联合应用骨髓间充质干细胞与促肝细胞生长素治疗大鼠急性肝衰竭

目的 评价骨髓间充质干细胞(MSCs)和促肝细胞生长素(pHGF)联合治疗大鼠急性肝衰竭(ALF)的可行性.方法 将ALF造模成功的37只SD大鼠分为4组:CC14组(A组,9只),CCl4/MSCs(B组,9只),CC14/pHGF组(C组,9只)和CC14/pHGF+ MSCs组(D组,10只).用携带hrGFP基因的慢病毒感染MSCs(hrGFP-MSCs),并以hrGFP示踪.对C、D组大鼠给予CCl4,同时腹腔注射pHGF;B组和D组造模12h后肝内注射hrGFP-MSCs悬液.对各组大鼠分别在造模后24 h、72 h、7天检测血清丙氨酸转氨酶(ALT)水平,造模后72 h检测血清IL-6、IL-10、TNF-α炎症因子水平,造模后24 h取肝组织行HE染色,造模72 h行PCNA免疫组化染色检测肝细胞增殖情况.造模后72 h后取注射部位肝组织行冰冻切片观察移植hrGFP-MSCs肝内分布情况,对非注射部位肝组织进行实时荧光定量RT-PCR检测hrGFP基因含量.结果 联合治疗后,ALF大鼠外周血ALT水平降低,IL 6、IL-10、TNF-α炎症因子水平下调,肝脏病理组织学损害减轻,肝细胞增殖率增高.造模后72 h,B组与D组注射部位肝组织内可见移植的hrGFP-MSCs,非注射部位肝组织未见hrGFP荧光阳性细胞,但用实时荧光定量RT-PCR技术可在非注射部位肝组织内检测到hrGFP基因,且D组含量高于B组.结论 pHGF与MSCs联合治疗大鼠ALF的疗效优于MSCs或pHGF单独治疗,可能与pHGF促使其肝内定植MSCs增多有关.     

多模态成像对间充质干细胞移植后活体示踪的应用研究

目的 探讨生成像（BLI）和MRI相结合对移植入大鼠损伤肝脏内的间充质干细胞（MSCs）的实时、无创示踪的能力及移植的MSCs对大鼠损伤肝脏的功能修复作用。方法 对急性肝损伤大鼠模型移植标记了荧光素酶和SPION的MSCs,分析大鼠的血浆转氨酶水平及肝脏病理学改变,观察MSCs对肝功能损伤的修复作用,并利用BLI和MRI观察MSCs在大鼠肝脏内积累的时间进程,最后进行大鼠肝脏组织切片的原位免疫组织化学染色。结果 MSCs移植后的第5、9、10天检测的大鼠血浆天冬氨酸转氨酶和丙氨酸转氨酶水平较对照组显著降低（P均<0.05）。MSCs移植后第1天,大鼠肝脏区域的生物发光强度显著增强,T2^*WI标准化信号强度明显减低,于移植后第10天才逐渐恢复到移植前水平。并且随着大鼠肝脏生物发光强度的逐渐下降,荧光素酶阳性的MSCs数目亦随时间进程逐渐减少。结论 移植MSCs对大鼠损伤肝脏的功能具有良好的修复作用;且将BLI和MRI相结合可实时、无创示踪移植入大鼠损伤肝脏内的MSCs。     

