原發(fā)性肝癌患者細(xì)胞因子水平的變化與轉(zhuǎn)移和復(fù)發(fā)的關(guān)系

1、原發(fā)性肝癌患者細(xì)胞因子水平的變化與轉(zhuǎn)移和復(fù)發(fā)的關(guān)系         08-07-14 16:53:00     編輯：studa20          作者：滕木儉 王明海 潘樂玉    【關(guān)鍵詞】肝腫瘤白細(xì)胞介素1白細(xì)胞介素4干擾素型腫瘤轉(zhuǎn)移腫瘤復(fù)發(fā)，局部       The relationship be

2、tween immunoregulatory cytokines and intrahepatic metastasis and recurrence in primary hepatocellular carcinoma    【ABSTRACT】Objective:To observe the expression of immunoregulatory cytokines including interleukin4(IL4,Th2)、 interferon(IFN,Th1)、interleukinbetal (IL1) in the tissues of

3、hepatocellular carcinoma(HCC),and to investigate the relationship between the changes of immunoregulatory cytokines in patients with hepatocellular carcinoma and intrahepatic metastasis and recurrence.Methods:The immunoregulatory cytokines:IFN(Th1)、IL4(Th2)、IL1 were analyzed by RTPCR.Results:34 case

4、s with HCC were compared with the ones with liver cirrhosis,the expression rates and density of IFN(Th1) in the tissues of hepatocellular carcinoma were much lower than that in the liver cirrhosis,they are 0.92±0.36(18/34,53.9%) and 1.63±0.64(12/15,80.0%),respectively.There is significant

5、difference between them (P0.01).The expression density and levels of IL4(Th2) in the tissues of hepatocellular carcinoma increased clearly,compared with the control group.They are 1.76±0.49 and 1.01±0.51,respectively.Their difference showed statistical significance (P0.01).The expression r

6、ates and levels of IL1 in the tissues of hepatocellular carcinoma and liver cirrhosis were 1.01±0.32(30/34,88.2%) and 0.35±0.30(4/15,26.7%),respectively.There was significant difference between them (P0.001).Above three immunoregulatory cytokines in patients with hepatocellular carcinoma s

7、howed five different expression styles.Conclusion:There is severe deficiency of immune responses against tumors in patients with hepatocellular carcinoma,the balance between those above immunoregulatory cytokines was destroyed,so that immune responses against tumors were weaker and in further suppre

8、ssion,which may enhance the incidence rates of intrahepatic metastasis and recurrence of hepatocellular carcinoma after operation.    【KEY WORDS】Liver neoplasmsInterleukin1Interleukin4Interferon typeNeoplasm metastasisNeoplasm recurrence,local    我國肝癌患者大多合并嚴(yán)重肝硬化，機(jī)體免疫狀態(tài)已

9、發(fā)生某些改變，并且容易發(fā)生肝內(nèi)及遠(yuǎn)處轉(zhuǎn)移。肝臟局部、全身免疫狀態(tài)和抗腫瘤免疫與原發(fā)性肝癌復(fù)發(fā)及轉(zhuǎn)移關(guān)系的研究已取得一些進(jìn)展13。腫瘤免疫研究占據(jù)重要位置46。本研究應(yīng)用RTPCR法對肝癌組織細(xì)胞因子進(jìn)行檢測，以探討肝細(xì)胞癌患者免疫狀態(tài)改變及其與肝癌轉(zhuǎn)移和復(fù)發(fā)的關(guān)系。    1  資料與方法    1.1  一般資料  肝細(xì)胞性肝癌（HCC）組織均來源于我院2000年3月到2002年3月手術(shù)切除標(biāo)本，均經(jīng)術(shù)后病理證實(shí)，肝硬化組織來源于同期門脈高壓癥手術(shù)肝活檢。其中肝細(xì)胞性肝癌組（肝病組）34例，男23例，女

10、11例，年齡3571歲，中位年齡43歲；門脈高壓癥肝硬化組（對照組）15例，男11例，女4例，年齡2963歲，中位年齡37歲。    1.2  方法    提取總RNA：（1）100mg標(biāo)本剪碎后加500l CIT變性液，在勻漿器內(nèi)勻漿，若為細(xì)胞標(biāo)本，吹打即可，加入50l 2M NaOAC、500l水飽和酚、100l氯仿，每步均需要混勻10s，冰浴515min。（2）410000g離心10min，小心吸取上清移入新Eppenderf管中，切忌吸入蛋白，加入等體積的異丙醇，-20放置1h。（3）412000g離心10min，棄

11、上清，沉淀使用100l CIT變性液重溶，加入等體積異丙醇混勻，-20放置1h。（4）412000g離心10min，棄上清，用75%乙醇洗一遍，至此可加入1ml無水乙醇于-70儲存?zhèn)溆?，亦可直接涼干，溶于適量無Rnase的去離子水中。（5）紫外分光光度計測出A260及A280的值，計算所提總RNA的含量。    逆轉(zhuǎn)錄（RT）反應(yīng)：DEPC處理0.5ml于Eppendorf管中，加入逆轉(zhuǎn)錄反應(yīng)體系試劑，快速離心混勻，37，1h，95，10min滅活MMLV，快速離心沉淀。    PCR反應(yīng)：在上述20l RT產(chǎn)物的0.5ml Eppendorf管中，繼續(xù)加入PCR反應(yīng)體系試劑，快速離心混勻，加入100l液體石蠟。循環(huán)條件：94 1min，72 1min，循環(huán)35次，末次延長7min，電泳鑒定。定量分析：用凝膠成像分析系統(tǒng)判定陽性細(xì)胞因子和actin表達(dá)強(qiáng)度。見表1。表1  RTPCR檢測相關(guān)細(xì)胞因子的引物序列（略）    2  結(jié)