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.

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.

Tumor radioimmunoimaging of chimeric antibody in nude mice with hepatoma xenograft

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Effects of targeting magnetic drug nanoparticles on human cholangiocarcinoma xenografts in nude mice

BACKGROUND: Targeting is a new therapeutic tool for malignant tumor as a result of combining nanotechnology with chemotherapeutics. The aim of our study was to investigate the effects of magnetic nanoparticles enveloping a chemotherapeutic drug on human cholangiocarcinoma xenografts in nude mice. METHODS: The human cholangiocarcinoma xenograft model was established in nude mice with the QBC939 cell line. The nude mice were randomly assigned to 7 groups. 0.9% saline or magnetic nanoparticles, including high (group 2), medium (group 4) and low (group 5) dosages, were given to nude mice through the tail vein 20 days after the QBC939 cell line was implanted. Calculations were made 35 days after treatment in order to compare the volumes, inhibition ratios and growth curves of the tumors in each group. Mice in each group were sacrificed randomly to collect tumor tissues and other organs for electron microscopy and pathological examination. RESULTS: The high and medium dosage groups were significantly different from the control group (P<0.05). The tumor inhibition ratios for the high, medium and low dosage groups were 39.6%, 14.6% and 7.9%, respectively. The tumor growth curve of groups 5, 4, and 2 changed slowly in turn. The high and medium groups showed cell apoptosis under an electron microscope. CONCLUSION: Magnetic nanoparticles can inhibit the growth of human cholangiocarcinoma xenografts in nude mice.

Antitumor effects of artesunate on human breast carcinoma MCF-7 cells and IGF-IR expression in nude mice xenografts

Purpose： The objective of this study was to investigate the anti-tumor effects and analyze the mechanism of artesunate （ART） action on breast cancer in vivo using tumor transplanted nude mice. Methods： The human breast tumor cell line MCF-7 was transplanted into nude mice, and the animals were treated with various doses of ART alone or in combination with cyclophosphamide （CTX） or normal saline （NS）. The tumor inhibitory effects were observed and compared, and the ultrastructural morphology of the transplanted tumor cells was observed by electron microscopy. The apoptosis rates and cell cycle status were detected by flow cytometry （FCM）. The expression of apoptosis-related proteins p53, Bcl-2, Bax and Caspase-3 were detected by immunohistochemistry and IGF-IR was detected by western blot. The expression correlation for these proteins was also analyzed. Results： The tumor inhibition rates in the low dose ART group, high dose ART group, CTX group and combined drug therapy group were （24.39±10.20）%, （40.24±7.02）%, （57.01±5.84）% and （68.29±5.1）%, respectively. The cell cycle was arrested in phase G0/Gt after treatment with ART. The expression of Bcl-2 was significantly reduced, and the expression levels of Bax and Caspase-3 were significantly increased in the ART group compared to the negative control saline group. There was no significant difference detected in p53 expression. The Bcl-2 level was negatively related to Bax and Caspase-3. The western blotting results showed IGF-IR downregulation. Conclusions： ART inhibits the growth of MCF-7 breast tumor cell xenografts in nude mice. The anti-tumor mechanism of ART for human breast carcinoma in nude mice might be correlated with the alteration of apoptosis related protein expression, which may further induce apoptosis and inhibit cell proliferation.

Functional Mechanism of Resveratrol in Inhabiting Growth of Cells ls174t and Its Mechanism in Subcutaneously Transplanted Tumor of Nude Mice

To explore the functional mechanism of Resveratrol against colon cancer cells ls174t and the growth of colon cancer tissue of tumor-bearing mice, MTT method was used to observe the functions of resveratrol for inhibition against cells ls174t in vitro. Transmission electron microscope was used to observe the cell apoptosis. FCM assay was performed to measure the change of the cell apoptosis rate and of cell cycle. RT-PCR method was used to detect the expressions of bcl-2 and bax mRNA. Western blot method was used to detect the expressions of bcl-2 and bax protein. Ceils ls174t were transplanted subcutaneously to nude mice to observe the effect of resveratrol on the growth of subcutaneously transplanted tumor, RT-PCR method was used to detect the expressions of bcl-2 and bax mRNA in the tumor tissue. Western blot method was used to detect the expressions of bcl-2 and bax protein in the tumor tissue. Resveratrol has an effect of inhibiting proliferation of cells ls174t in vitro（P〈0.01）. It is able to induce the apoptosis of cells ls174t, causing the decrease in the expression of bcl-2 and the increase in the expression of bax. Resveratrol could inhibit the growth of subcutaneously transplanted tumor of nude mice（P〈0.05）, causing the decrease in the expression of bcl-2 and the increase in the expression of bax. Resveratrol can inhibit the growth of cells 174t and the growth of subcutaneously transplanted tumor. The mechanism is possibly related to the induction of the cell apoptosis and the regulation of bcl-2/bax expression.

