A STUDY OF THE BIOLOGICAL CHARACTERISTICS OF HUMAN HEPATOMA XENOGRAFTS IN NUDE MICE AFTER IRRADIATION

Cell growth kinetics and changes in AFP in nude mice with human hepatoma xenografts were evaluated using the flow cytometry method. After receiving 10 Gy of radiation, the mice showed a marked delay in tumor growth; approximately 1 Gy of radiation caused a tumor growth delay of one day. Irradiation altered various phases of the cell cycle. An acute and temporary block of G2 cells was characteristic; FCM measurements demonstrated that about 58% of cells were blocked in the G2 phase and this blocking effect lasted 90 hours after an irradiation of 10 Gy. This indicated that human hepatoma xenografts in nude mice were quite sensitive to irradiation. It was also noted that the AFP decreased for 96 hours after irradiation. Changes in G2 cells after irradiation may be closely related to changes in AFP.

ABCG2-overexpressing S1-M1-80 cell xenografts in nude mice keep original biochemistry and cell biological properties

S1-M1-80 cells,derived from human colon carcinoma S1 cells,are mitoxantrone-selected ABCG2-overexpressing cells and are widely used in in vitro studies of multidrug resistance(MDR).In this study,S1-M1-80 cell xenografts were established to investigate whether the MDR phenotype and cell biological properties were maintained in vivo.Our results showed that the proliferation,cell cycle,and ABCG2 expression level in S1-M1-80 cells were similar to those in cells isolated from S1-M1-80 cell xenografts(named xS1-M1-80 cells).Consistently,xS1-M1-80 cells exhibited high levels of resistance to ABCG2 substrates such as mitoxantrone and topotecan,but remained sensitive to the non-ABCG2 substrate cisplatin.Furthermore,the specific ABCG2 inhibitor Ko143 potently sensitized xS1-M1-80 cells to mitoxantrone and topotecan.These results suggest that S1-M1-80 cell xenografts in nude mice retain their original cytological characteristics at 9 weeks.Thus,this model could serve as a good system for further investigation of ABCG2-mediated MDR.

Tumour Localization of A Mitomycin C-Monoclonal Antibody MGb_2 Conjugate in Nude Mice with Human Gastric Carcinoma Xenografts

MGb2 is a monoclonal antibody againstgastric cancer.It can be well localized in thetumour tissue and has been successfully used inthe radioimmunoimaging in the patients withgastric cancer.The conjugate of MGb2 withchemotherapeutic drug mitomycin C（MMC）al-so shows highly selective cytotoxic effect uponhuman gastric cancer cells in vitro.

Experimental study on effect of recombinant human growth hormone combined with chemotherapy on stomach neoplasms implanted in nude mice

Objective: To investigate the effect of different doses of recombined growth hormone (rhGH) on stomach neo- plasms implanted in nude mice, and its efficacy in combining with chemotherapy (flurouracil, 5-FU). Methods: Human stom- ach neoplasms model was established in nude mice. The nude mice were divided into control group, moderate-dose of rhGH group, low-dose rhGH group, 5-FU group, moderate-dose rhGH/5-FU group, and low-dose rhGH/5-FU group. The results of each group were observed after ten days. Results: After therapy, the body mass of rhGH groups was significantly increased compared with control group (P<0.05), the body mass of rhGH/5-FU groups was significantly increased compared with 5-FU group (P<0.05), but it was no significant difference between rhGH/5-FU groups and control group (P>0.05). The average tumor mass and volume of rhGH groups were not significantly increased compared with control group (P>0.05), but they were significantly reduced in 5-FU group and rhGH/5-FU groups (P<0.05). They were no significant difference between rhGH/5- FU groups and 5-FU group (P>0.05). After treatment, the percentages of S, G0/G1 and G2/M phases and proliferation index (PI) were not significantly changed in rhGH groups compared with control group (P>0.05), and the same with rhGH/5-FU groups compared with 5-FU group (P>0.05). The difference caused by dose of rhGH was not significant. Conclusion: rhGH enhances body mass, does not stimulate tumor growth, and has no adverse effects on tumor bearing nude mice. Combined with flurouracil, rhGH does not influence the efficacy of chemotherapy, and has no effect on tumor cell cycle kinetics.

