Experimental study on antitumor effect of arsenic trioxide in combination with cisplatin or doxorubicin on hepatocellular carcinoma

INTRODUCTIONThe main component of a traditional Chinese drug 'Pishuang'. arsenic trioxide (As2O3), has obviously selective anti-tumor effect on human hepatocellular carcinoma (HCC)in both in vitro and in vivo studies[1-5]. Due to limited effectiveness when any anti-carcinogen is used alone and obviously increased toxicity when the dose is raised, there is no exception for As2O3. Furthermore, combined chemotherapy contributes to improve therapeutic effectiveness, disperse toxicity and surmount drug-resistance,in which the combination of traditional Chinese and modern medicine has more advantages and characteristics. As a result,we made an experimental study on anti-tumor effect of As2O3in combination with cisplantin (PDD) or doxorubicin (ADM)on HCC. to investigate the possibility of AS2O3 in combination with PDD or ADM and nature of interaction between them,and to provide experimental basis for clinical application.

Inhibitory effects of two oligosaccharideson murine melanoma experimental liver metastasis

AIM To observe the effects of a chemically synthesized tetrose and a natural yeast mannan on experimental liver metastasis of mouse melanoma. METHODS After treated with 4mg tetrose (tetrose group) or 4mg mannan (mannan group) for 30 minutes at 37℃, 0 5ml 1×10 6 B16 MBK melanoma cells were injected into the spleen of mice. Fifty five days later, melanoma metastatic nodes on the surface of the liver and in other organs as well as mouse survival time were observed. RESULTS Of the 6 mice in control (B16 cell+PBS) group, 4 died naturally within 55 days, and 2 were killed on the 55th day. All of the 6 mice had metastases in livers, the total number of the melanoma nodes on each liver surface ranged from 2 to 30, with the largest one merging into the whole liver. One mouse had a neoplasm in the remnant site of injection, and 3 had metastases in lungs. In contrast, of the 6 mice in tetrose group, only one died on the 50th day after injection, with 3 metastases in the liver, the largest being 10mm in diameter, the other 5 mice survived until being dissected on the 55th day after injection and had no liver metastasis, but 3 of them had neoplasms in their remnant sites of injection. In mannan group, all of the 6 mice survived and no metastasis was seen except for 2 liver nodes in one mouse with the largest diameter of 1mm. Neither tetrose nor mannan group had metastasis out of the liver, and the weight of liver in the two groups was significantly lower than those in the control group. CONCLUSION Both tetrose and mannan had the effects of preventing melanoma cells from experimental metastasis to and out of the liver, and prolonging the survival time of the mouse.

不同双能CT虚拟单能谱图像衍生影像组学特征的可重复性:实验研究

目的比较3种双能CT(DECT)系统在不同条件下所获VX2兔肝肿瘤模型虚拟单能谱图像(VMI)增强CT影像组学特征(RF)的可重复性及其与诊断效能的关系。方法将15只VX2兔肝肿瘤模型随机均分为3组,分别接受采用双源DECT(dsDECT)、快速kV切换DECT(rsDECT)或双层探测器DECT(dlDECT)于不同体积CT剂量指数(CTDIvol)(6、9及12 mGy)下腹部增强CT扫描,于40~140 keV内每间隔10 keV重建单能图像并提取RF;以组内相关系数(ICC)计算RF可重复性,以ICC≥0.80为可重复RF,比较不同CT仪配对之间及不同CTDIvol下可重复RF占比(R),以及相同CTDIvol下不同CT仪配对可重复RF数量(N)最大值相应重建能量水平;绘制受试者工作特征(ROC)曲线,计算曲线下面积(AUC),比较最佳可重复条件下可重复RF与其他RF的诊断效能,计算RF的ICC与相应AUC的Spearman相关系数。结果R rsDECT-dsDECT[6.45%,95%CI(2.36%,8.87%)]高于R dlDECT-dsDECT[0.72%,95%CI(0.15%,1.79%)]及R rsDECT-dlDECT[1.43%,95%CI(0.60%,4.06%)](校正P均<0.05);R_(9mGy)[3.70%,95%CI(1.31%,5.73%)]及R_(12mGy)[2.63%,95%CI(0.60%,6.69%)]高于R_(6mGy)[1.31%,95%CI(0.12%,1.55%)](校正P均<0.05)。6、9及12 mGy下,RF最佳可重复重建能量水平集中于50~70 keV,可重复RF的AUC均高于其他RF(校正P均<0.05),且RF可重复性与诊断效能相关(r s=0.102~0.516,P<0.05)。结论兔肝肿瘤模型VMI CT增强图像RF可重复性与DECT设备、CTDIvol水平及重建能量水平相关,可重复RF可能诊断效能更佳。