CM-DiI标记大鼠脂肪干细胞的效力

背景:CM-DiI是DiI的衍生物,因具有一定的水溶性,且含有氯甲基化活性巯基部分,使其更易嵌入、弥散并稳固地结合到整个细胞膜上,使染色更快捷、均匀、持久。目的:进一步验证荧光染料CM-DiI标记大鼠脂肪干细胞的可行性。方法:切取大鼠腹股沟区皮下脂肪组织,采用胶原酶消化法体外分离培养脂肪干细胞,取第3代细胞分为实验组和对照组,用质量浓度4mg/L的CM-DiI对实验组细胞进行标记,于6,12,24,48h观察CM-DiI标记脂肪干细胞体内示踪效果。结果与结论:荧光显微镜下CM-DiI标记细胞的胞浆、胞膜均显红色荧光,标记的脂肪干细胞呈梭形,保持了良好的正常形态,CM-DiI标记阳性率为100%,标记早期细胞形态呈荧光环状,48h后细胞中荧光颗粒增多,荧光增强,细胞核未染荧光。与对照组比较,CM-DiI标记的细胞增殖后形态、上清液乳酸脱氢酶含量及MTT值均无明显变化(P>0.05)。细胞移植后4h后,在心、肺组织可见到发出红色荧光的标记细胞。提示CM-DiI能有效标记体外培养的脂肪干细胞,并在细胞内稳定表达,且标记细胞形态良好,对活体细胞无毒性。体内移植后,有良好的示踪效果。     

GFP标记干细胞修复椎间盘退变中的示踪研究

目的探讨绿色荧光蛋白(GFP)示踪时效及GFP标记的骨髓间充质干细胞(MSCs)在移植后参与退变修复过程中的细胞增殖情况。方法使用针吸髓核法建立兔的椎间盘退变模型,体外培养MSCs,并用含有绿色荧光蛋白标记的腺病毒载体(Adeno-XTM-GFP)转染,成为GFP标记的MSCs(GFP-MSCs),于造模2周后植入造模的椎间盘内。在移植后第2、4、6、8周用激光共聚焦显微成像技术,观察MSCs的增殖情况,对比GFP本身荧光和免疫荧光的示踪时间。结果非免疫荧光组GFP本身荧光强度能够持续到4周,后较微弱甚至难以辨别;免疫荧光组用FITC荧光二抗标记的GFP-MSCs在8周内都能够持续稳定的显现荧光,4个时间点GFP-MSCs的阳性率的可信区间分别是:2周(14.64±2.05)%;4周(21.85±2.45)%;6周(31.03±4.03)%;8周(36.37±4.42)%。结论GFP-MSCs在椎间盘微环境中能够稳定增殖。若示踪研究需要持续超过4周,则应对GFP行免疫荧光染色,可以使有效示踪时间持续至少8周。     

实时二维剪切波弹性成像评价急性肝衰竭大鼠肝脏硬度

目的 探讨实时二维剪切波弹性成像（2D-SWE）对于检测大鼠急性肝衰竭（ALF）和指导预防性用药治疗的应用价值。方法 将28只雄性SD大鼠分为4组，即对照组（8只）、模型组（8只）、乳果糖组（6只）和美常安组（6只）。实验第1~8天，对美常安组予以美常安稀释后灌胃，乳果糖组予以乳果糖灌胃，对照组和模型组予以等量生理盐水。实验第6、7天，对模型组、乳果糖组和美常安组大鼠均腹腔注射硫代乙酰胺，对照组注射生理盐水。实验第8天，采用2D-SWE技术测量4组大鼠肝脏硬度值（LSM）。构建ROC曲线，根据AUC选出准确率最高的LSM进行组间比较。实验第9天，取大鼠肝组织行病理检查。结果 模型组主要病理表现为炎症、坏死；美常安组及乳果糖组炎症坏死程度较模型组减轻；对照组表现为正常肝组织。根据模型组LSM构建的ROC曲线中，平均值诊断效能最佳；以8.44 kPa为截断值，其AUC、敏感度和特异度分别为0.768、100%和62.50%。模型组LSM较其他3组明显升高（P均<0.05）。结论 ALF大鼠LSM增高。2D-SWE对检测ALF及预防性用药后疗效评估具有一定价值。     