Effects and possible anti-tumor immunity of electrochemotherapy with bleomycin on human colon cancer xenografts in nude mice

AIM: To evaluate the anti-tumor effects and possible involvement of anti-tumor immunity of electrochemotherapy (ECT) employing electroporation and bleomycin in human colon cancer xenografts in nude mice, and to establish the experimental basis for clinical application of ECT.METHODS: Forty nude mice, inoculated subcutaneously human colon cancer cell line LoVo for 3 wk, were allocated randomly into four groups: B+E+ (ECT), B+E- (administration of bleomycin alone), B-E+ (administration of electric pulses alone), and B-E- (no treatment). Tumor volumes were measured daily. The animals were killed on the 7th d, the weights of xenografts were measured, and histologies of tumors were evaluated. Cytotoxicity of spleen natural killer (NK) and lymphokine-activated killer (LAK) cells was then assessed by lactic dehydrogenase release assay.RESULTS: The mean tumor volume of group B+E+ was statistically different from the other three groups after the treatment (F= 36.80, P＜0.01). There was one case of complete response, seven cases of partial response (PR) in group B+E+, one case of PR in group B+E- and group B-E+ respectively, and no response was observed in group B-E-. The difference of response between group B+E+ and the other three groups was statistically significant (χ2 = 25.67, P＜0.01). Histologically, extensive necrosis of tumor cells with considerable vascular damage and inflammatory cells infiltration were observed in group B+E+. There was no statistical difference between the cytotoxicity of NK and LAK cells in the four treatment groups.CONCLUSION: ECT significantly enhances the chemosensitivity and effects of chemotherapy in human colon cancer xenografts in nude mice, and could be a kind of novel treatment modality for human colon cancer.The generation of T-cell-dependent, tumor-specific immunity might be involved in the process of ECT.

Inhibitory effects of Curcuma aromatica oil on proliferation of hepatoma in mice

AIM To reveal the inhibitory effects ofCurcuma aromatica oil(CAO)on cellproliferation of hepatoma in mice.METHODS Two tumor inhibitory experimentsof CAO on hepatoma in mice were conducted.The inhibitory effects of CAO on proliferation ofhepatoma in mice were evaluated by DNA imagecytometry and immunohistochemical staining ofproliferating cell nuclear antigen(PCNA).RESULTS The tumor inhibitory rates of CAOwere 52% and 51% in two experiments,respectively.Compared with those of the saline-treated control groups,both differences werestatistically significant(P【0.01).In the groupof mice treated with CAO,the cellular nuclearDNA OD value(249±70),areas(623 μm^2±228 μm^2)and DNA(2.38±0.67)index of hepaticcarcinomas were significantly lower than thoseof the control group(430±160,1073 μm^2±101 μm^2 and 4.48±0.71).CAO also couldincrease diploidy cell rates(29.00%±9.34% vs2.97%±5.69%,P【0.01)and decreasepentaploidy cell exceeding rate(30.04%±15.10% vs 70.89%±14.94%,P【0.01).In thegroup of mice treated with CAO,the labelingindexes of proliferating cell nuclear antigen (PCNA-LI)were 30%±4%,which weresignificantly lower than 40%±6% of the controlgroup(P【0.01).CONCLUSION The inhibition of CAO on thegrowth of hepatoma in mice might be associatedwith its depression on cellular proliferativeactivity.

Growth-inhibiting effects of taxol on human liver cancer in vitro and in nude mice

