| Part I:Establishment and MRI Comprehensive Evaluation of a Rabbit Model Beared LiverVX2 TumorObjectiveTo establish the animal model of VX2 hepatic tumor in rabbits, then to evaluate VX2 hepatic tumor imaging features using magnetic resonance imaging (MRI) which set up the experimental basis for this model.Material and methodsVX2 tumor were implanted into the liver lobe with for establishment of animal models in Ten New Zealand rabbits. The tumors were allowed to grow for two weeks, then the animal model was scanned by MRI every two weeks for 2 times. Among them, five rabbits were sacrificed randomly after they were scanned by MRI. Their livers were harvested for histopathology examination, comparing the results of histopathology with MRI results.ResultsAll of 10 rabbits were successfully implanted with VX2 tumor in the livers by paunching. While MRI scans, the tumors were seen as hypointensity on T1WI, and hyperintensity on T2WI or DWI, and the profile of the tumor appears very clear on DWI. The regional necrosis of the tumor was observed as trait of intensity as free water on T1WI, T2WI and DWI. The tumors were slightly hyperintense on SWI, and have clear boundary. The draining veins around or within the tumor and tumor hemorrhage can be seen. PWI performance:the tumor parenchyma showed high perfusion on the relative hepatic blood volume (rHBV) map. Time-signal intensity curve of the tumor area was rapid downhill and ascending type, while the curve of normal liver parenchyma showed decline slowly over time. The tumors demonstrated itself inhomogeneous enhancement on arterial phase T1WI, with apparent ring-like enhancement of the edge, no enhancement in necrotic areas. The lesion is reduced to a low signal on the portal venous phase T1WI.ConclusionThe rabbits VX2 liver tumor model can be duplicated by the highly successful rate. MRI can accurately evalueate the growth, draining veins, bleeding and other histopathology of the tumors. The model can be used to investigating diagnostic imaging and treatment of liver cancer. MRI evaluation is very useful in monitoring and screening experimental animal. Part II:Experimental Study of transcatheter hepatic arterial infusion using 131I-anti-CD147-McAb in rabbitsObjectiveTo investigate CD 147 expression of VX2 liver cancer in rabbit. Then, we evaluate the technical feasibility of transcatheter hepatic arterial infusing 131I-anti-CD147-McAb and biodistribution of 131I after the procedures in a rabbit liver cancer model.Material and methodsEighteen rabbits with tumor diameter 2cm to 3cm was selected. Among them, three rabbits were randomly selected to investigate the expression of CD 147 antigen in VX2 liver tumor. Mouse anti-human CD 147 monoclonal antibody was used to stain the VX2 tumor tissue. Other 15 rabbits were infused 131I-anti-CD147-McAb by transcatheter hepatic arterial infusion. Radionuclide biodistribution was observed by SPECT-CT scanning in the 15 experimental rabbits at 24 hours,72 hours and 7 days after procedures respectively. Five rabbits were randomly selected to be sacrificed after SPECT-CT scanning at each time point. The tumor, liver, lung, spleen, kidney, heart, thyroid and a vertebrae (lumbar spine) were completely cut and crushed respectively.2ml sample of each tissue was used to detect the 131I biodistribution in the corresponding organ or tissue by y counter. The first test was performed in the background, all samples were counted for 30 seconds. The tumor/non-tumor (T/NT) ratio was calculated after the data were gotten, and three set of data were analyzed.ResultsMouse anti-human CD147 monoclonal antibody was used to stain the tumor tissue and normal liver tissue. Results showed that the VX2 tumor highly expressed CD147 antigen, while the normal liver tissue no clear signs of CD147 expression. Fifteen rabbits were successfully performed tanscatheter hepatic arterial infusion with 131I-anti-CD147-McAb. At 24 hours after the procedure, the rabbits were scanned by SPECT-CT. High radioactivity was discovered in VX2 tumor, and slightly higher radioactivity was showed in the thyroid, liver, heart, gastrointestinal tract, while the radionuclide was gathered little or no in other tissues and organs. At 7 day after procedure, the lesions still appeared slightly higher radioactivity, but other tissues and organs were not show the performance. At 24 hours after procedure,γscintillation counting mean value was up to 15244.2±1159.37 in tumor tissue, and the mean value was 11463.2±823.60 and 5966.4±914.54 respectively at 72 hours and 7 days after the procedure. The radionuclide was absorbed little relatively into Bone, lung, kidney, spleen, heart. The T/NT ratio of the liver was an average of 3.98 at 24 hours after procedure, while the mean was up to 6.35 after 7 days. The T/NT ratios of the remaining major organs was all more than nine, the highest ratio was up to 31.17 in bone tissue.ConclusionRabbit VX2 liver cancer highly expresses CD 147 antigen. Transcatheter hepatic arterial infusion with 131I-anti-CD147-McAb is feasible in a rabbit model. I-anti-CD147-McAb