大鼠自体移植脾组织再生的形态学研究

目的：探讨大鼠自体移植脾组织血管神经再生过程、VEGF及KDR表达的变化。方法：采用Wistar大鼠105只，行脾切除自体脾组织大网膜内移植术，分别于术后7、14、30、60、90、120、180d，采用免疫组化VEGF、KDR、NPY^ 神经纤维抗体染色方法，应用光镜、透射电镜、图像分析观测自体移植脾组织。结果：自体脾组织移植术后7d即有血管长入移植脾组织，180d再生血管接近正常；术后30d神经开始再生，180d趋于正常；术后7、14dVEGF、KDR阳性染色细胞密度迅速升高，60d达高峰，180d趋于正常；术后30d出现：NPY^ 神经纤维，120、180d NPY^ 神经纤维广泛分布。结论：移植脾组织再生血管神经来源于大网膜；移植脾组织内VEGF、KDR表达量早期升高，血管再生完成时恢复正常水平；移植脾组织血管再生早于神经再生。     

大鼠自体移植脾组织GAP-43+神经再生的实验研究

目的 研究大鼠自体脾组织移植术后移植脾组织GAP-43^ 神经纤维再生及分布规律。方法 健康Wistar大鼠105只，随机分为自体脾组织移植组和对照组，于术后7、15、30、60、90、120、180d取移植脾组织标本，应用免疫组化方法显示GAP-43^ 神经纤维，并用图像分析系统，对不同时相点免疫组化GAP-43^ 染色区域进行图像定量分析。结果 自体脾组织移植术后30d，移植脾组织周围大网膜内可见GAP-43^ 神经纤维，并向移植脾组织内伸展；自体脾组织移植术后60～120d，移植脾组织内再生神经纤维逐渐增多；术后180d，移植脾组织中GAP-43^ 神经纤维分布及密度接近正常。结论GAP-43^ 神经纤维于脾移植术后60d开始出现，术后180dGAP-43^ 神经纤维接近正常脾。     

不同自体脾组织移植量抗肺炎球菌感染的实验研究

随着人们对脾脏各种功能的认识以及脾切除后严重感染和心血管疾病的发生.脾损伤后采用各种保脾术越来越多。当严重脾损伤无法保留脾脏时,很多学者仍主张将自体脾组织再移植,但移植后再生的脾组织结构及功能怎样?能否防止发生“OPSI”?尤其是对脾组织的临界移植量问题,无论在临床或是动物实验方面都存在较大争议。本实验采用大鼠进行不同量脾组织网膜内移植,然后进行抗感染能力和实验研究,试图阐明移植脾组织的抗感染作用。移植多少脾组织才是抗感染的理想量,从而为临床处理严重脾损伤提供理论和实践依据。     

小肠浆肌膜腔内自体脾组织移植动物模型的建立与意义

目的探讨增加自体脾组织移植量的方法,提高自体移植脾组织的功能。方法贵州小型香猪3头,经肌注麻醉后剖腹切除脾脏,取脾脏的1/2约150g,切成1mm×1mm×1mm小块备用。在距Treitz韧带50cm处,切取保留肠系膜血管的空肠30cm旷置,再吻合肠管恢复肠腔通道,缝闭肠系膜裂口。旷置空肠等分2段,去除肠粘膜,缝闭一端,将约50g脾组织块分别植于2段肠浆肌膜腔内,缝闭另一端口,妥善固定于肠系膜上。大网膜内移植30g脾组织作为对照观察。结果4个月饲养期间内无肠梗阻发。3段肠浆肌膜腔内脾组织再生良好,组织结构与大网膜内移植脾组织无明显差别,2段再生较差,1段形成脓肿。结论肠浆肌膜腔内移植脾组织的量较大,可作为自体脾组织的移植术式。     

自体脾组织移植血供变化的实验研究

目的：探讨自体移植脾组织再血管化的形态构筑特点及其变化规律，同时阐明其血供与功能的关系。方法：采用健康Ｗｉｓｔａｒ大鼠，行脾切除半脾大网膜内移植术，定期采用微血管铸型扫描电镜，观察微血管构筑；通过碳素墨水混悬液灌注后脾组织切片，观察再血管化的演变过程；借助ＭＡＳ图像分析测定微血管的面积分数（Ａａ）。结果：自体脾移植术后１８０ｄ，微血管构筑的主要特征是：中央动脉不够清楚，白髓减少，边缘窦微血管网形态不规则，呈球囊状扩张，红髓内静脉窦扩张尤为显著，形成大的“血管湖”。组织学和图像分析Ａａ观察到移植脾组织呈动态变化，经历一个坏死再生、再血管化的过程。结论：自体移植脾组织的再生及再血管化在术后１８０ｄ基本完成，符合形态与功能相一致的规律。     

