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 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.

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

AIM： To study effects of recombinant human growth hormone （rhGH） on growth of a human gastric carcinoma cell in vivo. METHODS： Experimental mice were divided into control group, rhGH group, oxaliplatin （L-OHP） group and rhGH＋L-OHP group. Cultured human gastric carcinoma cells BGC823 were inoculated into right axilla of nude mice and carcinoma xenograft model was established successfully. Inhibitory rate of xenograft tumor growth was estimated by measuring tumor volume; expression of proliferating cell nuclear antigen （PCNA）, Bax and Bcl-2 proteins of xenograft tumor was detected using immunohistochemical S-P method. RESULTS： Tumor growth inhibitory rate, the positive expression rate of PCNA, Bax and Bcl-2 were 49.3%, 58.2%, 65.2% and 59.2% in rhGH＋L-OHP group respectively; 46.6%, 62.5%, 59.7% and 64.7% in L-OHP group; 5.0%, 82.7%, 23.2% and 82.2% in rhGH group and 0, 77.8%, 23.5% and 80.3% in control group. There was significant difference between rhGH＋L-OHP group （or L-OHP group ） and control group or rhGH group （P 〈 0.05）, whereas there were no significant differences （P 〉 0.05） between L-OHP group and rhGH＋L-OHP group and between rhGH group and control group. CONCLUSION： rhGH does not accelerate the proliferation of human gastric cancer cell in vivo.

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.

Inhibitory effect of parvovirus H-1 on the formation of colonies of human hepatoma cell line in vitro and its tumors in nude mice

The inhibitory effect of parvovirus H-1 on the colonyforming ability in vitro of QGY-7703, a cultured human hepatoma cell line, and on the formation and growth of its tumors in nude mice was studied. With higher multiplicity of infection (MOI) of H-1 given, survival of the QGY-7703 cells was found to be decreased. H-1 DNA amplification level at 30 h postinfection(p.i.) was detected to be 7.4 times higher than that at 2 h by dispersed cells assay, while the cells were delayed to enter into S phase.Plaques were formed in the indicator cells (new-born human kidney cell line, NBK) by progeny H-1 virus particles released from the infected QGY-7703 cells by infectious cell center assay. The formation of tumors in nude mice by QGY-7703 cells which were injected s c at 2 h postinfection was observed to be prevented in 2 groups with given MOI 25 and 50. The tumor growth of MOI 10 group occurred at a lower exponential rate than that of control,after a 20 d latent period. It was evident that parvovirus H-1 exhibited a direct inhibitory effect on the formation and growth of human hepatoma cells in vivo as well as in vitro.

ANTI-HUMAN LUNG GIANT CELL CANCER (PG) EFFECT OF HUMAN LAK CELLS IN VITRO AND IN NUDE MICE

Human LAK cells were prepared by culturing normal human peripheral blood mononuclear cells (PBMC) with or without rIL-2 and assayed for T cell surface markers as well as anti-tumor activity against PC in vitro and in nude mice. Although the percentages of T3, T4, and T8 positive cells in rIL-2-activated cells did not differ significantly from those of control cells in vitro, the former showed stronger cytotoxicity than control cells to PG tumor cells in vitro. In vivo, LAK cells completely inhibited the growth of PG tumor in nude mice, whereas PBMC control cells were to be of no effect. The anti-tumor effect of human LAK cells in nude mice may offer a useful model to study the role of human LAK cells against human tumor in vivo.

RESPONSE OF HUMAN RENAL CANCER TO UFT IN NUDE MICE

A primary human renal cell carcinoma was developed as a xenograft (NT-25) and maintained by serial transplantation in nude mice. The effect of UFT on this neogrowth was tested and evaluated as well its distribution in the animal tissues. The concentration of UFT was higher in tumor tissues than that in other tissues and in the animal experimentation UFT was found to be effective on human renal cell carcinoma.