诊断超声介导微泡提高肝纤维化通透性及促进基因传递

目的 探讨诊断超声联合微泡对肝纤维化组织通透性的影响及其介导基因转染肝纤维化大鼠的有效性。方法 采用二甲基亚硝胺(DMN)法建立大鼠肝纤维化模型,80只大鼠在建模第4周末随机分为:模型对照组、单纯微泡组、单纯超声组和诊断超声联合微泡组。分别进行肝纤维化微血管通透性实验和基因转染实验,采用激光共聚焦显微镜观察伊文思蓝(EB)在肝纤维化组织内分布情况,同时定量检测肝纤维化组织内EB的含量,评估不同分组微血管通透性。荧光显微镜下观察含增强型绿色荧光蛋白报告基因的质粒转染大鼠肝纤维化模型的基因表达情况。结果 激光共聚焦显微镜显示诊断超声联合微泡组纤维化肝实质内可见明显的EB红色荧光。诊断超声联合微泡组纤维化肝组织中EB含量明显高于其他3组(P<0.05)。荧光显微镜下观察,相比其余3组,诊断超声联合微泡组增强型绿色荧光蛋白最多,基因转染效率最高。结论 诊断超声联合微泡在提高纤维化肝脏微血管通透性的同时可促进基因传递。     

肝纤维化指数与大鼠肝炎症分级的相关性

目的 探讨肝纤维化指数(LF Index)与大鼠肝炎症分级的相关性。方法 对45只雄性SD大鼠尾静脉注射猪血清诱导免疫性肝炎,7只雄性SD大鼠注射生理盐水为正常对照组。于造模的第2、4、6、8、10、12和14周,从造模组与正常对照组分别随机取6只和1只大鼠进行实时组织弹性成像(RTE),得到LF Index。随后处死大鼠,取肝组织进行病理学检查,并进行肝炎症分级。依照炎症分级将大鼠分为正常对照组、G0亚组、G1亚组、G2亚组、G3亚组和G4亚组,比较各组的LF Index。结果 42只大鼠造模成功,其中炎症分级为G0级、G1级、G2级、G3级和G4级的各有6、12、10、10和4只。LF Index与炎症分级具有相关性(r=0.768,P<0.05)。以LF Index绘制肝炎症分级的ROC曲线,≥G1、≥G2、≥G3、G4的曲线下面积分别为0.851、0.911、0.928 和0.853。结论 炎症分级与LF Index具有相关性,炎症分级会影响LF Index评价肝纤维化程度的准确性。     

携带人肝细胞生长因子的腺病毒转染人骨髓间充质干细胞及超顺磁性氧化铁标记细胞体外MR成像

目的 评价携带人源化绿色荧光蛋白(hrGFP)-人肝细胞生长因子(hHGF)的腺病毒载体对人永生化骨髓间充质干细胞UE7T-13生物学特征的影响,并探讨对超顺磁性氧化铁(SPIO)标记细胞进行体外MR成像的可行性。 方法 构建和包装携带hrGFP-hHGF基因的腺病毒载体;以hrGFP-hHGF腺病毒转染UE7T-13细胞,检测细胞中hrGFP表达阳性率、hHGF mRNA及细胞上清液中hHGF水平;检测hrGFP-hHGF腺病毒对H₂O₂诱导细胞凋亡的影响。以SPIO标记UE7T-13细胞,检测标记后细胞内铁含量、细胞增殖及分化能力;对不同铁浓度SPIO标记的细胞行MRI。结果 成功构建hrGFP-hHGF腺病毒载体,将其转染UE7T-13细胞48 h后hrGFP阳性表达率达93.17%,hHGF mRNA表达提高3075.63倍,细胞上清液中hHGF水平显著升高,随后下降,于第14天仍高于hrGFP腺病毒转染细胞。hrGFP-hHGF腺病毒可抑制H₂O₂介导的细胞凋亡。SPIO标记后细胞铁染色阳性率达100%,细胞铁含量明显高于未标记细胞;SPIO标记不影响细胞增殖及分化能力;T2WI信号随标记铁浓度增高而降低。 结论 hrGFP-hHGF腺病毒载体可抑制H₂O₂介导的细胞凋亡;SPIO能高效标记细胞,不影响细胞增殖及分化能力,可用于细胞体外MR成像。     

