缺血性腦卒中微血管網(wǎng)絡(luò)重塑的同步輻射三維成像實(shí)驗(yàn)研究
發(fā)布時(shí)間：2019-05-22 07:40
【摘要】：目的 腦生理功能的維持依賴于微血管網(wǎng)絡(luò)結(jié)構(gòu)的完整。缺血性腦卒中后,機(jī)體啟動(dòng)瀑布式級(jí)聯(lián)反應(yīng)致微血管網(wǎng)絡(luò)穩(wěn)態(tài)失調(diào),神經(jīng)血管單元發(fā)生結(jié)構(gòu)與功能重塑。腦缺血可引發(fā)自體血管新生,然而其發(fā)生機(jī)制、發(fā)展進(jìn)程以及具體形式等目前尚不明確。傳統(tǒng)的血管可視化研究手段如組織病理學(xué)方法等,對(duì)微血管空間結(jié)構(gòu)的展示尚停留在二維平面層次,無(wú)法精確展示血管網(wǎng)絡(luò)的時(shí)空分布。常規(guī)影像學(xué)手段如MRA、 CTA、DSA等,由于有限的時(shí)間空間分辨率,尚無(wú)法探測(cè)直徑在50μm以下的微小血管。新興的同步輻射技術(shù)突破傳統(tǒng)光學(xué)成像方法的衍射極限,憑借亞微米至微米量級(jí)的極高分辨率能夠清晰實(shí)現(xiàn)對(duì)物質(zhì)顯微結(jié)構(gòu)的無(wú)損、三維成像。本研究應(yīng)用上海光源同步輻射相位襯度成像進(jìn)行正常和缺血性腦卒中模型的血管成像研究,明確腦微血管高分辨、三維成像的可行性和有效性,探索腦缺血后血管網(wǎng)絡(luò)重塑的動(dòng)態(tài)發(fā)展過程,力求以新的視角為缺血性卒中血管損傷與修復(fù)的研究提供豐富的3D影像學(xué)信息和形態(tài)計(jì)量學(xué)依據(jù)。 方法 調(diào)整成像參數(shù),使用濾波反算法及成像軟件多角度多平面重建正常大鼠全腦血管3D視圖。在此基礎(chǔ)上,對(duì)血管網(wǎng)絡(luò)進(jìn)行基于結(jié)構(gòu)信息學(xué)的量化分析。同時(shí)體式顯微鏡、HE染色檢測(cè)的腦組織樣品進(jìn)行比較。隨后圍繞大鼠缺血性腦卒中模型,利用同步輻射相位襯度成像觀察缺血后不同時(shí)點(diǎn)自發(fā)性血管新生動(dòng)態(tài)發(fā)展過程,探索定量分析血管新生的方法,同時(shí)結(jié)合免疫組織化學(xué)技術(shù),從3D/2D可視化角度綜合評(píng)估缺血性腦卒中的血管網(wǎng)絡(luò)重塑進(jìn)程。 結(jié)果 1.利用同步輻射相位襯度成像建立了從投影圖、二維slice到全腦血管網(wǎng)絡(luò)3D圖像的完整可視化體系。使用5.9μmCCD探測(cè)器,可分辨的最小血管直徑約為9.4μm,接近毛細(xì)血管水平。二維平片與HE染色比較,無(wú)需造影劑和繁瑣切片制備程序,即可清晰顯示海馬區(qū)域的血管走行。腦體表面的3D渲染圖像和Microfil血管鑄型劑灌注后體式顯微鏡原位攝片結(jié)果高度一致,軟腦膜的多級(jí)血管吻合支均能清楚辨析。 2.進(jìn)一步建立基于空間維度變換的全腦水平位、冠狀位、矢狀位虛擬切片和數(shù)字化3D腦血管斷層視圖,并對(duì)其功能區(qū)的血供配布進(jìn)行解析,從2D、3D視角構(gòu)建出一系列數(shù)字化的腦血管3D解剖圖集。 3.血管網(wǎng)絡(luò)骨骼提取結(jié)果顯示,大鼠全腦血管網(wǎng)絡(luò)主要是在20μm以下的微小血管骨架基礎(chǔ)上構(gòu)建而成的�；�3D網(wǎng)絡(luò)的形態(tài)計(jì)量學(xué)分析,進(jìn)一步明確了正常大鼠腦血管容積、血管密度、血管分支數(shù)、節(jié)點(diǎn)數(shù)、相鄰血管間距、直徑分布頻數(shù)等各項(xiàng)生理參考指標(biāo)。 4.此后圍繞大鼠缺血性腦卒中模型的一系列同步輻射光源2D、3D成像研究結(jié)果顯示：缺血后2小時(shí)、4小時(shí)、6小時(shí)微血管數(shù)目持續(xù)增高,密集分布在缺血區(qū)周緣,由缺血半暗帶向梗死區(qū)延伸包繞。然而隨著缺血時(shí)間的延長(zhǎng),缺血1天、3天后新生血管的分布范圍和密度均有所減小,微血管形態(tài)扭曲,形成不規(guī)則血管襻。缺血7天梗死灶液化成蜂窩狀囊腔,膠質(zhì)瘢痕已逐漸形成,缺血18天病灶側(cè)腦體積明顯減小,腦室受到擠壓,皮質(zhì)和紋狀體梗死灶發(fā)生機(jī)化,形成高密度的瘢痕組織和無(wú)實(shí)質(zhì)組織的“中風(fēng)囊腔”。 5.定量分析研究顯示：隨著缺血時(shí)間的持續(xù)增加,病灶側(cè)腦體積自3天后逐漸減小,其中缺血后18天病灶側(cè)腦體積較對(duì)照組顯著縮小(P0.05)。缺血后2小時(shí)、4小時(shí)、6小時(shí)血管密度均較對(duì)照組升高,差異有統(tǒng)計(jì)學(xué)意義(P0.05),而缺血后7天、18天血管密度逐漸降低(P0.05)。缺血后2小時(shí)、4小時(shí)、6小時(shí)血管分支數(shù)、節(jié)點(diǎn)數(shù)目較對(duì)照組顯著升高(P0.05),隨著時(shí)間的推移,缺血后3天、7天、18天血管分支、節(jié)點(diǎn)數(shù)目逐漸降低(P0.05)。缺血后2小時(shí)、4小時(shí)、6小時(shí)10-20μm區(qū)段的微小血管和對(duì)照組相比,頻數(shù)明顯增多,差異具有統(tǒng)計(jì)學(xué)意義(P0.05)。隨著時(shí)間的推移,頻數(shù)逐漸下降,缺血7天、18天頻數(shù)顯著減少(P0.05)。缺血后2小時(shí)、4小時(shí)、6小時(shí)、1天、3天的血管迂曲度明顯增高(P0.05)。 6.CD31免疫組化結(jié)果缺血后2小時(shí)、4小時(shí)、6小時(shí)紋狀體CD31陽(yáng)性染色微血管持續(xù)增多,密集排布缺血邊緣區(qū)域,血管呈點(diǎn)狀、彎曲短狀或圓管狀,形態(tài)不一。隨著時(shí)間推移,缺血1天、3天、7天,新生微血管數(shù)目逐漸減少。以上微血管的變化趨勢(shì)與同步輻射成像結(jié)果一致。 結(jié)論 1.基于同步輻射的腦微血管可視化體系的建立,可多平面多角度解析全腦和局部靶區(qū)域的3D血供配布情況,并實(shí)現(xiàn)血管網(wǎng)絡(luò)空間結(jié)構(gòu)參數(shù)的量化分析。 2.缺血性損傷導(dǎo)致微血管網(wǎng)絡(luò)發(fā)生結(jié)構(gòu)與功能重塑,血管新生在缺血后極早期即可啟動(dòng),并迅速達(dá)到高峰,新生血管迂曲度增加,分支增多,形成繁復(fù)的血管網(wǎng),對(duì)局部血流有重要的代償意義。 3.同步輻射相襯成像可同時(shí)從3D和2D角度,提供血管新生動(dòng)態(tài)發(fā)展的影像學(xué)證據(jù)。
