| Objective To evaluate the diagnostic efficacy in non-alcoholic fatty liver disease(NAFLD)quantitative diagnosis by multi-modality imaging technologies.Methods(1)40 male SD rats were randomly divided into two groups:the control group of 10 rats,fed standard rat feed;fat-fed group of 30 rats,fed with high fat diet.5 rats were randomly selected both in the normal control group who feed in 4 weeks and 10 weeks and in fat-fed group who reared in 4,6,8,10,12 and 16 weeks.Each of the secleted rat was performed ultrasound scan,the Virtual Touch tissue quantification(VTQ)scan,CT and 1HMRS scan.Shear wave velocity(SWV)value,CT value and psoas CT value were meassured and the liver psoas muscle density ratio was calculated.1HMRS scan to obtain intrahepatic contant of lipid(IHCL).After the completion of all imaging,aminotransferase(AST),alanine aminotransferase(ALT),cholesterol(TC)and triglycerides(TG)were examined.Finally,all rats were sacrificed,routine hematoxylin-eosin staining to make a histological diagnosis according to Brunt’s histologic diagnosis criteria.(2)Referring to China Working Group on Obesity(WGOC)recommended "Chinese children and adolescents are overweight,obese BMI value classification standards" and NAFLD treatment guidelines recommended diagnostic criteria in 2006.100 cases of obese children were divided into: non-NAFLD group,the simple fatty liver(NAFL)group and the NASH group,24 normal children were served as the control group.Physical parameter and biochemical tests were measured.General and VTQ liver ultrasound scanning,the degree of fatty liver was semi-quantitative judged and SWV value was measured.SPSS 19.0 statistical applications and Med Calc for Windows,version 15.10 statistical analysis software.Results(1)The method of fat-fed induced of NAFLD rat model is simple,molded high,but there is a long time modeling shortcomings;(2)The accuracy of conventional ultrasound in diagnosis fatty liver degree is poor;(3)The diagnostic performance of VTQ technology,CT and 1H MRS technique in diagnosis severe fatty liver and NASH are all good.The diagnostic efficacy of VTQ technology in diagnosis mild to moderate fatty liver is inferior to CT,the success rate of 1H MRS scan is low and time-consuming;(4)The diagnostic performance of VTQ technology in diagnosis NASH of obese children is good.Conclusions The diagnostic efficacy of VTQ technology and CT in diagnosis both severe fatty liver and NASH are similar;VTQ technology helps to diagnose NASH of obese children in clinical,which has some clinical significance.IntroductionThe liver is the main organ of body to intake,transport,metabolism and excretion lipids,where the function is abnormal in any one part of it can lead to triglyceride accumulation in parenchymal cells of liver.The formation of fatty liver is when hepatocyte steatosis is more than 5% per unit area in the light microscope or the accumulation of fat content in liver is more than 5% to 10%.When the quality and duration of fat accumulation in liver cells exceed,hepatocellular necrosis,hepatocellular ballooning,Mallory bodies,inflammation and fibrosis can be observed which leades to the formation of steatohepatitis hepatitis and liver fibrosis.Nonalcoholic fatty liver disease(NAFLD)including three types which are nonalcoholic simple fatty liver(NAFL),nonalcoholic steatohepatitis(NASH)and nonalcoholic fatty liver fibrosis.Histological examination is the gold standard of staging and grading diagnosis in fatty liver,which has a sampling error and invasive issues.Currently,conventional ultrasound,CT and MRI are the main imaging methods used for diagnosis of fatty liver,but there is a certain degree defect of each.Elastic of tissue(or hardness)is closely related to its biological characteristics,that is,pathological composition and structure of tissue in organ changes leades to the change in elastic properties accordingly.Virtual Touch tissue quantification(VTQ)technology launches short duration,high-energy focused pulsed excitation to thrust tissue by ultrasonic probe,tissue displaced and generate shear waves(SWV),get the SWV of ROI and measure the value of SWV,which could determine the elasticity(hardness)of tissue.Currently,the application of VTQ technology in diagnosis of liver fibrosis has been more positive results,but to reverse liver fibrosis is