Establishment of cell clones with different metastatic potential from the metastatic hepatocellular carcinoma cell line MHCC97

AIM: To establish clone cells with different metastatic potential for the study of metastasis-related mechanisms. METHODS: Cloning procedure was performed on parental hepatocellular carcinoma (HCC) cell line MHCC97, and biological characteristics of the target clones selected by in vivo screening were studied. RESULTS: Two clones with high (MHCC97-H) and low (MHCC97-L) metastatic potential were isolated from the parent cell line. Compared with MHCC97-L, MHCC97-H had smaller cell size (average cell diameter 43 microm vs 50 microm) and faster in vitro and in vivo growth rate (tumor cell doubling time was 34.2h vs 60.0h). The main ranges of chromosomes were 55-58 in MHCC97-H and 57-62 in MHCC97-L. Boyden chamber in vitro invasion assay demonstrated that the number of penetrating cells through the artificial basement membrane was (37.5 +/- 11.0) cells/field for MHCC97-H vs (17.7 +/- 6.3)/field for MHCC97-L. The proportions of cells in G0-G1 phase, S phase, and G2-M phase for MHCC97-H/MHCC97-L were 0.56/0.65, 0.28/0.25 and 0.16/0.10, respectively, as measured by flow cytometry. The serum AFP levels in nude mice 5wk after orthotopic implantation of tumor tissue were (246 +/- 66) microg.L(-1) for MHCC97-H and (91 +/- 66) microg.L(-1) for MHCC97-L. The pulmonary metastatic rate was 100% (10/10) vs 40% (4/10). CONCLUSION: Two clones of the same genetic background but with different biological behaviors were established, which could be valuable models for investigation on HCC metastasis.

Metastatic human hepatocellular carcinoma models in nude mice and cell line with metastatic potential

Metastatic human HCC model is needed for the studies on mechanism and intervention of metastatic recurrence. By using orthotopic implantation of histologically intact tissues of 30 surgical specimens, a patient-like metastatic model of human HCC in nude mice (LCI-D20) and a low metastatic model of human HCC in nude mice (LCI-D35) have been established. All mice with transplanted LCI-D20 tumors exhibited extremely high metastatic ability including spontaneous metastasis to liver, lungs, lymph nodes and peritoneal seeding. Remarkable difference was also found in expression of some of the invasiveness related genes and growth factors between the LCI-D20 and LCI-D35 tumors. PAI-1 increased gradually following tumor progression in LCI-D20 model, and correlated with tumor size and AFP level. Phasic expression of tissue intercellular adhesion molecule-1 in this model was also observed. Using corneal micropocket model, it was demonstrated that the vascular response induced by LCI-D20 tumor was stronger than that induced by LCI-D35 tumor. Similar report on metastatic human HCC model in nude mice and human HCC cell line with metastatic potential was rarely found in the literature. This LCI-D20 model has been widely used for the studies on intervention of metastasis, including anti-angiogenesis,antisense approach, metalloproteinase inhibitor, differentiation inducer, etc. It is concluded that the establishment of metastatic human HCC model in nude mice and human HCC cell line with metastatic potential will provide important models for the in vitro and in vitro study of HCC invasiveness, angiogenesis as well as intervention of HCC recurrence.

