A Serial Studies On Detecting The Hypoxia And Angiogenesis And Evaluating The Radiosensitivity Of Nasopharyngeal Carcinoma Xenografts In Nude Mice By Magnetic Resonance Diffusion Kurtosis Imaging

Part ?.Research of Detecting the hypoxia and radiosensitivity of human nasopharyngeal carcinoma xenografts in nude mice by magnetic resonance diffusion kurtosis imagingOBJECTIVE:The therapeutic effect of nasopharyngeal carcinoma is closely related to the radiosensitivity of cancer tissue,and tumor hypoxia is considered to be an important factor affecting the radiosensitivity of nasopharyngeal carcinoma.This study was designed to investigate the correlation between DKI parameters and the level of hypoxia-related biomarkers in nasopharyngeal carcinoma xenografts before and after fraction irradiation,and to explore the potential mechanism of DKI in detecting the hypoxia and evaluating the radiosensitivity of nasopharyngeal carcinoma xenografts.MATERIALS AND METHODS:Thirty nude mice were implanted with CNE-1 and CNE-2 nasopharyngeal carcinoma cells,and the xenografts were obtained.Then,the cells were irradiated separately?10Gy per time,30Gy in total,irradiation was carried on every other day?.Each serial nude mice were randomly divided into non-irradiated group?G0?,10Gy group?G1?,20Gy group?G2?,30Gy group?G3?and 3 days after the whole dose irradiation?G4?,and with 6 rats in each group.DKI scan was performed directly without fraction irradiation in G0 group;on the next day after irradiation in G1-G3 groups and on the third day after whole irradiation in G4 group.Each group of nude mice was sacrificed after magnetic resonance scanning was completed.Expressions of hypoxia-inducible factor-1??HIF-1??in the xenografts were detected by PCR,Western-blot and immunohistochemistry.One-way ANOVA is used to verify the differences between multiple parameter variables.The correlation between DKI parameters D and K and HIF-1?was analyzed by Pearson correlation analysis or Spearman rank correlation analysis.Significant level?was be chosen as0.05.RESULTS:There was no significant differences of the volume between the two groups?p=0.296?before fraction irradiation.And the volumes appear to shrink to varying degrees after irradiation,the overall change was statistically significant?p<0.001?.While the differences of xenografts`long diameters between the two cell lines were no statistically significant?p>0.05?.The differences between the D values and the K values were statistically significant?P<0.001?.The overall differences of the expression level of HIF-1?mRNA between the two-line xenografts tissues were statistically significant?p<0.001 and p<0.001?.Immunohistochemistry showed that the overall differences between the scores of HIF-1?expression in CNE-1 were statistical significant?p=0.041?.There was no statistically significant difference between the scores of CNE-2 xenografts?p=0.534?.The correlation study showed that,the change of D value before and after fraction irradiation of CNE-1 xenograft was significantly negatively correlated with the change of tumor volume?r=-0.631,p<0.001?;and significant negatively with that of HIF-1?mRNA?HIF-1?mRNA??r=-0.375,p=0.024?;was not correlated with immunohistochemical expression of HIF-1??r=0.178,p=0.347?.There was a significant positive correlation between the change of K value and the change of tumor volume?r=0.842,p<0.001?,a significant negative correlation with the change of HIF-1?mRNA?r=0.592,p<0.001?,a significant moderate negative correlation with the immunohistochemical expression?r=-0.526,p=0.003?.The change of D value in CNE-2 nasopharyngeal carcinoma xenografts was negatively correlated with the change of tumor volume?r=-0.856,p<0.001?,not significantly correlated with the expression of HIF-1?mRNA?r=-0.098,p=0.570?,but negatively correlated with the expression of mRNA in the first three groups?r=-0.668,p=0.002?,not significantly correlated with the expression of immunohistochemical exam.There was a significant positive correlation between the change of K value and the change of tumor volume?r=0.964,p<0.001?,no correlation with the change of HIF-1?mRNA?r=0.079,p=0.646?,but a significant moderate negative correlation with the expression of mRNA in the last two groups?r=-0.628,p=0.005?,no correlation with the change of immunohistochemical expression?r=-0.078,p=0.683?.Conclusion:The dynamic changes of DKI parameters can effectively identify xenografts with different radiosensitivity,have the potential ability to detect hypoxia and evaluate the radiosensitivity of nasopharyngeal carcinoma xenografts.It may be related to the potential mechanism by which HIF-1?expressed in tumor tissue is involved in regulating the radiosensitivity of tumors.Part ?.Detection of Angiogenesis in Nasopharyngeal Carcinoma Xenografts by Magnetic Resonance Diffusion Kurtosis ImagingOBJECTIVE:The resistance of nasopharyngeal carcinoma to radiation not only relates to tumor hypoxia,but also to the ability of tumor angiogenesis.Previous researches have shown that aquaporin 1?AQP-1?and vascular endothelial growth factor?VEGF?are the main factors that mediate tumor angiogenesis.Our study was to investigate the relationship between DKI parameters and the expression of angiogenesis-related biomarkers in nasopharyngeal carcinoma xenografts in mice during fraction irradiation,and to explore the potential mechanism of DKI in detecting angiogenesis in xenografts.MATERIALS AND METHODS:Construct the model of CNE-2 nasopharyngeal carcinoma xenograft in nude mice,and then fraction irradiation was performed?10Gy per time,30Gy in total,irradiation was carried on every other day?.Nude mice were randomly divided into non-irradiated group?G0?,10Gy group?G1?,20Gy group?G2?,30Gy group?G3?and 3,5 days after the whole dose irradiation?G4,G5?,and with 6rats in each group.DKI scan was performed directly without fraction irradiation in G0group;on the next day after irradiation in G1-G3 groups and on the third and fifth day after whole irradiation in G4 and G5 group.Each group of nude mice was sacrificed after magnetic