The Expression Of SALL4 In The Primary Liver Cancer And Construction And Application Of Its Nanofluorescent Biosensing Systems

Background:To study the role of human Saplt-like 4 in the occurrence of liver cancer,we developed a new type of high-efficiency aggregation induced emission organic fluorescent dye,and constructed a sensor system,to achieve high sensitivity,high selectivity,non-labeled fluorescent sensing and detection of active molecules.Ultimately,it can better guide early clinical diagnosis and treatment options.Objects and methods:(1)Study population: 40 patients diagnosed with primary liver cancer in recent years were selected from the tissue specimen library of our hospital.(2)Data collection: retrospectively analyzed the clinical data of the enrolled patients,and made record of demographic data(age,gender,etc.),laboratory data(aspartate aminotransferase,alanine aminotransferase,etc.),and pathological data(tumor size,tumor stage,and presence or absence of vascular infiltration,etc.).(3)We performed hematoxylin-eosin staining on the above specimens to reassess the pathology and identify cancerous tissues and adjacent tissues.SALL4 monoclonal antibody was used as the main immunohistochemical staining antibody for staining to determine the expression of SALL4 in primary liver cancer.(4)Relying on the State Key Laboratory of Supramolecular Structure and Materials of Jilin University,we designed and developed new AIE molecules.A molecule with AIE characteristics was used as a light-emitting element to couple with a hydrophobic group and a hydrophilic group with good biocompatibility to prepare a new type of organic polymer with AIE characteristics.Based on the synthetic highefficiency AIE fluorescent dye,SALL4 antibody was selected for modification,combined with an amphiphilic group,as a recognition unit,to complete the detection in the cell.At the same time,we assessed the toxicity of nanoparticles.(5)We performed immunofluorescence staining on the specimens of the aforementioned cases again,collected images under a fluorescence microscope with the same exposure time,calculated the fluorescence intensity,and performed statistical analysis on the results.Result:(1)The SALL4 positive staining classification is defined as: 1)Weak positive: 1-30% of cells are stained,2)Medium positive: 31-50% of cells are stained,3)Strongly positive: 51-80% of cells are stained,4)Very strong positive: >80% of the cells are stained.Immunohistochemical staining showed a significant difference in the expression of SALL4 between liver cancer tissues and adjacent tissues(P <0.05).(2)In immunohistochemical staining,there was no statistical correlation between SALL4 positive grade and age,gender,etiology,alanine aminotransferase level,aspartate aminotransferase level,cholinesterase level,serum albumin level,total bilirubin level,prothrombin time level,platelet level,creatinine value,model for endstage liver disease score,alpha fetoprotein level,maximum tumor diameter,tumor number,T stage,histological differentiation,whether there is a vessel invasion and nerve invasion,and the presence or absence of liver cirrhosis.(3)Based on the amphiphilic polymer DSPE-PEG-NHS and aggregation-induced emission NDSA,nanoparticles were prepared by self-assembly method.The SALL4gene-encoded protein antibody was further modified on the surface to obtain nanoparticles with bright orange fluorescence NDSA@ SALL4.The nanoparticle has strong fluorescence emission in water,the emission peak is located at 559 nm,and the fluorescence quantum yield is 2.89%.(4)Nanoparticles NDSA@SALL4 can not only target Hep G2 liver cancer cells,but also can effectively bind to the protein encoded by the SALL4 gene,enrich in the nucleus,and emit bright orange-yellow fluorescence.(5)After the nanoparticle NDSA@SALL4 is processed by immunofluorescence staining,the targeted liver cancer tissue can be seen under a confocal microscope.In immunofluorescence staining,there were significant differences in the expression of SALL4 between liver cancer tissues and adjacent tissues(P<0.05).(6)There was no statistical correlation between SALL4 expression level and age,gender,etiology,ALT level,AST level,CHE level,ALB level,TBi L level,PT level,PLT level,creatinine value,MELD score,AFP level,maximum tumor diameter,number of tumors,T staging,histological differentiation,vascular invasion and nerve invasion,and the presence or absence of liver cirrhosis.(7)Immunofluorescence staining has better sensitivity and specificity than immunohistochemical staining when judging tissue types.Conclusion:(1)Compared with adjacent tissues,SALL4 is highly expressed in liver cancer tissues.(2)There is no statistical difference between SALL4 expression level and age,gender,etiology,ALT level,AST level,CHE level,ALB level,TBi L level,PT level,PLT level,creatinine value,MELD score,AFP level,tumor maximum diameter,tumor number,T staging,histological differentiation,whether there is vascular invasion,nerve invasion,and whether there is cirrhosis.(3)Nanoparticles NDSA@SALL4 have excellent luminescence characteristics and good biocompatibility.After the surface of the nanoparticles is modified with the SALL4 antibody targeted by liver cancer cells,the fluorescent nanoparticles can specifically target liver cancer cells and liver cancer tissues,and are enriched in the nucleus,providing the possibility for early detection of liver cancer cells.(4)The sensitivity and specificity of nanoparticle NDSA@SALL4 for immunofluorescence staining of primary liver cancer tissue is better than that of immunohistochemical staining.