Background:Patient selection is one of the key factors influencing prognosis after liver transplantation (LT), and a number of selection criteria have been proposed and broadly applied. The benefit of LT is debatable when patients exceed these criteria in light of liver worldwide donor shortage. This study aims to identify key prognostic indicators predicting patients undergoing LT beyond Hanzhou criteria, to guide LT candidate selection. Methods:The clinical data and pathological findings of patients who underwent LT in the Liver Transplantation Center of General Hospital of Southern Theater Command from September 2003 to August 2017 were collected for retrospective analysis, only patients exceeding the Hangzhou criteria were enrolled. Patients were followed up until December 1, 2023; disease-free survival (DFS) and overall survival (OS) was analyzed; and predictive models of DFS and OS were constructed and validated. Results:A total of 161 patients were included in the analysis. A tumor diameter ≥6 cm [hazard ratio (HR) =2.01; P=0.01], alpha fetoprotein (AFP) levels >1,000 ng/mL (HR =2.68; P<0.001), lack of metformin administration (HR =2.98; P=0.008), operative blood loss >2,000 mL (HR =1.75; P=0.01), and female donor gender (HR =3.71; P=0.004) were independent risk factors for lower DFS, while a tumor diameter ≥6 cm (HR =2.05; P=0.001), AFP level >1,000 ng/mL (HR =1.54; P=0.04), and female donor gender (HR =4.45; P<0.001) were independent predictors of worse OS. Patients with a preoperative AFP level ≤1,000 ng/mL or a history of metformin administration had a significantly better prognosis after LT. Conclusions:For patients exceeding the Hangzhou criteria, those with an AFP level ≤1,000 ng/mL can still achieve acceptable long-term prognosis after LT. Administration of metformin has a strong positive association with longer recurrence-free survival. The patient criteria for selection for LT can be tentatively expanded.

Background The use of grafts from donation after circulatory death (DCD) overcomes the inadequate donor organ supply. Our team developed a transportable dual hypothermic oxygenated machine perfusion (DHOPE) device, which initiates DHOPE at a recipient center to reduce static cold storage (SCS) time and the risk of graft failure in DCD liver transplantation. Methods Six porcine livers per group with 30 min of warm ischemia exposure were preserved via SCS or DHOPE for 6 h and then reperfused for 12 h with whole blood to mimic transplantation. Hepatocellular and biliary function and injury were assessed in perfusate and bile samples. Molecular biomarkers and histology were compared between groups. Results Reperfusion portal vein pressure, in a flow-constant manner, and alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and gamma-glutamyltransferase (γ-GGT) release were significantly lower in the DHOPE group than in the SCS group at all time points. Higher bile production paralleled the lower levels of ALP and γ-GGT in the DHOPE group. The DHOPE group secreted more total bilirubin (TBIL) in bile, resulting in decreased TBIL in the perfusate, and livers preserved with DHOPE exhibited better cholangiocellular function. Furthermore, improvements in hypoxia, the inflammatory response, cell-free microRNAs and energy metabolism were observed in the DHOPE group. There were fewer apoptotic cells and TGF-β1-positive cells in the liver parenchyma and extrahepatic bile duct in the DHOPE group than in the SCS group. Conclusions This study demonstrates the efficacy of local 4 °C DHOPE to protect porcine liver grafts from 30-min warm ischemia damage.

Conventional static cold storage (SCS) exacerbates ischemic injury in the DCD liver, leading to severe complications for transplant recipients. To address this issue, clinical application of MP technology for donor liver preservation is underway. Simultaneously, efforts are focused on the development of various MP instruments, validated through relevant animal model experiments. Effective large animal trials play a pivotal role in clinical applications. However, challenges persist in the ex vivo preservation of DCD livers and the transplantation procedure in pigs. These hurdles encompass addressing the prolonged preservation of donor livers, conducting viability tests, alleviating ischemic injuries, and shortening the anhepatic phase. The use of a variable temperature-controlled MP device facilitates the prolonged preservation of DCD livers through sequential Dual Hypothermic Oxygenated Machine Perfusion (DHOPE) and Normothermic Machine Perfusion (NMP) modes. This protocol enhances the porcine OLTx model by improving the quality of DCD livers, optimizing the anastomosis technique, and reducing the duration of the anhepatic phase.