resonance scanning was completed.Expressions of AQP-1 and VEGF in xenografts were detected by PCR,Western-blot and immunohistochemistry.One-way ANOVA is used to verify the differences between multiple parameter variables.The correlations between DKI parameters D and K and AQP-1 and VEGF were analyzed by Pearson correlation analysis or Spearman rank correlation analysis.Significant level?was be chosen as 0.05.RESULTS:Xenografts`D value increased and K value decreased during the fraction irradiation,and the whole changes of the two parameters was statistically significant?P<0.001 and P<0.001?;expression of AQP1 and VEGF mRNA in xenografts decreased after fraction irradiation,and the difference was statistically significant?p<0.001 and p<0.038?;cell density?CD?decreased and the proportion of tissue necrosis increased after fraction irradiation,and the difference in overall variability was also statistically significant?p<0.001 and p<0.001?.The correlation study showed that,the change of D value during fraction irradiation of xenograft was significantly negatively correlated with the change of tumor volume?r=-0.856,p<0.001?;significantly negatively correlated with the expression of AQP-1 mRNA?r=-0.856,p<0.001?;significantly moderate negatively correlated with the expression of VEGF mRNA?r=-0.504,p=0.002?;significantly highly negatively correlated with cell density?r=-0.918,p<0.001?;significantly high positively correlated with necrosis rate of xenografts?r=0.908,p<0.001?;significantly low negatively correlated with immunohistochemical expression of AQP-1?r=-0.381,p=0.022?and significantly low negatively correlated with immunohistochemical expression of VEGF?r=-0.545,p=0.001?.The change of K value during fraction irradiation of xenograft was significantly highly positively correlated with the change of tumor volume?r=0.964,p<0.001?;significantly high positively correlated with the expression of AQP-1 mRNA?r=0.955,p<0.001?;significantly high positively correlated with the expression of VEGF mRNA?r=0.891,p<0.001?;significantly high positively correlated with cell density?r=0.888,p<0.001?;significantly high positively correlated with necrosis rate of xenografts?r=-0.930,p<0.001?;significantly low positively correlated with immunohistochemical expression of AQP-1?r=0.330,p=0.049?and significantly low positively correlated with immunohistochemical expression of VEGF?r=0.594,p<0.001?.CONCLUSION:DKI parameters D and K have the potential ability to detect the expression of angiogenic related biomarkers AQP-1 and VEGF in nasopharyngeal carcinoma xenografts,and the parameter K appears to be more dominant than the parameter D.The two parameters can be used to evaluate the effect of radiotherapy on xenografts,which may be related to their potential ability to detect changes in AQP-1and VEGF expression in nasopharyngeal carcinoma after fraction irradiation.Part ?.Magnetic resonance diffusion kurtosis imaging in evaluating the radiographic efficacy of human nasopharyngeal carcinoma xenografts in nude miceOBJECTIVE:There are still about 10%-20%of patients with poor prognosis or recurrence and distant metastasis after radiotherapy of nasopharyngeal carcinoma.However conventional MRI cannot find this part of patients in time.If this part of the patients could be detected before the morphological changes,doctors can early change the treatment program to improve the efficacy of this part of patients.In this study,we compared the changes of DKI and DWI parameters before and after fraction irradiation,in order to explore the value of DKI in early detecting radiotherapy of nasopharyngeal carcinoma xenografts.MATERIALS AND METHODS:High and low radiosensitivity?CNE-1 and CNE-2?of nasopharyngeal carcinoma xenografts in nude mice had been constructed and performed fraction irradiation?10Gy,30Gy,irradiated once every other day?.Nude mice were randomly divided into non-irradiated group?G0?,10Gy group?G1?,20Gy group?G2?,30Gy group?G3?and the 3rd and 5th day after the whole dose irradiation?G4,G5?,each group have 6 rats.DKI scan was performed directly without fraction irradiation in G0 group;on the next day after irradiation in G1-G3 groups and on the third and fifth day after whole irradiation in G4 and G5 group.Independent sample t test was used to compare the two samples;Pearson correlation analysis was used to analyze the correlation between the samples.ROC curve was used to analyze the value of each parameter in predicting the value of radiographic efficacy of xenografts.Diagnostic accuracy of DWI and DKI parameters were determined according to AUC?Area Under the Curve?.Significant level?was be chosen as 0.05.RESULTS:The volume of the two-line xenografts showed a decreasing trend after fraction irradiation,and the overall changes were statistically significant?p<0.001and p<0.001?.The D value and ADC value of the two-line xenografts showed an increasing trend after irradiation,and there was no significant difference between the two values before irradiation?G0??P>0.05?and the G3 group also showed no differences?P>0.05?.The differences of D and ADC values in other time points were statistically significant.D value and ADC value of CNE-2 system were significantly higher than CNE-1?p<0.019?.The K value of the two-line xenografts were statistically different?p<0.009?,and K values of CNE-2 were higher than CNE-1before irradiation and dereased rapidly than CNE-1 and became lower than CNE-1after the second irradiation?G2?.There were statistically significant differences in the image parameter changes?D,?K and?ADC of the two-line xenografts,and the change of CNE-2 was greater than that of CNE-1.ROC curve analysis showed that D_G1`s AUC with the value of 0.972 was the highest,followed by 0.944 of?K2,which was higher than 0.861 of ADC_G2,which was the highest AUC in ADC value.D_G1 had a sensitivity of 100%and a specificity of 83.3%when the diagnostic threshold was1.04×10^-3mm²/s.Conclusion:DKI and DWI parameters have the application value in early evaluation of radiographic effect of xenografts,and D value is better than ADC value.