The Study Of Magnetic Resonance Functional Imaging Of Hepatic Fibrosis Of Rats And MR Tracing Experiment About The Repair Of Hepatic Injury By Transplanting BMSCs Labeled With Magnetic Particles

Chapter I The Experimental Study of Early Diagnosis and Quantitative Analysis of Hepatic Fibrosis of rats with Magnetic Resonance Functional ImagingIn diffuse liver diseases, the diagnosis of hepatic fibrosis and earlier period of liver cirrhosis is the tough problem of early diagnosis of imageology, because the changes of the diseases in morphology are not obvious, and the valuable diagnosis information cannot be provided by the traditional imaging methods which reflect anatomic structure mainly. At present ,it is low that the sensitivity and specificity of the noninvasive diagnostic methods of hepatic fibrosis which are applied clinically;the biopsy of hepatic tissue is thought to be the"gold standard"to diagnose hepatic fibrosis, but it is an invasive method which is limited in clinical application .And the early prediction and intervention is helpful to prevent the development of the process. Thus ,it is urgent to develop new noninvasive methods which can predict hepatic fibrosis early and estimate the degree of the diseases .This study is to study the progression of hepatic fibrosis dynamically and discuss the parameter and the best imaging sequence of magnetic resonance functional imaging by establishing the rat model with liver cirrhosis and applying magnetic functional imaging which include diffusion-weighted imaging,spectrum imaging,perfusion imaging and the application of hepatocyte-specific contrast media); to analyse the appearance of magnetic functional imaging in different degrees and stages of hepatic fibrosis, explore the quantization diadynamic criteria of magnetic functional imaging of hepatic fibrosis and evaluate the value of magnetic functional imaging in the diagnosis of hepatic fibrosis and the early period of liver cirrhosis.Part I The Establishment of Rat Model with hepatic fibrosisObjective:To establish animal models with hepatic fibrosis with SD rats by injecting carbon tetrachloride subcutaneously to provide different stages of rat models with hepatic fibrosis for magnetic resonance functional imaging of hepatic fibrosis and magnetic resonance tracer study of cellular transplantation . Methods:The rats of experimental group received subcutaneous injection of 40% carbon tetrachloride oil solution 3ml/kg twice per week and the initial dose is 5ml/kg , and 10% Alcohol is the only drinking water. The rats of control group received subcutaneous injection of physiologic saline , and the dosage and usage is similar with the experimental group, but the drinking water is purified water. Since 2 weeks after injection, 4 rats of experimental group and a rat of control group were collected randomly per week to perform magnetic resonance functional imaging (DWI/MRS/PWI) of liver; after MR imaging but in 4 hours these rats were killed and HE staining,Masson trichrome stain,reticular fiber staining and transmission electron microscope(TEM) examination of liver were performed and the stages of hepatic fibrosis were assessed with light microscope. Results:94 rats of experimental group were successful and 46 were death. The death rate was 33%.The symptoms of different degree of nutritional disturbance and chronic liver disease presented in all the rats of experimental group . The histopathologic examination of liver showed that there was inflammatory cell infiltration, hepatic cell necrosis , collagen and reticular fibre hyperplasia , sinusoidal capillarization ,et al.The pathology stages of hepatic fibrosis was shown as follow: 28 rats with 0 stage,19 rats with 1 stage,27 rats with 2 stage,25 rats with 3 stage and 15 rats with 4 stage. The rats of control group were survival totally and had no corresponding symptoms. Conclusions:The composite factor (CCL4+Alcohol) could induce rats to generate hepatic fibrosis ,and had the advantages of high achievement ratio and short cycle of establishing models .Models of different pathology stages of hepatic fibrosis could be established to provide ideal experimental model for the study of hepatic fibrosis with the use of the composite factor. Part II The Magnetic Resonance Diffusion-weighted Imaging (DWI) of Hepatic Fibrosis of RatObjective:To discuss the value of DWI in early diagnosis,quantization analysis and staging of hepatic fibrosis by analyzing the signal intensity of DWI,the value of ADC and the change of EADC of different degree of hepatic fibrosis . Methods: During establishing rat model with hepatic fibrosis in part I, 4 rats of experimental group and 1 rat of control group were selected randomly per week to perform MR diffusion-weighted imaging. The sequence was SE-EPI and the gradient factor b were 0s/mm2,300s/mm2,600s/mm2,800s/mm2 and 1000s/mm2 respectively .ADC diagram and EADC diagram were achieved according to different value of b, then the signal intensity of DWI of different b values were determined and the value of ADC and EADC were calculated to compare with pathology stages . Results: (1) The signal intensity of DWI of the rats of experimental group were in the tendency of increasing with the increase of staging of hepatic fibrosis .The signal intensity of DWI was uneven because the development of each lobe was different. (2)When b values were 300,600,800 and 1000 s/mm2 ,SNR were(x—±S)36.30±23.25,28.11±12.48,25.71±11.82 and 15.23±6.54 respectively, and the picture quality was in the tendency of decreasing. When b values were 600 or 800 s/mm2, the picture quality were superior to that when b values were 1000 s/mm2,and P<0.05. (3)The analysis of ADC: the value of ADC of control group,grade 1of hepatic fibrosis,grade 2 of hepatic fibrosis,grade 3 of hepatic fibrosis,grade 4 of hepatic fibrosis were[(x—±S)×10-3(]1.542±0.299)×10-3,(1.334±0.268)×10-3,(1.108±0.198)×10-3,(0.978±0.169)×10-3,(0.680±0.260)×10-3 respectively , and there was a trend of decreasing . There were significant differences between control group and other groups,grade 1 of hepatic fibrosis and grade 2,grade 1 and 3,grade 1and 4,grade 2 and 3 ; but there was no significant difference between grade 2 and 3 . The result of correlation analysis of ADC and staging of hepatic fibrosis was : r=-0.766(p﹤0.001).(4) The analysis of EADC: the value of EADC of control group,grade 1,grade 2,grade 3 and grade 4 of hepatic fibrosis were[(x—±S)×10-3](0.315±0.068)×10-3,(0.345±0.081)×10-3,(0.411±0.074)×10-3,(0.465±0.056)×10-3 and(0.595±0.106)×10-3 respectively , and there was a tendency of increasing . There were significant differences between control group and grade 2,control group and grade 3 control group and grade 4,grade 1 and 2,grade 1 and 3,grade 1 and 4,grade 2 and 4,grade 3 and 4 stage ; but there was no significant difference between control group and grade 1,grade 2,grade 3 . The result of correlation analysis of EADC and staging of hepatic fibrosis was : r=0.753(p﹤0.001). Conclusions:(1)The signal intensity of DWI was increasing with the increase of staging of hepatic fibrosis and it is different that the degree of fibrosis of each lobe. (2) The better b values of DWI of liver were 600s/mm2 or 800s/mm2, because it could avoid the influence of perfusion and had higher SNR. (3) The hepatic fibrosis could be staged with ADC and EADC ,and there was better dependability between them .Part III The Magnetic Resonance Spectrum Imaging (1H-MRS) of Hepatic Fibrosis of RatObjective:To discuss the value of MRS in early diagnosis,quantization analysis and staging of hepatic fibrosis by analyzing the metabolites peaks,peak areas and the change of the ratio between them of different degree of hepatic fibrosis. Methods: During establishing rat model with hepatic fibrosis in part I, 4 rats of experimental group and 1 rat of control group were selected randomly per week to perform MRS with 3D PRESS multi-voxel 1H-MRS sequence. Then different metabolites of spectrogram were hand-marked, the peak height and area were generated automatically with the software. Then the peak height of metabolites and lipid and the peak area ratio(Cho/lip,Glx/lip,Lac/lip,Cr/lip)were calculated respectively to compare with pathology staging . Results: There were 5 main peaks of MRS of liver of control group, and the lipid peak of rats of experimental group was lower than that of control group ,but the peaks of other metabolites were higher. The ratio of the peak height of metabolites and lipid were as follows: (1)the ratio of Cho and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x—±S) 0.052±0.034,0.212±0.225,0.117±0.122,0.403±0.299 and 0.438±0.295 respectively .There were significant differences between control group and grade 3,control group and grade 4 (P<0.05), but there was no significant difference between control group and grade 1,control group and grade 2,grade 1 and other groups,grade 2 and other groups (P>0.05). (2) the ratio of Glx and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x±S) 0.150±0.132,0.406±0.650,0.656±0.551,0.750±0.452 and 0.763±0.517 respectively . There were significant differences between