超低微浓度菲立磁-硫酸鱼精蛋白复合物标记骨髓间充质干细胞的方法

目的 探索简单、无细胞外铁产生的超低微浓度菲立磁-硫酸鱼精蛋白标记骨髓间充质干细胞(MSCs)方法。 方法 贴壁法培养大鼠MSCs。待3代细胞汇合至80%~90%时,更换无血清培养液,根据菲立磁和硫酸鱼精蛋白的不同浓度分为4组:A组[(7.50∶1.00)μg/ml]、B组[(10.00∶1.20)μg/ml];C组[(15.00∶1.80)μl/ml]和空白对照组,加入培养液,混匀,5% CO₂孵育15 min,补加血清后孵育至次日。检测细胞标记率、细胞内外铁、细胞活力和标记细胞MR信号。 结果 B组可有效标记大鼠MSCs,普鲁士蓝染色阳性率100%,无细胞外铁产生,铁颗粒分布于溶酶体内。4组间台盼蓝拒染率差异无统计学意义(P>0.05);体外MR GRE T2^*WI序列可检测到1×10⁴个标记细胞。 结论 使用超低微浓度菲立磁10.00 μg/ml与鱼精蛋白1.20 μg/ml可有效标记大鼠MSCs,体外MR可检测到1×10⁴个标记细胞。     

光磁双模态成像显示大鼠脑组织间液引流路径及物质清除规律

目的 采用荧光示踪成像和磁示踪成像观察大鼠脑组织间液(ISF)引流路径及物质清除规律。方法 将33只雄性SD大鼠随机分为荧光示踪组(F组,n=18)和磁示踪成像组(MRI组,n=15),再细分为丘脑亚组、海马亚组和尾状核亚组。对F组大鼠注射伊文斯蓝并行心脏灌流,取脑组织制成冰冻切片,观察各亚组示踪剂引流路径;对MRI组大鼠分别于注射钆喷酸葡胺(Gd-DTPA)前、后行MR扫描,对比各亚组脑区ROI信号强度,计算单位信号比值并观察其变化趋势。结果 F组大鼠丘脑、海马、尾状核IFS引流分别存在优势路径,其到达相邻脑区及全脑的时间存在均差异。MRI组注射Gd-DTPA后4 h内丘脑、海马、尾状核示踪剂清除方向性及速率有所不同,表现为丘脑及海马区示踪剂向同侧皮层及侧脑室方向引流、尾状核区示踪剂向皮层和中脑方向扩散,且相邻引流脑区示踪剂信号达峰时间存在差异。结论 光磁双模态成像显示,大鼠丘脑、海马及尾状核ISF优势引流路径及其内小分子物质清除速率均存在差异。     

异位辅助肝移植治疗急性肝功能衰竭的实验研究

目的：探讨异位辅助性部分肝移植治疗急性肝功能衰竭的可行性。方法；制作息性肝功能衰竭犬动物模型26只。分为两组：移植组20只，对照组6只。移植组切除脾脏后，于脾窝处移植同系异体60％部分肝脏，手术成功17只。观察两组实验动物的存活时间、血液生化、残肝磁共振（MRI）检查、残肝和移植肝的组织细胞形态改变。结果：对照组和移植组实验动物72h存活率分别为16．7％和82．7％。移植术后2周残肝细胞明显增生，肝功能恢复近正常，而辅助肝渐萎缩，术后5周辅助肝完全纤维化。结论：急性肝功能衰竭时，异位辅助性部分肝脏移植可为病肝提供暂时性功能支持，为残肝细胞再生、功能恢复提供机会，同时移植肝渐萎缩、纤维化。     

The liver as A self-organizing system. I

Summary The liver is described as a composite system consisting of a set of operative creodic microunits open to a continuous flow of matter, energy and informations. Its dynamics depend on two interactive and interrelated subsystems with actions described ashomopoiesis andhomeorhesis, making it anautoisodiasostic system. The system’s emergent (equifinal) or emergence states, operative potential, diffusion and reaction phenomena and compensation states are also formally described. For readers not familiar with the language of general system theory, of system dynamics and of categorical analysis, a glossary of some terms is provided. This paper forms part of research on pre-cancer states of the liver system, financed by theAssociazione Italiana per la Ricerca sul Cancro (AIRC), Milan and by theOspedale Maggiore — Istituto Scientifico di Ricovero e Cura — Milan.     

