Image Monitoring Of The Anti-inflammatory Effects Of Atorvastatin In A Rabbit Model Of Atherosclerosis

Section I:The establishment of a rabbit atherosclerotic model and identification of tolerant dosage of atorvastatinObjective:To establish atherosclerotic model in rabbits by high fat diet plus balloon injury and explore the feasibility of morphology evaluation on AS by a clinical 3T MR system.In addition, identification of the tolerant dosage of atorvastatin for the rabbits was conducted.Methods:40 male New Zealand white rabbits were divided into 2 groups, one was atherosclerotic group by balloon injury plus high fat diet (24 rabbits,Group A),and another one was dosage identification group by atorvastatin and high fat diet (16 rabbits,group B). Weight measurement and blood sample from ear marginal vein for lipid profiles were taken before intitiation of high fat diet in all rabbits,then 1) Group A:before balloon injury to right iliac artery and high fat diet,3T MR scan and ultrasonic scan were made in Group A rabbits. Weight,blood sample and ultrasonic scan were taken at week 4 and 8 respectively after balloon injury, and at week 8,a 2nd MRI was performed.Within 24 hours of MRI completed at week 8, euthanasia was applied to this group to collect left and right iliac artery sample for HE and immunohistological staining. Morphological comparison between HE sample and MR measurement was conducted.The diameters and blood flow velocities of both arteries by ultrasonic scan were compared.2) Group B:16 rabbits were divided into high dosage and low dosage group with 8 equally in each group, 1mg/kg and 2.5mg/kg per day of atorvastatin (Ator) were administrated in low dosage group,each for 4 rabbits.Similarly,5mg/kg and 10mg/kg per day of atorvastatin were administrated in high dosage group. Weight and blood sample for blood lipid and liver function were taken at week 4 and 8. Chow consumption and survival were observed and histological comparisons of livers were made.Results:1. An atherosclerotic model was successfully eastablished in the right iliac artery in Group A rabbits featuring rich lipid deposition and large amount of infiltration of macrophages inside the plaque.The lumen area of the right iliac artery(RIA) was significantly smaller than that of the left iliac artery(LIA), while the thickness and area of the vessel wall were larger in the RIA than the LIA. High fat diet-only didn't lead to a marked atherosclerosis in the LIA.The thickness of RIA wall measured by MR was closely correlated with that of histological measurement.A higher SSI at week 8 than baseline suggested inflammatory response within the vessel wall of RIA. The inner diameter of the RIA was progressively narrowed with time going along and the blood flow velocity was markedly increased at week 8 indicated by ultrasonic scan.2. In group B, all 8 rabbits in the low dosage subgroup were alive at week 8 while only 2 rabbits in the high dosage subgroup (both were fed with 5mg/kg/d ator) were alive.The high dosage of Ator could significantly reduce the elevation of blool lipid,but at the unacceptable expense of live function damage with hepatic necrosis observed by pathological study.Conclusion:1. Atherosclerotic plaque in a rabbit model can be induced with balloon endothelial injury plus high fat diet.Infiltration of large amounts of macrophages was identified inside the plaque,which offered a good platform for evaluation of the vulnerability of the plaques2. The anatomical evaluation of atherosclerotic artery by high resolution 3T MR system was highly correlated with that of pathological assessment.3. Ultrasonic scan might have a good evaluation over the narrowness and hemodynamic change of injurious RIA.4. Rabbits in our study had poor tolerance with high dose atorvastatin (especially with 10mg/kg per day)Section 2:Experimental study of the anti-inflammatory effects of statin in the rabbit atherosclerotic model by USPIO enhanced MRI.Objective:To evaluate the effects of anti-inflammatory effects with atorvastatin therapy by USPIO enhanced MRI in an atherosclerotic rabbit model.Methods:30 male New Zealand white rabbits (weight 2-2.5kg) were randomly divided into 2 groups:Ator group (n= 15,atorvastatin 2.5mg/kg/d) vs control group(n=15).All rabbits underwent balloon endothelial injury in the right iliac artery (RIA)plus high fat diet. Before balloon injury,all rabbits were scanned by a clinical 3T MR system with a 8 channel kneel coil.At week 8, an MR scan before USPIO injection were conducted.24 hours and 72 hours after USPIO injection,a 2nd and 3rd MR scan were conducted.Within 24 hours of 3rd MR completed,an iliac angiography under X ray was performed and then the rabbits were euthanized.Histological,immunohistological and Perl's stain were conducted on the RIA slice.Blood sample for lipid, liver function and CRP was collected at baseline,week 4 and week 8. Parts of the RIA sample were assessed by electronic scanning and transmission microscopy.Results:Both blood lipids as well as CRP were significantly increased in control group compared with those in the Ator Group. MRI indicated there be significant inflammation on RIA with higher T2WI signal intensity and edema around arteries in control group in comparison with Ator group at week 8. The baseline MR scan indicated a smooth inner wall with clear interstitial space in both groups.8 weeks after injury,the SSI of control group after USPIO injection were significantly lower than that in the Ator group which was suggestive of higher inflammation response in the plaque and was supported by the more infiltration of macrophages and positive Perl's stain.The number of vulnerable plaques were significantly higher in the control group than that in Ator group.The luminal area from HE slice was much smaller in the control group while inner/medial area ratio was much higher compared with Ator group.There was no significant differences in the occurrence of severe stenosis of RIA in both groups identified by X ray angiography or MRA.Electronic transmisstion microscopy indicated iron deposition in macrophage inside atherosclerotic plaques of control groupConclusion:1. USPIO was swallowed by the macrophages inside the atherosclerotic plaque and induced the decline of T2WI SSI on MR image, which was helpful to evaluate the stability of the plaque2. X ray angiography and MRA could identify the stenosis of the artery in both groups, however its value on assessement of plaque stability was limited3. Atorvastatin therapy could help stabilize the atherosclerotic plaque,part of whose mechanisms might be its anti-inflammatory effects such as inhibition of macrophagesSectionâ…¢:Preliminary assessment with USPIO enhanced MRI on the protective effects with atorvastatin pretreatment for acute vascular endothelial balloon injury.Objective:To explore the role of USPIO enhanced MR in the evaluation of protective effects with atorvastatin pretreatment for acute vascular injury by balloonMethods: 16 rabbits were randomized into 2 groups:Atrovastatin pretreatment group(n=8,5mg/kg per day) vs control group.Both groups were fed with high fat diet for 2 weeks.After that, both groups underwent balloon endothelial injury of RIA.Before balloon injury,a baseline MR scan was performed for both groups.At the end of the injury surgery, USPIO was injected.24hours and 72 hours after USPIO injection,2nd and 3rd MR scan were conducted. Within 24hours of the 3 rd MR scan completed, an iliac angiography under X ray was performed,after which euthanasia was performed for all rabbits and RIA samples were harvested for histological,immunohistological and Perl's stain.Results:With over 2 weeks of 5mg/kg per day atrovastatin pretreatment, the elevation of blood lipid and CRP was much lower than that of control group. Both groups had an SSI elevation by MRI 24 hours after injury,however the elevation of ator groups was significantly lower than the control group.The SSI went down a bit in both groups in 72 hours after injury, however it continue to be higher in the control group. The immunohistological stain indicated, infiltration of macrophage and positive Perl's reaction in the control group.Conclusion:1. Pretreatment with atorvastatin of 5mg/kg per day for over 2 weeks could relieve the inflammatory reaction caused by balloon endothelial injury2. USPIO-enhanced MR might identify the acute endothelial injury by balloon with infiltration of macrophage and iron presence causing relatively focal decline of T2WI signal intensity.3. Angiography and MRA could not evaluate the protective role by atorvastatin pretreatment on acute vascular injury except for the identification of embolism.