干细胞,可诱导一定的免疫耐受。移植前输注供者骨髓干细胞形成的嵌合体与免疫耐受有关。     

配偶供肾移植后微嵌合体发生与急性排斥反应

背景：微嵌合作为移植物与受者之间的双相细胞移动的标志,在移植免疫耐受中的作用日益受到重视。 目的：探讨夫妻生活与嵌合体的发生,与肾移植后急性排斥反应及其他相关性的研究。 方法：将接受肾脏移植的女性受者(有过生育史的女性除外)分为丈夫活体供肾组、无关男性尸体供肾组,并设立接受妻子活体供肾的对照组。STR方法检测女性受者体内男性供者来源的Y染色体反映微嵌合体的存在,与急性排斥反应发生的关系,并比较配偶间供肾效果的差异。 结果与结论：尽管配偶间供肾移植存在供者年龄偏大以及人类白细胞抗原错配率较高的因素,但与接受无关男性尸肾移植的女性受者相比,接受丈夫活体供肾移植的女性更易检测出微嵌合体,而且肾移植后恢复情况好,急性排斥反应发生率低。而与接受妻子供肾的丈夫相比,接受丈夫供肾的妻子肾移植效果好。说明夫妻间长期相处导致女性接受丈夫体液的机会多,由此产生免疫耐受对于肾移植后人/肾的相容性好,急性排斥反应小。     

供体骨髓输注在肾脏移植耐受中的疗效和机制

同种移植免疫耐受的诱导和维持是控制移植后排斥反应 ,提高移植物存活的关键。动物实验及临床资料表明 :骨髓输注具有诱导免疫耐受的作用 ,其机制包括诱导T细胞无能 ,克隆清除 ,提供可溶性HLA抗原、抑制细胞及否决细胞 ,调节细胞表面分子及细胞因子 ,促进嵌合体的形成。因此 ,供体骨髓输注在诱导移植免疫耐受中具有相当的应用前景     

反向骨髓移植诱导异种造血嵌合体的研究

目的应用大鼠-小鼠骨髓移植模型,探讨反向骨髓移植诱导特异性免疫耐受的作用机制。方法首先诱导形成嵌合体S大鼠,再将此嵌合体大鼠的骨髓反向移植给Balb／c小鼠,监测存活率,GVHD发病率,做混合淋巴细胞培养,检测脾细胞增殖及TNF-α、INF-γ与IL-4产量,监测移植后小鼠嵌合率的变化。结果对照组小鼠观察期间出现严重GVHD表现,反向移植组小鼠仅出现轻度GVHD,生存期明显延长,TNF-α与INF-γ产量明显降低,而IL-4升高,移植后嵌合率较稳定,与对照组相比有显著性差异。结论有受者嵌合体的骨髓移植可以减轻急性GVHD,诱导出较长时间存在的异种造血嵌合体。     

移值免疫耐受研究的现状

探索组织器官移植后的排斥反应,提高移植物的存活率是移植免疫致力攻克的堡垒。在众多的研究中,诱导受体产生对同种的特异性免疫耐受则是最理想的途径。本文将对主动免疫诱导同种耐受,阻断同种免疫应答诱导对移植物的耐受及建立受体体内微嵌合体诱导移植耐受三方面的研究作一综述。     

供体骨髓输注在肾脏移植耐受中的疗效和机制

同种移植免疫耐受的诱导和维持是控制移植后排斥反应，提高移植物存活的关键。动物实验及临床资料表明：骨髓输注具有诱导免疫耐受的作用。其机制包括诱导T细胞无能。克隆清除。提供可溶性HLA抗原，抑制细胞及否决细胞。调节细胞表面分子及细胞因子，促进嵌合体的形成。因此，供体骨髓输注在诱导移植免疫耐受中具有相当的应用前景。     

移植免疫耐受研究的现状

探索组织器官移植后的排斥反应，提高移植物的存活率是移植免疫致力攻克的堡垒。在众多的研究中，诱导受体产生对同种的特异性免疫耐受则是最理想的途径。本文将对主动免疫诱导同种耐受，阻断同种免疫应答诱导对移植物的耐受及建立受体内微嵌合体诱导移植耐受三方面的研究作一综述。     

Differential dynamics of donor DC and non-DC peripheral blood mononuclear cell microchimerism in lung transplantation

Donor cell microchimerism induces tolerance in animal models and may increase graft survival in man. Since dendritic cells (DC) are critical for induction of both tolerance and alloreactivity we developed a method to quantitate microchimerism in donor DC and non-DC in peripheral blood mononuclear cells (PBMC) after lung transplantation. Longitudinal analysis of donor cell microchimerism in eleven sex mismatched lung transplant recipients (LTR) up to 12 months post-transplant used Y chromosome based real-time PCR on sorted cells. Total DC or a proportion of DC subsets in PBMC did not change but there were heterogeneous and dynamic changes in microchimerism in DC and non-DC. Analysis of changes in DC using a mixed model analysis showed significantly less reduction in DC compared to non-DC over time (0.49, p = 0.001). Preferential DC persistence compared to non-DC may indicate tolerance induction but future studies are required to determine if DC microchimerism after transplantation affects clinical outcomes.     

