| In this study,we used magnetic resonance diffusion kurtosis imaging(DKI)to observe the evolution and pathophysiology of ischemic penumbra in permanent middle cerebral artery occlusion(MCAO)model and reperfusion MCAO model.At the same time,we observed the correlation between the migration of NMDAR and the evolution of ischemic penumbra,and evaluated the neuronal damage of ischemic penumbra(IP).It aimed to provide imaging and theory evidence for clinical diagnosis and treatment.This study were divided into two parts.Part 1 Using DKI to observing the IP region of acute infarction of ratBackground and purpose We observed the developing of DKI parameters-mean diffusion coefficient(MD),mean kurtosis(MK),axial diffusion coefficient(Da),axial kurtosis(Ka),radial diffusion coefficient(Dr)and radial kurtosis(Kr),also used the corresponding percentage of ischemic region of in DKI parameters-mapping to define the IP region in permanent MCAO model of rat.It aimed to supply experimental evidence for diagnosis of IP region by using DKI.Materials and methods We manufactured MCAO model of rat in 4 groups and scanned apart using DKI at 1h,3h,6h and 24 h after infarction.DKI mappings of each time were divided into 3groups:1.Matching study-MD map and MK map.Lay out MD value of the ischemic area in MD map,MD value of the ischemic area in MK map,MD value of mismatch area between the ischemic area in MD map and MK map,MD value of the ischemic area in core and MD value of relatively normal area around ischemic area of MD map.At the same time,we lay out MK value of the ischemic area in MD map,MK value of the ischemic area in MK map,MK value of mismatch area between the ischemic area in MD map and MK map,MK value of the ischemic area in core and MK value of relatively normal area around ischemic area of MD map.We named them respectively:MD1,MD2,MD3,MD4,MD5,MK1,MK2,MK3,MK4,MK5;2.Matching study-Da map and Ka map.Lay out Da value of the ischemic area in Da map,Da value of the ischemic area in Ka map,Da value of mismatch area between the ischemic area in Da map and Ka map,Da value of the ischemic area in core and Da value of relatively normal area round ischemic area of Da map.At the same time,we lay out Ka value of the ischemic area in Da map,Ka value of the ischemic area in Ka map,Ka value of mismatch area between the ischemic area in Da map and Ka map,Ka value of the ischemic area in core and Ka value of relatively normal area round ischemic area of Da map.We named them respectively:Da1,Da2,Da3,Da4,Da5,Ka1,Ka2,Ka3,Ka4,Ka5;3.Matching study-Dr map and Kr map.Lay out Dr value of the ischemic area in Dr map,Dr value of the ischemic area in Kr map,Dr value of mismatch area between the ischemic area in Dr map and Kr map,Dr value of the ischemic area in core and Dr value of relatively normal area round ischemic area of Dr map.At the same time,we lay out Kr value of the ischemic area in Dr map,Kr value of the ischemic area in Kr map,Kr value of mismatch area between the ischemic area in Dr map and Kr map,Kr value of the ischemic area in core and Kr value of relatively normal area round ischemic area of Dr map.We named them respectively:Dr1,Dr2,Dr3,Dr4,Dr5,Kr1,Kr2,Kr3,Kr4,Kr5.We measure the percentage of ischemic region at different time points on DKI related parameters-graphs and analyze them using one-way ANOVA and LSD-pairwise comparison.After scanning,we sacrifice the rats by cervical dislocation and use the cerebrum to carry on TTC.We choose the maximum cross-section of infarction to calculate the percentage of ischemic areas and analysis the difference with the percentage of ischemic areas on DKI related parameters graphs and on DWI using one-way ANOVA,LSD-pairwise comparison and independent samples t-test.? =0.05,P < 0.05 was considered statistically significant.Results1.The mismatch area between the ischemic area in MD map and MK map,the mismatch area between the ischemic area in Da map and Ka map and the mismatch area between the ischemic area in Dr map and Kr map existed in IP1 h and PI3 h,then they disappeared in IP6 h and PI24 h.2.The value of diffusivity(MD?Da?Dr)decreased with time,and The value of diffusion kurtosis(MK?Ka?Kr)increased over time.MD,MK,Da,Ka,Dr and Kr in the same site at different time points(except relatively normal area round ischemic area)and in different parts of the same time points analyze respectively using one-way ANOVA and the differences were statistically significant difference,P <0.05.3.In groups of PI1 h,PI3h,PI6 h and PI24 h,the percentage of ischemic region in MD map,Da map and Dr map showed no difference with LSD-pairwise comparison.At same time,the percentage of ischemic region in MK map,Ka map and Kr map showed no difference with LSD-pairwise comparison.4.In groups of PI1 h and PI3 h,the percentage of ischemic region in TTC(20.53%(3.21%,25.92%(4.14%)showed statistically significant difference with the percentage of ischemic region in MD map(30.87%±5.19%,34.83%±3.05%),Da map(29.54%±3.08%,33.34%±3.15%)and Dr map(30.16%±3.73%,32.83%±3.27%)by LSD-pairwise comparison,P < 0.05.In groups of PI6 h and PI24 h,the percentage of ischemic region in MD map,MK map,Da map,Ka map,Dr map and Kr map showed no difference with the percentage of ischemic region in TTC byANOVN and LSD-pairwise comparison.5.In group of PI24 h,the percent change of diffusivity-MD(51.4%±2.3%),Da(57.6%±3.0%)and the percent change of diffusion kurtosis-MK(109.4%±4.8%)?Ka(149.8%±5.1%)were significant differences.In groups of PI1 h and PI3 h,the percentage change of diffusion kurtosis(MK,Ka)showed similar changes with diffusivity(MD,Da)in the the mismatch areas between the ischemic areas.Conclusions1.There were no difference in defined IP range using matching study-MD map and MK