Molecular mechanism about lymphogenous metastasis of hepatocarcinoma cells in mice

AIM: To investigate the correlation between lymphogenous metastasis and matrix metalloproteinases (MMPs) activity and the expression of Fas ligand of tumor cells in lymph nodes. METHODS: Fifty-six inbred 615-mice were equally divided into 2 groups and inoculated with Hca-F and Hca-P cells. Their lymph node metastatic rates were examined. Growth fraction of lymphocytes in host lymph nodes was detected by flow cytometry. The Hca-F and Hca-P cells were cultured with extract of lymph node, liver or spleen. The quantity of MMPs in these supernatants was examined by zymographic analysis. The expression of Fas ligand, PCNA, Bcl-2 protein of Hca-F and Hca-P cells in the mice were examined by immunohistochemistry. The apoptosis signals of macro-phages in lymph nodes were observed with in situ DNA fragmentation. RESULTS: On the 28th day post-inoculation, the lymph node metastatic rate of HcaF was 80%(16/20), whereas that of Hca-P was 25%(5/20). The growth fraction of lymphocytes was as follows: in the Hca-F cells, the proliferating peak of lymphocytes appeared on the 14th day post inoculation and then decreased rapidly, while in HcaP cells, the peak appeared on the 7th day post inoculation and then kept at a high level. With the extract of lymph node, the quantity of the MMP-9 activity increased (P【0.01) and active MMP-9 and MMP-2 were produced by both Hca-F and Hca-P tumor cells, which did not produce MMPs without the extract of lymph node or with the extracts of the liver and spleen. The expression of Fas Ligand of Hca-F cells was stronger than that of Hca-P cells (P 【0.01). The expressions of PCNA and Bcl-2 protein of Hca-F cells in the tumors of inoculated area were the same as that of Hca-P cells. In situ DNA fragmentation showed that the positive signals of macrophages were around Hca-F cells. CONCLUSION: Secretion of MMPs which was associated with metastatic ability of Hca-F and Hca-P tumor cells depends on the environment of lymph nodes. The increased expression of Fas ligand protein of Hca-F tumor cells with high lymphogenous metastatic potential in lymph nodes may help tumor cells escape from being killed by host lymphocytes.

Purification of heat shock protein 70-associated tumor peptides and their antitumor immunity to hepatoma in mice

AIM:To purify the heat shock protein (HSP) 70-associated tumor peptides and to observe its non-MHC-I molecule restrictive antitumor effect.METHODS:By ConA-sepharose affinity chromatography,ADP-agarose affinity chromatography, and DEAE anion exchange chromatography, we were able to purify HSP70-associated peptides from mouse hepatoma (HCaF) cells treated in heat shock at 42℃. Specific active immunization and adoptive cellular immunization assay were adopted to observe the immunoprotective effect elicited by HSP70-associated peptide complexes isolated from HcaF.RESULTS: The finally purified HSP-associated peptides had a very high purity and specificity found by SDS-PAGE and Western blot. Mice immunized with HSP70-associated peptide complexes purified from HCaF cells were protected from HCaF living cell challenge. This effect was dose dependent.Adoptive immunization of immune spleen cells of mice immunized with HSP70-associated peptide complexes could elicit immunity against HCaF challenge, and the tumor-free mice could resist repeated challenges. This effect could be continuously enhanced by repeated challenge with HCaF living cells. The tumor-free mice could tolerate the challenge for as high as 1×10^7 HCaF cells. The mice immunized once with spleen cells pulsed with HSP70-associated peptide complexes in vitro could also result in a certain adoptive immunity against HCaF.CONCLUSION:High purity and specificity of HSP70-associated peptides could be achieved from tumor cells by the low-pressure affinity chromatography method used in this study. HSP70-associated peptide complexes derived from the HCaF can elicit non-MHC-I molecule restrictive immunoprotective effect against HCaF.This effect can be transferred by adoptive immunization to mice and enhanced by repeated challenge with HCaF live cells.

