Application Research Of Fluorescence/photoacoustic Imaging Of Gallbladder Cancer Based On Carboxyl/amino Functionalized Fluorescent Polymer Nanoprobe

Gallbladder cancer is a common cancer of the gastrointestinal carcinoid,which leads a threat to human health due to year-on-year morbidity and mortality.Gallbladder cancer has not found its specific tumor markers,early clinical and imaging signs are not obvious,which objectively restricts the early diagnosis of gallbladder cancer,leading to low early diagnosis rate,poor prognosis,high mortality rate.It is necessary to discover new imaging techniques to improve the early diagnosis rate of gallbladder cancer.Fluorescence imaging technology has the advantages of high sensitivity,non-invasiveness and operability,but its penetration depth is not high,while photoacoustic imaging technology has good penetration depth and high spatial resolution.If combine high-sensitivity fluorescence imaging with good penetration depth photoacoustic imaging will greatly increase the sensitivity of the detection and obtain more accurate and detailed imaging information.Conjugated polymer dots are a new class of nanoprobes developed in recent years,it has significant advantages such as low toxicity,good biocompatibility,high brightness,functional design and good stability.It has been used in fluorescence imaging,photoacoustic imaging and magnetic resonance imaging.Nanoparticles as tumor imaging probes are still limited by the distribution of non-characteristic biological tissues,and nanoprobes with high-efficiency tumor cell targeting capabilities should be actively explored.Therefore,we have modified conjugated polymer dots to improve cell internalization and targets and accumulates in the tumor by combining folic acid targeting tumors,long blood cycle time of negatively charged PS-PEG-COOH,and MMAHN₂ with positively charged amino terminal.The study mainly contents are as follows:1.The carboxyl/amino functionalized MEH-PPV-COOH-NH₂（2:1）conjugated polymer dots and the conjugated polymer dots doped with NIR775 near-infrared dye were synthesized by nano-coprecipitation method.Among them,carboxyl/amino functionalized MEH-PPV-COOH-NH₂（2:1）conjugated polymer dots can effectively improve cell internalization and tumor targeting,and conjugated polymer dots doped with NIR775 near-infrared dye has photoacoustic imaging Potential.2.Eight primary gallbladder cancer cells were screened by flow cytometry and laser confocal microscopy,it was proved that folic acid has a targeting effect on gallbladder cancer cells,and screening for QBC-939,NOZ,FRH-0201,and GBC-SD four gallbladder cancer cells with high uptake of folate-functionalized conjugated polymer dots.Subsequently,four gallbladder cancer cells were screened by flow cytometry,and it was demonstrated that carboxyl/amino functionalized MEH-PPV-COOH-NH₂（2:1）polymer dots can achieve higher internalization of cells and the targeting of tumors.The conjugated polymer dots was detected to have low biotoxicity by using a CCK-8 kit.3.Establish a mouse model of subcutaneous tumor of gallbladder carcinoma,and demonstrate in vivo that carboxyl/amino functionalized MEH-PPV-COOH-NH₂（2:1）conjugated polymer dots has good tumor targeting and tumor accumulation by fluorescence imaging and photoacoustic imaging.The same effect was obtained by accumulating in the tumor for a long time ex vivo fluorescence imaging and at the level of histology.Using Probe-Based Confocal Laser Endomicroscopy（pCLE）imaging,it was found that the blood vessels around the tumor were abundant,and the conjugated polymer dots entered the tumor through the blood vessels,and provided evidence for the effective accumulation of conjugated polymer dots to the tumor.Therefore,these work results provide a new way for the development of imaging applications for gallbladder cancer.