Donor-derived alloantigen-presenting cells persist in the liver allograft during tolerance induction

The predictive value of chimerism was evaluated in three different transplantation models in the rat without immunosuppression: small bowel- (SBTx), liver- (LTx), and liver/small bowel transplantation (LSBTx) were performed in the Brown Norway (BN)-to-Lewis- (LEW) strain combination. Immunohistochemistry and flow cytometry were used to identify donor cells in the recipient's spleen. Their number did not change significantly during transient rejection or tolerance after LTx and LSBTx. However, the amount of donor-derived nonparenchymal cells within the liver allograft including antigen-presenting cells (APCs), such as dendritic and Kupffer cells, clearly mirrored the recipient's immune status: as expected, their number decreased during rejection, but recovered considerably during and after tolerance induction. We conclude that donor cells in the periphery of the recipient correlate with the presence of the allograft, but do not seem to influence graft acceptance actively. However, the kinetics of the detected donor APC population in the liver suggests their important role in modifying the recipient's immune response towards tolerance. Received: 29 March 1999/Revised: 19 August 1999/Accepted: 3 November 1999     

Dynamic changes of indoleamine 2,3-dioxygenase of Kupffer cells in rat liver transplant rejection and tolerance

Aims

To study the dynamic changes and the immunologic role of indoleamine 2, 3-dioxygenase (IDO) in Kupffer cells (KCs) after rat liver transplantation.

Methods

Animals were randomly divided into two groups: a rejection group (REJ; LEW to BN) and a tolerance group (TOL; BN to LEW). Liver morphological changes were observed optically with hematoxylin/eosin staining. KCs were isolated from recipients. mRNA and protein expressions of IDO were detected by real-time polymerase chain reaction and Western blotting at 1, 3, 5, and 7 days after transplantation.

Results

The levels of IDO mRNA and protein in KCs of TOL groups were similar to those in REJ groups at day 1 posttransplantation. However, the expression of IDO mRNA and protein time-dependently increased to much higher levels in the TOL than the in REJ groups at 3, 5, and 7 days posttransplantation (P < .05). The peak was observed at 7 days.

Conclusions

The IDO level of KCs was closely associated with immune tolerance induction. IDO-mediated immune modulation appears to be an attractive means to assess transplant tolerance induction.     

枯否细胞极化状态在肝移植免疫耐受中的作用

随着外科手术技术的成熟,肝移植手术成功率逐渐提高,然而术后长期免疫耐受的建立仍然面临着许多问题。枯否（Kupffer）细胞是一种组织驻留型巨噬细胞,常驻于肝脏当中,其可在肝移植术后向着不同方向极化,形成M1型Kupffer细胞和M2型Kupffer细胞。M1型Kupffer细胞具有促炎功能,M2型Kupffer细胞具有免疫调节功能。通过抑制M1型Kupffer细胞数量和功能,或者促使M2型Kupffer细胞数量增加和功能增强,有助于免疫耐受的建立。Kupffer细胞的极化受到诸多细胞因子和信号的调节,这为通过干预Kupffer细胞极化来建立肝移植免疫耐受的疗法提供了机会。本文将就Kupffer细胞极化状态与肝移植免疫耐受的关系、Kupffer细胞极化机制进行综述,旨在为建立肝移植免疫耐受提供参考。     

脂质体包裹氯膦酸二钠剔除大鼠肝脏枯否细胞作用的研究

目的探讨脂质体包裹氯膦酸二钠剔除大鼠肝脏枯否细胞的作用。方法实验组大鼠给予脂质体包裹的氯膦酸二钠,对照组给予等量生理盐水。给药后不同时间点计数ED1、ED2阳性细胞数目;尾静脉注射印度墨汁判断枯否细胞吞噬碳素颗粒的情况;RT-PCR检测枯否细胞受体mRNA的表达情况。结果给药后2d大鼠肝脏吞噬碳素颗粒的枯否细胞消失,ED1、ED2阳性细胞基本消失,PCR检测不到肝脏枯否细胞受体mRNA的表达。给药后第8天ED1阳性细胞开始明显增多,至第11天其数目基本恢复正常。ED2阳性细胞至第11天开始增加,但平均每中倍视野仍仅有(4.8±1.7)个细胞,明显少于对照组的(17.7±2.1)个细胞,差异具有统计学意义(P0.01)。结论静脉单次注射脂质体包裹氯膦酸二钠至少在1周内能有效剔除肝脏枯否细胞。     

Kupffer cell blockade prevents induction of portal venous tolerance in rat cardiac allograft transplantation

