肝癌化疗栓塞术后肝细胞周期的研究

目的　探讨经动脉化疗栓塞术 (TACE)后 ,肿瘤周围正常肝组织的细胞周期变化及其变化特点。材料与方法　VX2肝癌模型兔 5 0只 ,分为A～E 5组 ,每组 10只 ,分别行TACE和肝癌肝叶切除术 (PH)。术后第 1、第 3天取兔肝组织行流式细胞术 (FCM) ,进行细胞周期分析 ,测定S期细胞比率 (SPF) ,增殖指数 (PI)的变化 ,并测定有丝分裂指数 (MI)。结果　TACE术后第 1天兔MI、SPF、PI较对照组均明显升高 (P <0 .0 0 1) ;术后第 3天 ,所有的指标较对照组均无明显差异 (P >0 .0 5 )。并且术后第 1天 ,TACE术后肝细胞增殖弱于肝癌肝叶切除术 ,各项指标差异明显 (P <0 .0 5 )。结论　TACE术后兔肝组织存在一定程度的DNA复制与细胞增殖再生 ,但弱于肝叶切除术后的肝细胞增殖再生 ,其原因可能与碘油的选择性栓塞和化疗药物作用有关。     

人肝再生增强因子的cDNA克隆与序列分析

利用大鼠肝再生增强因子核苷酸序列，从人胎肝ｃＤＮＡ文库中筛选到人肝再生增强因子ｃＤＮＡ克隆，该克隆含有１．５ｋｂ的外源插入片段，核苷酸序列测定表明含３７５ｂｐ的读码框架，能编码１２５个氨基酸的蛋白质；与大鼠肝再生增强因子相比，核苷酸序列测定表明含３７５ｂｐ的读码框架，能编码１２５个氨基酸的蛋白质；与大鼠肝再生增强因子相比，核到序列同源性达８７％，氨基酸序列同源性达８４．８％。     

肝癌经皮酒精注射术后肝再生的研究

目的：研究经皮酒清注射术（PEI）后，周围叶的正常肝脏组织是否存在肝细胞再生，有其再生的特点。方法：VX2肝癌动物模型兔50只，分别行经皮酒精注射术和肝癌肝叶切除术（PH），分别检测术后24h、72h的兔肝组的有丝分裂指数（MI），以流式细胞术（FCM）行DNA含量分析，测量S期细胞比率（SPF），DNA指数（DI）的变化。结果：PEI要后24小时兔肝有丝分裂指数、S期细胞比率、DNA指数较对照组均明显升高（P＜0．001）；术后72h，有丝分裂指数、S期细胞比率较对照组均无明显差异（P＞0．05），DNA指数较对照组升高（t＝2．78，P＞0．05）。并且术后24h，PEI要后肝细胞增殖明显弱于肝癌肝叶切除要，各项指标差异明显。结论：PEI要后兔肝组织存在一定程度的再生，弱于肝叶切除要后再生，其原因可能与酒精的局限性扩散和肝内酒精浓度升高作用有关。     

人肝再生增强因子的cDNA克隆、高效表达及生物学活性研究

人肝再生增强因子的ｃＤＮＡ克隆、高效表达及生物学活性研究１００８５０北京军事医学科学院放射医学研究所杨晓明，谢铃，贺福初，宫锋，何浩，邱兆华，吴祖泽关键词细胞活性因子；肝疾病中国图书资料分类号Ｑ２６人胎肝组织中存在一种热稳定性、特异性促进肝细胞增殖的...     