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

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Effect of Quercetin on Breeding and Apoptosis of Cervical Cancer HeLa Cell and on Growth of Transplanted Tumor in Nude Mice

Effect of quercetin on HeLa cell system of cervical cancer was studied by methods of MTT and Annexin V-FITC/PI. The results show that quercetin has functions of inhibiting breeding of HeLa cells and inducing apoptosis of the cells. The total apoptosis rate is positively proportional to reaction duration and concentration of quercetin used. The maximum apoptosis rate being （88.76±2.35）% was obtained when the concentration was 50.0 μmol/L and the cells were treated with quercetin for 72 hours. Based on establishing a model of tumor of cervical cancer transplanted into nude mice, quercetin of different concentrations was injected into abdominal cavity of nude mice and situation of tumor growth was reviewed. The result showed that with quercetin concent＇ration increasing from 0 to 100.0 μmol/L, the transplantation volume and weight of the tumors decreased from （279.59±70.58） mm^3 and （0.145±0.019） g to （128.72±36.12） mm^3 and （0.089± 0.019） g respectively, while apoptosis rate of the transplanted tumor increased from （9.63±1.85）% to （34,98±0.47）%, which proved that quercetin inhibited increment of volume and weight of transplanted tumor in nude mice bodies.

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

Antitumor Activities and Apoptosis-regulated Mechanisms of Fermented Wheat Germ Extract in the Transplantation Tumor Model of Human HT-29 Cells in Nude Mice

Objective A subcutaneous transplantation tumor model of human HT-29 cells in nude mice was established to evaluate anticarcinogenic activities, and the apoptosis-regulated mechanism effect of aqueous extract of fermented wheat germ with Lactobacillus plantarum dy-1 （LFWGE）. Methods The HT-29 cells were transplanted via subcutaneous injection of 1×10^7cells into the right flank of each nude mouse. Then, nude mice were treated for 30 d with LFWGE （high-dose 2 g/kg/d; low-dose 1 g/kg/d） and for 7 d with 5-fluorouracil （5-FU, 25 mg/kg/d） by gavage and intraperitoneal injection, respectively. An inhibition of tumor growth was observed. Results Tumor volume and weights decreased significantly in both groups of nude mice treated with LFWGE. In addition, the cell apoptosis rate of the LFWGE group （2 g/kg/d, 60.2%＋4.4%; 1 g/kg/d, 58.6%＋6.9%） was significantly higher than that of the control group （11.5%＋1.6%） and 5-FU group （32.1%＋3.5%） as measured by the TUNEL assay. Moreover, the real-time fluorescent quantitative PCR and Western blot method further confirmed these enhancing apoptosis and growth inhibition effects. The involvement of LFWGE in inducing apoptosis was confirmed by the expression of Bax, Bcl-2, Caspase-3, and CyclinD1. Conclusion The results showed that LFWGE could induce subcutaneous transplantation tumor apoptosis in nude mice and could be as a natural nutrient supplements or chemopreventive agent in the treatment of human colon cancer.

Antitumor Activities and Apoptosis-regulated Mechanisms of Fermented Barley Extract in the Transplantation Tumor Model of Human HT-29 Cells in Nude Mice

Objective A subcutaneous transplantation tumor model of human HT-29 cells was established in nude mice to study the anticarcinogenic activities and apoptosis-regulatory mechanistic effect of aqueous extract of fermented barley with Lactobacillus plantarum dy-1 （LFBE）. Methods HT-29 cells were transplanted via subcutaneous injection of 1 × 107cells into the right flank of each nude mouse. Then, nude mice were treated for 30 days with LFBE （high-dose 2 g·kg-1·d-1; low-dose 1 g·kg-1·d-1） and for 7 days with 5-fluorouracil （5-FU, 25 g·kg-1·d-1） by gavage and intraperitoneal injection, respectively. Results Tumor volume and weight decreased significantly in both groups of nude mice treated with LFBE. In addition, the cell apoptosis rate of the LFBE group was significantly higher than that of the control group and 5-FU groups as measured by the TUNEL assay. Moreover, the real-time fluorescent quantitative PCR and Western blot methods further confirmed these apoptosis-enhancing and growth-inhibiting effects. The involvement of LFBE in inducing apoptosis was confirmed by the expression of Bax, Bcl-2, caspase-3, and cyclin D1. Conclusion The results showed that LFBE could induce subcutaneous transplantation tumor apoptosis in nude mice and could be used as a natural nutrient supplement or chemopreventive agent in the treatment of human colon cancer.