Pretransplant portal venous (pv) administration of donor antigen induces allospecific partial tolerance. Although the involved mechanism has not been defined, antigen presentation by Kupffer cells (KC) in the liver is considered to be critical. We evaluated the effect of KC blockade on this pv tolerance induction in Buffalo (RT1b) rats receiving Lewis (RT1(1] cardiac heterotopic allografts. Control rats received no treatment, while experimental animals received 25 X 10(6) ultraviolet B-irradiated (12,000 J/m2) donor spleen cells via either the iv (systemic intravenous) or the pv routes 7 days before transplantation. Gadolinium chloride (GdCl3), a rare earth metal known to inhibit KC phagocytosis, was given (7 mg/kg) 1 and 2 days before pv preimmunization. Cardiac graft prolongation was obtained by pv (MST = 13.3 +/- 1.9 days, n = 6, vs control = 7.3 +/- 0.5 days, n = 6; P less than 0.001) but not by iv preimmunization (7.7 +/- 0.7 days, n = 6, NS vs control). KC blockade abolished the pv tolerance, as indicated by abrogation of graft prolongation (PV + GdCl3 = 8.0 +/- 0.8 days, n = 6, NS vs control). These findings suggest that effective alloantigen uptake by KC in the liver is essential for the induction of pv tolerance in rat cardiac transplantation.     

FK 506 inhibits tolerance induction in mice liver transplantation

BACKGROUND: In the orthotopic mouse liver transplantation model, allografts are accepted without immunosuppression, and donor-specific tolerance is induced upto 40 days. Although FK 506 is a well-known immunosuppressive agent, its influence on tolerance induction is not known. In this study, we examined the influence of FK 506 on tolerance induction in a mouse liver transplant model. METHODS: Orthotopic liver transplantation was performed from B10.BR (H-2K) to B10.D2 (H-2D mice). In the experimental group, FK 506 (1 mg/kg/d) was given subcutaneously to the recipients from day 0 to day 21, whereas the control group received a placebo (1 mg/kg/d). On day 40, donor skin grafts were transplanted to the recipients to examine the survival times of the recipients and the skin grafts. On day 14, donor-type cells in recipient's blood, spleen, kidney, thymus, and lymph nodes were examined by RT-PCR using specific donor-type MHC class I and II primers. RESULTS: All recipients survived for more than 100 days. The mean survival time of skin grafts in the experimental group was significantly reduced compared to that of controls. On day 14, either donor-type MHC class I- or class II-positive cells were detected in the control group, whereas donor-derived MHC class II-positive cells disappeared in the experimental group. CONCLUSIONS: In the early period after mouse liver transplantation, FK 506 inhibits tolerance induction paradoxically. Some donor-derived MHC class II-positive cells might play an important role in tolerance induction.     

Kupffer cells participate in rejection following liver transplantation in the rat

Abstract Recognition of foreign antigens involves macrophages which release mediators such as immunoactive interleukins, and in the liver, the resident macrophages (Kupffer cells) are activated following transplantation. Therefore, we evaluated the hypothesis that Kupffer cells participate in the rejection reaction following transplantation. Orthotopic liver transplantation was performed between different syngenic rat strains. Livers from Lewis rats were stored in lactated Ringer's solution for 1 h to minimize cold ischemic injury and transplanted into PVG recipients. At 24 h postoperatively, transaminases (AST) were elevated to values around 2000 U/l, total bilirubin was increased to values around 20 μmol/l, and five of six rats died within 3 days. Macroscopic and histological examination showed large areas of necrosis without cellular infiltration, characteristic of rejection. When donor rats were treated with gadolinium chloride (GdCl₃, 10 mg/kg i.v. 24 h before storage of the liver) to inactivate the Kupffer cells, AST levels only rose to around 700 U/l, and the total bilirubin level was in the normal range (<4 μmol/l). Survival was improved significantly by GdCl₃, with five of seven rats surviving more than 1 month ( P < 0.05) and four of seven rats surviving for at least 100 days without immunosuppressive drug therapy. Rejection was not totally prevented, however, since the surviving rats had elevated AST and bilirubin levels, and cellular infiltration in portal areas along with proliferation of bile canaliculi was observed. These data are consistent with the hypothesis that Kupffer cells participate in mechanisms of early rejection following liver transplantation.     

人枯否细胞在同种肝移植免疫中作用机制初步探讨

目的 探讨肝脏枯否细胞(KC)在肝移植后早期免疫反应中的可能作用.方法 将KC和(或)异体PBMC共培养,收集细胞上清液,培养结束时分别收获培养的KC和PBMC.检测HLA-G在细胞表面的表达;测定上清液中NO、IFN-γ、IL-10和TGF-β1的浓度;MTT试验观察KC对淋巴细胞增殖的影响.结果 实验组及对照组中KC和PBMC表面均未检测到HLA-G的表达.与不含KC实验组相比,含KC实验组中,NO、IL-10和TGF-β1的产量显著升高,而IFN-γ呈相对偏低趋势;对照组中未能检测到IL-10和IFN-γ的分泌,仅含KC的对照组中含少量NO及TGF-β1,且显著低于实验组.MTT实验发现,不含KC实验组OD值显著高于含KC实验组及对照组.结论 体外KC接触异体PBMC后早期,各细胞膜表面均无HLA-G表达,但参与了NO及Th2/Th3样细胞因子的分泌调节,并能抑制淋巴细胞增殖反应,可能促进肝脏移植早期免疫耐受的形成.     