difficult,which leades to poor efficacy clinical intervention treatment.The histological character in NASH includes hepatic cell necrosis on the basis of steatosis,hepatocyte ballooning,Mallory bodies,inflammatory cell infiltration and fibrosis,liver fibrosis must not be in NASH preparation conditions.Pathology composition and structure of NASH is different from NAFL,the elastic(hardness)of tissue also changes accordingly,which is the theoretical basis for diagnosis liver fibrosis in NASH with or without fibrosis by ultrasound elastography techonology.In summary,conventional ultrasound,VTQ technology,CT and 1H MRS technique are belong to different kind of imaging technologies,and their advantages and limitations exist respectively.The experiments is intended to apply conventional ultrasound,VTQ technology,CT and 1H MRS in studying NAFLD rats,to explore the value of multi-modal imaging techniques for quantify diagnosis of NAFLD.PART Ⅰ A study of establishment fat diet induced rat model with nonalcoholic fatty liver diseaseObjective Establish a high fat diet induced animal model of nonalcoholic fatty liver disease in rat.Methods 40 SD rats,male,50 days of age,weighing 170-180 g,1 week after adaptation feeding were randomly divided into 2 groups: normal control group of 10 rats fed standard rat feed;fat-fed group,30 rats were given high fat diet,both of two groups free access to water.45% fat diet fatty liver animal feed,which raw material composed as follows: rat breeding material,lard,cholesterol,bile salts,sugar,casein and so on;which energy is composed as follows: 45% fat;crude 18% protein,37% carbohydrate.5 rats were randomly selected in the normal control group who fed in 4 weeks and 10 weeks and in fat-fed group who reared in 4 weeks,6,8,10,12 and 16 weeks.Serum aspartate aminotransferase test(AST),alanine aminotransferase(ALT),cholesterol(TC),and triglycerides(TG)and other biochemical were examined in each rat.At the end of the experiment,the animals were sacrificed and the wet weight of liver was weighed to calculate the liver index(HI)= liver wet weight(LW)/ body weight(BW)x100%.Finally,pathological analysis is processed,routine hematoxylin-eosin staining to make a histological diagnosis of liver histologic criteria according to Brunt’s diagnosis.Results:(1)40 rats were involved in the successful completion of the pilot,molding was 100%(40/40);(2)according to Brunt’s histologic diagnostic criteria,classification diagnosis of steatosis level: 11 cases of mild fatty liver group(F1),8 cases of moderate fatty liver group(F2),12 cases of severe fatty liver group(F3);12 rats were consistent with the diagnostic criteria for steatohepatitis,fibrosis is not be observed in all rats;(3)TC,TG,ALT and AST in severe fatty liver group were significantly higher than mild to moderate liver fatty liver group and normal group(P <0.05),consistent with liver enzymes and dyslipidemia characteristics in NAFLD;(4)liver wet weight and HI were significantly higher than mild to moderate liver fatty liver group and normal(P <0.05).Conclusions The modeling method of high-fat induced NAFLD rat model is simple,low cost,low mortality rate,the pathological and pathophysiological changes is in line with changes in characteristics and progression of NAFLD.The model is a kind of ideal model for studying NAFLD,but the shortcomings of the model is modeling for a long time,pending further study of to seek a improved modeling methods to study NAFLD is more desirable.PART Ⅱ Assessment of Nonalcoholic fatty liver disease in Rats using Virtual Touch tissue quantification(VTQ)technologyObjectives The objective of this study is to investigate the diagnostic efficacy of VTQ technology for diagnosis NAFLD in rat.Methods 40 male SD rat were randomly divided into 2 groups: normal control group of 10 rats fed standard feed;fat-fed group,30 rats were given high-fat diet.5 rats were randomly selected both in the normal control group who feed in 4 weeks and 10 weeks and in fat-fed group who reared in 4,6,8,10,12 and 16 weeks.Each of secleted rats was to be performed routine ultrasound and VTQ scan by using Siemens ACUSOA 2000 color Doppler ultrasound,9L4 linear array probe.Conventional ultrasound was observed in rat liver,renal cortex echo intensity,intrahepatic vascular,envelope and display clarity of the diaphragm,according