Magnetic Particle Markers Between Bone Marrow Mesenchymal Stem Cells Via Artery Transplantation Therapy Dogs Cerebral Infarction Of The Mr Imaging Studies

Part one: Establishment of cerebral infarction and its MRevaluation in a canine modelSection Ⅰ: Magnetic resonance imaging for assessing a novelembolic stroke resembling lacunar infarction after proximal middlecerebral artery occlusion in a canine modelPurpose: We hypothesized that lacunar infarction could be induced by the proximalMCA occlusion involving the orifices of lenticulostriate arteries in animal models,which had abuntdant distal cerebral collateral anastomosis.This study established aproximal MCA occlusion model using thrombi (1.7mm in diameter and5mm inlength), investigated the MRI characterics of this model and discussed the potentialmechanisms of lacunar infarction.Materials and Methods: The left proximal MCA was embolized with an autologousthrombus in six beagles. Diffusion-weighted imaging (DWI) and T2-weightedimaging (T2WI) were performed every half hour during the first six hours afterocclusion, followed by three time points at12hours,24hours, and one week.Perfusion-weighted imaging (PWI) and magnetic resonance angiography (MRA)were carried out at six hours,24hours and one week. The PWI-DWI mismatch ratiowas defined as (PWI-DWI)/DWI ischemic volume.Results: Solitary or multiple small ischemic lesions resembling lacunar infarctionwere induced by MCA occlusion. They located in the left caudate nucleus, internalcapsule and/or deep white matter, with the diameter of6.38±1.56mm at24hour. Allthe lesions could be detected within two hours by DWI. Lesion volume on DWIincreased in a time dependent manner, from (87.19±67.16) mm~3at one hour up to (368.98±217.05) mm~3at24hours (P=0.009), while that on PWI gradually decreasedfrom (7315±2054.38) mm~3at six hours to (4900.33±1319.71) mm~3at24hours and(3334.33±1195.11) mm~3at one week (P=0.002). The mismatch ratio was41.93±22.75at six hours after ischemia, showing “extensive mismatch”, anddecreased to18.10±13.74at24hours (P=0.002). No MCA recanalization wasobserved within24hours after MCA occlusion.Conclusions: Lacunar infarction could be induced by proximal MCA occlusion incanine model due to abundant collateral anastomosis. It could be detected early byDWI and was characterized by extensive PWI-DWI mismatch. Section Ⅱ:MR evaluation of canine stroke models established bycombination of middle cerebral artery occlusion and ipsilateralinternal carotid artery blockagePurpose: To establish two canine stroke models by combination of MCA occlusionand temporal blockage or consistent occlusion of ipsilateral internal carotid artery andto further compare these two models by MR examination.Materials and Methods: Twelve beagle dogs were enrolled and randomly separatedinto two groups. Group A: left proximal MCA was embolized with autologous clot(1.7mm in diameter,5mm in length), the left ICA was then temporally blocked for2hours using5F vertebral catheter. Group B: left proximal MCA was embolized withautologous clot. The left ICA was also embolized by gelatin sponge. DWI, T2WI andPWI were performed at24hours and one week after embolization. The infarction volumes were measured at24h-DWI,24h-T2WI, and1w-T2WI. rrCBF was assessedfrom24h-and1w-PWI. The neurological scoring at24hours and one week afterembolization were also compared between the two groups.Results: The ischemic infarctions were mainly located in the left basal ganglia andtemporal lobe in group A. The volume measured on24h-DWI and24h-T2WI were2154.00±1034.35mm~3and1417.57±926.83mm~3, respectively. rrCBF was0.61±0.05.One week later, the infarction volume on T2WI decreased to841.58±582.42mm~3while rrCBF increased to0.82±0.11. The neurological scores were6.53±1.58at24hours and4.80±0.98at one week after embolization, respectively. In group B, largecerebral infarctions involving the left MCA territory were observed. The volumemeasured on24h-DWI and24h-T2WI were8928.10±1515.52mm~3and6523.94±1460.33mm~3, obviously larger than those of group A (P <0.01). rrCBFsharply decreased to0.21±0.09, lower than that of group A (P <0.01). Theneurological scores at24hours was8.67±0.52, higher than that of group A (P <0.05).One