肝切除术后肝脏再生的研究进展

肝切除术是目前治疗各种肝脏良恶性疾病的有效方法,但扩大性肝切除术后易出现肝衰竭,其治疗仍是世界性难题。虽然肝移植是有效治疗途径,但供体严重不足限制了肝移植的广泛开展。现有的生物人工肝技术仍不能完全替代肝脏功能,肝细胞移植体内后增殖有限,也限制了其治疗效果。肝脏与其他器官不同,具有巨大的潜在再生能力。肝切除术后,剩余肝脏内实质细胞、非实质细胞及体内各种细胞因子、生长因子,还有各类肝干细胞等能够迅速做出回应,并很快使肝脏体积恢复至原来大小,肝功能恢复正常。肝脏再生大致分为三个阶段,即启动阶段、增殖阶段、终止阶段。肝脏再生的各个阶段机制复杂,影响因素多,至今仍未完全明了。探索肝细胞再生机制可为临床促进肝再生、防治肝衰竭奠定理论基础。肝切除术后肝脏再生已成为近年的研究热点,本文就近年来肝切除术后肝脏再生的研究进展做一综述。     

肝叶(段)切除治疗肝内胆管结石

近１３年手术治疗肝内胆管结石１０５例，其中３８例作了肝叶（段）切除术。左肝叶（段）切除３４例，右肝叶（段）切除４例。２０例同时作了胆管空肠Ｒｏｕｘ－Ｙ吻合术。术后发生并发症８例，其中死亡１例。３７例获随访。治疗效果优良者为８７％（３３／３８例）。本文对肝叶（段）切除治疗肝内胆管结石的适应证，手术方法等进行了讨论。作者认为，肝叶（段）切除术是治疗肝内胆管结石的一种有效的方法。     

肝脏损伤肝叶切除术的适应证是什么?

解答:肝脏损伤肝叶切除术的适应证为:(1)局部毁损伤,切除损伤的肝组织有利于修复;(2)肝后静脉伤,需要切除部分肝组织以利显露;(3)肝损伤出血,用各种方法难以止血;(4)肝左外叶严重损伤。     

胃肠外营养对肝切除后肝脏再生的影响

目的：探讨胃肠外营养对肝脏再生的影响．方法：将大鼠随机分成４组：Ⅰ组不手术，自由摄食；Ⅱ组６５％肝切除后用１０％葡萄糖治疗；Ⅲ组术后用支链氨基酸强化的胃肠外营养治疗；Ⅳ组术后用标准胃肠外营养治疗．实验第三、六天观察肝脏再生反应．结果：Ⅱ组肝重、肝重／体重、肝脏再生率（ＲＬＲ）、肝细胞有丝分裂率（ＭＩ）和ＤＮＡ合成率明显降低，肝细胞脂肪变性和水样变性明显且伴有灶性坏死；Ⅲ、Ⅳ组ＲＬＲ、ＭＩ及ＤＮＡ合成率明显增加，肝细胞脂肪变性和水样变性减轻，其中Ⅲ组明显优于Ⅳ组．结论：肝切除后营养摄入不足抑制肝脏再生反应，适当的营养支持能促进肝脏再生     

创伤性肝外胆道损伤的诊断和治疗

目的 探讨创伤性肝外胆道损伤的诊治方法。方法 回顾分析本组17例创伤性肝外胆道损伤的诊治资料。结果 17例均于术中确诊。10例胆囊破裂均行胆囊切除术；4例胆总管部分破裂行缝合修补术并“T”管引流；2例胆总管横断行胆总管空肠Roux-en-Y吻合术；1例胰腺段胆管断裂行胰十二指肠切除术，术组17例均治愈。结论 创伤性肝外胆道损伤术前诊断困难，常于术中确诊，治疗可采用胆囊切除术，一期缝合修补术，胆肠吻合术和胰十二指肠切除术等手术治疗，包括单纯左，右单侧肝管结扎术。     