Influence of hepatic arterial blockage on blood perfusion and VEGF,MMP-1 expression of implanted lver cancer in rats

AIM: To investigate the influence of hepatic arterial blockage on blood perfusion of transplanted cancer in rat liver and the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase-1 (MMP-1), and to explore the mechanisms involved in transarterial embolization (TAE)-induced metastasis of liver cancer preliminarily. METHODS: Walker 256 carcinosarcoma was transplanted into rat liver to establish the liver cancer model. Hepatic arterial ligation (HAL) was used to block the hepatic arterial blood supply and simulate TAE. Blood perfusion of tumor in control, laparotomy control, and HAL group was analyzed by Hoechst 33342 labeling assay, the serum VEGF level was assayed by ELISA, the expression of VEGF and MMP-1 mRNA was detected by in situ hybridization. RESULTS: Two days after HAL, the number of Hoechst 33342 labeled cells which represent the blood perfusion of tumor directly and hypoxia of tumor indirectly in HAL group decreased significantly compared with that in control group (329+/-29 vs 384+/-19, P【0.01). The serum VEGF level in the HAL group increased significantly as against that of the control group (93 ng.L(-1)+/-44 ng.L(-1) vs 55 ng.L(-1)+/-19 ng.L(-1), P【0.05). The expression of VEGF and MMP-1 mRNA in the tumor tissue of the HAL group increased significantly compared with that of the control and the laparotomy control groups (P【0.05). The blood perfusion data of the tumor, represented by the number of Hoechst 33342 labeled cells, showed a good linear inverse correlation with the serum VEGF level (r=-0.606, P【0.05) and the expression of VEGF mRNA in the tumor tissue ( r =-0.338, P【0.01). CONCLUSION: Blockage of hepatic arterial blood supply results in decreased blood perfusion and increased expression of metastasis-associated genes VEGF and MMP-1 of transplanted liver cancer in rats. Decreased blood perfusion and hypoxia may be the major cause of up-regulated expression of VEGF.

Effect of a cancer vaccine prepared by fusions of hepatocarcinoma cells with dendritic cells

AIM: To prepare a cancer vaccine (H(22)-DC) expressing high levels of costimulatory molecules based on fusions of hepatocarcinoma cells (H(22)) with dendritic cells (DC) of mice and to analyze the biological characteristics and induction of specific CTL activity of H(22)-DC. METHODS: DCs were isolated from murine spleen by metrizamide density gradient centrifugation, purified based on its characteristics of semi-adhesion to culture plates and FcR-,and were cultured in the medium containing GM-CSF and IL-4. A large number of DC were harvested. DCs were then fused with H(22) cells by PEG and the fusion cells were marked with CD11c MicroBeads. The H(22)-DC was sorted with Mimi MACS sorter. The techniques of cell culture, immunocytochemistry and light microscopy were also used to test the characteristics of growth and morphology of H(22)-DC in vitro. As the immunogen, H(22)-DC was inoculated subcutaneously into the right armpit of BALB/C mice, and their tumorigenicity in vivo was observed. MTT was used to test the CTL activity of murine spleen in vivo. RESULTS: DC cells isolated and generated were CD11c+ cells with irregular shape, and highly expressed CD80, CD86 and CD54 molecules. H22 cells were CD11c- cells with spherical shape and bigger volume, and did not express CD80, CD86 and CD54 molecules.H(22)-DC was CD11c+ cells with bigger volume, being spherical, flat or irregular in shape, and highly expressed CD80, CD86 and CD54 molecules, too. H(22)-DC was able to divide and proliferate in vitro, but its activity of proliferation was significantly decreased as compared with H(22) cells and its growth curve was flatter than H(22) cells. After subcutaneous inoculation over 60 days, H(22)-DC showed no tumorigenecity in mice, which was significantly different from control groups (P【0.01). The spleen CTL activity against H(22) cells in mice implanted with fresh H(22)-DC was significantly higher than control groups (P 【 0.01). CONCLUSION: H(22)-DC could significantly stimulate the specific CTL activity of murine spleen, which suggests that the fusion cells have already obtained the function of antigen presenting of parental DC and could present H(22)specific antigen which has not been identified yet, and H(22)-DC could induce antitumor immune response; although simply mixed H(22) cells with DC could stimulate the specific CTL activity which could inhibit the growth of tumor in some degree, it could not prevent the generation of tumor. It shows that the DC vaccine is likely to become a helpful approach in immunotherapy of hepatocarcinoma.