[Abstract]:Purpose The maintenance of the physiological function of the brain depends on the completion of the microvessel network In the whole, after the ischemic stroke, the body starts the waterfall type cascade reaction to cause the steady state disorder of the microvessel network, and the structure and function of the neurovascular unit Plasmoplastic. Cerebral ischemia can trigger the neogenesis of the autograft. However, the mechanism, the development process, the specific form and so on are not yet known. It is true that the traditional methods of vascular visualization, such as the method of histopathology and the like, can not accurately show the time and space of the vascular network in the two-dimensional plane of the display of the microvessel space structure. The conventional imaging means, such as MRA, CTA, DSA, etc., can not detect the minute blood of diameter below 50 & mu; m due to the limited time-space resolution the new synchronous radiation technology breaks through the diffraction limit of the traditional optical imaging method, and can clearly realize the non-destructive and three-dimensional formation of the material microstructure by virtue of the extremely high resolution of the sub-micron to the micron order, Objective: To study the feasibility and effectiveness of brain microvessel high resolution and three-dimensional imaging, and to explore the dynamic development of vascular network remodeling after cerebral ischemia by using the phase contrast imaging of the synchrotron radiation phase contrast imaging of Shanghai light source to study the blood vessel imaging of the normal and ischemic stroke model. In order to provide a rich 3D imaging and morphometric method for the study of vascular injury and repair of ischemic stroke with a new angle of view. It was reported. Methods: The imaging parameters were adjusted, and the whole brain blood of the normal rats was reconstructed from multi-angle and multi-plane by using the filtering inverse algorithm and the imaging software. The 3D view of the tube is based on the structural information of the vascular network. Quantitative analysis of the brain tissue of the body microscope and HE staining The model of the ischemic stroke of the rat was then used to observe the dynamic development of the spontaneous blood vessel at the same time after the ischemia, and the method of quantitative analysis of the new blood vessel was explored, and the immune group was combined. Comprehensive evaluation of vascular network of ischemic stroke from 3D/ 2D visual angle winding weight Results 1. Using the synchrotron radiation phase contrast imaging, the three-dimensional slice from the projection, the two-dimensional slice to the full-vascular network is established. The complete visualization system of the image. A 5.9. m u.M CCD detector is used, with a resolution of a minimum vessel diameter of about 9.4. m And is close to the capillary level. The two-dimensional plain film is compared with the HE staining, and the preparation procedures of the contrast agent and the complex slice are not needed, so that the blood capillary level can be clearly displayed. The blood vessels in the hippocampus of the hippocampus. The 3-D rendered image on the surface of the brain and the microfil blood vessel cast-in-place microscope after-perfusion of the body-type microscope in situ were highly consistent, and the multi-stage blood vessels of the dura mater 2. The whole-brain horizontal position, the coronal, the sagittal and the digital 3D cerebral angiography were further established, and the blood supply and distribution of the functional areas were analyzed, and a series of numbers were constructed from the 2D and 3D perspectives. 