ObjectivesSalvage liver transplantation (sLT) is considered an effective method to treat hepatocellular carcinoma (HCC) recurrence. This multicenter research aimed to identify the prognostic factors associated with recurrence-free survival (RFS) and overall survival (OS) after sLT.Material and MethodsA retrospective analysis of 114 patients who had undergone sLT for recurrent HCC between February 2012 and September 2020 was performed. The baseline and clinicopathological data of the patients were collected.ResultsThe 1-, 3-, and 5-year RFS rates after sLT were 88.9%, 75.2%, and 69.2%, respectively, and the OS rates were 96.4%, 78.3%, and 70.8%. A time from liver resection (LR) to recurrence < 1 year, disease beyond the Milan criteria at sLT and macrotrabecular massive (MTM)-HCC were identified as risk factors for RFS and were further identified as independent risk factors. A time from LR to recurrence < 1 year, disease beyond the Milan criteria at sLT and MTM-HCC were also risk factors for OS and were further identified as independent risk factors.ConclusionsCompared with primary liver transplantation (pLT), more prognostic factors are available from patients who had undergone LR. We suggest that in cases of HCC recurrence within 1 year after LR, disease beyond the Milan criteria at sLT and MTM-HCC patients, sLT should be used with caution.

目的 比较不同供体来源的同种异体血管在长时间深低温保存后出现的病理学差异.方法 将深低温保存时间超过3年的同种异体髂动脉分为脑死亡器官捐献(donation after brain death,DBD)组和心死亡器官捐献(donation after circulatory death,DCD)组,每组随机选取10例,复温后取样,通过苏木精-伊红(hematoxylin and eosin,HE)染色以及Masson染色观察血管的病理特点.结果 与DBD组对比,DCD组供体经历的热缺血时间较长(P＜0.01).经过长时间深低温保存后,DCD组血管在HE染色下的血管中膜损伤评分、外膜损伤评分和总体损伤评分高于DBD组(P＜0.05),其中血管外膜和中膜损伤评分与热缺血时间呈正相关.在Masson染色下,DCD组血管胶原纤维光密度值低于DBD组(P＜0.05),且该值与热缺血时间呈负相关.结论 DBD供体来源的同种异体髂动脉超3年深低温保存的质量优于DCD供体来源血管.

Introduction: For end-stage liver diseases, liver transplantation remains the most suitable therapy for its treatment. Nonetheless, the current gap between the patients on the waiting list and the graft supply, as well as the gold standard preservation method (Static Cold Storage or SCS), limits its usefulness. Extended Criteria Donors (ECD) has been proposed as a potential solution to increase the donor pool, although these have a greater susceptibility to ischemia-reperfusion injury (IRI), especially after SCS, causing delayed graft function and primary non-function. These side effects have fueled research into more suitable methods such as ex-situ normothermic machine perfusion (NMP). Methods: Sixteen healthy pig livers were subjected to normothermic perfusion using blood as a perfusion solution. in 5 experiments, taurocholic acid was administrated 6 hours after perfusion initiation. Perfusate and bile were sampled before, during and after perfusion to assess the hepatocellular injury and liver function and bile duct injury and bile production. Liver samples were collected before and after perfusion to perform analysis of general damage, stellate cell activation, oxidative stress and LSEC response. Results: AST and LDH levels increased progressively for the first 12 hours (P= 0.02 and P=0.001 respectively). Afterward, both parameters stabilized. Glucose and lactate levels decreased rapidly during the first 4 hours (p=0.003 for glucose and p=0.04 for lactate) and reached the lowest within 8 (p<0.001) and 12 hours (p<0.001) respectively. Overall, blood pH remained steady (p=0.10). Serum levels of ALKP, GGT and Bil didn’t suffer any changes during all the perfusion (p=0.99, p=0.75 and p=0.13 respectively). Bile production started within 1 hour after ex-situ perfusion. Livers supplied with taurocholic acid had a higher bile production rate, 9±3 mL/h, compared to those not supplied, 4±3 (p=0.005). No structural alterations were detected in liver samples after the Suzuki injury score assessment (p=0.33) and stellate cell activation quantification by both gene (p=0,64) and protein expression (p=0,79) and percentage of the stained area (p=0.15). Although eNOS gene and protein expression increased (p=0.01 and p=0.008 respectively), histological analysis of CD31 endothelial staining showed no differences between the baseline, 35,7±9,1 %, and the 24-hour perfusion 36,3±7,2%, of the sinusoidal area (p=0.87). This increase was associated with higher levels of MDA after the perfusion (p=0.006) as the KLF2 gene and protein expression remained invariable (p=0.80 and p=0.26 respectively). Conclusion: Our results show that NMP, as a preservation method, is capable of maintaining the normal hepatic structure and, at the same time, supporting normal metabolic activity. However, further studies should be conducted to demonstrate post-transplant liver viability.

