Hepatocyte transplantation for total liver repopulation

Hepatocyte transplantation (HT) is an attractive therapeutic alternative to liver transplantation. A number of experiments have shown the feasibility of total liver parenchymal cell replacement by transplanted hepatocytes. In this review, we would like to highlight researches and clinical reports of HT for liver repopulation. Cellular source of clinical HT should be safety. Immortalized cells, hepatic stem cells, and other stem cells have been used for an experimental model for HT. The exact mechanism of the cell engraftment after HT has not been completely understood, although there were some markers to detect and investigate transplanted cells. In order to achieve liver repopulation following HT, a mild hepatic damage may need to facilitate cell engraftment and replace the host liver by transplanted cells. Hormonal factor may use for the same purpose. Despite the results of preclinical studies promising clinical benefits for cell therapy, the clinical experience of HT has been disappointing, except in a few cases. HT may become an alternative for liver transplantation in the future; however, many efforts should made before establishing an effective method for HT and liver replacement therapy.     

Portal vein thrombosis after intraportal hepatocytes transplantation in a liver transplant recipient

Hepatocytes transplantation is viewed as a possible alternative or as a bridge therapy to liver transplantation for patients affected by acute or chronic liver disorders. Very few data regarding complications of hepatocytes transplantation is available from the literature. Herein we report for the first time a case of portal vein thrombosis after intraportal hepatocytes transplantation in a liver transplant recipient. A patient affected by acute graft dysfunction, not eligible for retransplantation, underwent intraportal infusion of 2 billion viable cryopreserved ABO identical human allogenic hepatocytes over a period of 5 h. Hepatocytes were transplanted at a concentration of 14 million/ml for a total infused volume of 280 ml. Doppler portal vein ultrasound and intraportal pressure were monitored during cell infusion. The procedure was complicated, 8 h after termination, by the development of portal vein thrombosis with liver failure and death of the patient. Autopsy showed occlusive thrombosis of the intrahepatic portal vein branches; cells or large aggregates of epithelial elements (polyclonal CEA positive), suggestive for transplanted hepatocytes, were co-localized inside the thrombus.     

Nodular regenerative hyperplasia: a controversial indication for orthotopic liver transplantation

Nodular regenerative hyperplasia of the liver is an uncommon cause of portal hypertension. Patients with nodular regenerative hyperplasia have signs and symptoms of portal hypertension, without evidence of hepatocellular failure or encephalopathy. We report the case of a 44-year-old woman with recurrent esophageal bleeding and refractory ascites who had a history of hemosiderosis, hepatitis C, and chronic renal allograft rejection. Our preoperative diagnosis was cirrhotic end-stage liver disease and end-stage renal disease for which the patient underwent combined hepatic and renal transplantation. Her portal hypertension symptoms resolved, and her renal function has been normal for 18 months of follow-up. Histologic examination of the liver revealed nodular regenerative hyperplasia, and a review of the literature regarding the surgical management of patients with nodular regenerative hyperplasia revealed that various shunting procedures are generally recommended. After the failure of medical management in patients with nodular regenerative hyperplasia, portosystemic shunting may be indicated before proceeding to hepatic transplantation.     

Changes in liver regenerative factors in a case of living-related liver transplantation

Liver regeneration in a patient with fulminant hepatic failure (FHF) who underwent living-related partial liver transplantation (LRLT) was investigated regarding hepatic growth factors. The patient was a 16-yr-old Japanese male who developed severe subacute FHF. LRLT was performed using an extended left lobe of the ABO matched patient's mother. In the recipient, the pre-transplant levels of both plasma hepatocyte growth factor (HGF) and transforming growth factor (TGF)-beta were extremely high and rapidly decreased following the liver replacement. The liver volume evaluated using a CAT scan increased 195% after 2 wk in graft liver and 110% after 2 wk in the hepatectomized donor. The explanted liver (FHF liver), the liver from donor (normal liver), and the graft liver [the 3rd post-transplant day (POD 3)] were all investigated immunohistochemically. FHF liver: No liver regeneration was observed [proliferative cell nuclear antigen (PCNA) labeling index (L.I.): 0%]. In the liver, both HGF in the hepatocytes and c-met on the membrane of the hepatocytes were positive. TGF-beta was positive in the hepatocytes and no apoptosis was detected by the TUNEL method. Donor liver (POD 0): Few PCNA stained hepatocytes were detected. No HGF was detected but c-met was clearly detected on the cell membrane of the hepatocytes. Neither TGF-beta nor apoptosis was detected. Graft liver (POD 3): The PCNA L.I. was conspicuous at 40%. HGF was positive in non-parenchymal cells and c-met was positive in the cytoplasm of the hepatocytes. TGF-beta was negative while apoptosis was positive in the zone 3 hepatocytes. In conclusion, these findings suggested that the liver of the patient with FHF did not respond to liver regenerative stimulus, in part, through involvement of inhibitor TGF-beta. On POD 3, the transplanted graft was in a vigorous regenerative status in comparison to that in the hepatectomized donor. The HGF/c-met system is thought to be involved in the mechanism of regeneration. Intrahepatic apoptosis was detected in the graft on the 3rd post-transplant day probably due to transient ischemia in the liver, which was not related to the Fas/Fas-ligand system.     