少见胰腺、胃壁、肝、脾联合伤1例

病例女，12岁。患者全身多处被垮塌房屋砸伤，当时疼痛难忍，其家人急送我院进行救治。CT 检查诊断为：（1）脾破裂；（2）双肺挫伤、左侧肋骨骨折、双侧胸腔积液；（3）胰腺挫伤；（4）胃壁挫伤；（5）腹膜后血肿；（6）左侧锁骨骨折。立即行脾切除术、左侧胸腔闭式引流术，术后给予肌注破伤风抗毒素、输血600 ml、止血、抗炎及补液及对症治疗，患者术后病情稳定。外伤后3 d 胸腹部 CT 检查示：（1）双上肺挫伤；右肺下叶不张，右下叶支气管轻度狭窄；左肺下叶血肿；双侧少量胸腔积液；左侧多根肋骨骨折，左侧胸壁软组织挫伤并血肿；左侧锁骨骨折。（2）肝右叶条片状低密度影，考虑：挫伤；（3）胰腺挫伤，较上次 CT 片有所好转；（4）脾脏术后改变；（5）胃底后壁挫伤，较上次 CT 片有所好转；（6）双肾体积增大，皮髓界限不清，考虑：肾挫伤可能；（7）腹膜后血肿。外伤后12 d 胸腹部 CT 复查示：（1）右肺下叶不张有所好转；左肺下叶血肿有所好转；左侧肋骨及锁骨骨折无明显变化。（2）肝右叶挫伤，范围有所增大；（3）胰腺、胃壁、双肾形态、密度未见异常；（4）腹膜后局限性血肿，较上次 CT 片有所缩小，边缘更清晰。经过治疗，患者病情明显好转。外伤后14 d 行上腹部 CT 复查示：（1）肝右叶迟发性血肿；（2）腹膜后血肿明显缩小。     

三氧化二砷逆转乳腺癌细胞株MCF-7/ADM多药耐药的研究

目的探讨三氧化二砷（As2O3）体外逆转人乳腺癌细胞多药耐药性的作用及机制。方法采用MTT法检测As2O3的细胞毒作用和处理前后耐药细胞对化疗药物的敏感性,用流式细胞仪检测细胞内阿霉素浓度,通过RT-RCR检测MDR1基因的表达。结果 As2O3在0.25mg/L以下剂量时对MCF-7和MCF-7/ADM耐药细胞株的抑制率均小于15%,半数抑制率（IC50）分别为1.01m/L和1.28mg/L,无细胞毒剂量0.2mg/L的As2O3能部分逆转MCF-7/ADM细胞对阿霉素的耐药性。同时无细胞毒剂量0.2mg/L的As2O3能使MCF-7/ADM细胞内阿霉素浓度明显增加,MDR1表达下降。结论 As2O3具有体外部分逆转人乳腺癌细胞多药耐药性的作用,可能与增加细胞内药物积累、下调MDR1表达有关。     

ABCG_2食管癌耐药细胞系Eca109/ABCG_2的建立

目的 探讨腺苷三磷酸结合转运蛋白G超家族成员2(ABCG_2)在介导食管癌多药耐药中的作用.方法 扩增ABCG_2基因,与pcDNA3.1载体连接,构建pcDNNA3.1-ABCG_2重组质粒,转染Eca109细胞后用G418筛选稳定转染细胞.采用流式细胞术检测细胞内阿霉素的含量,评价细胞对药物的外排作用.MTT法检测阿霉素对Eca109/ABCG_2、Eca109/pcDNA3.1、Eca109细胞生长的抑制作用,并计算半数抑制浓度(IC_(50))和细胞的耐药指数.采用Western blotting和RT-PCR法检测细胞中ABCG_2蛋白及mRNA的表达量.结果 成功建立转染ABCG_2基因的Eca109/ABCG_2细胞.Eca109/ABCG_2细胞中ABCG_2的mRNA和蛋白表达量升高.阿霉素对Eca109/ABCG_2、Eca109/pcDNA3.1、Eca109细胞的IC_(50)值分别为18.61±3.94、4.18±0.14、4.69±0.88μg/ml.Eca109/ABCG_2细胞对阿霉素的耐药指数为3.97,且外排阿霉素的作用增强.结论 Eca109/ABCG_2细胞具有ABCG_2的耐药表型,能够稳定表达ABCG_2蛋白,可作为研究ABCG_2介导的多药耐药相关机制的细胞模型.     