AIM To investigate the effects of taxol onSMMC-7721 human hepatoma and itsmechanisms.METHODS In vitro cell growth was assessedby trypan blue exclusion method.Experimentalhepatoma model was established by seedingSMMC-7721 cells subcutaneously into Balb/c(nu/nu)nude mice.In vivo tumor growth wasdetermined by measurement of tumor diameterwith Vernier calipers.The syntheses of DNA,RNA and protein were analyzed by incorporationof ~3H-thymidine,~3H-uridine and ~3H-leucinerespectively.Using light and electronmicroscopes to observe the morphologicalchanges of cells including mitosis andapoptosis.RESULTS Taxol was effective against SMMC-7721 human hepatoma cell growth in the rangesof 2.5 nmol/L-10 nmol/L with mitotic arrestand apoptosis in vitro.DNA,RNA and proteinsyntheses in cells were also obviouslysuppressed by in vitro treatment of taxol for72 h.Taxol at 2.5 nmol/L reduced ~3H-thymidineuptake to about 34% of the control value(P【0.05).Increasing the dose of taxol to20 nmol/L resulted in a greater decrease in ~3H-thymidine incorporation to 60% of the controlvalue(P【0.01).At a concentration of 20 nmol/L,the ~3H-uridine and ~3H-leucine uptakeswere reduced to 52%(P【0.05)and 63%(P【0.01),respectively.In vivo,taxolsignificantly inhibited SMMC-7721 tumor growthat 10 mg/kg,i.p.,once daily for 10 d.A morethan 90% decrease in tumor volume wasobserved by day 11(P【0.01)similarly withmitotic arrest and cell apoptosis.CONCLUSION Taxol has a marked anticanceractivity in SMMC-7721 human hepatoma both invitro and in nude mice.Its mechanisms might beassociated with mitotic arrest,subsequently,apoptosis of the hepatoma cells.No obvioustoxicity was observed with in vivoadministration of taxoi.

The Inhibitory Effects of Rh-endostatin(YH-16) in Combination with Radiotherapy on Lung Adenocarcinoma A549 in Mice and the Underlying Mechanisms

In order to investigate the inhibitory effects of Endostar(rh-endostatin,YH-16)in combination with radiotherapy on lung adenocarcinoma A549 in mice and the interaction mechanisms of combined therapy,the transplantation tumor models of A549 lung adenocarcinoma were established.When the largest diameter of tumor reached 1.0cm,all nude mice were randomly divided into 4 groups:Endostar group,radiotherapy group,radiotherapy plus Endostar(combined treatment)group,and control group(n=6 in each group).The largest d...

^(99m)Tc-labeled HAb18 McAb Fab fragment for radioimmunoimaging in nude mice bearing human hepatocellular carcinoma

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Study of primary leiomyosarcoma induced by MNNG in BALB/C nude mice

INTRODUCTIONIt has been well known that MNNG is one of thestrong and multipotential carcinogens that havebeen frequently reported inducing malignant peptictumors.We have successfully induced rat and doggastric adenocarcinomas,squamous cell carcinomasof rat forestomach and gastric leiomyosarcoma

Immuno-protective effect of tumor cell vaccine on Kunming mice bearing Ehrlich ascites tumor

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Tumor radioimmunoimaging of chimeric antibody in nude mice with hepatoma xenograft

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Effects of endostatin on expression of vascular endothelial growth factor and its receptors and neovascularization in colonic carcinoma implanted in nude mice

AIM: To investigate the antiangiogenic effects of endostatin on colonic carcinoma cell line implanted in nude mice and its mechanism. METHODS: Nude mice underwent subcutaneous injection with LS-174t colonic carcinoma cell line to generate carcinoma and were randomly separated into two groups. Mice received injection of vehicle or endostatin every day for two weeks. After the tumor was harvested, the tumor volumes were determined, and the expressions of CD34, VEGF and FIk-1 were examined by immunohistochemical method. RESULTS: Tumor volume was significantly inhibited in the endostatin group (84.17%) and tumor weight was significantly inhibited in the endostatin group (0.197±0.049) compared to the control group (1.198±0.105) (F=22.56, P=0.001), microvessel density (MVD) was significantly decreased in the treated group (31.857±3.515) compared to the control group (100.143±4.290) (F=151.62, P<0.001). Furthermore, the expression of FIk-1 was significantly inhibited in the treated group (34.29%) compared to the control group (8.57%) (x^2=13.745, P=0.001). However no significant decrease was observed in the expression of vascular endothelial growth factor (VEGF) between these two groups (x^2=0.119, P=0.730). CONCLUSION: Endostatin can inhibit tumor growth and angiogenesis by blocking Vegf/FIk-1 pathway. This experiment provides the theory basis for developing a new anti-carcinoma drug through studying the properties of anti-angiogenesis inhibitors.