control group and grade 2,grade3,grade 4 (P<0.05), but there was no significant difference between grade 1 and other groups,grade 2 and 3,grade 2 and 4,grade 3 and 4 (P>0.05). (3) the ratio of Lac and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x±S) 0.139±0.128,0.262±0.178,0.251±0.344,0.355±0.446 and 0.233±0.185 respectively ,and there was no significant difference between these groups (P>0.05), but there was a tendency of increasing with the increase of staging. (4) the ratio of Cr and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x—±S) 0.136±0.274,0.767±0.902,0.638±0.960,0.917±0.576 and 0.778±0.856 respectively . There were significant differences between control group and grade 3 (P<0.05), but there was no significant between other groups. The results of correlation analysis between the ratio of the peak height of metabolites and lipid and staging of hepatic fibrosis were as follows : Cho/lip(r=0.503 p﹤0.001),Glx/lip(r=0.388 p﹤0.05),Lac/lip(r=0.124 p﹥0.05),Cr/lip(r=0.235 p﹥0.05). The peak area ratio of main metabolites and lipid of liver were as follows: (1) the ratio of Cho and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis wer(x—±S)e 0.115±0.133,0.257±0.316,0.167±0.187,0.185±0.328 and 0.468±0.372 respectively. There were significant differences between control group and grade 4(P<0.05), but there was no significant difference between other groups (P>0.05). (2) the ratio of Glx and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x±S) 0.045±0.039,0.540±0.318,0.448±0.364,0.482±0.402 and 0.531±0.336 respectively. There were significant differences between control group and other groups (P<0.05), but there was no significant difference between other groups(P>0.05) . (3) the ratio of Lac and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x—±S) 0.062±0.069,0.258±0.266,0.277±0.320,0.170±0.314 and 0.274±0.312 respectively. There was no significant difference between these groups(P>0.05) ,but there was a tendency of increasing with the increase of staging. (4) the ratio of Cr and Lip of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x±S) 0.109±0.231,0.481±0.614,0.704±0.797,0.465±0.525 and 0.810±0.706 respectively. There were significant differences between control group and grade 4 (P<0.05), but there was no significant difference between other groups(P>0.05) . The results of correlation analysis between the peak area ratio of main metabolites and lipid of liver and staging of hepatic fibrosis were as follows : Cho/lip(r=0.282 p﹥0.05),Glx/lip(r=0.313 p﹤0.05),Lac/lip(r=0.135 p﹥0.05),Cr/lip(r=0.267 p﹥0.05). Conclusions:(1)The ratios of peak height of Cho/lip,Glx/lip,Cr/lip were important for the staging of hepatic fibrosis . The correlation between Cho/lip,Glx/lip and staging of hepatic fibrosis were better than others . (2) The peak area ratios of Cho/lip,Glx/lip,Cr/lip were important for the staging of hepatic fibrosis (Cho/lip,Cr/lip is important for grade 4 and Glx/lip for grade1-4). The correlation between Glx/lip and staging of hepatic fibrosis was better than others .(3)The ratio of peak height and peak area ratio of Lac/lip were not important for the staging of hepatic fibrosis , but there was a tendency of increasing with the increase of staging.Part IV The Perfusion Imaging (PWI) of Hepatic Fibrosis of RatObjective:To discuss the value of PWI in early diagnosis,quantization analysis and staging of hepatic fibrosis by analyzing the changes of perfusion parameters of PWI of different degree of hepatic fibrosis . Methods: During establishing rat model with hepatic fibrosis in part I ,,4 rats of experimental group and 1 rat of control group were selected randomly per week to perform MR perfusion imaging with single shot SE-EPI sequence. After bolus injection of Gd-BOPTA (0.2mmlo/kg , 2ml/s) through vena caudalis of rats, 40 dynamic state were scanning continuously to cover the whole liver . The time-signal intensity curve(TIC)was generated automatically with Perfusion software ,then some correlated parameters were calculated as follows: (1) maximal signal reduction ratio (SRRmax);(2)time to peak (TTP);(3)mean transit time(MTT). The perfusion parameters of PWI were analyzed to compare with pathology staging. Results: (1) The time-signal intensity curve(TIC)of liver parenchyma : the curve of control group descended quickly, then recovered slowly after peak value ,and the recovery extent was larger and recovery course was shorter ; the curve of experimental group descended slowly and the extent was smaller , the time to peak was longer , the recovery extent after peak value was smaller and recovery course was longer, and the peak was lenity. And with the development of hepatic fibrosis ,the changes were more obvious.