MR imaging of traumatic hepatic laceration: Evaluation and course of healing after surgery

Six patients with hepatic laceration underwent magnetic resonance imaging (MRI) at 0.5T. Acute hepatic laceration was slightly hypointense on T1-weighted spin-echo (SE) image, and hyperintense on T2- and proton-weighted SE images. Subacute laceration was heterogeneously intense on T1-weighted image and hyperintense on T2- and proton-weighted images. Consistent changes in signal intensity of postoperative hepatic laceration were observed. On Tl-weighted image, the signal intensity at first increased and then decreased from periphery to the center. On the T2- and proton-weighted images, the laceration was uniformly hyperintense relative to the liver prior to the appearance and growth of a hypointense ring at its periphery. The appearance of the above changes in signal intensity was also observed in postoperative recurrent hemorrhage. The postoperative biloma had none of the above changes in signal intensity. Our cases show that MRI is effective in the evaluation of hepatic laceration and in the assessment of the course of healing after operation.     

Hepatocellular carcinoma with or without cirrhosis: A comparison of CT and angiographic presentations in the United States and Japan

We compared the computed tomographic (CT) and angiographic presentations of hepatocellular carcinoma (HCC) with or without cirrhosis in the United States and Japan. Tumors in the United States were advanced and less frequently associated with liver cirrhosis (association of cirrhosis: United States) 56.2%, Japan 91.0%. In patients with cirrhosis, the size of the tumor tended to be smaller, and nodular tumors (single or multiple) were frequent. In early stage of HCCs with cirrhosis, tumors were hypovascular without a capsule. In advanced stage, tumors were hypervascular and a capsule was frequently observed around the tumor both with CT and angiography. HCCs without cirrhosis were seen in younger patients. These tumors were large at the time of diagnosis. A massive or diffuse mass without a capsule was frequently seen. Most tumors were hypodense on precontrast CT and hypervascular on angiography. Lymph node enlargement was significantly frequent. The radiological characteristics of HCC in both countries were significantly different depending upon associated cirrhosis, as well as the time of the diagnosis.     

Does CTAP prior to hepatic resection improve patient survival rates?

The purpose of our study was to compare survival rates of colon carcinoma patients who had undergone attempted curative hepatic resection based on liver staging by computed tomographic angiography (CTA) or portography (CTAP) with previously reported survival rates of patients who underwent similar surgery without preoperative CTAP evaluations. A total of 404 CTAP studies performed at three institutions were reviewed. Of this group, 197 had colon carcinoma. Sixty-nine of the colon patients went to surgery. Actuarial adjusted yearly survival rates were calculated for the prior CTAP colon group and compared to historical controls. The control survival data were taken from reports published prior to the CTAP era. Our study demonstrated no difference in the 1-year survival data between the groups. However, the CTAP patients had greater survival in years 2–4. This greater survival may be multifactorial but in part due to better surgical selection caused by CTAP.An invited commentary on this article follows on pp. 320-324.     

Liver manifestations in COVID-19 patients: A review article

The coronavirus disease 2019 (COVID-19) initially presented as a disease that affected the lungs. Then, studies revealed that it intricately affected disparate organs in the human body, with the liver being one of the most affected organs. This review aimed to assess the association between COVID-19 and liver function, shedding light on its clinical implication. However, its exact pathophysiology remains unclear, involving many factors, such as active viral replication in the liver cells, direct cytotoxic effects of the virus on the liver or adverse reactions to viral antigens. Liver symptoms are mild-to-moderate transaminase elevation. In some patients, with underlying liver disease, more serious outcomes are observed. Thus, liver function should be meticulously considered in patients with COVID-19.     