Fetal-maternal microchimerism: impact on hematopoietic stem cell transplantation

Reciprocal cell traffic between mother and fetus during pregnancy gives rise to postpartum fetal-maternal lymphohematopoietic microchimerism, which is frequently detected in blood or tissue from healthy individuals. Although such microchimerism has been implicated in the pathogenesis of autoimmune diseases and tissue repair, recent clinical experiences have suggested the association of microchimerism with acquired immunologic hyporesponsiveness to non-inherited maternal HLA antigens (NIMAs) or inherited paternal HLA antigens (IPAs); T cell-replete HLA-haploidentical hematopoietic stem cell transplantation from a microchimeric IPA/NIMA-mismatched donor confers relatively lower incidence of severe graft-versus-host disease. The underlying mechanisms by which fetal-maternal microchimerism contributes to IPA/NIMA-specific tolerance are still elusive, although emerging experimental evidence suggests an involvement of the central deletion of IPA/NIMA-reactive T cells, the induction of peripheral regulatory T cells, and affinity-dependent modulation of NIMA-reactive B cells.     

Gender as risk factor for autoimmune diseases

Most autoimmune diseases occur significantly more frequently in women than men. This female preponderance for abnormal autoimmune function has largely gone unexplained. Many investigations have concentrated on the effects of female and male sex hormones on immune function, by suggesting that estrogens favor the antibody production-enhancing Th2 response and, by doing so, possibly, increase the risk towards abnormal autoimmune function. Others have suggested that women are genetically predisposed towards abnormal autoimmune function, possibly because the X chromosome may confer susceptibility towards tolerance breakdown. Recent developments have, however, opened new research avenues. The possible association between persistent fetal-maternal microchimerism and the development of autoimmune diseases has attracted special interest. Since, in analogy to allogeneic organ transplantation, fetal-maternal (and maternal-fetal) microchimerism may play an important role in the immunologic tolerance of the fetal semi-allograft, female preponderance for autoimmune diseases may be understood as a consequence of increased allogeneic cell traffic in females (in comparison to males), increased risk for long-term microchimerism and, therefore, as a consequence of the former two, the development of abnormal autoimmunity. Under an evolutionary view point the occurrence of autoimmune diseases, in general, can be seen as the price to be paid for successful reproduction. In view of increased exposure to cell traffic, women, of course, would be expected to pay a higher price, reflected in more autoimmunity.     

Impact of fetal-maternal tolerance in hematopoietic stem cell transplantation

Allogeneic hematopoietic stem cell transplantation (HSCT) is known to cure various hematological disorders; however, its widespread use is limited due to a lack of histocompatible donors. Reciprocal cell traffic between the mother and fetus during pregnancy gives rise to postpartum fetal-maternal lymphohematopoietic microchimerism, which is frequently detected in the blood or tissue of healthy individuals. Studies in clinical and experimental transplantation provide evidence that exposure to non-inherited maternal antigens (NIMAs) during pregnancy may result in long-lasting fetomaternal microchimerism and tolerance induction. Studies of HLA-mismatched HSCT have suggested a relatively lower incidence of severe graft-versus-host disease (GVHD) after transplantation from a NIMA-mismatched donor. Studies using a mouse model have also demonstrated a “child-to-mother” bone marrow transplantation from an NIMA-exposed donor to reduce the morbidity and mortality of GVHD in an antigen-specific manner while preserving the graft-versus-leukemia effects and favoring the immune reconstitution, thus resulting in a marked improvement in outcome after HSCT. Prospective clinical studies are therefore warranted to confirm these beneficial effects of fetal-maternal tolerance in allogeneic HSCT.     

Maternal microchimerism—Allogenic target of autoimmune disease or Normal Biology?

Chimerism is the state of cells from two distinct individuals living within one body. Fetal cells pass into a mother during pregnancy, where they may persist at low levels for years, creating a state of fetal microchimerism. At the same time, maternal cells pass into the fetus, leading to maternal microchimerism that can persist into adulthood. Hematopoietic stem cell transplantation also creates a state of chimerism, and can lead to a complication of chronic multi-organ inflammation called graft-versus-host disease, (GVHD). The similarities between GVHD and some autoimmune diseases like scleroderma, lupus and myositis suggest that chimerism may be involved in the pathogenesis of both. Maternal and fetal microchimerism in the blood and in tissues have been associated with autoimmune diseases. However, many healthy individuals harbor maternal and fetal cells. Human and animal studies have begun to elucidate the mechanisms for normal tolerance to maternal and fetal microchimeric cells, and how this tolerance may be broken in states of chronic inflammatory disease.     