Amelia Hessheimer, Xiran He, Mingju Liang, Javier Muñoz, Josep M. Sanahuja, Xiaoyu Tan, Marina Vendrell, Felipe León, I sabel Mora, Joaquím Albiol, Li Lin, Feng Huo, Constantino Fondevila Hospital Clínic i Provincial, Barcelona, Spain; Guanguong Shunde Innovative Design Institute, Guangdong, People’s Republic of China; General Hospital of Southern Theater Command of PLA, Guangdong, People’s Republic of China; DTI Foundation, Barcelona, Spain.

随着器官捐献数量不断增加和扩大标准供者(ECD)供肝定义不断拓展,供肝质量必然成为影响肝移植高质量发展的突出问题,也是相关领域的研究重点.最大限度解决器官短缺和推动器官移植高质量发展是我国器官捐献与移植事业发展的方向.近年来,利用机械灌注(MP)对供肝进行灌注、保存、评估及修复,已成为当前国际上提高肝移植质量的研究热点.本文针对国内外ECD供肝的不同应用情况,结合国际上MP研究进展和本中心有关研究经验,探讨整合器官保护技术建设器官重症监护室(ICU)的可行性,拟在推动我国器官移植高质量发展同时,进一步丰富器官捐献与移植"中国模式"的技术内涵.

目的:探讨自主研发的离体肝脏双泵双氧合机械灌注设备DEVOCEAN-LIVER 2000在离体猪肝脏常温机械灌注上的安全性与可操作性.方法:从45～50 kg雌性巴马小型猪获取供肝,使用DEVOCEAN-LIVER 2000,采用恒压模式对动脉和门静脉进行常温机械灌注6 h,使用雅培iSTAT-300G血气分析仪对每小时的灌注液进行分析,同时利用微纳芯Pointcare V2对灌注液进行生化检测.实时记录并比较灌注过程中动脉和门静脉流量、动脉血液pH值、胆汁生成量、转氨酶、血糖、乳酸、总胆红素的变化.灌注前后,切取肝脏组织制作病理切片,观察肝脏组织形态的变化.采用SPSS 23.0统计学软件进行实验数据分析.结果:灌注过程中,动脉流量及门静脉流量随灌注时间平稳增长;动脉血液pH值一直维持在7.2～7.5;胆汁持续性生成,最高峰可达0.0085 mL/[h·g(肝重)];灌注液中的谷丙转氨酶(ALT)和谷草转氨酶(AST)小幅增加;血糖总体呈现平稳下降状态;乳酸水平整体呈下降趋势,灌注前与灌注6 h的乳酸水平相比差异有统计学意义(P<0.05);灌注前后胆红素变化不明显(P>0.05).灌注结束后,肝脏外观无明显瘀斑,显微镜下肝小叶结构完整.结论:DEVOCEAN-LIVER 2000能为肝脏提供6 h稳定的体外常温机械灌注,使肝脏在体外仍然保持活性,并且有效改善肝脏微循环,具有良好的安全性与可操作性.