Effects of recombinant human growth hormone on growth of human gastric carcinoma xenograft model in nude mice

瞄准：在人的胃的癌房间在活体内的生长上学习 recombinant 人生长激素(rhGH ) 的效果。方法：试验性的老鼠被划分成控制组， rhGH 组， oxaliplatin (L-OHP ) 组和 rhGH+L-OHP 组。有教养的人的胃的癌房间 BGC823 被接种进裸体老鼠的正确腋下，癌异种皮移植模型成功地被建立。异种皮移植肿瘤生长的禁止的率被测量肿瘤体积估计；增殖的房间的表示原子抗原(PCNA ) ， Bax 和异种皮移植肿瘤的 Bcl-2 蛋白质用免疫被检测组织化学的 S-P 方法。结果：肿瘤生长禁止的率， PCNA 的积极表示率， Bax 和 Bcl-2 分别地在 rhGH+L-OHP 组是 49.3% ， 58.2% ， 65.2% 和 59.2% ；46.6% ， 62.5% ， 59.7% 和 64.7% 在 L-OHP 组织；5.0% ， 82.7% ， 23.2% 和 82.2% 在 rhGH 组织， 0， 77.8% ， 23.5% 和 80.3% 在控制组织。在 rhGH+L-OHP 组之间有有效差量(或 L-OHP 组) 并且控制组或 rhGH 组(P 【 0.05 ) ，而没有有效差量(P 】 0.05 ) 在 L-OHP 之间，组和 rhGH+L-OHP 组织并且在 rhGH 之间组织并且控制组。结论：rhGH 不加速人的胃的癌症房间在活体内的增长。

Hepatitis B virus x gene and cyanobacterial toxins promote aflatoxin B_1-induced hepatotumorigenesis in mice

瞄准：估计黄麴毒素的结合的角色 B1 (AFB1 ) ， cyanobacterial 毒素(cyanotoxins ) ，和肝炎 B (HBV ) 在 hepatotumorigenicity 的 x 基因。方法：( ip )带 HBV x 基因和他们的野类型的同窝出生的人的 One-week-old 动物 intraperitoneally 与也被注射单个剂量的 AFB1 [ 6 mg/kg 体重( bw )]，重复剂量 cyanotoxins ( microcystin-LR 或在榴状， 10 microg/kg bw 每周一次为 15 wk )， DMSO (车辆控制)独自一个，或 AFB1 一个星期以后由 cyanotoxins 列在后面，并且在 24 和 52 wk 被牺牲处理以后。结果：AFB1 在 29 中的 13 个导致了肝肿瘤(44.8%) 在 52 wk 的转基因的老鼠处理以后，显著地比在野类型的老鼠(13.3%) 更经常。这有效差量没在 24-wk 学习被显示出。与 AFB1 暴露相比独自一个， MC-LR 并且在榴状在导致的 AFB (1 ) 的发生产出近似 3 褶层和 6 褶层增加在在 24 wk 的野类型的动物的肝肿瘤分别地。HBV x 基因进一步没提高与合作暴露联系到 AFB1 和 cyanotoxins 的风险。与一个 MC-LR-dosed 野类型的鼠标的异常，没有肝肿瘤在在 24 wk 独自与 cyanotoxins 对待的鼠标被观察。既不对待 DMSO 的转基因的老鼠也不他们的野类型的同窝出生的人在甚至多达 52 wk 有与 hepatotumorigenesis 相关的病理学的改变。结论：HBV x 基因并且在榴状支持导致的 AFB (1 ) 的发展肝肿瘤。到 AFB1 和 MC-LR 的合作暴露趋于相对暴露在 24 wk 提高肝肿瘤的风险到他们之一。hepatotumorigenesis 上的 AFB1， cyanotoxins 和 HBVx 的结合的效果在 24 wk 是弱的。

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 Iow 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-Iincreased gradually following tumor progression in LCID20 model, and correlated with tumor size and AFP level,Phasic expression of tissue intercellular adhesion molecule-I 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 antiangiogenesis, 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 vivo and in vitro study of HCC invasiveness, angiogenesis as well as intervention of HCC recurrence.