(2)The relationship of the perfusion parameters of liver and the staging of hepatic fibrosis: 1) the values of SRRmax of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were[(x±S)×100%] 0.754±0.073,0.674±0.137,0.632±0.154,0.603±0.201 and 0.535±0.135 respectively . There were significant differences between control group and grade 3,grade 4 (P<0.05) ; but there was no significant difference between other groups(P>0.05). 2) the values of TPP of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were([x—±S)s]14.175±4.845,18.433±7.293,26.789±3.621,31.755±7.308 and 35.213±6.322 respectively . There were significant differences between control group and grade 2,control group and grade 3,control group and grade 4, grade 1 and 2,grade 1 and 3,grade 1 and 4,grade 2 and 4(P<0.05) ; but there was no significant difference between control group and grade 1,grade 2 and 3,grade 3 and 4 (P>0.05). 3) the values of MTT of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were [(x—±S)s] 24.620±5.577,28.945±2.758,32.502±4.268,35.861±4.651 and 35.203±5.674 respectively . There were significant differences between control group and grade 2,control group and grade 3,control group and grade 4,grade 1 and 3,grade 1 and 4 ; but there was no significant difference between control group and grade 1 , grade 1 and 2,grade 2 and 3,grade 2 and 4,grade 3 and 4 (P>0.05). The results of correlation analysis of the perfusion parameters of liver and staging of hepatic fibrosis were : SRRmax (r=-0.439 p﹤0.05),TTP(r=0.798 p﹤0.001),MTT(r=0.647 p﹤0.001) . Conclusions:(1) The time-signal intensity curve(TIC)of liver parenchyma of rats of hepatic fibrosis was characterized by slow- washin and slow-washout, and the peak was low and lenity; (2)The analysis of perfusion parameters of SRRmax,TPP and MTT was helpful to the staging of hepatic fibrosis . The correlation was good between these perfusion parameters and the staging of hepatic fibrosis, especially TTP and MTT . Part V The Magnetic Resonance Imaging of Hepatic Fibrosis of Rat with Hepatocyte-specific Contrast MediaObjective:To discuss the value of the method of delayed - contrast enhanced scanning with Gd-BOPTA in early diagnosis,quantization analysis and staging of hepatic fibrosis by observing rat models with hepatic fibrosis dynamically after the injection of Gd-BOPTA and analyzing the characteristics of dynamic contrast-enhanced MR imaging and enhancement pattern to compare with pathology staging . Methods: During establishing rat model with hepatic fibrosis in part I , ,4 rats of experimental group and 1 rat of control group were selected randomly per week to perform contrast-enhanced MR imaging with hepatocyte-specific contrast media . After bolus injection of Gd-BOPTA (0.2mmlo/kg) through vena caudalis of rats, delayed scans were performed with TSE-T1WI-TRA sequence after 60min,120min and 180min after contrast media were injected . Then the signal intensity and relative enhancement ratio of liver parenchyma of different time were measured respectively ; and the changes of signal intensity of bile duct and blood vessel of different time of different stages of hepatic fibrosis were observed . Results: The relationship of relative enhancement ratio of liver parenchyma in different time and the staging of hepatic fibrosis were as follows : (1)the value of RER1 of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x—±S) 1.436±0.374,1.487±0.477,1.476±0.440,1.489±0.431 and 1.476±0.436 respectively .There was no significant difference between these groups(P>0.05) ,but there was a tendency of increasing with the increase of the staging of hepatic fibrosis .(2) the value of RER2 of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x—±S)1.220±0.370,1.292±0.387,1.344±0.367,1.371±0.388 and 1.405±0.370 respectively .There was no significant difference between these groups(P>0.05) ,but there was a tendency of increasing with the increase of the staging of hepatic fibrosis . (3) the value of RER3 of control group,grade1,grade 2,grade 3 and grade 4 of hepatic fibrosis were(x±S) 0.844±0.275,0.910±0.380,1.041±0.399,1.209±0.299 and 1.241±0.398 respectively. There were significant differences between control group and grade 3,grade 4 (P<0.05) ; but there was no significant difference between other groups(P>0.05). The results of correlation analysis of relative enhancement ratio of liver parenchyma in different time and staging of hepatic fibrosis were : RER1(r=0.039 