3. The results of the blood vessel network skeleton extraction show that the whole blood vessel network of the rat is mainly below 20. m Based on the morphometric analysis of the 3D network, the cerebral vascular volume, the vessel density, the number of vessel branches, the number of nodes, the adjacent blood vessel spacing, the diameter, 4. The results of 2D and 3D imaging of a series of synchronous radiation sources around the ischemic stroke model in rats showed that the number of microvessels increased continuously at 2 hours,4 hours and 6 hours after the ischemia, and the number of microvessels in the ischemic area was densely distributed at the periphery of the ischemic area. However, with the prolongation of the time of the ischemia, the distribution and the density of the new blood vessels decreased after 3 days. The shape of the vessel was distorted and the irregular blood vessel was formed. The ischemic 7-day infarction was liquefied into a honeycomb-like capsule, and the glial scar was gradually formed. The volume of the lateral brain of the focal side of the lesion was significantly reduced in the 18-day-after-ischemia. The cerebral ventricle was compressed, and the cortical and striatal infarction foci were developed to form a high-density scar tissue and a non-parenchymal tissue-free> "apoplex. Quantitative analysis showed that with the increase of the time of the ischemia, the volume of the lateral brain of the lesion was gradually decreased after 3 days, of which the focal side of the lesion was 18 days after the ischemia. The volume of blood vessel was significantly reduced in the control group (P0.05). The blood vessel density of 2 hours,4 hours and 6 hours after the ischemia was higher than that in the control group (P0.05). The blood vessel density was gradually decreased in 8 days (P0.05). The number of blood vessels in 2 hours,4 hours and 6 hours after the ischemia was significantly higher than that in the control group (P0.05). The blood vessel score was 3 days,7 days and 18 days after the ischemia. The number of the branches and the number of the nodes decreased gradually (P0.05). The frequency of the small blood vessels and the control group of 2 hours,4 hours,6 hours and 10-20. m The difference was statistically significant (P0.05). The frequency gradually decreased with the lapse of time and the ischemia was 7 days. The frequency of 18 days was significantly decreased (P0.05). After the ischemia,2 hours,4 hours,6 hours,1 day and 3 days The blood vessel tortuosity of CD31 was significantly higher (P0.05). The vessel is in the form of a punctiform, curved, short or round tube with varying forms. Over time, ischemia 1 The number of newly-born microvessels was gradually reduced on day,3 and 7 days. microblood Conclusion 1. The development trend of the tube is consistent with the results of the synchronous radiation imaging. Conclusion 1. The 3D blood supply of the whole brain and the local target area can be resolved by multi-plane multi-angle based on the establishment of the visualization system of the brain microvessel based on the synchronous radiation. 2. The structure and function remodeling of the microvessel network are caused by the ischemic injury, and the angiogenesis can be started at the early stage of the ischemia, and the peak is rapidly reached, the tortuosity of the new blood vessel is increased, and the branch and the synchronous radiation phase contrast imaging can
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R743.3
本文編號(hào)：2482781
