Overexpression of HAb18G/CD147 promotes invasion and metastasis via α3β1 integrin mediated FAK-paxillin and FAK-PI3K-Ca2+ pathways

Mechanism of HAb18G/CD147 underlying the metastasis process of human hepatoma cells has not been determined. In the present study, we found that integrin α3β1 colocalizes with HAb18G/CD147 in human 7721 hepatoma cells. The enhancing effect of HAb18G/CD147 on adhesion, invasion capacities and matrix metalloproteinases (MMPs) secretion was decreased by integrin α3β1 antibodies (p<0.01). The expressions of integrin downstream molecules including focal adhesion kinase (FAK), phospho-FAK (p-FAK), paxillin, and phospho-paxillin (p-paxillin) were increased in human hepatoma cells overexpressing HAb18G/CD147. Deletion of HAb18G/CD147 reduces the quantity of focal adhesions and rearranges cytoskeleton. Wortmannin and LY294002, specific phosphatidylinositol kinase (PI3K) inhibitors, reversed the effect of HAb18G/CD147 on the regulation of intracellular Ca²⁺ mobilization, significantly reducing cell adhesion, invasion and MMPs secretion potential (p<0.01). Together, these results suggest that HAb18G/CD147 enhances the invasion and metastatic potentials of human hepatoma cells via integrin α3β1-mediated FAK-paxillin and FAKPI3K-Ca²⁺ signal pathways. Received 5 June 2008; received after revision 16 July 2008; accepted 23 July 2008     

木瓜凝乳蛋白酶对卡铂杀伤鼠肝癌细胞的增强作用

在不同培养时间下,将不同浓度的木瓜凝乳蛋白酶、卡铂(CBP)、木瓜凝乳蛋白酶 CBP增加到鼠肝癌细胞hepa-6培养液中,采用MTT法检测它们对hepa-6的细胞毒性．结果表明,木瓜凝乳蛋白酶对hepa-6细胞的半数抑制浓度(IC50)约为1200μg／mL,随浓度增加,酶对细胞的生长抑制率增加;且均能增强CBP对鼠肝癌细胞hepa-6的杀伤作用．在hepa-6细胞加药培养至48h时第二次加入酶,能继续增强CBP对hepa-6的杀伤作用．     

6-F硫色烯并[4,3-c]吡唑啉的体内抗肿瘤活性

用小鼠移植性肿瘤法对6-F硫色烯并[4,3-c]吡唑啉(Ja)进行体内抗肿瘤活性研究.通过检测其对小鼠H22肝癌细胞、S180肿瘤细胞的体内抑瘤作用,计算肿瘤抑制率和生命延长率,并与阳性药顺铂做比较.结果表明,该化合物可抑制H22肝癌细胞皮下移植瘤的生长,在剂量为1,2,4 mg/(kg·d)时,腹腔注射给药抑瘤率分别为13.93%,37.49%,47.45%;而以剂量为5,10,20 mg/(kg·d)灌胃给药时,抑瘤率分别为31.53%,43.77%,50.73%;另外,该化合物可明显延长S180腹水瘤小鼠存活时间,以剂量为5,10,20 mg/(kg·d)灌胃给药时实验测得生命延长率分别为30.4%,64.7%,88.2%.该化合物有较强的体内抗肿瘤活性,对其抗癌作用机制值得进行深入研究.     

Effect ofP15 INK4b/MTS2 on the proliferation of human hepatoma cells SMMC-7721

The full length cDNA coding forP15 ^INK4b, which is a cyclin-dependent kinase inhibitor, was cloned to plasmid PXJ41-neo (Eco R I /Xho I site) and the new constructed plasmid pXJp15 was obtained. pXJp15 was transferred into the human hepatoma SMMC-7721 cetls by lipofectine reagent. After G418 setection, a series of cetl lines stably expressing high levets ofP15 (named SHT) and the clone containing vector PXJ41-neo only (named SVXJ) were obtained by Northern and Western analysis. The results showed that the proliferation of SHT cells is inhibited compared with that of SVXJ cetls. Cell cycle analysis indicated that overexpressing ofP15 inhibited the growth of SHT cetls by decreasing progrssion of cetls from G1 to S and G2 to M phases. The levets ofc-Myc andc-Fos were obviously decreased in SHT cells compared with control cetls by Western blotting. The decreased expression of oncogene may be one of the molecular mechanisms of the effect ofP15 on the proliferation of in SHT cetls.     

Increased activity of dipeptidyl peptidase IV in serum of hepatoma-bearing rats coincides with the loss of the enzyme from the hepatoma plasma membrane

Summary The specific activity of dipeptidyl peptidase IV (DPPIV E.C. 3.4.14.-) in the plasma membrane of Morris hepatoma 9121 or hepatoma 7777 was 3.5% and 2.9%, respectively, of that in the plasma membrane of rat liver. The enzyme activity in the serum of hepatoma-bearing rats was 141% (hepatoma 91219) and 162% (hepatoma 7777) of the normal value. cytochemical investigation showed that the DPP IV activity was almost completely absent from the hepatoma cell plasma membrane and was not sequestered within these cells. Indirect immunofluorescence staining with a polyclonal antibody directed against DPP IV indicated that the loss of activity was due to the absence of DPP IV molecules in the plasma membrane. The possibility that the enzyme is transferred from the membrane into the serum as a result of structural alterations is discussed.     