Systemic study on the safety of immuno-deficient nude mice treated by atmospheric plasma-activated water

Cold atmospheric-pressure plasma is a new technology, widely used in many fields of biomedicine,especially in cancer treatment. Cold plasma can selectively kill a variety of tumor cells, and its biological safety in clinical trials is also very important. In many cases, the patient’s immune level is relatively low, so we first studied the safety assessment of plasma treatment in an immunocompromised animal model. In this study, we examined the safety of immuno-deficient nude mice by oral lavage treatment of plasma-activated water, and studied the growth status, main organs and blood biochemical indexes. Acute toxicity test results showed that the maximum dose of plasma treatment for 15 min had no lethal effect and other acute toxicity. There were no significant changes in body weight and survival status of mice after 2 min and 4 min of plasma-activated water(PAW)treatment for 2 weeks. After treatment, the major organs, including heart, liver, spleen, lung and kidney, were not significantly changed in organ coefficient and tissue structure. Blood biochemical markers showed that blood neutrophils and mononuclear cells were slightly increased, and the others remained unchanged. Liver function, renal function, electrolytes, glucose metabolism and lipid metabolism were not affected by different doses of PAW treatment. The above results indicate that PAW treatment can be used to treat immuno-deficient nude mice without significant safety problems.

^(131)I-labelled anti-hepatoma monoclonal antibodies in targeting therapy of hepatoma xenografts in nude mice

Anti-hepatoma monoclonal antibodies HAb8,HAb18 IgG and HAb18 F（ab’）2were labelled with[131I]according to chloramine T method.The labelling rate andradioactivity were 51% and 9.58×104Bq/mg,respectively.After administration of the[131I]-labelled conjugates through mouse tail vein,hepatoma xenografts were clearly im-aged,Localization index was 2.15,3.09,and 6.99 for HAb8,HAb18 IgG and HAb18F（ab’）2,respectively.A total of 16 hepatoma-bearing nude mice（SMMU-LTNM）were di-vided into 4 groups.When xenografts reached 0.3cm in diameter,the treatment was be-gun by injection of the[131I]-conjugates through mouse tail vein.After 2 weeks of treat-ment,obvious inhibitory or killing effects on the xenografts were found.The tu-mor-bearing mice were killed during the period from 14 to 21d after treatment.Xenografts as well as other organs were sampled and viewed under light microscope ac-cording to routine procedure.The vital organs,such as heart,liver,spleen,lung,kidneyand brain showed no pathological lesions.In hepatoma xenografts,some tumor cellmembranes were found,with obscure cytoplasmic structure and increased eosinophilia,representing necrotic alterations.

Tumor localization of anti-gastric cancer monoclonal antibody MGb_2-daunomycin conjugate in nude mice bearing human gastric carcinoma SGC-7901

In the present study,an anti-gastric cancer monoclonal antibody MGb2-daunomycin（DM）conjugate was prepared by cis-aconitic anhydride method.The distribution of the conjugatein nude mice bearing human gastric carcinoma SGC-7901 was investigated.Four to five moleculesof DM were introduced into each molecule of antibody while the antigen-binding capacity ofthe antibody was well-retained.This conjugate could be satisfactorily localized in the tumortissue.Ninety-six hours after intraperitoneal injection of 125I-MGb2-DM conjugate,the ratio ofradioactivity per milligram of tumor to that per milligram of blood reached 203,the result beingquite similar to that of unmodified MGb2.

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.

NEOPLASTIC CELL APOPTOSIS IN NUDE MICE TRANSPLANTS WITH NASOPHARYNGEAL CARCINOMA CELL LINES

Objective: To observe the morphological features of neoplastic cell apoptosis developed in nude mice transplants with nasopharyngeal carcinoma (NPC) cell lines, CNE-1 and CNE-2, and to investigate the roles of p53, bcl-2 and bax playing in the process of apoptosis. Methods: CNE-1 and CNE-2 cell lines were inoculated and passed in nude mice for 3 generations. The cell apoptosis was detected on H & E and TUNEL staining slides. The expression of p53, bcl-2 and bax were detected by using immunohistochemistry. p53 gene alteration was assayed in cell lines and transplants by PCR-SSCP. Results: A considerable number of neoplastic cells underwent apoptosis in CNE-1 and CNE-2 transplant tissues. The “shrinkage necrosis” and apoptotic bodies were the main appearances of apoptosis. The p53 alteration was detected in exon 8 by PCR-SSCP and p53 protein accumulation observed in the cell smears and nude mice transplant tissue sections. All the transplant tissue sections of 3 passages showed bcl-2 negativity and bax overexpression. Conclusion: The neoplastic cells of CNE-1 and CNE-2 transplants underwent death mainly taking the way of apoptosis. The “shrinkage necrosis” and apoptotic bodies were the main morphological features of apoptosis seen in those transplants. The apoptosis in CNE-1 or CNE-2 nude mice transplant is highly probable through a p53-independent and bax-mediated pathway.

