Reichenspurner H, Soni V, Nitschke M, Berry G J, Brazelton T R, Shorthouse R, Huang X, Reitz B A, Morris R E
Transplantation Immunology and Department of Cardiothoracic Surgery, Stanford University School of Medicine, California 94305, USA.
Transplantation. 1997 Aug 15;64(3):373-83. doi: 10.1097/00007890-199708150-00001.
The purpose of this study was to investigate whether obliterative bronchiolitis might occur after xenogenic pulmonary transplantation. A model for obliterative airway disease (OAD) after tracheal allograft transplantation in the rat undergoes tracheal obliteration with histologic features characteristic of obliterative bronchiolitis in human lung transplant recipients. Using this model, the pathogenesis of OAD and its prevention with immunosuppressive drugs was studied in rat recipients of hamster tracheal grafts.
Tracheae from 30 hamsters (xenografts) or 23 Brown-Norway rats (allografts) were implanted and wrapped in the greater omentum of untreated Lewis rats. The grafts were removed on day 1, 3, 7, 14, 21, or 28 after transplantation and stained with hematoxylin and eosin and Masson's trichrome and by immunohistochemistry and immunofluorescence (IFL) techniques. In addition, 25 recipients were treated with cyclosporine (CsA, 10 mg/kg p.o.), leflunomide (LFM, 20 mg/kg p.o.), or rapamycin (RPM, 6 mg/kg i.p.) for 14 or 21 days (5 animals per treatment group). Visual and morphometric analyses were used to evaluate the extent of airway obliteration, luminal coverage by respiratory or flattened cuboidal epithelium, and extent and density of peritracheal cellular inflammation.
In all xenografts, a neutrophilic infiltration of the mucosa and submucosa was observed from day 1 until day 14 and was associated with complete loss of tracheal epithelium by day 14. A marked peritracheal mononuclear cellular infiltrate mixed with plasma cells and eosinophils was seen on days 7 and 14. Both the extent of peritracheal inflammation and the density of the mononuclear cell infiltrate were significantly increased in xenograft tracheae when compared with the allografts. Tracheal obliteration began on day 14 and reached a maximum of 43% on day 21 with evidence of intraluminal fibrosis. In contrast to IFL of allografts, IFL of xenografts demonstrated marked deposition of rat immunoglobulin in the peritracheal tissue on days 7 and 14. The effects of treatment with immunosuppressive drugs on tracheal graft narrowing and protection of respiratory epithelium were as follows: After 14 days of treatment, the percentage of tracheal graft narrowing was 12%, 23%, and 19% in the no treatment, CsA, and LFM groups, respectively; the percentage of respiratory epithelium at 14 days was 0%, 21%, and 95%. After 21 days of treatment, the percentage of tracheal graft narrowing was 43%, 49%, 12%, and 5% for the no treatment, CsA, LFM, and RPM groups, respectively; the percentage of respiratory epithelium at 21 days was 0%, 39%, 86%, and 0%. Using computerized morphometry, the extent and densities of the peritracheal cellular infiltrates were significantly reduced in LFM- and CsA-treated groups when compared with untreated xenograft controls. LFM and RPM, but not CsA, significantly reduced the degree of luminal obliteration compared with no treatment (P<0.05). LFM and, to a lesser extent, CsA were able to prevent the loss of normal respiratory epithelium. Analysis by IFL revealed a marked decrease in rat immunoglobulin deposition in xenografts from LFM- and RPM-treated groups compared with xenografts from CsA-treated or untreated rats.
(1) OAD occurs not only after tracheal allotransplantation but also after xenotransplantation. (2) Subepithelial infiltration of neutrophils and the appearance of plasma cells and eosinophils in the peritracheal infiltrates distinguished the histology of rejected xenografts from allografts. (3) Antibody deposition was detected by IFL only in xenografts. (4) Treatment with LFM or RPM significantly decreased the severity of luminal obliteration. Importantly, LFM also prevented the loss of respiratory epithelium.
本研究旨在调查异种肺移植后是否会发生闭塞性细支气管炎。大鼠气管同种异体移植后闭塞性气道疾病（OAD）的模型会出现气管闭塞，其组织学特征与人类肺移植受者的闭塞性细支气管炎相似。利用该模型，对仓鼠气管移植大鼠受者的OAD发病机制及其免疫抑制药物预防进行了研究。
将30只仓鼠的气管（异种移植物）或23只棕色挪威大鼠的气管（同种移植物）植入未处理的Lewis大鼠的大网膜并包裹。移植后第1、3、7、14、21或28天取出移植物，用苏木精-伊红染色、Masson三色染色以及免疫组织化学和免疫荧光（IFL）技术进行染色。此外，25只受者用环孢素（CsA，口服10mg/kg）、来氟米特（LFM，口服20mg/kg）或雷帕霉素（RPM，腹腔注射6mg/kg）治疗14或21天（每个治疗组5只动物）。采用视觉和形态计量分析来评估气道闭塞程度、呼吸或扁平立方上皮的管腔覆盖情况以及气管周围细胞炎症的程度和密度。
在所有异种移植物中，从第1天到第14天观察到黏膜和黏膜下层有中性粒细胞浸润，到第14天时气管上皮完全丧失。在第7天和第14天可见明显的气管周围单核细胞浸润，并伴有浆细胞和嗜酸性粒细胞。与同种移植物相比，异种移植气管的气管周围炎症程度和单核细胞浸润密度均显著增加。气管闭塞在第14天开始，第21天达到最大值43%，伴有管腔内纤维化的证据。与同种移植物的IFL不同，异种移植物的IFL显示在第7天和第14天大鼠免疫球蛋白在气管周围组织中有明显沉积。免疫抑制药物治疗对气管移植物狭窄和呼吸上皮保护的影响如下：治疗14天后，未治疗组、CsA组和LFM组气管移植物狭窄百分比分别为12%、23%和19%；14天时呼吸上皮百分比分别为0%、21%和95%。治疗21天后分别为43%、49%、12%和5%；未治疗组、CsA组、LFM组和RPM组21天时呼吸上皮百分比分别为0%、39%、86%和0%。使用计算机形态计量学，与未处理的异种移植对照组相比，LFM和CsA治疗组的气管周围细胞浸润程度和密度显著降低。与未治疗相比，LFM和RPM（但不是CsA）显著降低了管腔闭塞程度（P<0.05）。LFM以及程度较轻的CsA能够防止正常呼吸上皮的丧失。IFL分析显示，与CsA治疗或未治疗大鼠的异种移植物相比，LFM和RPM治疗组的异种移植物中大鼠免疫球蛋白沉积明显减少。
（1）OAD不仅发生在气管同种异体移植后，也发生在异种移植后。（2）中性粒细胞的上皮下浸润以及气管周围浸润中浆细胞和嗜酸性粒细胞的出现区分了异种移植物与同种移植物排斥的组织学表现。（3）仅在异种移植物中通过IFL检测到抗体沉积。（4）LFM或RPM治疗显著降低了管腔闭塞的严重程度。重要的是，LFM还防止了呼吸上皮的丧失。