99mTc glucarate high-resolution imaging of drug sensitive and drug resistant human breast cancer xenografts in SCID mice

BACKGROUND AND AIM: Previous studies have showed that 99mTc labelled glucarate (GLA) might be an agent for non-invasive detection of breast tumours. In xenografted BT20 breast tumours, GLA was found to have higher uptake than 99mTc sestamibi (MIBI). It is unclear whether GLA can localize in all cell line breast cancer xenografts, as well as breast tumours with multidrug resistance (MDR). The present study aimed to investigate the properties of GLA in detecting drug sensitive and drug resistant MCF7 breast cancer xenografts in mice by using dynamic single photon emission computed tomography (SPECT) imaging. METHODS: MCF7/S cells are drug sensitive breast carcinoma cells. MCF7/D40 cells are 40-fold more resistant to doxorubicin compared to MCF7/S. Subcutaneous tumours were grown in SCID mice for 10-14 days after injection of 1 x 10(6) cells into the right thigh. Anaesthetized mice with MCF7/S (MIBI, n=9; GLA, n=8) and MCF7/D40 (MIBI, n=6; GLA, n=5) tumours were imaged using a high-resolution SPECT system called FASTSPECT. Dynamic images were acquired for 2 h after intravenous injection of GLA or MIBI. Expression of MDR P-glycoprotein (Pgp) in the tumours was demonstrated in the MCF7/D40 tumours by western blotting, not in the MCF7/S tumours. RESULTS: The xenografted tumours were visualized unequivocally within 10-30 min in GLA images and remained detectable for at least 2 h after injection. Drug resistant tumours, from which MIBI was rapidly expelled, retained GLA as readily as did drug sensitive tumours. The biodistribution data of GLA demonstrated significantly higher accumulation (%ID/g) compared to MIBI. CONCLUSION: MCF7 tumour xenografts can be detected by 99mTc glucarate imaging. More importantly, 99mTc glucarate can potentially localize drug resistant breast tumours.     

人肝癌多药耐药细胞系 Bel-7402／DOX 两种模型比较

目的：比较体外浓度梯度递增诱导和裸鼠肝脏移植诱导两种方法建立人肝癌多药耐药细胞系Bel-7402/多柔比星（ Doxorubicin ,DOX）模型的生物学特性。方法分别采用体外浓度梯度递增诱导和裸鼠肝脏移植诱导建立人肝癌多柔比星多药耐药细胞亚系Bel-7402/DOXV 和Bel-7402/DOXL 后,利用相差显微镜观察细胞,MTT法检测两种细胞的耐药性,细胞计数法绘制生长曲线并用公式法计算倍增时间,流式细胞仪测定细胞DOX的摄入和外排以及P糖蛋白（ P-gp）、多药耐药相关蛋白（MRP）、谷胱甘肽硫转酶系统（GSH/GST）的表达。结果两组耐药细胞Bel-7402/DOXV 和Bel-7402/DOXL 对DOX、CD-DP均产生了交叉耐药性,较亲本Bel-7402 IC50值差异有统计学意义（P＜0．01）;较亲本细胞的倍增时间明显延长,分别为65 h和46 h;较亲本的DOX外排率明显升高,分别为81．06％、66．56％;两组耐药细胞P-gp、MRP表达较亲本细胞显著提高（ P＜0．01）,而GSH/GST的表达无明显变化。结论两种方法建立的耐药细胞系模型均有稳定的耐药性。肝脏移植法更能高度模拟人肝癌,它是具有近似人肝癌生物学和抗癌药物动力学特征的较理想模型。     

Influence of multidrug resistance on <Superscript>18</Superscript>F-FCH cellular uptake in a glioblastoma model