Donor cell chimerism permitted by immunosuppressive drugs: a new view of organ transplantation

On line of thought in organ transplantation feels that immunosuppressive drugs can lead to tolerance induction by allowing a previously unrecognized common mechanism of cell migration and microchimerism to occur perist, and in some cases, become drug independent. It has been recognized that there is a spectrum of susceptibility of different organs to cellular rejection and that the variable ability of these organs to induce donor-specific nonreactivity reflects their comparative content of migratory leukocytes. Here, Thomas Starzl and colleagues discuss how many of the enigmas of transplantation immunology can be explained by this chimerism.     

Microchimerism in Human Health and Disease

During pregnancy some cells traffic between the fetus and the mother. Recent investigative work indicates a low level of fetal cells commonly persists in the maternal circulation for years, or even indefinitely, after pregnancy has been completed. The term microchimerism refers to one individual harboring DNA or cells at a low level that derive from another individual. Chronic graft-versus-host disease (cGvHD) shares similarities with some autoimmune diseases and is an iatrogenic form of chimerism, occurring as a complication of hematopoietic stem cell transplantation. The HLA genes of the donor and the host are known to be of central importance to the development of cGvHD. When also considered in light of the female predilection to autoimmunity, these series of observations led to the hypothesis that microchimerism and HLA genes of host and non-host cells are involved in some autoimmune diseases. The hypothesis can also apply to men, children, and women who have not been pregnant because there are other sources of microchimerism. Persistent microchimerism can follow a blood transfusion, or can occur from transfer between twins in utereo. Additionally, maternal cells have recently been found to persist in her immune competent progeny. A number of studies have investigated a potential role of microchimerism in human diseases including systemic sclerosis (SSc), primary biliary cirrhosis (PBC), Sjögren's syndrome, polymorphic eruption of pregnancy, myositis, and thyroid disease. While some studies lend support to the concept that microchimerism is involved in the pathogenesis of selected autoimmune diseases, studies also indicate microchimerism is not uncommon in other human conditions and in healthy individuals.     

Gene therapy in the transplantation of allogeneic organs and stem cells

Most of the current hematopoietic stem cell (HSC) -directed gene therapy applications have focused on the replacement of defective or deficient genes in an autologous setting. More recently HSC gene therapy applications have also included the enhancement or improvement of HSC features. Allogeneic HSCs have been used to facilitate and improve allogeneic transplantation and to achieve tolerance to transplanted cells, tissues or organs. Different gene transfer approaches addressing a variety of immunomodulatory mediators contributing to graft tolerance or immunological ignorance may have a critical role in improving long-term graft survival. Allogeneic tissues are frequently recognized by allospecific T cells as foreign and are rapidly rejected in the absence of immunosuppression. The higher susceptibility to cancer and infectious diseases of immunosuppressed patients led to investigation of new therapies to induce graft-specific tolerance. Peripheral tolerance to allogeneic grafts can be achieved by a variety of mechanisms including clonal deletion, suppression caused by regulatory T cells and anergy induction associated with microchimerism effect. In the last decades, potential candidates to confer allograft protection were identified. In this review, we summarize ongoing strategies and developments in genetic manipulation of cells, tissues and organs for allogeneic transplantation including modulating the effector arm of the immune response.     

Microchimerism in human health and disease

During pregnancy some cells traffic between the fetus and the mother. Recent investigative work indicates a low level of fetal cells commonly persists in the maternal circulation for years, oreven indefinitely, after pregnancy has been completed. The term microchimerism refers to one individual harboring DNA or cells at a low level that derive from another individual. Chronic graft-versus-host disease (cGvHD) shares similarities with some autoimmune diseases and is an iatrogenic form of chimerism, occurring as a complication of hematopoietic stem cell transplantation. The HLA genes of the donor and the host are known to be of central importance to the development of cGvHD. When also considered in light of the female predilection to autoimmunity, these series of observations led to the hypothesis that microchimerism and HLA genes of host and non-host cells are involved in some autoimmune diseases. The hypothesis can also apply to men, children, and women who have not been pregnant because there are other sources of microchimerism. Persistent microchimerism can follow a blood transfusion, or can occur from transfer between twins in utereo. Additionally, maternal cells have recently been found to persist in her immune competent progeny. A number of studies have investigated a potential role of microchimerism in human diseases including systemic sclerosis (SSc), primary biliary cirrhosis (PBC), Sj?gren's syndrome, polymorphic eruption of pregnancy, myositis, and thyroid disease. While some studies lend support to the concept that microchimerism is involved in the pathogenesis of selected autoimmune diseases, studies also indicate microchimerism is not uncommon in other human conditions and in healthy individuals.