目的:初步探索离体肝脏双泵双氧合机械灌注设备在离体猪肝脏低温灌注中的可操作性以及稳定性.方法:从45～50 kg雌性巴马小型猪获取供肝,利用灌注设备同时对肝动脉和门静脉实施低温携氧灌注8 h.肝动脉采用压力恒定模式控制,门静脉采用流量恒定模式控制,观察灌注过程中的流量、压力、血气指标、炎症因子以及病理组织学评分的变化.采用IBM SPSS Statistics 23.0统计学软件进行实验数据分析.结果:灌注过程中,肝动脉压力稳定维持在24.09～24.65 mmHg(1 mmHg=133.32 Pa),流量从初期的0.04 mL/[min·g(肝重)]逐渐升高至0.09 mL/[min·g(肝重)];门静脉的流量稳定维持在0.47～0.50 mL/[min·g(肝重)],压力呈降低趋势,从初期的2.79 mmHg下降至1.84 mmHg.灌注液pH值、葡萄糖(Glu)以及乳酸(Lac)水平维持稳定.肝组织的炎症因子白介素6(IL-6)和肿瘤坏死因子α(TNF-α)亦维持稳定.在组织病理学评价方面,肝组织的空泡化水平呈下降趋势,而肝组织的坏死、水肿以及肝窦扩张水平则维持稳定.结论:该离体肝脏双泵双氧合机械灌注设备能够无故障地运转8 h,同时维持肝脏的低代谢状态,降低肝内灌注阻力和改善肝细胞缺氧,具有安全性和稳定性.

There is a dearth of effective parameters for selecting potentially transplantable liver grafts from expanded-criteria donors. In this study, we used a nuclear magnetic resonance (NMR) relaxation analyzer-based assay to assess the viability of ex vivo livers obtained via porcine donation after circulatory death (DCD). Ex situ normothermic machine perfusion (NMP) was utilized as a platform for viability test of porcine DCD donor livers. A liver-targeted contrast agent, gadolinium ethoxybenzyl diethylenetriamine pentaacetic acid (Gd-EOB-DTPA), was injected into the perfusate during NMP, and the dynamic biliary excretion of the Gd-EOB-DTPA was monitored by measuring the longitudinal relaxation time (T1). The longitudinal relaxation rate (R1) of the bile was served as a parameter. The delay of increase in biliary R1 during early stage of NMP indicated the impaired function of liver grafts in both warm and cold ischemia injury, which was correlated with the change of alanine aminotransferase. The preservative superiority in cold ischemia of dual hypothermic oxygenated machine perfusion could also be verified by assessing biliary R1 and other biochemical parameters. This study allows for the dynamic assessment of the viability of porcine DCD donor livers by combined usage of ex situ NMP and NMR relaxation time based assay, which lays a foundation for further clinical application.

目的 构建离体肝脏常温机械灌注设备,探讨灌注实验过程对红细胞的影响.方法 获取健康雌性巴马小型猪肝脏和血液,利用常温机械灌注设备对离体肝脏持续灌注8 h,温度设定37℃,动脉压力设定80/60 mmHg,门静脉设定恒流模式(0.5 mL/minrg),记录灌注过程中灌注液温度、肝动脉和门静脉流量等数据,每小时化验红细胞及其他相关指标,包括红细胞压积(Hematocrit,HCT)、血红蛋白(Hemoglobin,HGB)、总胆红素(Total Bilirubin,TBIL)、间接胆红素(Indirect Bilirubin,IBIL)、溶血率、乳酸(Lactate,Lac)以及血糖(Glucose,Glu).结果 灌注过程中灌注液温度稳定维持37℃,肝动脉流量从最初的(54.64±11.94)mL/min,上升到(249.34±93.01)mL/min,门静脉流量平均值稳定在(442.67±79.19)mL/min.灌注开始和结束时HCT和HGB分别为(27.67±2.42)％、(9.40±0.81)g/dL和(28.00±2.45)％、(9.52±0.81)g/dL.分别比较灌注开始和结束时HCT、HGB数值,均无统计学差异.灌注前后TBIL和IBIL水平比较,差异无统计意义.灌注过程中每小时检测溶血率均为0.Lac从灌注开始的(2.52±0.39)mmol/L下降至灌注结束的(0.35±0.07)mmol/L(P＜0.05).Glu在第2小时达到最高值(21.37±5.96)mmol/L,灌注结束时下降至(9.78±2.63)mmol/L(P＜0.05).结论 离体肝脏常温机械灌注设备能够维持HCT和HGB稳定,不会导致溶血;过程中Lac和Glu持续消耗,间接反映了灌注过程红细胞携氧能力可以改善离体肝脏功能.