p﹥0.05),RER2(r=0.174 p﹥0.05),RER3(r=0.420 p﹤0.05) . The appearance of the visualization of bile duct and blood vessel were as follows: the delayed phase of contrast enhanced MRI showed that the tree-like vessels of liver were low signal while the bile duct was high signal in T1WI. The courser of portal vein and bile duct of liver of rats of experimental group were winding and the changes of caliber were not regular, and the bile duct was enhanced in the delayed phase. Conclusions:(1)It was helpless that measuring the relative enhancement ratio of liver parenchyma when 60 min or 120min after injection of Gd-BOPTA to the staging of hepatic fibrosis . It was helpful that measuring the relative enhancement ratio of liver parenchyma when 180 min after injection of Gd-BOPTA to the staging of hepatic fibrosis (especially grade3 and 4), but it was helpless to grade 1 and 2.(2) The morphological changes and enhancement in delayed phase of bile duct of rat model with hepatic fibrosis were a cue of reconstitution of biliary tree and damage of hepatocyte function .Chapter II The Magnetic Resonance Tracing Experiment in vivo about the Repair of Hepatic Injury by Transplanting BMSCs Labeled with Magnetic ParticlesLiver function failure is the main cause of death of various chronic liver diseases. Orthotopic liver transplantation is the most effective method to treat terminal stage of liver diseases at present , but the donor liver are too famine to satisfy clinical requirement . The cellular transplant provides a new therapy strategy for cell reconstitution of the lesion liver and functional recovery of failure liver to make it possible to make use of self-bone marrow stem cell as seed to repair the affection and injury of autologous tissue. The stem cellular transplant tracer study is very important to monitor the disposition,immigration and differentiation of transplanted cells in vivo dynamically and evaluate the effective of cellular transplant ,but this tracer technique is always a puzzle in medical scientific research and traditional tracer technique of transplanted cells must be carry out with histology observation ex vivo . Therefore, it is urgent to establish a transplanted cell tracer technique in vivo which is safe,sensitive,effective and fit to clinic . Close attention was paid to magnetic resonance tracer technique in vivo in the study of stem cell increasingly, and it is a new technique of observing the survival and distribution of transplanted cells in vivo to perform MRI examination after cellular transplant while the cell is labled with contrast media as molecular probe which is sensitive to MR. The MRI tracer technique of transplanted cell which is applied in the study of cardiac muscle and central nervous system is more , but that applied in the study of bone marrow stem cell in hepatic fibrosis is seldom . This study was focused on: (1)Separation,proliferation and identification of BMSCs from rats.(2) Establishing rat model with hepatic fibrosis. (3) Transplanting BMSCs labeled with SPIO and bromodeoxyuridine (BrdU) to treat the rats with hepatic fibrosis . (4) Monitoring the labeled cells after transplantation into the hepatic fibrosis rats with MR scanning dynamically in vivo and contrasting with pathohistology . This study was to discuss the feasibility of the technique of separation,proliferation and identification of BMSCs,MR tracer technique of BMSCs in vivo which was labeled with SPIO as molecular probe,the optimum scanning parameters and imaging sequence,the feature of distribution and immigration of transplanted cells in liver and the rule of the MR signal intensity changes , and to explore the feasibility of the application of 1.5T MR in stem cell tracer study to establish the foundation of clinical application of stem cell transplant MR tracer study in vivo .Part I :In Vitro Culture of BMSCs from SD Rats and label of BMSCs with Brdu and SPIOObjective To develop the in-vitro culture condition of BMSCs from SD rats and to establish the foundation of clinical application of stem cell transplant with MR tracer study in vivo. Methods SD rats (weight 60~90g) were sacrificed. BMSCs were isolated from their bone marrows. Cultured BMSCs were observed daily under phase-contrast microscope and the cell purity were further confirmed by FACS analysis. Results 93.1% of the passage 3 cells were CD90 (one of the BMSCs'surface markers) positive. On the other hand, CD45 + cells were dropped from 71.2% in primary culture to 3.9% in the passage 3. 