大网膜内植入自体脾组织与原位脾组织的结构比较

目的 :为临床应用自体脾组织植入术提供实验研究资料。方法 :大鼠分为实验组和对照组 ,前者切取 1 /2脾脏去包膜后切成 1mm× 1mm× 1mm大小均匀组织块 ,植入大网膜囊袋内。饲养 6个月后取 2组脾组织制片 ,光镜和电镜定性观察组织结构变化 ,计算机图像分析系统比较血管、红髓、白髓及胶原纤维的面密度 ;免疫组化法结合计算机图像分析测定神经肽 (NPY)阳性神经纤维密度。结果 :神经和边缘窦内皮细胞结构恢复较好 ,血管 ,白髓的面密度值较对照组减少 ,红髓与对照组相当 ,胶原纤维面密度增加。结论 :大网膜内植入的自体脾组织通过再生能恢复脾脏的主要组织结构 ,但不能完全恢复正常。     

外伤脾切除后自体脾组织移植的临床应用

近年来,脾脏外科有了长足进展,主要标志之一是对脾脏功能有了新的认识,对脾脏功能认识程度的逐步加深又促进了脾脏外科的发展,两者相得益彰,不可或缺。自体脾组织移植(以下简称脾组织移植)在国内外已被接受为脾破裂进行保脾手术的主要措施之一。我院自1999年1月至2000年4月,因外伤脾破裂无法修补,行全脾切除术后自体脾组织移植32例,术后随访观察均获得满意疗效。     

自体移植脾组织内GAP-43神经的生成机制

为探索自体移植脾组织内GAP-43神经的再生机制,将Wistar大鼠42只随机分为实验组和对照组。实验组切除脾脏以后,切取1／2脾脏,切成1 mm×1 mm×1 mm大小的组织块植入大网膜内,术后7、14、30、60、90、120、180 d取脾组织标本通过原位杂交检测GAP-43 mRNA、NGF mRNA和TrkA mRNA,同时进行免疫组化染色观察GAP-43神经。对照组手术松动脾脏,在术后相同时间点取脾组织作对照观察。结果显示:术后30 d检测到移植脾组织内有GAP-43 mRNA、NGF mRNA和TrkA mRNA表达,90 d达高峰后开始下降;术后60 d明显可见GAP~43染色阳性神经纤维,90 d密度最大,主要存在于血管周围,以后无明显改变。结果提示:自体移植脾组织内GAP-43神经再生与内源性NGF和TrkA表达密切相关。     

脾切除和自体脾组织移植对肺吞噬功能的影响

随着免疫学的进展,脾脏抗感染免疫功能日臻重要。脾切除患者对病原易感性增加,有可能发生凶险性感染(OPSI)。据统计其中半数以上是由肺炎球菌通过呼吸道引起的,而脾脏的去留对肺脏抗感染能力的影响怎样,文献上研究得很少,尤其外伤性脾切除后自体移植脾组织能否维持肺的正常防卫能力,从而起到保留脾的作用,国内未见报道。本文就脾切除和自体脾组织移植对肺吞噬功能的影响进行动态观察,以确定其应用价值,为临床提供参考依据。     

外源性VEGF对自体移植脾组织血管再生的影响

目的探讨应用外源性VEGF促进自体脾组织移植后血管再生与结构重建的影响,为VEGF用于自体移植脾组织的实验研究和临床应用提供理论参考依据。方法采用216只昆明种小白鼠随机分为脾切除自体脾组织移植组和假手术组,再将脾移植组分为实验组和对照组。实验组小鼠分别于术后7、14、30、60 d于尾静脉内注射VEGF,注射后7、15、30、60、90 d取脾组织标本。进行组织学观察、血管面密度测定、免疫组化染色方法KDR的检测、KDR阳性细胞密度测定、组织原位杂交技术KDR mRNA检测。结果术后7、15、30 d注射VEGF组注射VEGF后较对照组的血管数量明显增多;术后60 d注射VEGF组注射VEGF后与对照组各时相点的组织学特征没有明显差别;术后7、15、30 d注射VEGF组KDR面积密度值高于对照组,术后60 d注射VEGF组KDR面密度值与对照组比较无显著差别;术后7、15、30 d注射KDR mRNA表达阳性细胞密度高于对照组,术后60 d注射VEGF组KDRmRNA表达阳性细胞密度与对照组比较没有明显差异。结论外源性VEGF能够促进移植脾组织内血管生长,自体脾组织移植术后7~15 d开始静脉应用VEGF是促进血管再生的较理想时间;外源性VEGF可能是通过与移植脾组织内所表达的KDR结合发挥作用。     