肝移植是治疗终末期肝病唯一有效的方法,然而供肝短缺严重制约了临床上的应用.为了解决供肝短缺问题,以往认为风险较大的心脏死亡器官捐献(donation after cardiac death,DCD)供肝,越来越多地用于临床移植[1-3].DCD在心脏停跳过程中不可避免要经历低血压、休克、缺氧等热缺血时间(warm ischemia time,WIT)[4],WIT过长会影响供肝质量,降低供肝利用率,增加移植肝功能延迟恢复、缺血性胆病(ischemic cholangiopathy,IC)等并发症发生率[5-6],严重者甚至出现原发性移植物无功能(primary graft nonfunction,PNF)[7].有报道称DCD供肝移植术后上述并发症发生率可高达50%[8],随着评估和应用经验增加,DCD肝移植疗效有了明显的改善,但仍是影响DCD肝移植预后的主要问题[9-10].

目的 探讨自主研发的常温机械灌注(Normothermic Machine Perfusion,NMP)设备对不同热缺血时间下供肝的保护作用.方法 选取雌性巴马猪作为血液和肝脏供体,分为DCD30min组和DCD60min组,利用NMP设备灌注12 h,观察灌注过程中两组的灌注参数、血气生化指标以及胆汁相关指标的水平.结果 两组供肝不同时间点的肝动脉流量比较,差异无统计学意义(P＞0.05);DCD 30 min组的门静脉压力低于DCD 60 min组,差异有统计学意义(P＜0.05);两组的酸碱平衡、乳酸、血糖、ALP、γGGT和TBIL的水平比较,均无统计学意义(P＞0.05);DCD 60 min组的尿素氮水平更高,DCD 30 min组的转氨酶水平更低,均具有统计学差异(P＜0.05);两组肝脏均能持续合成胆汁,胆汁的pH、碳酸氢根及葡萄糖水平均无统计学差异(P＞0.05);而胆汁胆红素和胆汁酸浓度具有统计学差异(P＜0.05).结论 NMP12h,两组不同热缺血时间的供肝都能进行有氧代谢和持续合成胆汁,提示自主研发的NMP设备可有效保护有热缺血损伤的供肝.

为了推动动物实验教学的发展,提供巴马小型猪胆汁和血液生理生化指标等相关参考数据.选取健康合格的巴马小型猪17只,体重45～50 kg,使用血气分析仪和全自动生化分析仪检测麻醉状态下巴马小型猪术前的胆汁和动脉血气、生化等生理生化指标,并运用血液指标与相关文献和人类正常参考值作比较,使用SPSS23.0统计软件进行统计学分析.结果检测数据均呈正态分布,经比较,更多的血液生理生化指标在人的正常参考范围值内,表明实验数据具有一定的参考意义.

目的 探讨在体常温局部灌注对猪心脏死亡器官捐献(donation after cardiac death,DCD)肝脏的修复作用.方法 45 kg巴马小型猪(n=3)在麻醉后,分别经颈静脉、髂动脉插入导管并与体外膜肺氧合(extracorporeal membrane oxygenation,ECMO)连接.静脉注射氯化钾溶液引发心跳骤停,30 min后置入球囊导管阻断胸主动脉,开始运转设备进行在体常温(37℃)局部灌注4 h,静脉血氧维持在60%～70%,每隔1 h检测其血气、生化指标,灌注前、后取肝脏组织,进行病理观察.结果 4 h灌注过程中,设备运转稳定,乳酸显著下降(P<0.05),肝脏持续有胆汁生成,丙氨酸转氨酶、谷氨酰转肽酶及总胆红素浓度变化无统计学差异(P>0.05),天冬氨酸转氨酶在正常范围内,病理切片提示灌注前、后的病理评分差异极其显著(P<0.01).结论 本研究证明在体常温局部灌注对猪DCD30 min肝脏具有修复作用.

近年来,随着国家器官移植改革不断推进不断深化,我国器官捐献与移植事业取得了举世瞩目的成就.根据2019年12月发布的《中国器官移植发展报告(2015 —2018)》,自2015年1月1日 —2018年12月31日,我国公民逝世后器官捐献累计完成18294例,2018年完成公民逝世后器官捐献6302例,器官移植手术20201例,捐献数量和移植例数均位居亚洲首位,全球范围仅次于美国.我国每百万人口器官捐献率(per million population,PMP)从2015年的2.01上升至2018年的4.53.随着器官捐献理念不断宣传,庞大人口基数前提下的PMP增长,我国完全具备成为全球器官移植第一大国的潜力.