100% of BMSCs were labeled with SPIO- PLL.Conclusion The passage 3 cells could be used to cell transplantation because of its high purity of BMSCs. It was effective to label BMSCs with SPIO- PLL.PartⅡ:MRI Study of Bone Marrow Stromal Cells of Rats Labeled with Magnetic Particle in VitroObjective:To discuss the changes of MR signal intensity of different numbers of labeled BMSCs and the most sensitive sequences of MR tracing after stromal cells were labeled with magnetic particle to establish the foundation for MR tracing in vivo and provide theoretical evidence . Methods:BMSCs which were labeled with Brdu and SPIO and that which were not labeled were suspended with 1% agarose solutions . The subjects were divided into four groups and put into different EP ducts, including 2×106labeled cells,1.0×106 labeled cells,5×105 labeled cells and 2×106 unlabeled cells .MR scanning sequences included coronal and axial TSE-T1WI,TSE- T2WI and FFE-T2WI.Signal intensity of different sequences of the same object were measured and the MR signal intensity changing rate were compared . Results : The MR signal intensity changing rate of BMSCs which were labeled and not labeled were as follows : (1)the MR signal intensity changing rate of TSE-T1WI of 5×105 labeled cells,1×106 labeled cells and 2×106labeled cells were [(x±S)%] -4.19±0.788,-16.35±1.228,-22.80±1.053 respectively . (2) the MR signal intensity changing rate of TSE-T2WI of 5×105 labeled cells,1×106 labeled cells and 2×106 labeled cells were([x±S)%] -14.15±1.366,-35.09±1.391,-53.02±1.299 respectively . (3)the MR signal intensity changing rate of FFE-T2WI of 5×105 labeled cells,1×106 labeled cells and 2×106 labeled cells were[(x±S)%] -44.98±0.4562,-69.38±0.820,-87.24±0.818 respectively . There were significant differences among labeled cells groups(P<0.001) and significant differences among MRI sequences(P<0.001) .Conclusion : The signal intensity changes of FFE-T2WI sequence is the most apparent when the BMSCs which were labeled with magnetic particle were in the same concentration , and with the number of the labeled cells increasing ,the signal intensity changes were more visible. FFE-T2WI sequence was the ideal sequence for MR tracing of cells labeled with magnetic particle. PartⅢ:The Homogeneous Transplantation of BMSCs of Rat into LiverObjective:To transplant BMSCs into liver with two ways -direct injection and intravenous injection via portal vein . To discuss the feature of the two transplant ways to establish the foundation of comparing the effective of MR tracing and feature of signal intensity changes of them and understanding the rule of distribution and immigration of transplanted cells in the target organ .Methods:20 rat models with hepatic fibrosis which were established in part I of chapter I were selected and divided into four groups , including 1 group :control group (normal sodium which contained no BMSCs were transplanted , and in the group , the number of intra-portal-venous injection and direct intra-liver injection was 2 and 2 respectively); 2 group: BMSCs control group (2×106 BMSCs which were not labeled with Brdu and SPIO were transplanted , and in the group , the number of intra-portal-venous injection and direct intra-liver injection was 2 and 2 respectively ) ; 3 group : labeled cells transplanted via portal vein (2×106 BMSCs which were labeled with Brdu and SPIO were transplanted via portal vein .n=6) ; 4 group : labeled cells transplanted into liver by direct injection (2×106 BMSCs which were labeled with Brdu and SPIO were transplanted into liver by direct injection.n=6).The transplanted cells were taken suction with microinjector and injected into the main portal vein or liver slowly. Results :In the process of transplanted cells into liver by direct injection , the operative procedure was convenient and the cellular transplant was successful . In the process of transplanted cells via portal vein , the transplant operation of 3 rats were difficult because the separation of main portal vein was difficult which was resulted from adherence of abdominal membrane and mesenterium in abdominal cavity , and the operation of other rats were successful . Because the main portal vein of experimental rats was thickening in different degree , the puncturation of main portal vein was successful . After cells or normal saline were transplanted into liver slowly, there was no paradoxical reaction occurred in the rats. The healing of operative incision was well. Conclusion: Transplant stromal cells into liver via portal vein and direct injection into liver were safe and effective. And the choice of transplant way