Purpose  Multidrug resistance, aggressiveness and accelerated choline metabolism are hallmarks of malignancy and have motivated the development of new PET tracers like ¹⁸F-FCH, an analogue of choline. Our aim was to study the relationship of multidrug resistance of cultured glioma cell lines and ¹⁸F-FCH tracer uptake. Methods  We used an in vitro multidrug-resistant (MDR) glioma model composed of sensitive parental U87MG and derived resistant cells U87MG-CIS and U87MG-DOX. Aggressiveness, choline metabolism and transport were studied, particularly the expression of choline kinase (CK) and high-affinity choline transporter (CHT1). FCH transport studies were assessed in our glioblastoma model. Results  As expected, the resistant cell lines express P-glycoprotein (Pgp), multidrug resistance-associated protein isoform 1 (MRP1) and elevated glutathione (GSH) content and are also more mobile and more invasive than the sensitive U87MG cells. Our results show an overexpression of CK and CHT1 in the resistant cell lines compared to the sensitive cell lines. We found an increased uptake of FCH (in % of uptake per 200,000 cells) in the resistant cells compared to the sensitive ones (U87MG: 0.89 ± 0.14; U87MG-CIS: 1.27 ± 0.18; U87MG-DOX: 1.33 ± 0.13) in line with accelerated choline metabolism and aggressive phenotype. Conclusions  FCH uptake is not influenced by the two ATP-dependant efflux pumps: Pgp and MRP1. FCH would be an interesting probe for glioma imaging which would not be effluxed from the resistant cells by the classic MDR ABC transporters. Our results clearly show that FCH uptake reflects accelerated choline metabolism and is related to tumour aggressiveness and drug resistance.     

Photodynamic Therapy with PLGA-Lipid Hybrid Nanoparticles to Overcome the Diverse Mechanisms of Multidrug Resistance in Lung Cancer Cells

Multidrug resistance (MDR) continues to be a critical hurdle to cancer therapy. Two main drug resistance mechanisms have been attributed to MDR, drug-selected MDR can be induced after receiving chemotherapy, and metastasis-associated MDR can be developed by adaptation of the cell in response to changing microenvironmental conditions during metastasis. The objective of the present study was to evaluate the efficacy of nanoparticle-mediated photodynamic therapy (PDT) to overcome MDR of lung cancer cells. The nanoparticles used consist of biodegradable poly(D,L-lactide-co-glycolide) (PLGA) as polymeric core encapsulating a photosensitizer, 5,10,15,20-Tetrakis(4-hydroxy-phenyl)-21H,23H-porphine (pTHPP), wrapped with a mixture of lecithin and PEGylated phospholipids as lipid shell. The resulting nanoparticles had an average particle size of 70.4 ± 1.4 nm and zeta potential value of -39.2 mV with monodisperse distribution. The PDT effect of the nanoparticles was evaluated using two different MDR models established from the same cell line, the A549 human lung adenocarcinoma. An MDR cell line, A549RT-eto, established by continuous exposure to Etoposide, was used a model of drug-selected MDR. Metastasis-associated MDR cells were obtained by culturing A549 cells as floating cells under non-adherent conditions, which simulate metastatic floating cells in the blood or lymphatic circulations, without experience of drug exposure. Compared to the A549 parental cells, the A549RT-eto cells showed 17.4- and 1.8-fold increased resistance to Etoposide and Paclitaxel, respectively. In contrast, the A549 floating cells exhibited resistance to Etoposide (11.6-fold) and Paclitaxel (57.8-fold) compared to A549 attached cells. Both MDR cells were equally sensitive to the photocytotoxic effect of the PDT with pTHPP-loaded nanoparticles. The cellular uptake of pTHPP and light-induced superoxide anion generation were observed at similar levels in both MDR and parental cells. The PDT treatment with nanoparticles induced apoptosis in the two cell lines as detected by flow cytometry. This work suggests that PLGA-lipid hybrid nanoparticles are a potentially effective drug delivery system for using PDT to overcome MDR lung cancer, independent of the mechanisms for MDR.     