can bind specifically with its antigen in the VX2 tumor, and more 131I-anti-CD147-McAb can be gathered long time within the VX2 tumor with higher T/NT ratio. This results can be a basis to investigate the anti-tumor mechanism of 131I-anti-CD147-McAb and evaluate efficacy and safety of 131I-anti-CD147-McAb. PartⅢ:The Experimental Study on Treating Liver Cancer by Transcatheter Hepatic Arterial Infusion with 131I-anti-CD147-McAbObjectiveTo evaluate the efficacy and safety of treating liver cancer using 131I-anti-CD 147-McAb by transcatheter hepatic arterial infusion (TAI) in a rabbit liver cancer model, which provide the basic proof for the clinical application of 131I-anti-CD147-McAb.Material and methodsForty-five rabbits model with tumor diameter 2cm-3cm were randomly divided into three groups, fifteen rabbits in each group. All of them were performed transcatheter hepatic artery infusion under general anesthesia. Group A:control group, 2-3ml saline was infused into the hepatic artery. Group B:pure 131I solution was infused into the hepatic artery. Group C:131I-anti-CD147-McAb solution was infused into the hepatic artery. About 2ml vein blood was gotten from all rabbits ear vein for detecting liver and kidney function at pre-TAI and post-TAI 1 day,3 days,7 days,14 days,21 days. About 2ml vein blood were used to detect blood and thyroid function of all rabbits at pre-TAI and post-TAI on 7 days,14 days,21 days. The rabbits were scanned by SPECT-CT to monitor radionuclide biodistribution on 1 day,7 days,14 days after procedures in group B and C. The maximum diameter of each tumor was measured by CT for assessment of tumor size change at pre-TAI and post-TAI 7days, 14days,21 days. On 14 days after procedure, five rabbits were randomly selected to be sacrificed in every group. The liver, lung, spleen, kidney, and the metastatic lesions were cut completely for pathological and immunological investigations. The remaining rabbits continued to be fed, and survival rates were measured using the Kaplan-Meier method.ResultsThe TAI and SPECT-CT/CT procedures were successfully performed in all rabbits with the exception of one rabbit. The rabbit in Group A died of diarrhea on day 3 after TAI and it were excluded from analyses. Test results showed that AST and ALT levels tended to increase transiently 1 day after TAI. Mean values of AST and ALT were 3-4 times that before the procedures (P<0.05) 1 day after TAI, and significantly decreased 3 days after TAI. The liver function returned to preoperation levels (P> 0.05) 7 days after TAI. TBIL, BUN and Cr did not change significantly (P> 0.05) in any group. FT3 and FT4 mean values of rabbits in group B and C continued to decline 7 days after TAI, while TSH corresponding increase (P<0.05). WBC mean value of rabbits in group B and C declined slightly after procedures, but the differences were not statistically significant (P> 0.05).SPEC-CT imaging of rabbits shows that most of the radionuclide was gathered in the gastrointestinal tract and thyroid in Group B, but there was less radionuclide in liver lesion. After 131I-anti-CD 147-McAb was infused, radionuclide was concentrated mainly in the VX2 tumor lesions. There was little or no radionuclide imaging in other tissues and organs.14 days after procedures, radionuclide imaging of all rabbits disappeared in group B and C. The VX2 liver tumors increased rapidly after treatment in group A and B; but the tumors gradually reduced in group C. At 14 days after TAI, the number of metastatic lesions of rabbits in group C was significantly fewer than that in group A and B.14 days after TAI, the proportion of tumor necrosis in group C was significantly greater than that in groups A and B (P<0.001).14 days after treatment, TUNEL analysis suggested that more apoptosis bodies was displayed in the residual tumor tissue in group C than that in groups A and B. Immunohistochemistry results showed that MMP-2 and VEGF was highly expressed within the tumor tissue in groups A and B at 14 days after TAI, but the expression in group C compared with group A (P <0.001) and group B (P<0.001) was significantly reduced. MVD quantitative analysis:the MVD number of residual tumor in group C was less than groups A or B (P<0.001) 14 days after procedures.All animals died within 73 days after TAI. Median survival time of the rabbits in groups A, B, C was 22 days,26 days and 54 days respectively. Survival time of the rabbits in group C was significantly prolonged than the other two groups (P<0.001). ConclusionRadioimmunotherapy with 131I-anti-CD 147-McAb by TAI can inhibit the growth, invasion and metastasis of liver cancer, and prolonged the survival of experimental animals. Liver dysfunction is temporal in the procedure with block thyroid to uptake 131I, so it is a safe and effective treatment method for liver cancer. This therapy method is a potential clinical option for the treatment of liver cancer. This study will provide more fundamental basis for the clinical application of 131I-anti-CD147-McAb.
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