Heterotopic autologous splenic grafts in rat

Summary Splenic grafts autotransplanted at splenectomy into the omentum of 23 Porton strain rats were compared with spleens from 10 sham-operated controls. Six months after transplantation, the grafts weighed between 81 to 545 mg (median 166 mg) compared to control spleens which weighed 775 to 1,250 mg (median 995 g). Histoquantitation of the grafts revealed marked reduction of the splenic white pulp when compared to control spleens. Morphological examination showed not only a reduction of lymphocytes but also a striking architectural abnormality in all grafts. In 2 of the transplants, no lymphoid aggregates were observed; small subcapsular collections were present in 7, while in 8, isolated perivascular aggregates of lymphocytes with poorly formed marginal zones were observed. Only 6 transplanted spleens showed linkage of adjacent lymphoid aggregates but the number and size of these aggregates were clearly less than normal. These findings indicate that autotransplanted splenic tissue in rats does not regain histological normality. The implications of these observations for autotransplantation in splenectomized patients are discussed.M.T.F. Moore was a fifth year medical student at the time he undertook this work     

IgG-mediated phagocytosis in regenerated splenic tissue.

The risk of severe infections after splenectomy is well established. Operations such as auto-transplantation, splenic artery ligation or partial resection have been advocated for the retention or regeneration of splenic tissue following splenic trauma. The potential of such tissue to protect from infection is unclear. The ability of splenic tissue to phagocytose IgG opsonized syngeneic erythrocytes was measured in rats 6 months following splenectomy and splenic autotransplantation, splenic artery ligation, total or partial splenectomy, and compared with eusplenic controls. In eusplenic and partially splenectomized rats 71% of the label was cleared at 3 h, compared with approximately 50% in rats following total splenectomy, splenectomy and splenic autotransplantation or splenic artery ligation. The autotransplanted and the ligated splenic tissue cleared less than 10% compared with control spleen, but there was no difference between them when clearance was expressed as uptake per gram of tissue. Splenic autotransplants and ligated spleens were small and histologically abnormal, with an increase in the red pulp, significantly less white pulp and marginal zone, and the frequent absence of a central arteriole in the white pulp. The clearance of label was proportional to the amount of red pulp in the tissue, although the red pulp from the regenerated tissues was not as efficient at phagocytosis as control red pulp. The tissue which regenerated following autotransplantation or splenic artery ligation did not result in greater clearance of erythrocytes from the circulation than that which occurred in splenectomized rats.     

Ultrastructural study on endothelial cells in splenic autotransplantation

The splenic tissue of the rat was autotransplanted into the greater omentum to investigate the angiogenic process during splenic regeneration. Chronological observations showed the following two stages. Initially surviving subcapsular sinus endothelial cells migrated to form a preliminary vascular network, which anastomosed with ingrowing omental capillaries to form a circulatory system. In this way, splenic autotransplants would not undergo necrosis and would rapidly form endothelial and reticular cells. At a later stage, the preliminary vascular network differentiated into arteries (arterioles, capillaries), splenic sinuses, and veins. Marked morphologic changes occurred in the endothelial cells, such as vacuole formation, fenestration, and the condensation of intraendothelial microfilaments, resulting in interendothelial slits. These results showed that the formation of a preliminary vascular network caused distinct splenic regeneration and that splenic autotransplantation is a suitable model to study angiogenesis in vivo.     

Evaluation of immunity of sheep after splenectomy, splenic artery ligation and autotransplantation of splenic tissue

The studies aiming at evaluation of immunity after splenectomy, splenic artery ligation, and autotransplantation of splenic tissue were performed in sheep. It was found that splenectomy alters both humoral and cellular immunity. The phagocytic activity of granulocytes, reduced level of IgG, drop in T and ARF cells as well as bone marrow stimulation were noted. In animals subjected to splenic artery ligation or autotransplantation of splenic tissue no significant differences in parameters studied were observed. These results correlated with histological examinations indicate that in the case of spleen trauma two important steps should be followed: the spleen, if possible, should be saved or a spleen saving procedure should be employed; in case of extensive trauma or if contraindications for spleen saving exist, autotransplantation of splenic tissue should be performed.     

Autotransplantation of splenic fragments: lymphocyte subsets in blood, lymph nodes and splenic tissue.

Abnormal ratios of T helper-type to T suppressor-type lymphocytes in the blood of patients with replanted autologous splenic tissue led to the present study in rats. Lymphocyte subsets were studied in the blood, mesenteric lymph nodes and spleen after autotransplantation and compared to splenectomized and control rats. In the blood of transplanted rats the percentage of T and T helper-type lymphocytes was lower, in the spleen B lymphocytes higher and T lymphocytes and their subsets lower. Comparable changes were seen in the lymph nodes. The data of the mesenteric lymph nodes in autotransplanted rats did not differ from splenectomized animals. Even after 37 weeks the regenerated splenic tissue only reached 13% of the weight of control rats and the absolute lymphocyte number was only 2.5% of a normal spleen. Splenic autotransplantation results in a small hypocellular mass of splenic tissue with a different composition of lymphocyte subsets and does not correct the obvious effect of splenectomy on lymphocyte subpopulations in lymph nodes.