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.     

Functional, life-threatening disorders and splenectomy following liver transplantation

Splenectomy (SPL) in cirrhotic patients undergoing liver transplantation (LTx) may resolve specific problems related to the procedure itself, in case of functional and life-threatening clinical situations often occurring as a result of liver cirrhosis and portal hypertension. METHOD: A single-center experience of ten splenectomies in a series of 180 consecutive adult liver transplant patients over a period of 6 yr is reported. The mean patient age was 46.8 +/- 9.5 yr (range 25 57 yr). Indications for SPL were post-operative massive ascitic fluid loss (n = 3), severe thrombocytopenia (n = 3), acute intra-abdominal hemorrhage (n = 2), infarction of the spleen (n = 1), and multiple splenic artery aneurysms (n = 1). RESULTS: Extreme ascites production due to functional graft congestion disappeared post-SPL, with an improvement of the hepatic and renal functions. SPL was also effective in cases of thrombocytopenia persistence post-LTx, leading to an increase in the platelet count after about 1 wk. Bleeding episodes related to left-sided portal hypertension or trauma were also resolved. The rejection rate during hospitalization was 0%, and no other episodes were recorded in the course of the long-term follow-up. However, sepsis with a fatal outcome occurred in 4 patients, i.e. between 2 and 3 wk post-SPL in three cases and 1 yr after the procedure as a result of pneumococcal infection in the last case. Fatal traumatic cranial injury occurred 3 yr post-LTx in another case. Five patients (50%) are still alive and asymptomatic after a median follow-up period of 36 months. CONCLUSION: The lowering of the portal flow appears to resolve unexplained post-operative ascitic fluid loss as a result of functional graft congestion following LTx. However, because of the enhanced risk of SPL-related sepsis, a partial splenic embolization (PSE) or a spleno-renal shunt could be used as an alternative procedure because it allows us to preserve the immunological function of the spleen. SPL is indicated in case of post-transplant bleeding due to left-sided portal hypertension and trauma, spleen infarction, and to enable prevention of hemorrhage in liver transplant patients with multiple splenic artery aneurysms. Severe and persistent thrombocytopenia could be treated with PSE. Because the occurrence of fatal sepsis post-SPL is a major complication in LTx, functional disorders, such as ascites and thrombocytopenia, should be treated with a more conservative approach.     

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转化医学是近年来医学领域出现的新概念,强调的是基础研究成果向临床应用的转化,但基础研究的命题往往来源于临床中需要回答和解决的问题。对于肝移植后急性移植物抗宿主病这种低发生率但高死亡率的疾病而言,单单从临床工作上对其进行研究是极其困难的。作者近年来将临床中遇到的问题进行了总结和思考,再通过实验研究包括建立的大鼠肝移植后急性移植物抗宿主病模型,研究其中的免疫学机制,寻找治疗靶点和方法等,以期在今后的临床中加以应用和验证。这些工作充分体现了转化医学的概念。     

氯化钆抑制大鼠肝移植急性排斥反应的机理

目的　探讨氯化钆抑制大鼠肝移植急性排斥反应的机理。方法分别采用健康雄性Wistar和SD大鼠作为供、受者。随机分为A组(假手术对照):SD大鼠10只,开腹后不作任何处理,关腹结束手术;B组(氯化钆预处理 肝移植):受者1 5只,供者在移植前进行氯化钆预处理2 d,再获取供肝移植给受者;C组(生理盐水预处理 肝移植):受者15只,供者用生理盐水代替氯化钆预处理,其余处理同B组。分别检测各组的术后生存率、肝功能、肝脏病理组织学、肝组织和胆汁中细胞因子表达、枯否氏细胞核转录因子κB(NF-κB)以及细胞膜表面分子的表达情况。结果(1)B组术后1个月生存率明显高于C组(P<0.01)。(2)B组术后肝功能逐渐恢复正常,C组肝功能进行性恶化;B组肝组织病理学改变轻微,C组出现典型急性排斥改变。(3)B组肝组织和胆汁中γ-干扰素(IFN-γ)和白细胞介素2 (IL-2)较A组明显降低(P<0.05),IL-10较A组明显升高(P<0.05),IL-4无明显变化;C组出现与之相反的变化。(4)C组枯否氏细胞NF-κB活性明显高于A、B两组(P<0.01)。(5)B组枯否氏细胞膜表面主要组织相容性抗原复合物(MHC)、CD80、CD86分子明显低于A组(P<0.05),C组高表达上述膜表面分子。结论氯化钆能够有效地抑制枯否氏细胞的免疫活性,从而抑制大鼠同种肝移植后急性排斥反应的发生。     