week after embolization, the infarction volume on T2WI decreased to4059.87±916.49mm~3, but still much larger than that of group A (P <0.01). rrCBFand neurological score recovered to0.56±0.12and7.33±0.58, respectively. All thebeagles survived for one week in group A. But two beagles in group B sacrificedwithin one week.Conclusions: Beagle dogs presented moderate volume of cerebral infarction,moderate decreased rCBF and mild neuro-behavioral disfunction after combination ofleft proximal MCA embolization and temporal blockage of left ICA for two hours.This canine stroke model is more suitable for experimental research than the modelinduced by occluding both proximal MCA and ICA. Part Two: Magnetically labeling of canine bone marrowmesenchymal stem cells and MR imaging in vitroPurpose: To explore the optimal labeling procedure of canine BMSCs withhome-synthesized superparamagnetic iron oxide (SPIO) and to obtain the in vitro MRimages of the labeled BMSCs by3.0T MR.Materials and Methods: BMSCs isolated from canines were incubated with SPIOwith different concentrations (5,10,20,40,80μg/mL, respectively) at37℃5%CO2for24hours. The labeling ratio was assessed by Prussian blue staining andimmunofluorescence tests. Cell viability was evaluated by trypan-blue exclusion andAnnexin-FITC/PI apoptosis. Labeled-cells were imaged by3.0T MRI. MR signalattenuation of labeled BMSCs was monitored continuously after in vitro culture.Results:①When SPIO concentration went up to20μg/mL or above, the labelingratio could reach95%or more;②The apoptosis ratio was8.86±3.56%after BMSCswere labeled with20μg/mL SPIO for24hours, which was not significant differentfrom that of the unlabeled BMSCs;③As few as5×10~4BMSCs could be detected by3.0T MR imaging with T2*WI and SWI sequences. T2*value and BMSCs numberwere inversely correlated (P <0.05);④The labeling ratio and MR signal of BMSCswere attenuated with time after in vitro culture. MR imaging could only track labeledBMSCs efficiently within2weeks.Conclusions: BMSCs could be labeled with home-synthesized SPIO efficiently andsafely. T2*WI and SWI sequences could detected5×10~4BMSCs sensitively. However,the tracking time of SPIO-labeled BMSCs by3.0T MRI in vitro was short. Part Three: In vivo MR imaging of intraarterially deliveredmagnetically labeled bone marrow mesenchymal stem cellsin a canine stroke modelPurpose: This study aimed to evaluate the feasibility of intraarterial (IA) delivery andin vivo MR imaging of superparamagnetic iron oxide (SPIO)-labeled bone marrowmesenchymal stem cells (BMSCs) in a canine stroke model.Materials and Methods: BMSCs harvested from beagles’ bone marrow were labeledwith home-synthesized SPIO. Adult beagle dogs (n=14) were subjected to leftproximal middle cerebral artery (MCA) occlusion by autologous thrombus, followedby two-hour left internal carotid artery (ICA) occlusion with5French vertebralcatheter. One week later,12dogs were classified as three groups before transplantation:group A: complete MCA recanalization, group B: incomplete MCA recanalization,group C: no MCA recanalization.3×106labeled-BMSCs were delivered through leftICA. The left two dogs were distributed to the control group. Series in vivo MRIimages were obtained before cell grafting, one and24hours after transplantation andweekly thereafter until four weeks. MRI findings were compared with histologicalstudies at the time point of24hours and four weeks.Results: Home-synthesized SPIO was useful to label BMSCs without cell viabilitycompromise. BMSCs scattered widely in the left cerebral hemisphere in group A,while fewer grafted cells were observed in group B and no cell was detected in group Cat one hour after transplantation. A larger infarction on the day of cell transplantationwas associated with more grafted cells in the brain. Grafted BMSCs could be trackedeffectively by MRI within four weeks and were found in peri-infarction area byPrussian blue staining. Conclusions: It is feasible of IA BMSCs transplantation in a canine stroke model.Both the ipsilateral MCA condition and infarction volume before transplantation mayaffect the amount of grafted cells in target brain. In vivo MR imaging is useful fortracking IA delivered BMSCs after SPIO labeling.