[Abstract]:Purpose The maintenance of the physiological function of the brain depends on the completion of the microvessel network In the whole, after the ischemic stroke, the body starts the waterfall type cascade reaction to cause the steady state disorder of the microvessel network, and the structure and function of the neurovascular unit Plasmoplastic. Cerebral ischemia can trigger the neogenesis of the autograft. However, the mechanism, the development process, the specific form and so on are not yet known. It is true that the traditional methods of vascular visualization, such as the method of histopathology and the like, can not accurately show the time and space of the vascular network in the two-dimensional plane of the display of the microvessel space structure. The conventional imaging means, such as MRA, CTA, DSA, etc., can not detect the minute blood of diameter below 50 & mu; m due to the limited time-space resolution the new synchronous radiation technology breaks through the diffraction limit of the traditional optical imaging method, and can clearly realize the non-destructive and three-dimensional formation of the material microstructure by virtue of the extremely high resolution of the sub-micron to the micron order, Objective: To study the feasibility and effectiveness of brain microvessel high resolution and three-dimensional imaging, and to explore the dynamic development of vascular network remodeling after cerebral ischemia by using the phase contrast imaging of the synchrotron radiation phase contrast imaging of Shanghai light source to study the blood vessel imaging of the normal and ischemic stroke model. In order to provide a rich 3D imaging and morphometric method for the study of vascular injury and repair of ischemic stroke with a new angle of view. It was reported. Methods: The imaging parameters were adjusted, and the whole brain blood of the normal rats was reconstructed from multi-angle and multi-plane by using the filtering inverse algorithm and the imaging software. The 3D view of the tube is based on the structural information of the vascular network. Quantitative analysis of the brain tissue of the body microscope and HE staining The model of the ischemic stroke of the rat was then used to observe the dynamic development of the spontaneous blood vessel at the same time after the ischemia, and the method of quantitative analysis of the new blood vessel was explored, and the immune group was combined. Comprehensive evaluation of vascular network of ischemic stroke from 3D/ 2D visual angle winding weight Results 1. Using the synchrotron radiation phase contrast imaging, the three-dimensional slice from the projection, the two-dimensional slice to the full-vascular network is established. The complete visualization system of the image. A 5.9. m u.M CCD detector is used, with a resolution of a minimum vessel diameter of about 9.4. m And is close to the capillary level. The two-dimensional plain film is compared with the HE staining, and the preparation procedures of the contrast agent and the complex slice are not needed, so that the blood capillary level can be clearly displayed. The blood vessels in the hippocampus of the hippocampus. The 3-D rendered image on the surface of the brain and the microfil blood vessel cast-in-place microscope after-perfusion of the body-type microscope in situ were highly consistent, and the multi-stage blood vessels of the dura mater 2. The whole-brain horizontal position, the coronal, the sagittal and the digital 3D cerebral angiography were further established, and the blood supply and distribution of the functional areas were analyzed, and a series of numbers were constructed from the 2D and 3D perspectives. 