卵巢癌细胞株COC1／DDP多药耐药逆转对中性鞘糖脂表达的调控作用

对探讨逆转卵巢癌细胞株的多药耐药性（MDR）对细胞表面中性鞘糖脂（N-GSLs）表达的调控作用,以三苯氧胺（TAM）和维拉帕米（VRP）为逆转剂,用MTT法分析了TAM、VPP的逆转效应,采用改良的Hakamori方法提取,纯化了耐药亚株COC1/DDP MDP逆转前后及亲本株COC1细胞的N-GSLs,高效薄层层析（HPTLC）分析了N-GSLs的表达。结果显示,COC1/DDP和COC1的N-GSLs表达有显著差异,COC1/DDP的CMH表达较COC1强,经TAM、VRP作用后,COC1/DDP的耐药性发生逆转,CMH表达降低。说明卵巢癌的多药耐药性与中性鞘糖脂CMH相关,CMH可能为卵巢癌MDR相关鞘糖脂抗原。     

人卵巢癌拓扑替康耐药细胞株SKOV3/TPT的建立及生物学特性的研究

目的建立人卵巢癌拓扑替康（TPT）耐药细胞株（SKOV3／TPT）,研究其生物学特性和耐药特点。方法模拟临床用药特点,采用大剂量TPT（2160ng／ml）冲击建成人卵巢癌SKOV3／TPT耐药细胞株。MTT法检测多药耐药性,流式细胞仪测定细胞周期、胞内药物的积聚及P糖蛋白（Pgp）表达,从生长曲线、光镜下形态、超微结构等方面比较耐药细胞和SKOV3细胞某些生物学特性的差异。结果成功建立了SKOV3/TPT耐药株,耐药指数为10．67,对喜树碱、米托蒽醌及长春新碱有明显的交叉耐药,对紫杉醇、顺铂、5氟尿嘧啶、足叶乙甙和甲氨蝶呤敏感。与SKOV3细胞相比,SKOV3/TPT耐药细胞倍增时间延长;G2＋M期的比例明显增加（P〈0．01）;细胞内TPT浓度明显减少（P〈0．05）;无Pgp过表达。结论SKOV3/TPT细胞株呈中等水平耐药,耐药性稳定,呈非典型的多药耐药特征;对TPT耐药的主要原因可能与细胞内药物浓度降低和细胞周期改变及凋亡有关,与Pgp的过表达无关。     

Multidrug resistance mediated by ABC transporters in osteosarcoma cell lines: mRNA analysis and functional radiotracer studies

Drug resistance remains a significant impediment to successful chemotherapy and constitutes a major prognostic factor in osteosarcoma (OS) patients. This study was designed to identify the role and prognostic significance of multidrug-resistance (MDR)-related transporters, such as multidrug resistance protein 1 (MDR1), multidrug-resistance-associated protein (MRP1) and breast-cancer-related protein (BCRP), in OS using cationic lipophilic radiotracers. We evaluated the chemosensitivity of four OS cell lines (Saos-2, 143B, MNNG/HOS and U-2OS) to doxorubicin (DOX), cisplatin (CIS) and methotrexate. The expression of MDR-related transporters was analyzed at mRNA level by quantitative polymerase chain reaction and at functional level by 99mTc sestamibi and 99mTc tetrofosmin. The effectiveness of MDR modulators [cyclosporin A (CsA) and imatinib] on transporter inhibition and on the reversal of resistance was also assessed. MNNG/HOS and U-2OS cells expressing high levels of MDR1 were highly resistant to DOX and showed reduced accumulation and higher efflux for radiotracers. Although MRP1 was uniformly expressed in all cells, only U-2OS was resistant to CIS. CsA restored sensitivity to DOX and CIS, and enhanced the accumulation and efflux half-life of radiotracers in MDR1-expressing cell lines. The chemosensitivity of OS cells to DOX was strongly dependent on mRNA MDR1 expression and could be circumvented by adding CsA. The kinetic parameters of radiotracers correlated with MDR1 expression levels, hence predicting DOX resistance. We concluded that sensitivity to chemotherapy is strongly dependent on the expression of MDR1 transporter and that radiotracer studies could prove clinically useful in predicting chemotherapy response and in evaluating the efficacy of MDR-reversing agents.