Sequential ultrasound molecular imaging for noninvasive identification and assessment of non-alcoholic steatohepatitis in mouse models

Background and objective:Noninvasive non-alcoholic steatohepatitis(NASH)assessment is a clinical challenge to the management of non-alcoholic fatty liver disease.We aim to develop diagnostic models based on sequential ultrasound molecular imaging(USMI)for the noninvasive identification of NASH in mouse models.Methods:Animal experiments were approved by the Animal Ethics Committee of South China Agricultural University.Forty-nine C57BL/6 mice were divided into normal control,non-alcoholic fatty liver,NASH,and hepatitis groups.Sequential USMI was implemented using CD36-targeted microbubbles(MBs-CD36)and intercellular adhesion molecule-1(ICAM-1)-targeted microbubbles(MBs-ICAM-1)to visualize hepatic steatosis and inflammation.The targeting signal of USMI was quantified as the normalized intensity difference(NID)with the destruction-replenishment method.Correlation analysis was conducted between the NID-MBs-CD36 and pathological steatosis score and between the NID-MBsICAM-1 and pathological inflammation score.Finally,diagnostic models combining NID-MBs-CD36 with NID-MBs-ICAM-1 were established for NASH diagnosis.Results:MBs-CD36 and MBs-ICAM-1 were successfully prepared and used for sequential USMI in all mice.NID-MBs-CD36 values increased with the progression of steatosis,while NID-MBs-ICAM-1 values increased in parallel with the progression of inflammation.A strong positive correlation was identified between NID-MBs-CD36 and pathological steatosis grade(r_(s)=0.9078,P<0.0001)and between NIDMBs-ICAM-1 and pathological inflammation grade(r_(s)=0.9071,P<0.0001).Among various sequential USMI-based diagnostic models,the serial testing model showed high diagnostic performance in detecting NASH,with 95%sensitivity,97%specificity,95%positive predictive values,97%negative predictive values,and 96%accuracy.Conclusions:Sequential USMI using MBs-CD36 and MBs-ICAM-1 allows noninvasive grading of hepatic steatosis and inflammation.Sequential USMI-based diagnostic models hold great potential in the noninvasive identification of NASH.