Undifferentiated embryonal sarcoma of the liver treated with chemotherapy: CT imaging in four patients

Background: The purpose of this study was to describe the computed tomography (CT) findings of undifferentiated embryonal sarcoma after chemotherapy and to correlate the CT imaging findings with pathologic findings. Methods: Ten CT images obtained before and after chemotherapy in four patients with hepatic undifferentiated embryonal sarcoma were retrospectively reviewed and correlated with pathologic findings. Results: After chemotherapy, tumor volume decreased by 50–90% and initially nonresectable tumor or gross residual tumor was successfully excised in three patients. In all patients, enhancing peripheral solid portions and septations changed to low-attenuation areas, and in three patients increased or de novo calcifications were found at the periphery of the tumor. Resected pathologic specimen after chemotherapy showed well-encapsulated masses with central necrosis, fibrosis, and dystrophic calcifications. Conclusions: These CT findings will be useful in monitoring the treatment response of hepatic undifferentiated embryonal sarcoma during chemotherapy.     

三维MRA技术在肝脏血管成像中的应用研究

目的：探讨三维增强MRA（3D CEMRA）技术在肝脏磁共振血管成像中的应用价值。方法：52例行肝脏磁共振血管成像患者及30例对照组病人，利用自动透视触发技术进行肝脏血管成像，造影剂到达降主动脉起始段时启动3D扫描，扫描时相包括肝动脉期、门静脉期及腔静脉期，分离各组图像并与蒙片剪影，再进行最大信号强度投影（MIP）及多平面图像重建（MPR），评价各期图像质量。结果：采用3DCEMRA技术所获肝动脉期及门静脉期图像质量满意；造影剂到达降主动脉起始段时间与造影剂流速、总量之间呈线性相关；造影剂总量和流速与肝血管信号强度呈正相关。结论：3DCEMRA技术可准确把握造影剂到达靶血管时间，结合最佳的造影剂总量和注射速度，明显提高肝脏磁共振血管成像质量。     

人工肝支持系统在肝衰竭中的应用经验和进展

人工肝支持系统对肝衰竭的治疗作用已经得到公认,可以使部分患者病情得到恢复,还可以使患者有效地过渡到肝脏移植,分为非生物型人工肝、生物型人工肝和混合型3种,包括血浆置换、血液滤过、血液/浆灌流、连续性血液透析滤过、血液透析、白蛋白透析、血浆胆红素吸附和生物型人工肝等。目前常应用的方法有血浆置换、连续性血液透析滤过和血浆胆红素吸附,其作用机制就是通过机械性的方法祛除肝衰竭患者体内的毒性物质及代谢产物,补充部分活性物质,纠正凝血物质缺乏,使患者内环境改善,利于肝细胞再生,病情恢复。     

酒精性脂肪肝和肝硬化患者各种血清酶的检测及其临床评价

目的 探讨酒精性脂肪肝血清特异性的鉴别指标，并用其来判断病程，指导临床诊断和治疗。方法 对76例酒精性脂肪肝、62例酒精性肝硬化以及70例非酒精性脂肪肝患者的血清酶和血脂进行测定及分析。结果 与非酒精性脂肪肝相比，酒精性脂肪肝的谷草转氨酶（AST）和γ-谷氨酰转肽酶（GGT）的异常率以及谷草转氨酶／谷丙转氨酶（AST／ALT〉1）的构成比的增高，差异均具有统计学意义；与酒精性脂肪肝相比，酒精性肝硬化的AST／ALT比值，AST和GGT的水平都有明显增高。结论 AST、AST／ALT（〉1）、GGT等指标对鉴别酒精性脂肪肝以及判断其病程，具有一定的临床意义。