枯否细胞在肝移植脂肪供肝的缺血再灌注损伤中的作用研究进展

缺血再灌注损伤(ischemia-reperfusion injury,IRI)是肝移植的常见危险因素,与早期移植肝的无功能和功能障碍密切相关.当发生IRI时,会产生大量的氧自由基和炎症因子,引起肝脏损伤,枯否细胞(Kupffer cells,KCs)的激活与其密切相关.同时,由于肝移植供肝的需求不断增多,脂肪供肝在肝...     

问充质干细胞在诱导免疫耐受中的作用

问充质干细胞是中胚层来源多能非造血干细胞,其不仅有多向分化能力,还具有免疫调节能力.本文就问允质干细胞在诱导免疫耐受中的研究进展作一综述.     

Structural changes of Kupffer cells in rat liver following experimental thermal injury

Changes in plasma haemoglobin levels and morphology of Kupffer cells were studied in rats following lethal thermal injury. Severe haemolysis was observed immediately after thermal injury. The plasma haemoglobin levels rapidly increased to a maximum level 15 min after injury and then rapidly decreased with time. However, the values were still higher 5 h after injury than those found before injury. Soon after burning the Kupffer cells phagocytized not only circulating cell debris including erythrocyte membranes and degenerate leucocytes but also large amounts of haemoglobin. The degradation of phagocytized haemoglobin was relatively slow compared to that of other cell debris. This phagocytized haemoglobin is considered to inhibit the generation of bactericidal-free radicals and to depress Kupffer cell function. The number of Kupffer cells was markedly decreased 5 h after thermal injury, and probably relates to the persistent depletion of the reticuloendothelial system function which follows lethal thermal injury. In contrast to the decreased number of Kupffer cells, a small number of monocytic cells appeared in the sinusoidal spaces and adhered to the endothelial cells. These monocytic cells may transform into Kupffer cells.     

Kupffer cells infiltrate liver tissue early after ischemia-reperfusion and partial hepatectomy

Kupffer cells, ED2+macrophages of the liver, play an important role in liver damage and regeneration. It is proposed that Kupffer cells are stationary and regenerate after acute liver trauma by local proliferation. We analyzed their kinetics in three surgically relevant murine models of acute liver injury: partial liver resection, ischemia with reperfusion and sepsis. We found an early increase in ED2+cells after 0.5 h and a maximum after 12 h. These results suggest an infiltration of the cells early after the injury and a later local proliferation. These ED2+macrophages are localized predominantly periportally; nearly no macrophages are found pericentrally, except in the sepsis model. Therefore, a shifting of macrophages from portal to central seems to be unlikely, suggesting a hepatic zonation of homing factors.     

Inhibition of Kupffer cells reduced CXC chemokine production and liver injury

BACKGROUND: Cytokine production is a critical component of ischemia/reperfusion (IR) injury. In the liver, Kupffer cells produce cytokines and chemokines (i.e., cytokines with chemoattractant properties) that are important mediators in neutrophil recruitment and subsequent hepatocellular injury. Therefore, the role of Kupffer cells in chemokine production in hepatic IR injury was investigated. METHODS: Adult male C57BL/6 mice underwent 90 min of partial hepatic ischemia followed by various reperfusion times (i.e., 0, 1.5, 3, and 6 h). Gadolinium chloride (GC), which inhibits Kupffer cell activity, was administered to mice 48 and 24 h prior to ischemia. The control group received a corresponding volume of normal saline. Plasma levels of the cytokine macrophage inflammatory protein-2 (MIP-2), KC, and tumor necrosis factor (TNF)-alpha and liver mRNA were measured. Liver injury was assessed by plasma level of alanine transaminase (ALT) and histopathology. RESULTS: A reperfusion time-dependent liver injury occurred as indicated by increased levels of plasma ALT and histopathology. The injury was associated with increased plasma TNF-alpha, MIP-2, and KC and their hepatic mRNA expression and neutrophil infiltration into ischemic lobes of the liver. GC treatment significantly reduced the number of Kupffer cells as determined by the immunostained liver tissue sections. The extent of liver injury significantly decreased in GC-treated mice that were associated with decreased levels of plasma ALT, TNF-alpha, MIP-2, and KC and neutrophil infiltration. CONCLUSIONS: This study suggests that Kupffer cells are major contributors to cytokine production in hepatic IR and their modulation may serve as a potential target for therapeutic intervention.