Name: in vivo tracking of edema development and microvascular pathology in a model of experimental cerebral malaria using magnetic resonance imaging
Uploaded: 2017-06-08T12:30:00
Description: Watch this Scientific Journal Video about In Vivo Tracking of Edema Development and Microvascular Pathology in a Model of Experimental Cerebral Malaria Using Magnetic Resonance Imaging at JoVE.com

미리 암 레이 닉 (Miriam Reinig)의 전문가 기술 지원은 감사하게 생각합니다. AH는 하이델베르그 대학 (University of Heidelberg)의 의과 대학의 박사후 연구원으로부터 기금을 받았다. MP는 Else-Kröner-Fresenius Foundation의 기념비에 의해 지원됩니다. AKM은 독일 감염 연구소 (DZIF)의 DZIF 아카데미에 의해 출산 휴가를받습니다. JP는 분자 의학 (HRCMM) 경력 개발 펠로우쉽을위한 하이델베르그 연구 센터의 수령인이다. 우리는 또한 sporozoite 운동의 모범적 인 영화를 제공 한 Julia M. Sattler와 Friedrich Frischknecht에게 감사를 표합니다.

이 논문에서보고 된 모든 동물 실험은 실험 동물 학회 연맹 (FELASA) 카테고리 B 및 실험실 동물 과학 학회 (GV-SOLAS) 표준 지침에 따라 수행되었으며 카를 스루에 지역 독일 당국 (Regierungspräsidium Karlsruhe)의 승인을 받았습니다 , 독일). biosavety 레벨 2 모기 및 Plasmodium berghei ANKA sporozoite 작품에 적용됩니다. 1. 감염 <ol><li> 해리 동물 에 15 분 동안 먹이를 줌으로써 Anophel...

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이 기사에서는 실험적인 대뇌 말라리아의 변화를 묘사하기위한 전체 뇌 MRI 프로토콜에 대해 설명합니다. 우리는 지금까지 말라리아 연구에서 MRI가 제대로 활용되지 않았으며 우리의 프로토콜이 다른 수사관을 도울 것이라고 희망합니다. 도움이 될 수있는 몇 가지 추가 점을 설명하고자합니다. 심하게 아픈 생쥐가 이미징 된 경우, 위치 결정이 중요합니다. 두개 내압이 증가...

말라리아는 전 세계적으로 심각한 건강 문제입니다. 중증 말라리아는 부분적으로 뇌의 침범으로 특징 지어지며 흔히 예후가 나쁜 요인입니다. 대뇌 침범은 말라리아 전염이 심한 지역에서 5 세 미만의 어린이에게 흔히 발생하며 그 연령대에서 말라리아 관련 사망의 주요 원인을 나타냅니다. 공격적인 치료가 생명을 구할 수 있지만 특히 초기 단계의 뇌 말라리아의 검출은 어려울 수 있습니다. 뇌 말라리아와 관련된 병리학 적 과정은 심한 뇌의 부종으로 이어질 수있는 미세 혈관 파괴 및 뇌부종을 포함합니다. 이 기사에서는 실험적 대뇌 말라리아 (ECM)의 전체 뇌 생체 내 이미징을 허용하는 자기 공명 영상 (MRI) 프로토콜을 제시합니다. ECM이 중앙에서 어떻게 시작되는지에 대해서는 거의 알려지지 않았지만 전체 뇌 고해상도 이미징 방법은이 질병에서 널리 활용되지 못했습니다.신경계 또는 어떤 메커니즘이이 질병을 일으키는 지 확인하십시오. 생체 내 (in vivo) MRI는 전뇌를 포괄하며 ECM 병리를 더 잘 이해할 수있는 중요한 연구 도구입니다. MRI는 세계적 뇌 뇌 팽창을 평가할 수 있는데, 이는 최근 ECM뿐만 아니라 인간 대뇌 말라리아에서 중요한 사망 예측 인자로 인식되고 있습니다. 심한 뇌의 부종은 치명적인 질병에서 발생하며 ECM 모델과 인간 질병 (염증성 및 미세 혈관계 변화가 특징 인 질환) 사이의 여러 병리학 적 특징 중 하나를 나타냅니다. 4 치명적인 Plasmodium berghei ANKA 감염에 의해 CBA 또는 C57BL 생쥐에서 ECM을 유도 할 수 있습니다 . ECM의 발병은 전형적으로 감염 후 6 일에서 10 일 사이에 발생하며 피팅, 운동 장애, 호흡 곤란 및 혼수 상태를 초래하여 랩이드의 죽음. Rapid Murine Coma and Behavior Scale (RMCBS)은 ECM의 임상 증상을 평가하는데 도움이되는 점수입니다. 그것은 0에서 2까지 점수가 매겨진 10 개의 매개 변수로 구성되며 가능한 최대 점수는 20입니다. 6 최근에 ECM 마우스의 RMCBS 점수와 MRI로 입증 된 병리학 적 변화의 정도간에 좋은 일치를 보였습니다. 7 이 프로토콜에서 우리는 생쥐의 ECM 유도와 ECM이있는 마우스의 생체 내 자기 공명 영상을 설명합니다.