map,matching study-Da map and Ka map and matching study-Dr map and Kr map,Wherein the Ka value showed the most significant reaction in reflecting the degree of cell damage.2.The mismatch area between the ischemic area in MD map and MK map,the mismatch area between the ischemic area in Da map and Ka map and the mismatch area between the ischemic area in Dr map and Kr map might be IP region.Simultaneously,the percent change of diffusivity and the percent change of diffusion kurtosis in this region were similar,further verified that this area was IP area with cell low dysfunction.Part 2 DKI Observation the evolution mechanism of ischemic penumbra after ischemia-reperfusion in ratBackground and purpose Using DKI-related parameters to observe the evolution of IP region of acute ischemia-reperfusion in rat.At the same time,we analyze the correlation between DKI-related parameters and apoptosis,expression and migration of NMDAR2 A and NMDAR2 B in ischemic region and supply theoretical basis for diagnosis of IP region in acute ischemia-reperfusion by using DKI,and provide experimental evidence for the correlation between the changes of DKI-related parameters and pathological changes.Materials and methods We manufactured MCAO model in rat and unsheathed suture at 2 h after ischemia to reperfusion and divided into 5 groups: IR0 h,IR1h,IR3 h,IR6h and IR24 h.MRI-DKI scanning were carried out in each group at the corresponding time points.Site selection were same as the first part.We use 6 groups Data above-mentioned to analyze respectively the same site at different time points and different parts of the same time points with one-way ANOVA and LSD-pairwise comparison,P < 0.05 was considered statistically significant.After scanning,we removed the brain tissue in rats.Pathological examinations include immunohistochemistry,apoptosis,RT-PCR,Western Blotting which were executed and the got value for one-way ANOVA,LSD-pairwise comparisons and spearman correlation analysis.P < 0.05 was considered statistically significant.Results1.MD,Da and Dr show downward trend during IR0h~ IR6 h of acute ischemia-reperfusion in ischemic region and appear pseudonormalization during IR6 h ~ IR24 h.MK,Ka and Kr show upward trend over time.MD,MK,Da,Ka,Dr and Kr in the same site at different time points(except relatively normal region)and in different parts of the same time points analyze respectively using one-way ANOVA and the differences were statistically significant,P < 0.05.2.In IR0h~ IR6 h groups,the study found that the percent change between MD value and MK value and the percent change between Da value and Ka value were not significant diffrence;then the percent change of Kr value was significantly lower than Dr value that in the mismatch areas between the ischemic areas,LSD-pairwise comparison showed statistically significant difference,P < 0.05.In group of IR24 h,the percent change of diffusion kurtosis(MK?Ka?Kr)(98.52% ± 2.13%,145.63% ±1.19%,64.35% ± 2.19%)obviously higher than the percent change of diffusivity(MD?Ka?Kr)(52.13% ± 1.20%,62.16% ± 2.31%,47.78% ± 1.15%).3.The immunohistochemistry express of NMDAR2 A and apoptosis in ischemic region showed that they were statistically significant difference between each group with one-way ANOVA and LSD-pairwise comparison and showed upward trend over time;the immunohistochemistry expression of NMDAR2 B were statistically significant difference between each group with one-way ANOVA and LSD-pairwise comparison and showed downward trend over time.4.The expression of NMDAR2A-m RNA showed upward trend over time by RT-PCR,RQ values were from 1 of the control group to 4 of IR24 h group.They showed statistically significant difference between each group with one-way ANOVA,P < 0.05.The expression of NMDAR2B-m RNA showed upward trend over time by RT-PCR,RQ values were from 1 of the control group to 7 of IR24 h group.They showed statistically significant difference between each group with one-way ANOVA,P < 0.05.5.The expressions of Western Blotting after ischemia-reperfusion showed NMDAR2 A increased in homogenates(HOM),but it reduced in synaptic fraction(SYN),extrasynapticfraction(EXTRA)and light membrane(LM);Also,the expression of NMDAR2 B reduced in HOM,EXTRA and LM,but it decreased after the first upward trend in SYN.6.Spearman correlation analysis showed MD,Da and Dr appeared positive correlation with the expression of NMDAR2 A in immunohistochemistry,r Was 0.508,0.686 and 0.479;and they simultaneously showed negative correlation with the expression of NMDAR2 B of immunohistochemistry and apoptosis,r were-0.513,-0.657 and-0.449;-0.492,-0.672 and-0.367.MK,Ka and Kr appeared negative correlation with the expression of NMDAR2 A in immunohistochemistry,r were-0.562,-0.640 and-0.466;and they simultaneously showed positive correlation with the expression of NMDAR2 B in immunohistochemistry and apoptosis,r were 0.550?0.698?0.407;0.526?0.646?0.413.Conclusions1.The mismatch area between the ischemic area in MD map and MK map,the mismatch area between the ischemic area in Da map and Ka map and the mismatch area between the ischemic area in Dr map and Kr map existed in IR0h~IR6h and disappeared in IR24 h.Diffusion kurtosis is more accurately to reflect the changes within IR6h~IR24h.2.The expression changes of NMDAR2 A and NMDAR2 B demonstrated migration and redistribution in each sub-structure of the cell.3.MD value,MK value,Da value,Ka value,Dr value and Kr value showed well correlation with apoptosis and the expression of NMDAR2 A and NMDAR2 B.Da value and Ka value showed the most correlation,so they might be better to reflect the changes in the microstructure of cells after infarction... |
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