EFFECTS OF PSEUDOFYPE RETROVIRUS CONTAINING HUMAN N-RAS ANTISENSE GENE ON THE GROWTH OF HUMAN LIVER CANCER LTNM4 TRANSPLANTED IN NUDE MICE

An amphotropic pseudotype retrovirus containing human N-ras antisense gene was constructed and packaged with helper cells. It has been previously demonstrated that the virus did inhibit the growth of human hepatocarcinoma cell line PLC PRF/5 in vitro accompanied with the blockage of p21 expression. Based on these results, further study was carried on to examine the effect of these viruses on the growth of human hepatoma transplanted LTNM4 in nude mice. It has been shown that the retrovirus containing human antisense N-ras gene could inhibit the hepatoma in nude mice at a rate of 78% (P<0.05) as compared with saline control. No inhibition was observed in group treated with retrovirus which contained no N-ras sequence. These results in vivo lend further support that human N-ras antisense gene mediated by retrovirus could block the expression of the relevant oncogene and lead to the inhibition of cancer growth. It also provided the basis for further approaches of gene therapy for human cancer.

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.

MORPHOLOGICAL SURVEY ON ENDOGENOUS C-TYPE VIRUSES INFECTING A HUMAN LUNG SQUAMOUS CARCINOMA PASSAGED IN NUDE MICE

A human lung squamous carcinoma was transplanted and passaged in Swiss-DF nude mice, called LSX-83, for more than five years in our laboratory. The morphological characteristics of the original tumor were maintained in passages from 4 to 33. But from the 35th generation, an increasing amount of tonofilaments and nuclear segregation with typical features was found with electron microscopy. The C-type virus particles were first detected in extra cellular space after 40 passages. The viruses were observed in different stages of growth, but their distribution and number did not show apparent change up to 54 passages. Such findings suggest that LSX-83 cells probably possess certain barrier of resistance against C-type viruses. The relation between C-type viruses and the morphological changes of LSX-83 cells was discussed.

Orthotopical transplantation of human renal carcinoma tissue into nude mice and the establishment of a high metastatic cell line MRCC

Objective To establish a SOI model of human renal carcinoma and a high metastatic cell subline. Methods A human renal cell line RCC-9863 has been established by inoculating a human renal tumor tissue into nude mice s. c.. When RCC-9863 passaged for 20 times, the tissue from the same xemotransplant tumor were used to construct SOI model. Cultured the metastatic tissue in vitro, the tumor cell suspension was then injected orthotopically, The metastatic tissue obtained underwent the same procedure again. At last, the metastatic tumor was cultured in vitro and cloned. Results 15 days later, a tumor mass sized 1. 7 cm × 0. 6 cm in the nude mouse’s renal parenchyma was grown which lobulated, rude, and with multiply blood vessels and 55 days later later the mouse became moribund and metastases in the lungs were formed. The transplanted renal tumor in the SOI model grew fast and invasively and metastasized to lungs, lymphatic node and liver. A subline, MRCC, with metastatic ability to the lung was selected.

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.

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 role in the regulation of gastrointestinal tissue growth and development, and it can stimulate epithelial proliferation, cell differentiation and growth. It has been established that the EGF can promote gastric cytoprotection and ulcer healing. But the potential ability of EGF to regulate the gastric cancer growth is unknown. This study is to investigate the influence of EGF on human gastric cancer cell and the implanted tumor growth of nude mice. METHODS: The cell growth rates of human gastric adenocarcinoma cell lines MKN-28, MKN-45, SGC-7901 and normal human gastric epithelial cells 3T3 were assessed when 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 (gastric cancer tissue 1.5mm(3)) were implanted in the BALB/cA nude mice for 10 days.The EGF was given intraperitoneally (15, 30, 60 microg.kg(-1)) for 3 weeks. The body weights of the tumor-bearing animals and their tumor mass were measured afterwards to assess the mitogenic effect of rhEGF in the nude mice. RESULTS: Within the concentration range of 0.05-100mg.L(-1), rhEGF could increase the cell growth of normal 3T3 cells (cell growth rate 100% vs 102.8%, P【0.05), but partially restrain the gastric cancer cell growth. The latter effect was related to cell differentiation. In 15-60 microg/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 tumor mass of SGC-7901 cell were 1.53 g, 1.07 g, 1.20 g/NS group 1.07 g (P】0.05), and for MKN-45 cell, the tumor mass were respectively 1.92 g, 1.29 g, 1.77 /NS group 1.82 g (P】0.05). So rhEGF had no obvious effect on implanted MKN-28, SGC-7901 and MKN-45 tumor growth. CONCLUSION: EGF has no stimulating effect on the human gastric cancer cell growth neither in vitro nor in vivo.

