| ObjectiveOne of the purposes of tumor radiotherapy and chemotherapy is to induce apoptosis of tumor cells and achieve the purpose of treatment.Molecular imaging methods can be used to detect apoptosis at an early stage and evaluate the efficacy.A novel 99mTc labeled caspase-3polypeptide active imaging agent 99mTc-CP3-peptide was constructed.Non-invasive early detection of apoptosis of human lung squamous cell carcinoma H520 cells after radiotherapy in vitro and in vivo was conducted.MethodsThe precursors were labeled with 99mTc by bifunctional chelation method,and the primary products were purified and the radiochemical purity of the products before and after purification was determined.In vitro cell binding assay was performed in human lung squamous cell carcinoma H520 cells treated with radiotherapy?2 Gy or 4 Gy?,and the uptake rate of H520 cells to 99mTc-cp3-peptide was detected 24 h or 48 h after radiotherapy.Cell apoptosis rate was measured by flow cytometry to evaluate the correlation between99mTc-CP3-peptide radioactivity uptake rate and apoptosis rate.Imaging of 99mTc-CP3-peptide was performed in nude mice bearing H520 tumors.24 h after radiotherapy?0,4 and 8 Gy?of tumor-bearing nude mice,99mTc-CP3-peptide was injected through the tail vein of rats and SPECT imaging was performed after 1 h.Delineate the region of interest and measure the radiation ratio between the tumor and normal muscle tissue on the contralateral side.?T/NT?.The uptake rate of 99mTc-CP3-peptide in the tumor and contralateral muscle tissue was measured in vitro,and TUNEL?TdT-mediated dUTP Nick-End Labeling?was performed on the tumor tissue.SPSS 19.0 statistical software was used for statistical analysis.The measured data were expressed as mean plus or minus standard deviation??x±s?.The data were tested for normality and homogeneity of variance.Mean differences between groups were analyzed by one-way ANOVA and two-factor F variance test.Pearson linear correlation was used for correlation analysis.?=0.05 was the test level?both sides?,and P<0.05 was statistically significant.Results1.After purification,there was only one main peak on the HPLC analysis of99mTc-cp3-peptide.The product was a clear colorless solution with a pH value of 6.57.5.and the radiochemical purity at 1,2,4,6 h after the labeling was?99.30±0.38?%,?99.02±0.13?%,?97.33±0.12?%,?97.02±0.26?%,respectively.2.Significant binding of tracer was observed 24 h after radiotherapy for H520 cells.The uptake rates of tracer in the group of 2 Gy and 4 Gy were?0.29±0.01?%and 0.41±0.00)%,significantly higher than of the non-radiotherapy group?0.13±0.00?%,with significant difference?F=110.14,P=0.001?.The uptake rate of cells in doses of 2 Gy and 4 Gy at 48 h after radiotherapy was?0.38±0.02?%and?0.48±0.04?%,respectively,significantly higher than of the non-radiotherapy group?0.15±0.01?%,with significant difference?F=380.26,P=0.001?.The rate of uptake of tracer by cells and flow cytometry to detect apoptosis rate has a good correlation?r=0.96,P=0.002?.3.SPECT imaging showed that radiation concentration of tumor was observed in the radiotherapy group,and no significant radioactive uptake was observed in the non-radiotherapy group.In vitro tumor tissue detection,radiation dose 4 Gy,8 Gy tumor tissue uptake rate were 2.56±0.08%ID/g and 2.98±0.17%ID/g,is significantly higher than the non-radiotherapy group?0.58±0.03%ID/g,F=184.74,P=0.001?.The uptake rate of tumor tissue was positively correlated with the apoptosis rate detected by TUNEL?r=0.95,P=0.001?.ConclusionsThe 99mTc-CP3-peptide was successfully constructed.The radiochemical purity was high and remained stable within 6 h.The uptake of 99mTc-CP3-peptide by human lung squamous cell carcinoma H520 cells after radiotherapy was increased.With the increase of radiation dose,the uptake rate was significantly increased,and was significantly correlated with other apoptosis monitoring results.99mTc-CP3-peptide can detect tumor cell apoptosis noninvasively at early stage. |
PDF Full Text Request