应用SCID鼠筛选肝癌转移性亚克隆及M-H7402亚细胞系的建立

本研究应用人肝癌细胞SCID鼠转移模型，筛选和建立肝癌转移细胞系，旨在为肝癌转移研究提供实验材料．采用人肝癌细胞H7402，腋后背部皮下接种SCID鼠，接种后66天时，在荷瘤鼠赢弱时引颈处死，取其肺组织，进行原代细胞培养和传代培养，获得肝癌细胞H7402的亚细胞克隆，命名为M—H7402．然后，对M—H7402细胞进行了生物学鉴定，检测了细胞形态、染色体、细胞动力学、细胞周期、甲胎蛋白表达、癌基因表达和转移相关基因表达等指标．结果显示，M—H7402细胞的生长、增殖和形态等与亲本细胞H7402十分相近，其染色体形态仍为人类核型，众数维持在72～80之间，占75．0％．RT—PCR检测结果显示，M—H7402细胞甲胎蛋白表达为阳性．Western blot检测结果显示，与亲本细胞H—7402相比，癌基因C—myc表达水平较高，转移抑制基因nm23的表达水平明显下调．从肺组织中获得的肝癌转移细胞系M—H7402，在体外可连续传代培养，细胞形态不变．应用SCID鼠筛选获得的亚细胞克隆，与亲本细胞形成配对关系，可为肝癌细胞转移的研究提供新的实验材料。     

亚砷酸诱导肝癌Bel-7402细胞分化的研究

目的:探讨亚砷酸诱导肝癌Bel-7402细胞的分化作用。方法:通过体外细胞培养,观察肿瘤的生长特点;用Giemsa染色法观察细胞的分化形态特点,比色法测定酪氨酸-α-酮戊二酸转氨酶(TAT)活性的改变。结果:在亚砷酸(0 25、0 5、1 0mol/L)作用下,肝癌Bel-7402细胞连续培养4d,细胞生长抑制作用较明显,并呈剂量依赖关系(P<0 05);培养到第5d,细胞形态和结构向成熟阶段分化;在连续培养4d以后,酪氨酸-α-酮戊二酸转氨酶的比活性升高并呈时间依赖关系(P<0 05)。结论:亚砷酸具有明显的诱导肝癌Bel-7402细胞分化作用。     

沙棘提取物对癌细胞DNA、蛋白质合成及体内血浆中cAMP含量的影响

采用同位素示踪技术和蛋白质结合法研究了沙棘汁和沙棘油的抗癌机理。结果显示：沙棘汁对Ｌ＿（７７１２）白血病、Ｓ＿（１８０）肉瘤、腹水型肝癌等细胞的ＤＮＡ、蛋白质合成均无抑制作用。沙棘油浓度在０．００５～１．０ｍｇ／Ｌ时对Ｌ＿（７７１２）白血病细胞ＤＮＡ合成有抑制作用，且此时抑制率与浓度呈负相关。沙棘汁ｉｐ注射剂量为１／５ＬＤ＿（５０）时，沙棘汁对小鼠血浆ｃＡＭＰ含量略有提高。     

The effect of isoforms of the cell polarity protein, human ASIP, on the cell cycle and Fas/FasL-mediated apoptosis in human hepatoma cells

Human ASIP (hASIP) is expressed as numerous alternative splicing isoforms and there is an atypical protein kinease C (aPKC) phosphorylation site in exon 17b of the encoded sequence. We have identified an important role for exon 17b in cancer cells. Our results showed that hASIP-sa and sb had different effects on cell growth and Fas/FasL-mediated apoptosis in BEL-7404 human hepatoma cells. Human ASIP-sa modified the S phase of the cell cycle and might stimulate cell proliferation. Growth inhibition by hASIP-a antisense oligonucleotide-confirmed the positive action of hASIP-sa. Compared with hASIP-sa, hASIP-sb accelerated Fas/FasL-induced apoptosis, examined by sub-G1 accumulation, chromatin condensation, nuclear fragmentation, PARP cleavage, caspase-8 degradation and mitochondria- regulated cell death. Treatment with aPKC inhibitor could enhance Fas/FasL-mediated apoptosis in hASIP-sa-overexpressing cells, suggesting that hASIP-sa and its interaction with aPKC might contribute to the malignant growth and the blocking of Fas/FasL-mediated apoptosis, while hASIP-sb might function as an antagonist of hASIP-sa.Received 24 March 2005; received after revision 31 May 2005; accepted 21 June 2005