Protection of liver sinusoids by intravenous administration of human Muse cells in a rat extra-small partial liver transplantation model

Small-for-size syndrome (SFSS) has a poor prognosis due to excessive shear stress and sinusoidal microcirculatory disturbances in the acute phase after living-donor liver transplantation (LDLT). Multilineage-differentiating stress enduring (Muse) cells are reparative stem cells found in various tissues and currently under clinical trials. These cells selectively home to damaged sites via the sphingosine-1-phosphate (S1P)–S1P receptor 2 system and repair damaged tissue by pleiotropic effects, including tissue protection and damaged/apoptotic cell replacement by differentiating into tissue-constituent cells. The effects of intravenously administered human bone marrow-Muse cells and -mesenchymal stem cells (MSCs) (4 × 10⁵) on liver sinusoidal endothelial cells (LSECs) were examined in a rat SFSS model without immunosuppression. Compared with MSCs, Muse cells intensively homed to the grafted liver, distributed to the sinusoids and vessels, and delivered improved blood chemistry and Ki-67(+) proliferative hepatocytes and -LSECs within 3 days. Tissue clearing and three-dimensional imaging by multiphoton laser confocal microscopy revealed maintenance of the sinusoid continuity, organization, and surface area, as well as decreased sinusoid interruption in the Muse group. Small-interfering RNA-induced knockdown of hepatocyte growth factor and vascular endothelial growth factor-A impaired the protective effect of Muse cells on LSECs. Intravenous injection of Muse cells might be a feasible approach for LDLT with less recipient burden.     

The relationship of systemic hemodynamics and oxygen consumption to early allograft failure after liver transplantation

The early postoperative hemodynamic data of 88 patients who underwent primary liver transplantation between July 1989 and October 1990 at the University Health Center of Pittsburgh were analyzed to establish the relationship of systemic hemodynamics and oxygen consumption to perioperative allograft function. The 15 patients whose allografts failed within the 1 st month following transplantation were designated as group 1, while 73 patients who retained adequate graft function constituted group 2. Although the cardiac index and oxygen delivery did not differ significantly between the groups, group 1 consistently demonstrated a lower mean arterial pressure, oxygen consumption, arteriovenous oxygen content difference, and arterial ketone body ratio. The etiology of reduced oxygen consumption in group 1 patients is speculative, but the data support the notion that oxygen consumption is a useful, predictive indicator for liver allograft function after transplantation.     

肝移植术后侵袭性曲菌病的防治

目的探讨原位肝移植术后侵袭性曲菌病的防治。方法回顾性分析2000年1月至2005年1月完成的576例原位肝移植的临床资料,总结术后侵袭性曲菌病的预防和治疗经验。结果9例患者术后并发侵袭性曲菌病,发病率为1.74%(9/576),首发感染部位为肺部8例,中枢神经系统感染1例。6例患者停用免疫抑制治疗,3例患者将他克莫司(FK506)或CsA降低到最低有效血药浓度。6例患者选用两性霉素B脂质体(其中1例先试用氟康唑)、3例首选伊曲康唑进行治疗。5例肺部感染患者痊愈,2例因肺部感染无法控制死亡,2例因并发多器官曲霉菌感染死亡。结论早发现并及时调整免疫抑制治疗方案,早期、足量和足程使用抗真菌药物,积极行手术治疗是降低肝移植术后侵袭性曲菌病发病率和病死率的根本措施。     

The incidence and management of biliary complications following liver transplantation in children

Biliary complications following liver transplantation are a cause of significant morbidity and mortality. During the period 1988–1993 ten cases of biliary complications occurred after 98 transplantations in 78 children. The complications were four bile leaks, three intrahepatic biliary strictures (one with recurrent cholangitis), two anastomotic biliary strictures (one with recurrent cholangitis) and one recurrent cholangitis. All leaks occurred within 6 weeks of transplantation whereas all strictures and cholangitic episodes occurred after 3 months. Two biliary complications (20%) — one intrahepatic and one anastomotic stricture — developed secondary to hepatic artery thrombosis. The incidence of biliary complications was 13.2% with whole liver grafts as compared to 6.7% with partial liver grafts and it was 4.3% with duct-to-duct anastomosis as compared to 12.0% with Roux-en-Y hepatico-jejunostomy. Seven children required intervention for management of biliary complications and three were managed conservatively. There were no deaths related to the biliary complications.     