目的 利用离体肝脏常温灌注系统初步探索门静脉恒压控制模式与门静脉恒流控制模式对热缺血损伤肝脏的保存效果差异.方法 使用雌性巴马小型猪获取血液和供肝,所有供肝的热缺血时间为60 min,一组采用门静脉恒压控制模式(n=6),一组采用门静脉恒流控制模式(n=6),利用常温机械灌注设备灌注6 h,观察肝脏的灌注参数、血气生化指标以及组织病理学变化.结果 灌注前,两组的冷缺血时间差异无统计学意义(P>0.05).猪肝脏经过6 h的常温机械灌注,恒压组的肝动脉流量为(0.13±0.08)mL/min/g(肝重)比恒流组的(0.25±0.09)mL/min/g(肝重)更低(P<0.05).两组比较,灌注液的pH、乳酸水平、葡萄糖水平差异无统计学意义(P>0.05);尿素氮水平除了在第2小时,两组比较差异无统计学意义(P>0.05).对于肝细胞酶学和总胆红素水平,两组在灌注结束时亦差异无统计学意义(P>0.05).恒流组的胆汁生成量为(54.47±15.63)mL,较恒压组的(6.98±3.20)mL更多(P<0.05).两组在病理学评分的比较差异无统计学意义(P>0.05).结论 门静脉恒流模式更有利于肝动脉的供血,从而促进肝细胞进行胆汁合成,提示门静脉流量控制模式可能是更合适的灌注方式.但是这仍需要通过动物肝移植实验以及临床肝移植的数据进一步验证.

目的 探讨便携式在体机械灌注设备(IMPD)运行稳定性和保护无心跳大动物供器官的作用.方法 6只健康清洁巴马小型猪麻醉后开腹,分别经腹主动脉、下腔静脉插管并连接到自主研发的便携式IMPD,静脉注射氯化钾骤停心脏,观察5 min无复跳,启动设备对动物器官进行常温(37℃)机械灌注.调整离心泵转速和灌注流量以控制静脉血氧饱和度维持在60％ ～70％,持续灌注12 h,每2小时为观察时间点,记录设备管路流量、离心泵转速和流量/转速比值,检测肝、肾功能和血液内环境指标,灌注结束后获取胆总管、肝、肾和小肠组织,HE染色后光镜下评估病理改变情况.采用重复测量资料单因素方差分析比较灌注过程中各时间点运行参数、肝肾功能和血液内环境指标,组内两两比较采用LSD法,P<0.05为差异有统计学意义.结果 全部实验均完成12 h灌注,灌注过程中设备运行稳定,无故障或停机意外,管路流量、离心泵转速和流量/转速比值均稳定.ALT、总胆红素(TBIL)和谷氨酰转移酶(GGT)稳定,灌注开始至结束各时间点与术前相比,差异均无统计学意义(P均>0.05).灌注开始后AST缓慢上升,灌注第8、10和12小时AST分别为(73±21)、(90±30)和(114±48)(U/L),与术前相比,差异均有统计学意义(P均<0.05).在实验猪心脏停跳的5 min内,乳酸在灌注开始时便处于峰值,自灌注第8小时起,乳酸数值与峰值相比差异均有统计学意义(P均<0.05).K+浓度水平自灌注第2小时起各时间点与灌注开始时相比,差异均有统计学意义(P均<0.05).ALT、TBIL、GGT、血尿素氮、血清肌酐、血气分析指标、Na+、Ca2+和Cl-浓度水平稳定,灌注开始至结束各时间点数值与灌注开始时相比,差异均无统计学意义(P均>0.05).自灌注开始后持续有胆汁生成、尿量产生.供肝灌注12 h的过程中,肝脏始终保持外观红润、质地柔软、边缘锐利.灌注结束后,胆总管、肝、肾和小肠组织病理结果 示组织结构完整,无淤血和大面积坏死征象.结论 便携式IMPD可以稳定灌注无心跳猪供器官12 h,可以较好维护胆总管、肝、肾和小肠器官.