Fundamental Research On The Application Of Radionuclides And Fluorescein Sodium Mediated Intraoperative Navigation Imaging

ObjectiveRadical resection is the most effective method for the treatment of malignant tumors,but conventional imaging methods are currently unable to meet the needs of clinical navigation.As a new optical imaging method,Cerenkov luminescence imaging?CLI?has a good potential for clinical navigation.However,the weak signal intensity of Cerenkov luminescence?CL?and the long exposure time seriously restrict its further clinical application.The enhanced CL signal strength methods such as cerenkov radiation energy transfer?CRET?imaging and radiopharmaceuticals excited fluorescence imaging?REFI?are not clinically applicable due to the potential toxicity of the fluorescent medium.Therefore,this study selected the medical contrast agent fluorescein sodium as a fluorescent medium to enhance the CL signal intensity by CRET imaging and explore its CRET characteristics in vitro and in vivo experiments.On this basis,a new method of intraoperative navigation of CRET combined with confocal laser endomicroscopy?CLE?imaging was further proposed and its feasibility was verified in a nude mouse model of orthotopic liver tumor.Explore a new way of intraoperative navigation that can be applied clinically.Method1.Different concentrations of fluorescein sodium solution were mixed with three groups of ¹⁸F-FDG of 10?Ci,50?Ci and 100?Ci respectively,and CRET imaging was performed to analyze the relationship between signal intensity and the concentration of fluorescein sodium.2.The fixed concentration of fluorescein sodium and different activities of¹⁸F-FDG were mixed with CRET imaging,and the relationship between the intensity of the signal and the activity of ¹⁸F-FDG was analyzed.3.The optimal concentration of fluorescein sodium and ¹⁸F-FGD were mixed for spectral analysis.Exploring the emission spectra of CRET at the optimal concentration of sodium fluorescein.The spectral analysis of different concentrations of fluorescein sodium and¹⁸F-FDG was carried out to explore the emission spectra of CRET imaging of fluorescein sodium at different concentrations.4.The blackboard was placed between fluorescein sodium and ¹⁸F-FDG to study the mechanism of ¹⁸F-FDG-induced fluorescein sodium.The fluorescein sodium and¹⁸F-FDG were imaged at different separation distances to study the effects of separation distance on the excitation effect influences.5.In the 2mm deep phantom,a mixture of fluorescein sodium and¹⁸F-FDG or ¹⁸F-FDG of the same activity is added.CLI and CRET imaging are performed in full band and 540nm filter mode,and the difference between signal intensity and signal to noise ratio is studied.CLI and CRET imaging was performed in different depth phantoms with a mixture of fluorescein sodium and¹⁸F-FDG mixture or¹⁸F-FDG of the same activity.The penetration depth of CLI and CRET was compared.6.A pseudo tumor was constructed in the left side of nude mice after mixing¹⁸F-FDG with matrix glue.The same activity ¹⁸F-FDG plus fluorescein sodium and matrix glue were used to construct pseudotumor in the right side of nude mice.CLI and CRET imaging of both sides of the tumor were performed under the full band and 540nm filter mode,and the difference of signal intensity was studied.7.Three groups of 4T1 subcutaneous tumor bearing mice were injected with the same activity ¹⁸F-FDG after CLI and PET imaging,then injected 10%fluorescein sodium with different doses?0.1ml/kg,0.5ml/kg and1.0ml/kg?,then CRET imaging and fluorescence imaging were performed.The optimal dose of fluorescein sodium in body CRET imaging was investigated.Three groups of4T1 subcutaneous tumor bearing mice were injected into the same activity 18F-FDG for continuous observation and imaging.During this process,the best dose of fluorescein sodium was injected into group 1 and group 2 at 65min and 115min respectively,and group 3 as a control group without injection of fluorescein sodium.To explore the duration of sodium fluorescein in body CRET imaging.8.PET imaging and CLI imaging after laparotomy were performed after the injection of¹¹C-CHO into the orthotopic tumor model of nude mice.The best dose of fluorescein sodium was injected,and CRET imaging and fluorescence imaging were performed.According to the location of the tumor by CRET imaging,CLE imaging is used to accurately detect the tumor boundary and to guide the resection of the liver tumor.The tumor and the tumor bearing mice after the operation were then CRET imaging,and all the organs of the tumor bearing mice were dissected and CRET imaging were performed.Results1.When the concentration of sodium fluorescein is 0.05%,the intensity of CRET imaging signal is the strongest.The CRET signal is 5 times the intensity of the CL signal generated by the same activity ¹⁸F-FDG.2.The intensity of CRET imaging signal is linear with the activity of¹⁸F-FDG.3.The 0.05%concentration of fluorescein sodium could redshift the emission peak of CL to 540nm,and significantly enhance the signal intensity.The higher the concentration of fluorescein sodium,the more obvious the red shift of the emission peak of CL during CRET imaging.When the concentration of fluorescein sodium is 1%,the emission peak is near 580nm,but its optical signal intensity is obviously lower than that of 0.05%fluorescein sodium at 540nm emission peak.4.The black disk barrier Cherenkov light can significantly affect the signal strength of the signal intensity of fluorescein sodium.The signal intensity of sodium fluorescein is inversely proportional to the excitation distance of the radionuclide.5.In the phantom experiment,the signal intensity of CRET imaging in the mode of full band and 540nm filter is obviously higher than that of CLI,and the signal to noise ratio is obviously higher than that of fluorescence imaging.CRET imaging can increase the penetration depth of the CLI by 7mm.6.The signal intensity of PET imaging on bilateral pseudotumor in nude mice is basically the same.The optical signal of right pseudotumor on the left side is significantly higher than that on the left side,which is3.8 times that of the left side pseudotumor.7.In body application,When the injection amount is 0.5ml/kg of 10%fluorescein sodium,the enhancement of CL signal to the tumor site is the strongest,and the signal intensity is enhanced by 4.7 times.The best dose of fluorescein sodium can immediately increase the CL signal of the tumor site by3-5 times and continue to be 30-50min after the application of the fluorescein sodium.8.In the surgical navigation experiment of liver tumor in the tumor bearing mice,preoperative PET could detect tumors in the right lobe of the liver.The CLI and CRET after open surgery could clearly identify the size and location of the tumor,and the signal intensity and signal-to-noise ratio of the tumor sites were higher than those of CLI when CRET imaging.On the basis of CRET localization of the tumor,CLE accurately detected the tumor margin and guided the resection of the liver cancer.The postoperative CRET imaging proved to be a radical resection of the liver cancer.Conclusions1.Using experiments such as optimum concentration experiment,emission spectrum experiment,and imitative experiment,it was proved that sodium fluorescein is a non-toxic and effective CRET fluorescent medium,which can increase the CL signal of radionuclides by 5 times and redshift the emission peak band to 540 nm.Nearby,penetration depth increased by 7mm.2.The feasibility of radionuclide and sodium fluorescein CRET imaging has been validated for the first time.The imaging method can increase the CL signal intensity at the tumor site by 4.7-fold and last for 30-50 minutes,which has a better tumor detection effect than CLI.3.The radionuclide and sodium fluorescein-mediated PET/CRET/CLE multimode imaging can achieve accurate detection and boundary recognition of tumors.It is expected to become a new method for tumor detection and intraoperative navigation,and has a good prospect of clinical application of transformation.