Preliminary Investigation on Regulating Effects of Different TCM Treatments on Transcription of the Correlated Genes of Liver Cancer in Rats

The regulating effects of TCM treatments including clearing away heat and toxic materials,promoting blood circulation and removing blood stasis,and strengthening the spleen and regulating qi on the oncogene transcription were observed in the liver cancer model rats.The preliminary results indicated that the mRNA levels of H-ras N-ras and K-ras,and signal molecules correlated with the ras/MAPK signal transduction pathway were down-regulated by the different TCM treatments in varying degrees.Also,the regulating effects of the treatments on differently-displayed genes were discrepant.It is suggested that the molecular mechanisms of the TCM treatments for liver cancer was complex with different target genes.

Tumor radioimmunoimaging of chimeric antibody in nude mice with hepatoma xenograft

IM To study the radioimmunoimaging (RAII) using the human/mouse chimeric Ab to evaluate its targeting activity in animal models.METHODS To chimeric Ab was labeled with 131I. RAII was performed at different intervals after injection of radiolabeled Abs in nude mice with human hepatoma xenograft, and tissue distribution of radioactivity was measured. Comparison was made in the chimeric Ab between the single segment Ab and previous murine mAb against HBxAg.RESULTS The experimental objects developed tumorpositive image after 2 days of radiolabeled Abs injection, and the peak accumulation of radioactivity fell on the 7th day. The tumor/liver ratioactivity of the chimeric Ab, single segment Ab, antiHBx mAb, and the control group was 281±021, 244±016, 460±019, and 096±014, respectively.CONCLUSION The genetic engineering Abs have a considerable targeting activity which can be used as a novel humanized vector in the targeting treatment of liver cancer..

Inhibition effect of natural killer T cells on transplantation hepatocellular carcinoma in mice

Objective:The aim of this study was to investigate the inhibition effect of natural killer T(NKT) cells on transplantation hepatocellular carcinoma in mice.Methods:α-galactosylceramide(α-GalCer)-pulsed DC and Hep S were prepared as stimulus.Hepatoma xenograft model was established and mice were randomly divided into 4 groups(n=13 each group):(1) control group,intravenous injection of the same volume of saline.(2) mature DC group,intravenous injection of mature DC cells(4×106 cells).(3) α-GalCer-pulsed HepS group,intravenous injection of α-GalCer-pulsed HepS(4×106 cells).(4) α-GalCer-pulsed mature DC group,intravenous injection of α-GalCer-pulsed DC(4×106 cells).The changes of tumor volume in mice and survival period were measured every 2 days.Percentage of NKT cells in spleens and cytotoxicity of spleen cells were detected by flow cytometry.Tumor tissues were analyzed by histopathological examination.Results:In α-GalCer-pulsed Heps and DC groups,the average survival period was prolonged and tumor volume was markedly decreased,spleen cells and NKT cells were significantly increased,and tumor necrosis was evident,compared to the control group.Conclusion:α-GalCer-pulsed DC and HepS could activate NKT cells in vivo,also increase NKT cells cytotoxicity,inhibit the growth of hepatomas and prolong survival period.