Antihepatoma effect of alpha-fetoprotein antisense phosphorothioate oligodeoxyribonucleotides in vitro and in mice

AIM To evaluate antihepatoma effect ofantisense phosphorothioate oligodeo-xyribonucleotides (S-ODNs) targeted to alpha-fetoprotein (AFP) genes in vitro and in nudemice.METHODS AFP gene expression was examinedby immunocytochemical method or enzyme-linked immunosorbent assay. Effect of S-ODNson SMMC-7721 human hepatoma cell growth invitro was determined using microculturetetrazolium assay. In vivo antitumor activitiesof S-ODNs were monitored by measuring tumorweight differences in treated and control micebearing SMMC-7721 xenografts. Induction of cellapoptosis was evaluated by fluorescence-activated cell sorter (FACS) analysis.RESULTS Antisense S-ODN treatment led toreduced AFP gene expression. Specificantisense S-ODNs, but not control S-ODNs,inhibited the growth of heaptoma cells in vitro.In vivo. only antisense S-ODNs exhibitedobvious antitumor activities. FACS analysisrevealed that the growth inhibition by antisenseS. ODNs was associated with their cell apoptosisinduction.CONCLUSION Antisense S-ODNs targeted toAFP genes inhibit the growth of human hepatomacells and solid hepatoma, which is related totheir cell apoptosis induction.

Effects of epidermal growth factor on the growth of human gastric cancer cell and the implanted tumor of nude mice

AIM: Epidermal growth factor (EGF) plays an important rolein the regulation of gastrointestinal tissue growth anddevelopment, and it can stimulate epithelial proliferation,cell differentiation and growth. It has been established thatthe EGF can promote gastric cytoprotection and ulcerhealing. But the potential ability of EGF to regulate thegastric cancer growth is unknown. This study is toinvestigate the influence of EGF on human gastric cancercell and the implanted tumor growth of nude mice.METHODS: The cell growth rates of human gastricadenocarinoma cell lines MKN-28, MKN-45, SGC-7901 andnormal human gastric epithelial cells 3T3 were assessedwhen incubated with recombinant human EGF (rhEGF, 0.05, 0.1, 0.5, 1.0, 10, 50, 100 mg.L-1) using MTT method.The cells of MKN-28, MKN-45, SGC-7901 (gaatric cancertissue 1.5 mm3 ) were implanted in the BALB/cA nude micefor 10 days. The EGF was given intrapsritoneally (15, 30, 60μg. kg-1) for 3 weels. The body weights of the tumor-bearing animals and their tumor mass were measuredafterwards to assess the mitogenic effect of rhEGF in thenude mice.RESULTS: Within the concentration range of 0.05-100 mg.L-1 , rhEGF could increase the cell growth ofnormal 3T3 cells(cell growth rate 100 % vs 102.8 %, P＜0.05), but partiallyrestrain the gastric cancer cell growth. The latter effect wesrelated to cell differentiation. In 15-60μg/kg rhEGF groups,the mean implanted tumor mass of MKN-28 cell were 1.75 g,1.91 g, 2.08 g/NS group 1.97 g ( P＞ 0.05), the mean tumormass of SGC-7901 cell were 1.53 g, 1.07 g, 1.20 g/NS group1.07 g ( P ＞ 0.05), and for MKN-45 cell, the tumor masswere respectively 1.92 g, 1.29 g, 1.77 g/NS group 1.82 g( P＞ 0.05 ). So rhEGF had no obvious effect on implantedMKN-28, SGC-7901 and MKN-45 tumor growth.CONCLUSION: EGF has no stimulating effect on the humangastric cancer cell growth neither in vitro nor in vivo.

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.

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 Iigand of tumor cells in lymph nodes.METHODES 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 macrophages in lymph nodes were observed with in situ DNA fragmentation.RESULTS On the 28th day post-inoculation, the lymph node metastatic rate of Hca-F 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 HcaF 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.