Successful resolution of severe hepatopulmonary syndrome following liver transplantation

Hepatopulmonary syndrome (HPS) is a complication of portal hypertension, defined by the presence of liver disease, abnormal pulmonary gas exchange and evidence of intrapulmonary vascular dilatations producing a right‐to‐left intrapulmonary shunt. Liver transplantation (LT) is the treatment of choice; however, severe hypoxemia (PaO₂ < 50 mmHg on room air) is considered a contraindication to LT. This approach disadvantages some patients, particularly young patients with no intrinsic cardio‐respiratory disease. We discuss one such patient who improved with LT despite having extremely severe HPS (PaO₂ < 29 mmHg)     

Diaphragmatic nerve palsy in young children following liver transplantation

Diaphragmatic paralysis was identified in four children after liver transplantation. All presented with persistent right upper lobe atelectasis, pleural effusion and recurrent respiratory infections and could not be weaned from mechanical ventilatory support. Fluoroscopy and real-time ultrasound confirmed paradoxical right diaphragmatic movements. Diaphragmatic plication was undertaken and enabled rapid and sustained weaning from respiratory support in all four cases. Vascular clamping of the suprahepatic vena cava seems to be the cause. Diaphragmatic plication allows optimal recruitment of the respiratory muscles with a favourable impact on lung mechanics and gas exchange. Received: 31 December 1997 Received after revision: 27 March 1998 Accepted: 17 April 1998     

Cryopreserved iliac artery allograft for primary arterial revascularization in adult liver transplantation

Arterial allograft represents a material of choice for primary arterial revascularization in liver transplantation (LT) when interposition of a vascular conduit is required. In case of non-availability of such graft, the use of cryopreserved vessels should be an interesting option. Three patients were grafted using a cryopreserved iliac artery allograft (CIAA) previously harvested and stored at -140°C in a tissue bank. An auxiliary partial LT was performed in one patient for acute liver failure. During follow-up, an efficient regeneration of the native hemi-liver was observed while atrophy of the auxiliary graft occurred, leading to functional portal vein and hepatic artery thrombosis at six and nine months, respectively. Two other patients presented with celiac trunk compression because of arcuate ligament without available arterial allograft in the donor. Late arterial thrombosis occurred at six months in one patient without impairment of graft function. The last patient was alive and symptom free 29 months after LT with a patent cryopreserved arterial conduit. Our preliminary results suggest that CIAA might represent an efficient solution as vessel interposition for primary arterial hepatic revascularization in LT setting when no other suitable graft is available. However, long-term patency of CIAA remains questionable.     

Increase in natural killer cell activity following living-related liver transplantation

Abstract We monitored the serial changes of natural killer cell (NK) activity in eight recipients of living-related liver transplantation. The HLA types of all eight patients were haplotypically identical with those of their donors. Tacrolimus and methylprednisolone were used for immunosuppression. The NK activity before transplantation was 24.1 ± 20.2 % which is surprisingly low when compared with the value for normal individuals (67.7 ± 13.2%, P < 0.01) or a liver dysfunction group (49.4 ± 21.9%, P < 0.05). Serial changes in NK activity revealed a minimum of 6.1 ± 3.6% 1 week after transplantation, gradually increasing to 49.2 ± 12.5 % at 2 months after transplantation. These results suggest that the diseased liver might play an important role in the suppression of NK activity.     

Chronic renal dysfunction following liver transplantation

Abstract:  With most of the immunosuppressive protocols consisting of calcineurin inhibitors (CI), nephrotoxicity has become a major long-term complication often compromising outcome. In a single-center retrospective study, we reviewed 1173 liver transplantations to identify variables indicative for the occurrence of chronic renal dysfunction (CRD) (defined as ≥1 episode of serum creatinine increase ≥1.8 mg/dL ≥2 wk). Chronic renal dysfunction was found in 137 (11.7%) of all transplants [82 (7%) early (after 3–12 months), 55 (4.7%) late-onset (>12 months)]. Compared to 5-/10-yr survival rates in non-CRD transplants (84/74%) survival was significantly decreased in early (66/46%), but unchanged in late-onset CRD (98/86%). Rates of alcoholic cirrhosis and prior renal dysfunction were significantly increased in patients with CRD. In a multivariate logistic regression analysis, only cyclosporine A (CyA) as immunosuppression remained an independent risk factor. No correlations to age, gender, rejection/retransplantation or diabetes were found. Surprisingly, renal function (creatinine) showed no difference between patients on CI monotherapy (FK/CyA) compared to those who had mycophenolate mofetil (MMF) added. In liver transplantation, early onset CRD significantly compromises survival. CyA-based immunosuppression appears to have a stronger impact than FK. The fact that patients with long-term severe chronic renal dysfunction failed to improve under MMF rescue therapy emphasizes the importance of new diagnostic strategies to earlier identify at-risk patients.