碘油羟基磷灰石纳米粒对兔VX2肝肿瘤Paxillin和P130Cas表达的影响

目的研究碘油羟基磷灰石纳米粒(nHAP)经肝动脉灌注治疗对兔VX2肝肿瘤转移相关基因Paxillin(桩蛋白)和P130Cas表达的影响。方法将100只新西兰白兔肝内肿瘤种植后2周,随机分为5组,每组20只,设生理盐水组(A组)、nHAP组(B组)、单纯碘油组(C组)、阿霉素碘油组(D组)和碘油nHAP组(E组)。经肝动脉灌注给药后2周,采用免疫组织化学和Western blotting检测这两个基因在蛋白水平的表达。结果两种方法结果均提示:兔VX2肝肿瘤中Paxillin呈低表达而P130Cas呈高表达。与A组相比,C组和D组残余肿瘤区的Paxillin表达降低,而E组表达则显著升高(P<0.05);与A组相比,C组和D组残余肿瘤区的P130Cas表达升高,而E组表达则显著降低(P<0.05)。结论兔VX2肝肿瘤中Paxillin的低表达和P130Cas的高表达可能与其高转移性有关。单纯碘油及阿霉素碘油经肝动脉治疗可降低Paxillin的表达,而碘油nHAP可升高其表达;单纯碘油及阿霉素碘油经肝动脉治疗可升高P130Cas的表达,而碘油nHAP可降低其表达。提示碘油nHAP可能可以抑制VX2肿瘤的转移。

Effects of Ginkgo biloba extract on expression of biomarkers during aflatoxin B_1-induced hepatocarcinogenesis in Wistar rats

Objective： The aim of this study was to study the effect of Ginkgo biloba extract （EGb761） on metabolism of afiatoxin B1 （AFB1） in Wistar rats. Methods： Seventy one Wistar rats were assigned at random to groups A, B and C. Rats in groups A, B were injected with AFB1 （intraperitoneal, 100-200 ug/kg body weight, 1-3 times/week）. Group C was normal control. Rats in group B were fed in food with EGb761, while rats in groups A, C were given normal food. Blood samples were collected and liver biopsies were performed on the 14th, 28th and 42nd week. All the rats were sacrificed on the 64th week. The incidence of hepatocarcinoma was investigated. The hepatic phase I drug-metabolizing enzyme Cytochrome-P450 （CYP450） and phase II metabolizing enzyme glutathione S-transferase （GST） were analyzed with spectrometry. Serum AFB1- lysine adduct levels were assessed with high performance liquid chromatography （HPLC）. The expression of 8-hydroxydeoxy- guanosine （8-OHdG） was measured with immunohistochemistry. Results： The incidence of hepatocellular carcinoma （HCC） in group B was significantly lower than that in group A （26.92% vs 76.00%, P 〈 0.001）. No HCC developed in group C. EGb761 showed no effects on the activities of CYP450 and GST in rat liver tissues. The level of AFB1-lysine adduct reached the peak （4356.01 pg/mg albumin） at the 14th week in group A. EGb761 significantly inhibited the formation of AFB1-lysine adduct in serum by 13.07% at the 14th week （P = 0.033）, and 73.63% at the 42nd week （P = 0.002）. The expression of 8-OHdG protein in rat liver tissues in group B was significantly lower than that in group A at the 28th, 42nd, and 64th week （P 〈 0.05）. Conclusion： The main mechanism underlying the effect of EGb761 in blocking hepatocarcinogenesis induced by AFB1 may not be fully attributable to its influence on the activity of liver phase I and phase II metabolizing enzymes. EGb761 inhibits the production of AFB1-lysine adducts, decreases the expression of 8-OHdG protein, and finally alleviates the DNA oxidative injury, which may be one of the mechanisms for the effects of EGb761 in inhibiting or delaying AFB1-induced hepatocarcinogenesis.