to the NAFLD diagnosis and treatment guidelines to grade the degree of fatty liver.Then,on the basis of two-dimensional image observation,the ROI size of 5 mm × 5mm was placed in the right lobe of the liver location,away from the surface at about 1-1.5cm.SWV value was measured and repeated three times,to calculate the average value of SWV.After the completion of imaging,all rats were sacrificed and the liver tissue was made a histological diagnosis according to Brunt’s diagnosis criteria.SPSS 19.0 statistical applications and Med Calc for Windows,version 15.10 statistical analysis software.Results(1)depending on tne histopathological findings,all rats were divided into 4 groups: 9 cases normal liver group,11 cases mild fatty liver group(F1);8 cases moderate fatty liver group(F2)and 12 cases severe fatty liver group(F3);(2)the diagnostic accuracy of routine liver ultrasound scanning was 55%(22/40),conventional ultrasonic scanning underestimated the degree of fatty liver diagnosis in 45.5%(17/40)rats,overestimated by 2.5%(1/40)of rats the degree of hepatic diagnosis;(3)SWV in normal liver,mild fatty liver group,moderate and severe fatty liver fatty liver group were 1.02±0.12m/s,1.12±0.10m/s,1.21±0.07m/s 1.46±0.16m/s respectively,SWV in severe fatty liver group is higher than that in mild to moderate liver fatty liver group and normal group(P<0.05),the area under the receiver operating characteristic curve(AUROC)for = F3 was 0.993(optimal cut-off value,1.27 m/s;sensitivity,100%;specificity,92.9%;(4)based on liver histology,nine rats was in normal live group,19 rats in line with simple fatty liver diagnostic criteria,12 rats in line with NASH diagnostic criteria but no fibrosis,SWV in normal liver group,simple fatty liver and NASH without fibrosis group were 1.02±0.12m/s,1.16±0.10m/s and 1.46±0.16m/s,SWV values are significant differences among the three groups,the ROC(Az value)for =NASH was 0.993(optimal cut-off value,1.27 m/s;sensitivity,100%;specificity,92.9%;(5)the degree of fatty liver and SWV had a significant positive correlation coefficient of 0.863(P <0.05).Conclusion Application VTQ to detect tissue hardness in NAFLD rat liver had a high performance in diagnosis severe fatty liver and NASH,superior to conventional ultrasound.PART Ⅲ A experiment study of CT evaluation in NAFLD ratObjective To investigate the diagnostic efficacy of CT scan for quantitative diagnosis NAFLD rat.Methods 40 male SD rat were randomly divided into 2 groups: normal control group of 10 rats fed standard feed;fat-fed group,30 rats were given high-fat diet.5 rats were randomly selected both in the normal control group who feed in 4 weeks and 10 weeks and in fat-fed group who reared in 4,6,8,10,12 and 16 weeks to be performed CT scanning.CT scan was used by a Siemens company Somaton definition CT scanning machine.Band pressurized fixed abdomen to inhibit breathing exercise.Scanning parameters: tube voltage 120 k V,tube current of 50 m A,thickness: 2.4mm,scanning time 2.0s,scan range: from the top of the diaphragm to the pelvis.ROI which size is 10mmx10 mm placed in the right lobe of liver to measuring CT value and ROI which size is also 10mmx10 mm placed on right psoas muscle to measurse CT value.Both liver CT value and the right psoas muscle CT value were measured three times repeated,taking the average value of the measured 3 times.Calculation liver-psoas muscle density ratio.After the completion of imaging,all the rats were sacrificed and the liver tissue were stained by conventional hematoxylin-eosin to make a histological diagnosis according to Brunt’s histologic diagnosis criteria.SPSS 19.0 statistical applications and Med Calc for Windows,version 15.10 statistical analysis software.Results(1)Liver CT values were 71 ± 3.3HU,45 ± 11 HU,29 ± 11 HU and 17 ± 9HU in normal liver,mild fatty liver group,moderate and severe fatty liver fatty liver respectively;liver-psoas muscle density ratio,respectively 1.00 ± 0.04,0.67 ± 0.14,0.43 ± 0.17 and 0.26 ± 0.13,the difference among 4 groups was significant(P <0.05);(2)CT values of the liver and liver-psoas muscle density ratio ROC(Az value)in diagnosis ≥mild fatty liver(F1)were 1 and 0.980;≥moderate fatty liver(F2)were 0.965 and 0.956,= severe fatty liver(F3)were 0.949 and 0.943(3)the liver CT values in normal liver group,NAFL group,ASH group were 