Early Diagnosis Of Thyroid Carcinoma By Contrast-enhanced Ultrasound With SHP2 Targeted Nanoparticles Based On The SHP2 Expression Significance

Background: The current incidence of thyroid tumor is increasing and younger,because the mechanisms is unclear,make early diagnosis of thyroid carcinoma with challenge.Search for tumor-specific molecular markers,the development has the tumor targeted contrast agent,is of great significance to early diagnosis of thyroid tumor.SHP-2 is a bona fide oncogene.SHP-2 regulate several cellular processes and contribute to tumorigenesis.More and more researches reported that SHP2 is over-expressed in solid tumors and promotes the malignant biological behavior of tumor.The aim of this study is to establish an imaging model system for the evaluation of thyroid cancer in vivo for the early diagnosis of thyroid cancer.The main research contents are as follows:PART ?: Expression significance of SHP2 and effects on Malignant Biological Behaviors of thyroid cancer cell?Objective?The present study aimed to evaluate SHP2 expression in thyroid cancer cell lines and tissues,and clinical significance and effects on malignant biological behaviors of SHP2 over-expressed in human thyroid cancer.?Methods?SHP2 protein expression was investigated in thyroid cancer tissue,and paired adjacent normal tissue samples were collected..The detected the effect of SHP-2 on cell growth,cloned forming and anti-apoptotic ability.?Results?Increased SHP2 expression was detected n thyroid cancer patient samples and cell lines compared with that of the normal thyroid tissues and cell line(P<0.05).SHP2 expression was significantly correlated with poor tumour differentiation(P<0.05),late TNM stage(P<0.05)and lymph node metastasis(P<0.05),suggesting that SHP2 may represent a potential target for thyroid cancer therapy.After the expression block of SHP2,the ability of cell clones and tumor growth in a mouse xenograft model were significantly inhibited and the apoptotic cells increased obviously.These are shown that over-expression of SHP2 promotes cell proliferation and anti-apoptotic ability.?Conclusion?SHP2 is over-expressed in thyroid cancer cells lines and tissues,and promotes tumor growth and tumor growth in a mouse xenograft model.Part ? Preparation and Characterization of SHP2 targeted ultrasound contrast agent?Objective?To prepare SHP2 ultrasonic molecular probes with PLGA as the shell and liquid fluorocarbon PFP as the kernel targeting thyroid cancer cell,and to evaluate its characteristics.?Methods?SHP2 ultrasonic molecular probes are prepared with PLGA as the shell and liquid fluorocarbon PFP as the kernel by double emulsion solvent evaporation,surface targeting SHP2 antibody,namely SHP2-PLGA nanoparticles(NPs-SHP2).The morphology,structure,particle size,potential and stability were detected by fluorescence microscope and Malvern laser particle size measuring instrument.The emulsion is placed in a temperature-regulating board,and the temperature was set to 60 degrees,and the changes of the emulsion prepared by the optical microscope at high magnification were observed.?Results?Successful build a targeted thyroid cancer SHP2 US molecular probes,its morphological rules,spherical,particle size(765.0 + /-12.5 nm,size,uniform distribution,good dispersibility,no obvious aggregation,concave and convex surface porous structure.After temperature adjustment,the microbubbles with a diameter of about 1um-5um began to appear at 40 degrees.?Conclusion?SHP2 targeted ultrasound nano contrast agent was successfully prepared,and the morphological rules,uniform size and high stability of the contrast agent could promote the liquid-gas phase transition.Therefore,this contrast agent can realize the function of US target imaging and temperature phase transition,and may have the potential of enhanced ultrasound imaging.Part ? Research on the biocompatibility of NPs-SHP2?Objective?To evaluate the biocompatibility of SHP2 targeted contrast agent and provide an experimental basis for its further clinical research and translation.?Methods? The MTS experimental suggests that SHP2 targeted ultrasound contrast agents had no obvious toxicity to the thyroid cells;Countess observed that contrast agents had no effect on cell survival and mortality;LD50 detection targeting contrast agent has a wide range of safety;The expression of caspase-3,BAX and bcl-2 in thyroid cancer cells was detected by western-blot,indicating that the contrast agent had good bio-safety.?Results? The MTS experiment showed that the SHP2 targeted ultrasound contrast agent had no obvious toxic effect on thyroid cells.Countess observed that contrast agents had no effect on cell survival and mortality;LD50 detection targeting contrast agent has a wide range of safety;The expression of caspase-3,BAX and bcl-2 in thyroid cancer cells was detected by western-blot,indicating that the contrast agent had good bio-safety.?Conclusion?SHP2 targeted ultrasound contrast agent has good biological safety,can be further used in the diagnosis of tumor.Part ? Application research of SHP2 targeted ?ultrasound contrast agent in the early diagnosis of thyroid carcinoma?Objective?(1)cells in vitro experiment: The target combination efficiency of the contrast agent was tested.(2)the animal experiment: To explore the feasibility of ultrasonographic early targeting imaging of thyroid carcinoma by ultrasonography in the detection of target contrast agent in nude mice.?Methods? The experiment was divided into two groups: NPs-SHP2,non-targeted group: NPs-control,the specific target-binding ability of NPs-SHP2 was assessed in vitro and in vivo,and the effect on the enhancement of ultrasonic imaging was studied in vivo by LIFU induced.?Results?In vitro target test showed that the NPs-SHP2 group attachment per cell was notably higher than the average number of NPs-Control attached per cell.In human thyroid cancer xenografts in mice,the imaging signal was significantly enhanced in thyroid carcinoma specimens of the SHP2-targeted group by LIFU induced.? Conclusion ? This study provided a basis for preclinical exploration of NPs-SHP2 as a new ultrasound molecular imaging tool that can be applied for clinical thyroid nodule detection to enhance diagnostic accuracy.