Cyclopamine对肝癌细胞增殖和凋亡的影响及其机制的研究

目的:探讨sonic hedgehog(Shh)信号通路的特异性抑制剂环杷明(cyclopamine)对血清饥饿的人肝癌Hep3B细胞增殖、早期凋亡和白蛋白(albumin,ALB)、甲胎球蛋白(α-fetoprotein,AFP)表达的影响。方法:采用WST-8法、激光共聚焦扫描显微术、FCM法、RT-PCR和电化学发光免疫法,检测血清饥饿状态下cyclopamine引起的Hep3B细胞增殖抑制、线粒体膜电位改变,以及对细胞分泌ALB和AFP的影响。结果:血清饥饿状态下cyclopamine可明显抑制Hep3B细胞的增殖。半定量测量结果显示,Hep3B细胞平均红绿荧光吸光度比值有所降低;FCM法检测结果显示,6h后线粒体膜电位较0h时明显降低,差异有统计学意义(P<0.05)。AFP mRNA及蛋白分泌水平均明显降低,但对ALB mRNA及蛋白分泌影响不明显。结论:Cyclopamine对Hep3B细胞增殖以及AFP mRNA和蛋白的分泌有明显抑制作用,同时促进线粒体膜电位的降低,提示cyclopamine具有抑制肿瘤细胞生长和促进早期凋亡发生的作用。

In situ hybridization assay of androgen receptor gene in hepatocarcinogenesis

AIM To determine the correlation between expression of androgen receptor (AR) gene and hepatocarcinogenesis. METHODS Male SD rats were used as experimental animals and the animal model of experimental hepatocarcinoma was established by means of 3′ me DAB administration. Androgen receptor mRNA was detected by a non radioactive in situ hybridization assay in neoplastic and non neoplastic liver tissues. RESULTS The expression of androgen receptor mRNA was observed only in neoplastic cells and some atypical hyperplastic cells. In the liver tissue of control animal and the remaining normal liver cells adjacent to the carcinoma tissue, no positive signal was seen. CONCLUSION Androgen has an important correlation with hepatocarcinogenesis and the expression of androgen receptor gene might be a mark event during hepatocarcinogenesis.

High-intensity focused ultrasound extracorporeal ablation of liver tissuesin rabbits

ＨｉｇｈｉｎｔｅｎｓｉｔｙｆｏｃｕｓｅｄｕｌｔｒａｓｏｕｎｄｅｘｔｒａｃｏｒｐｏｒｅａｌａｂｌａｔｉｏｎｏｆｌｉｖｅｒｔｉｓｓｕｅｓｉｎｒａｂｂｉｔｓＣＨＥＮＧＳｈｕＱｕｎ１，ＺＨＯＵＸｉｎＤａ１，ＴＡＮＧＺｈａｏＹｏｕ１，ＹＵＹａｏ１，Ｂ...