3. The results of the blood vessel network skeleton extraction show that the whole blood vessel network of the rat is mainly below 20. m Based on the morphometric analysis of the 3D network, the cerebral vascular volume, the vessel density, the number of vessel branches, the number of nodes, the adjacent blood vessel spacing, the diameter, 4. The results of 2D and 3D imaging of a series of synchronous radiation sources around the ischemic stroke model in rats showed that the number of microvessels increased continuously at 2 hours,4 hours and 6 hours after the ischemia, and the number of microvessels in the ischemic area was densely distributed at the periphery of the ischemic area. However, with the prolongation of the time of the ischemia, the distribution and the density of the new blood vessels decreased after 3 days. The shape of the vessel was distorted and the irregular blood vessel was formed. The ischemic 7-day infarction was liquefied into a honeycomb-like capsule, and the glial scar was gradually formed. The volume of the lateral brain of the focal side of the lesion was significantly reduced in the 18-day-after-ischemia. The cerebral ventricle was compressed, and the cortical and striatal infarction foci were developed to form a high-density scar tissue and a non-parenchymal tissue-free> "apoplex. Quantitative analysis showed that with the increase of the time of the ischemia, the volume of the lateral brain of the lesion was gradually decreased after 3 days, of which the focal side of the lesion was 18 days after the ischemia. The volume of blood vessel was significantly reduced in the control group (P0.05). The blood vessel density of 2 hours,4 hours and 6 hours after the ischemia was higher than that in the control group (P0.05). The blood vessel density was gradually decreased in 8 days (P0.05). The number of blood vessels in 2 hours,4 hours and 6 hours after the ischemia was significantly higher than that in the control group (P0.05). The blood vessel score was 3 days,7 days and 18 days after the ischemia. The number of the branches and the number of the nodes decreased gradually (P0.05). The frequency of the small blood vessels and the control group of 2 hours,4 hours,6 hours and 10-20. m The difference was statistically significant (P0.05). The frequency gradually decreased with the lapse of time and the ischemia was 7 days. The frequency of 18 days was significantly decreased (P0.05). After the ischemia,2 hours,4 hours,6 hours,1 day and 3 days The blood vessel tortuosity of CD31 was significantly higher (P0.05). The vessel is in the form of a punctiform, curved, short or round tube with varying forms. Over time, ischemia 1 The number of newly-born microvessels was gradually reduced on day,3 and 7 days. microblood Conclusion 1. The development trend of the tube is consistent with the results of the synchronous radiation imaging. Conclusion 1. The 3D blood supply of the whole brain and the local target area can be resolved by multi-plane multi-angle based on the establishment of the visualization system of the brain microvessel based on the synchronous radiation. 2. The structure and function remodeling of the microvessel network are caused by the ischemic injury, and the angiogenesis can be started at the early stage of the ischemia, and the peak is rapidly reached, the tortuosity of the new blood vessel is increased, and the branch and the synchronous radiation phase contrast imaging can
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類號(hào)】：R743.3
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