Using ^99mTc-HYNIC-NGR Targeted Aminopeptidase N For SPECT Imaging To Evaluate The Preliminary Application Value Of Liver Fibrosis In Rats

Objective: 1.In vitro through human liver biopsy immunohistochemical staining method,research in the APN,explore the relationship between APN and liver fibrosis stage,preliminary validation on human liver biopsy in APN as a target for the possibility of molecular imaging.2.By pulling the APN immunohistochemical staining results with the results of immunohistochemical staining of ?-SMA compares,explore the pathology of liver fibrosis gold standard of ?-SMA and APN,possible mechanisms for APN in liver fibrosis in theory.3.A preliminary study of 99mTC-HYNIC-NGR in animals of the imaging results of in vitro and in vivo biological distribution,preliminary studies in biological metabolism in the body,the best scanning imaging conditions and parameters of 99mTC-HYNIC-NGR for providing theoretical basis for the follow-up related research,as well as at the same time express APN targeted imaging and ?v?3,even is targeted to provide basis for treatment.4.By using 99mTC-HYNIC-NGR targeted APN of normal rats and liver fibrosis model group of SPECT imaging in rats,the 99mTC-HYNIC-NGR in normal rats in vivo distribution and drug metabolism,and then compare normal rats and rat liver fibrosis model group differences,molecular imaging evaluation of its preliminary application in rat liver fibrosis.Methods: 1.The use of a liver biopsy pathology biopsy(provided by the department of pathology,the China medical university),according to the pathology,the first by the diagnosis of liver fibrosis,which can be divided into two groups,respectively in the early group(S1 – S2)of liver fibrosis and early cirrhosis group,use of APN antibodies and ?-SMA resistance as a resistance,through the Leica Bond-Max fully automatic mechanism for immunohistochemical staining,the liver tissue of the immunized group antigen after repair,phosphate buffer solution(PBS),instead of one as an alternative test.Application of the Image Pro Plus for early liver fibrosis and cirrhosis group average optical density of the dyeing results of analysis,research APN expression level changes in the progress of liver fibrosis and relations with ?-SMA.The stage of liver fibrosis was determined by HE staining.2.The rat liver fibrosis model group,through the use of thioacetamide(TAA)intraperitoneal injections modeling,simulation of hepatitis to liver fibrosis models,injected with normal saline control group is used for abdominal injection.Wistar rats with similar age and weight were selected.3.By adjusting the 99mTC-HYNIC-NGR process,different synthetic drug concentration,such as oxalic acid ethylenediamine(EDDA)using 2-25 mg/ml,respectively,three hydroxyl methyl glycine(tricine)respectively using 5 to 30 mg/ml,Sn Cl2 respectively using 5-25 mg/ml,2-10 m Ci Na99 mTc O4 mixed with water bath for 10 min,it is concluded that the best concentration of drug synthesis,synthetic drugs paper by thin layer chromatography(TLC)determine its radioactive chemical purity,finally choose the best condition of the chemical reaction.4.After editing probe,scan before using 10% chloral hydrate anesthesia,through the rat caudal vein injection,respectively to the normal group and liver fibrosis in rats rats for continuous dynamic scan,dynamic scanning image of SPECT for Wistar rats,respectively in main organs of rats,such as heart,liver,kidney,interested area(ROI)sketch,draw 99mTC-HYNIC-NGR in 30 min in distribution images of animals,.5.Using the normal control group and the image of Liver fibrosis model group,normal group and Liver fibrosis in rats of rat Liver and heart of radioactive count ratio(Liverto-heart ratios,L/H),the component data after statistical analysis,this paper compares and analyzes the application the Mann-Whitney U test to compare each time difference between two groups,the evaluation of rat Liver between the two groups of 99mTCHYNIC-NGR intake differences.Results: 1.The immunohistochemical staining results of in vitro experiments showed that APN did increase with the progression of liver fibrosis,and the expression of APN was positively correlated with hepatic fibrosis.2.Synthetic solution under different conditions are verified through the TLC,and it is concluded that 99mTC-HYNIC-NGR optimum synthesis conditions of each drug concentration,respectively,EDDA(5 mg/ml),tricine(10 mg/ml),Sn Cl2(15 mg/ml),use 2m Ci Na99 mTc O4,synthetic drug radioactive chemical purity of 95%.3.3.99mTC-HYNIC-NGR animals distribution is given priority to with renal metabolism,heart,liver and other organs is to absorb,and the metabolism of the urinary system can be quickly,in the heart of intake of stability,low intake of liver intake has obvious uptake curve,helps the follow-up clinical application.4.SPECT images showed that the time points L/H of the liver damage group images were higher than that of the normal group,and there was a statistical difference between the two groups of L/H in 2,5min(P < 0.05).Conclusion: The APN in the liver fibrosis lobular area of the main expression in liver cell membrane,and expressed in portal area more new capillaries,its in liver fibrosis,the expression of S1 and S2 phase positive rate is lower than the early liver cirrhosis group;APN had no correlation with the expression of alpha-sma.99mTC-HYNIC-NGR animal is mainly composed of renal metabolism,which can be applied to SPECT molecular imaging,and it is established in the laboratory to synthesize the ideal conditions and quality control scheme of 99mTC-HYNIC-NGR.The SPECT imaging of 99mTC-HYNIC-NGR targeted APN has the basic conditions and application potential for evaluating liver fibrosis in rats,and corresponding to 2-5min in scanning.