金合欢素对肝癌HepG2细胞增殖、凋亡和迁移的影响及机制研究

目的探讨金合欢素(Aca)通过调节PTEN诱导激酶1(PINK1)/E3泛素连接酶(Parkin)通路介导的线粒体自噬对肝癌HepG2细胞增殖、凋亡和迁移的影响。方法将肝癌HepG2细胞分为对照组(正常培养的HepG2细胞)、Aca组(10μmol/L Aca)、PINK1小干扰RNA阴性对照(si-NC)组(转染si-NC)、PINK1小干扰RNA(si-PINK1)组(转染si-PINK1)、Aca+si-NC组(转染si-NC后用10μmol/L Aca处理)、Aca+si-PINK1组(转染si-PINK1后用10μmol/L Aca处理)。CCK-8法检测细胞增殖;流式细胞术检测细胞凋亡;Transwell实验检测细胞迁移;透射电镜观察自噬小体的形成;Western blot检测细胞中自噬相关蛋白[Beclin-1、微管相关蛋白1轻链3(LC3)-Ⅰ、LC3-Ⅱ]及PINK1/Parkin通路相关蛋白表达。结果与对照组比较,Aca组HepG2细胞存活率、迁移细胞数目降低,凋亡率、自噬小体数量、Beclin-1、LC3-Ⅱ/LC3-Ⅰ、PINK1、Parkin蛋白表达水平升高(P<0.05),si-PINK1组HepG2细胞存活率、迁移细胞数目升高,凋亡率、自噬小体数量、Beclin-1、LC3-Ⅱ/LC3-Ⅰ、PINK1、Parkin蛋白表达水平降低(P<0.05);与Aca组、Aca+si-NC组比较,Aca+si-PINK1组HepG2细胞存活率、迁移细胞数目升高,凋亡率、自噬小体数量、Beclin-1、LC3-Ⅱ/LC3-Ⅰ、PINK1、Parkin蛋白表达水平降低(P<0.05)。结论Aca可能通过激活PINK1/Parkin通路介导的线粒体自噬抑制肝癌HepG2细胞增殖、迁移,促进细胞凋亡。

HDGF、VEGF表达与肝癌患者病灶浸润转移风险的相关性

【目的】探讨肝癌衍生生长因子(HDGF)、血管内皮生长因子(VEGF)表达与肝癌患者病灶浸润转移风险的相关性。【方法】回顾性分析2019年7月至2022年7月西安市第三医院西北大学附属医院收治的82例接受手术切除治疗的肝癌患者的临床资料,收集患者的肝癌组织标本和癌旁组织。采用免疫组化染色法检测肝癌组织及癌旁组织内HDGF、VEGF表达情况,比较不同临床病理特征肝癌患者HDGF、VEGF表达水平,采用多因素logistic回归分析影响肝癌组织HDGF、VEGF表达的危险因素。【结果】肝癌组织中HDGF、VEGF阳性表达率高于癌旁组织,差异有统计学意义(P<0.05)。不同AJCC分期、包膜完整性和肿瘤转移的肝癌组织HDGF、VEGF表达比较,差异有统计学意义(P<0.05)。多因素Logistic回归模型分析显示,AJCC分期、包膜完整性和肿瘤转移是肝癌组织HDGF、VEGF表达的影响因素(P<0.05)。【结论】肝癌组织中HDGF、VEGF呈高表达,且与AJCC分期、包膜完整性和肿瘤转移相关,对评估肝癌进展有一定的参考价值。

长链非编码RNA MEG3在原发性肝癌中的表达及其意义

【目的】探讨长链非编码RNA(LncRNA)MEG3在原发性肝癌(HCC)中的表达及其临床意义。【方法】本院收治的78例HCC患者均接受手术治疗,术中收集患者肝癌组织及癌旁正常组织;招募同期就诊于本科的肝血管瘤患者78例,收集术中标本。采用RT-PCR检测三种样本中LncRNA MEG3的表达水平并比较,评价LncRNA MEG3诊断HCC的价值,探讨LncRNA MEG3与HCC临床病理特征的相关性。【结果】LncRNA MEG3在肝癌组织中的表达为0.43±0.16,显著低于癌旁正常组织(1.18±0.35)及肝血管瘤组织(1.21±0.27)(均P<0.05);LncRNA MEG3诊断HCC的曲线下面积(AUC)(95%CI)为0.85(0.80~0.91),最佳诊断界值为0.91,敏感度89%,特异度76%;LncRNA MEG3与甲胎蛋白呈线性正相关(r=0.63,P=0.03);低表达LncRNA MEG3与HCC临床分期、分化程度以及远处转移有相关性(χ^(2)分别为3.93、6.59及4.43,均P<0.05)。【结论】LncRNA MEG3在HCC中呈低表达,低表达LncRNA MEG3与HCC病理特征有相关性,是潜在的HCC诊断指标及治疗靶点。