Evaluation of Tumor Formation of Three Bladder Cancer Cell Lines in Nude Mice

This study examined the differences in tumor formation of three bladder tumor cell lines （BIU-87, T24 and EJ） after subcutaneously transplanted into nude mice, in order to find the best technique for establishing in vivo bladder tumor model. BIU-87, T24 and EJ cells at logarithmic phase were re-suspended in serum-free medium. The cells suspensions of the identical concentration were subcutaneously transplanted into nude mice and then the success rate and tumor growth were compared among the three cell groups. The results of tumor formation were pathologically evaluated. Lung, liver and kidney tissues were also pathologically examined for distant metastasis. The proliferation of the three cells were determined by immunohistochemically detecting the PCNA expression in the tumors. The results showed that the success rates of EJ and T24 cells were significantly higher than that of BIU-87 cells and no distant metastasis was noted among the three groups. The proliferation levels of EJ and T24 cells was significantly higher than that of BIU-87. But at the later stage of tumor formation, as compared with T24 cells, EJ grew more vigorously, soon resulting in the central necrosis of tumor, which affected the measurement of the actual size of the tumors. Moreover, PCNA staining exhibited that the proliferation of EJ and T24 was significantly higher than that of BIU-87 cells. It is concluded that as compared with BIU-87 cells, EJ and T24 cells had higher success rates, with not significant differences in death rate and distant metastasis found among them. There existed no significant difference in tumor formation between EJ and T24 cells and T24 cells do not rupture easily, which makes it a better cell line for the establishment of in vivo bladder tumor model.

RESEARCHES ON THE ANTI-AUTOLOGOUS TUMOR EFFECTS OF HUMAN CANCER-KILLING CELLS IN NUDE MICE

An investigation of human splenic LAK cells killing autologous tumor in vivo has been performed in this study. Briefly, an ovary embryo carcinoma (OEC) removed surgically from patient was transplanted Into nude mice (s. c. ) and cultured in vitro, that would be used as targets. Lymphokine-activated killer (LAK) cells were generated from splenic lymphocytes of the OEC patient, who died two and half months after operation, by co- culture with recombinant human interleukin- 2 ( rIL- 2 ) in vitro. The results from Winn' s test in nude mice suggested that these LAK cells could effectively inhibit the tumorlgenicity of autologous tumor m vitro.

Localization and Biodistribution of Conjugate ATG－Dex－DNR in Nude Mice as Models for Human Leukemia

I-labelled anti-thymoglobuline (ATG), 131 I-labelled immunoconjugate ATG-Dex-DNR and 131 I-labelled Ts-MoAb as control antibody, respectively.were injected by intraperitoneal (i. p.) administration into nude mice used as models for human T-cell leukemia. SPECT imaging was performed from day 1 to day 8 following i. p. injection. The results showed that radioimmunoimaging of buman tumor xenografts was clearest day 3 after injection in both of ATG and ATG-Dex-DNR groups, whereas it's not the case in Ts-MoAb group. Nude mice were killed 8th day after injection with antibody or conjugate. The tumor, as well as different dissected normal organs including heart, liver, lungs, kidney, femur and intestine, were harvested, weighed precisely, and radioiodine-counted. T/NT ratios in experimental group was greater than 1. 0 (ranged from 1. 246-7. 865). and in control group they were less than 1. 0 (ranged from 0. 263-0. 757, except for tumor/femur ratio). Our results indicated that ATG and ATG-Dex-DNR had specific affinity to cell line of Tcell leukemia CEM.

INHIBITORY EFFECT OF DEMO ON THE GROWTH OF TRANSPLANTED HUMAN COLON CANCER IN NUDE MICE

A human colon cancer cell line Hce- 8693 was heterotransplanted in nude mice. Polyamine blosythesis Inhibitor a- dlfiuoromethylomithine (DFMO ) show a marked reproducible inhibition in this model. The size and weight of transplanted tumor In DFMO group were smaller than those of the control group and the average inhibition rate was 72.8% (P < 0.001) . DFMO showed higher tumor inhibitory rate than 5-Fu (35. 4%) (P<0. 001) . Furthermore. DFMO demonstrated less severe bone marrow inhibition in the nude mice than 5-Fu (20. 0% Vs 53. 2%. P<0. 001) .There was no synergistic action in these two drugs at the experimental dose. The concentration of putrescine and spermidine in the plasma and tumor tissue in the DFMO group were 70% lower than those of the control group (P<0. 001) . These results indicate that the anti-tumor effect of DFMO might be explained by the inhibition of polyamine biosynthesis and this study provides an experimental basis for future clinical application of DFMO.