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.

Targeting Effect Study of ￣(3) H-Mitoxantrone Nanosphereson Hepatocellular Carcinoma(HCC) Model in Nude Mice

The distribution of ￣(3)H-mitoxantrone polybutyl cyanoacrylate nanospheres(￣(3)H-DHAQ-PBCA-NS)in the viscera,muscle and tumors of human hepatocellular carcinoma (HCC)model in nude mice was studied with liquid scintillation counting techniique. The results showed that the ￣(3)H-DHAQ-PBCA-NS had remarkable liver targeting effect. The content of ￣(3)H-DHAQ-PBCA-NSin liver and heterotopic liver tumor was found to be 71.31±10. 49% of total amount of drug in animal body. It was also found that the content of ￣(3)H-DHAQ-PBCA-NS in liver was higher than that in liver tissue, and the content of ￣(3)H-DHAQ-PBCA-NS in annpit tumor was higher than that in armpit muscle tissue,but had no significant difference;It provides an ideal preparation for the DHAQ admini-stration.

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.

EFFECTS OF TNF ALONE OR IN COMBINATION WITH CHEMOTHERAPEUTIC AGENTS ON HUMAN OVARIAN CANCERS IN VITRO AND IN NUDE MICE

Using the tetrazolium (MTT) assay, we examined the cytotoxicities of recombinant human tumor necrosis factor (rhTNF) and five chemotherapeutic agents, namely CTX, 5-FU, VCR, DDP and KSM, on human ovarian cancer cell lines OVCAR3 and CAOV3. The results showed that the cytotoxicities of rhTNF at concentrations of 50-50 000 U / ml on OVCAR3 cell line and CAOV3 cell line exposed to rhTNF for 24 hours were from 14.2% ± 6.8% to 67.2%± 3.0% and from 8.2%± 4.3% to 60.9%±1.3%, respectively. The cytotoxicities of all five chemotherapeutic agents tested on the two cell lines were much lower than that of rhTNF. We also studied the combined antitumor potential of rhTNF with the five chemotherapeutic agents and the results showed that there were various degrees of synergism in cytotoxicities of rhTNF in combination with DDP or KSM on the two cell lines. Based on experiments in vitro, the in vivo antitumor activities of rhTNF, both alone and in combination with KSM, were examined in OVCAR3 cancer transplanted in nude mice. The results showed a considerable antitumor effect of rhTNF when it was used alone and a marked synergistic effect when it was used in combination with KSM on the xenograft tumors.

Human breast carcinoma xenografts in nude mice

OBJECTIVE: To investigate spontaneous metastasis, micrometastasis and genetic stability in human breast carcinoma xenografts in nude mice. METHODS: Intact tissue from surgical specimens from breast carcinoma patients was xenografted into nude mice and transplanted from generation to generation. Cells from the xenografts were cultured in vitro and retransplanted into nude mice. Microsatellite DNA in the genome of human breast carcinomas, xenotransplanted tumors and metastases in nude mice were analyzed at three microsatellite loci. RESULTS: The tumorigenicity of orthotopic xenotransplantation was 88.6% (31/35), with a metastatic rate of 41.9% (13/31). Cells from xenotransplants were successfully cultured in vitro. The taking rate of retransplantation into nude mice and the spontaneous lung metastasis rate were both 100% (10/10). Microsatellite DNA sequences in the genome of xenotransplanted tumors and metastases in nude mice were identical with that of the original human breast carcinoma at three microsatellite loci. CONCLUSIONS: Tumorigenicity and metastatic potential can be improved in human breast carcinoma xenografts using intact fresh tumor tissue and orthotopic grafts. Xenotransplanted tumors and tumors after serial passage maintained the genetic stability. The detection of microsatellite DNA may identify micrometastases in a nude mouse model.