was determined according to clinical or investigative purpose.The difficulty of the separation of main portal vein because of adherence of abdominal contents of subjects should be considered. PartⅣ: MR Tracing of the Homogeneous Transplantation of Bone Marrow Stromal Cells of Rat into Liver in VivoObjective:To discuss the effective of MR tracing and the changes of MR signal intensity of the two transplant ways (BMSCs were transplanted into liver which were labeled with Brdu and SPIO via portal vein and direct injection ); to understand the rule of distribution and immigration of transplanted cells in the target organ to establish the foundation of MR tracing in vivo . Methods:2 rats were selected from every group randomly after transplantation in partⅢ, and performed axial TSE-T2WI-SPAIR,TSE-T1WI and FFE-T2WI when 2hours,3days,7days and 2weeks after transplantation .The developing effect of transplanted cells of different sequences were compared. MR signal intensity,distribution range and rule of change of each group in different time were observed . After MR scanning ,1 rat was selected from erery group randomly and killed to obtain liver specimen,fraction of lung and spleen to perform HE staining,hemosiderin staining and Brdu immunohistochemistry staining .The distribution of hemosiderin-positive cells and Brdu immunohistochemistry-positive cells in liver,lung and spleen should be noticed . Results: (1)MR examination showed as followed :the imaging of FFE-T2WI sequence was the best of all sequences .1 group and 2 group showed no signal changes when 2hours,3days,7days and 2weeks after transplant .3 group showed that there were many hypo-intense lesions in hepatic hilar region when 2 hours after transplant , and with the time going on ,the hypo-intense lesions became smaller . The effect of MR tracing was worst when 2 weeks after transplant. 4 group showed that there were mass-like hypo-intense area in the area of cell injection, and with the time going on , the area was becoming larger , the edge was becoming ambiguous , and when 2 weeks after transplant , the hypo-intense area was still obvious .(2) The histopathologic examination showed as follows : 1) HE staining showed that 2 hours and 3 days after transplant , necrosis,inflammation and fibroplasia of liver of all groups were similar, but when 7days and 2 weeks after transplant , the necrosis and inflammatory cell infiltration of 2-4 group were improving and more obvious than control group . 2) Hemosiderin staining showed that 2hours,3days,7days and 2weeks after transplant ,there were no hemosiderin-positive cells in 1-2 group .The hemosiderin-positive cells of 3group were in the portal vein of porta hepatic 2 hours after transplant , in the small branches of portal vein,sinus hepaticus and around central veins of hepatic lobules 3 days after transplant , and in the liver parenchyma (especially in the area of lesion) 7 days and 2weeks after transplant . The histology change in different time was corresponding with the signal intensity changes of MRI. At the same time, there was a few positive cells in the lung and in the spleen . The hemosiderin-positive cells of 4 group were around the injection point, and with the time going on, the cells immigrated into the liver parenchyma nearby. The histology change in different time was corresponding with the signal intensity change of MRI. At the same time, there was no positive cells in the lung ,and a few in the spleen . 3) Brdu immunohistochemistry staining showed that the result was in coincidence with Hemosiderin staining. Conclusion: (1) FFE-T2WI sequence is the ideal sequence of liver MR tracing of transplanted cells labeled with magnetic particle in vivo. (2)The transplanted cells were scattered in the whole liver (especially in the area of lesion) with the time going on . It was helpful to repair the lesion of liver by transplanting BMSCs .But the time window of MR tracing was shorter when BMSCs were transplanted into liver via portal vein ;It is the ideal way to treat diffuse disease of liver when BMSCs were transplanted into liver via portal vein;but the distribution of transplanted cells were localized and the time window of MR tracing was longer when BMSCs were injected into liver directly. (3) The rule of MR signal intensity changes and the rule of distribution and immigration of transplanted cells in target organ were in good concordance.(4) The rule of MR signal intensity changes reflected the condition of distribution,immigration,proliferation and differentiation of transplanted cells . (5) The...