71±3.3HU,38±14HU and 17±9HU;liver-psoas muscle density ratio was 1.00±0.04,0.57±0.20 and 0.27±0.13;(4)CT values of the liver and liver-psoas muscle density ratio ROC(Az value)in diagnosis NASH were 0.935 and 0.933;(5)through Z test,the difference of SWV,liver CT value and the liver-psoas muscle density ratio ROC(Az value)in diagnosis =F3(severe fatty liver)and NASH were not statistically significant,P> 0.05.Conclusion The diagnostic performance between CT scan and VTQ technology in diagnosis severe fatty liver and NASH is similar,the diagnosis efficacy of CT scan in diagnosis mild to moderate fatty liver is superior to VTQ technology.PART ⅣA experimental Study of 1H MRS evaluation in NAFLD rat Objective To investigate the diagnostic efficacy of 1H MRS for quantitative diagnosis NAFLD in rat.Methods 40 male SD rat were randomly divided into 2 groups: normal group of 10 rats were fed standard feed;30 fat-fed group rats were given high-fat diet.5 rats randomly selected in the normal group fed in 4 weeks and 10 week were performed 1H MRS scanning.Besides,5 rats in fat-fed group reared in 4,6,8,10,12 and 16 weeks were also performed 1H MRS scanning.1H MRS scanning was performed by GE Signa HDx3.0T superconducting MR scanner,8-channel head coil.1H MRS scan sequences include: three-plane positioning MRI scan sequence and liver single voxel point-resolved selective spectroscopy(PRESS)imaging sequences.According to the first locate of the scanned imagel,select proposed spectrum scan on the T2 WI leve.Try to avoid place ROI in the area which include large blood vessels and duct.6 saturated zone was perimeter set around ROI which size was 7mm × 7mm × 7mm.The acquired original data was analyzed by SAGE 7.0 software to determine the area under water and TG peak within ROI.1H MRS postprocessing comprising: loading the data,Spectrum reconstruction,zoom panel,Zero filling,phase correction,obtained the area of water and fat under the peak,according to the formula to obtain intrahepatic contant of lipid(IHCL).At last,all the rats were sacrificed and took routine hematoxylin-eosin staining to make a diagnosis based on Brunt’s liver histological criteria.SPSS 19.0 statistical applications and Med Calc for Windows,version 15.10 statistical analysis software.Results(1)stable lipid peak and peak water lines were obtained in 12 rats,normal liver,mild fatty liver group(F1),moderate-fat group and severe fatty liver group(F3)succeeded to obtain a stable lipid peak and peak water lines was 22.2%(2/9),18.2%(2/11),25%(2/8)and 50%(6/12)respectively;(2)IHCL in normal liver group is 0 and the IHCL is 31.6% ± 10.8% in severe fatty liver group.The ROC(Az value)of IHCL in diagnosis =F3(severe fatty liver)is 0.861,corresponding IHCL cutoff > 20.85 %,the sensitivity was 83.3 % and specificity of 83.3%,suggesting 1H MRS has a high diagnostic performance in the diagnosis of severe fatty liver.Conclusion The diagnostic performance of 1H MRS in diagnosis severe fatty liver(F3)is better,but the 1H MRS scanning is time-consuming,tedious step,post-processing complex and is not a best choice as a diagnostic tool in NAFLD.PART ⅤApplication virtual touch tissues quantification in diagnosis nonalcoholic fatty liver disease in obese childrenObjective To discussion the feasibility of virtual touch tissues quantification(VTQ)technology diagnose of non-alcoholic fatty liver disease(NAFLD)in obese children.Methods 100 cases of obese children,including nothing more than a group of Non-NAFLD(n=43),NAFL group(n=22)and NASH group(n=35).24 healthy ageand gender-matched children served as control group.Physical parameter and biochemical tests were measured.General and VTQ liver ultrasound scanning,the degree of fatty liver was semi-quantitative judged and SWV value was measured.Results SWV in NASH group was significantly higher than the rest three groups,the difference was statistically significant(P <0.05);Corresponding application SWV value diagnose NASH was 1.21 m /s,with a sensitivity of 97.1% and a specificity of 96.6%.Conclusion SWV contribute to the diagnosis and clinical classification of NAFLD in obese children,and help clinicians combined with the history and the biochemical tests to identify NAFL and NASH to improve the diagnostic accuracy of NASH in obese children. |
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