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			<td>MATLAB&#160;</td>
			<td>The MathWorks, Inc.,</td>
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			<td>FDT toolbox&#160;</td>
			<td colspan="2">FMRIB&#39;s Software Library</td>
			<td><a href="http://www.fmrib.ox.ac.uk/fsl/fdt/index.html">http://www.fmrib.ox.ac.uk/fsl/fdt/index.html</a></td>
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in vivo tracking of edema development and microvascular pathology in a model of experimental cerebral malaria using magnetic resonance imaging

Cerebral malaria is a sign of severe malarial disease and is often a harbinger of death. While aggressive management can be life-saving, the detection of cerebral malaria can be difficult. We present an experimental mouse model of cerebral malaria that shares multiple features of the human disease, including edema and microvascular pathology. Using magnetic resonance imaging (MRI), we can detect and track the blood-brain barrier disruption, edema development, and subsequent brain swelling. We describe multiple MRI techniques that can visualize these pertinent pathological changes. Thus, we show that MRI represents a valuable tool to visualize and track pathological changes, such as edema, brain swelling, and microvascular pathology, in vivo.

C57BL / 6 마우스에서 P. berghei ANKA 스포르 조제 감염 후 6 일에서 10 일 사이에 ECM의 첫 임상 증상이 관찰 될 수 있습니다. ECM은 감염된 마우스의 60 ~ 80 %에서 발병하며 24 ~ 48 시간 내에 빠르게 혼수 상태 및 사망으로 진행됩니다. 반대로, ECM이 발생하지 않는 마우스는 과증식으로 인해 심한 빈혈로 감염 후 2 주 후에 사망합니다. 12 <p class="jove_content" f...

Watch this Scientific Journal Video about In Vivo Tracking of Edema Development and Microvascular Pathology in a Model of Experimental Cerebral Malaria Using Magnetic Resonance Imaging at JoVE.com

In Vivo Tracking of Edema Development and Microvascular Pathology in a Model of Experimental Cerebral Malaria Using Magnetic Resonance Imaging

We describe a mouse model of experimental cerebral malaria and show how inflammatory and microvascular pathology can be tracked in vivo using magnetic resonance imaging.

자기 공명 영상을 이용한 실험용 뇌 말라리아 모델에서의 부종 발달 및 미세 혈관 병리학 적 추적

Cerebral malaria is a sign of severe malarial disease and is often a harbinger of death. While aggressive management can be life-saving, the detection of ...

The overall goal of this method is to monitor pathological changes of experimental cerebral malaria in vivo. This method can help to answer key questions in the pathogenesis of experimental cerebral malaria as it allows us to visualize in vivo how pathology of the whole brain evolves in space and over time. The technique thereby allows a detailed assessment of brain pathology, and can be used to assess the efficacy of novel treatment strategies in cerebral malaria.Performing the MRI setup will be Manuel Fischer, our lab manager. Begin by collecting female mosquitoes from their cage, 17 to 22 days after the blood meal. Use forceps to place three to four mosquitoes on a glass slide, combined with a drop of cold RPMI medium.Then, place the slide under a microscope. Carefully stretch the mosquito between the head and body with forceps, and use a syringe and needle to isolate the salivary gland. Next, collect the salivary glands from the glass slide by sucking them up with a glass pipette, and transferring them into 1.5 milliliter centrifuge tube.Then, use a small plastic stick to smash the isolated salivary glands within the centrifuge tube for three minutes in order to isolate the sporozoites from the salivary gland tissue. Centrifuge for three minutes at 1, 000 times g and four degrees Celsius to purify the sporozoites from the remaining tissue. Pipette the supernatant, which contains the sporozoites, to a new centrifuge tube, and count the purified sporozoites in a Neubauer hemocytometer.Sporozoites show a typical counterclockwise movement. Next, adjust the concentration of the purified sporozoites to 10, 000 per milliliter by adding phosphate-buffered saline. Finally, place a C57BL/6 mouse in a restrainer, and put its tail into warm water to better visualize the tail vein.Then, inject a total of 1, 000 sporozoites, or 0.1 milliliters, into the tail vein to initiate infection. Begin by bringing the mouse to a 9.4T small animal Magnetic Resonance Imaging, or MRI scanner, that uses a volume resonator for radiofrequency transmission. Then, apply dexpanthenol eye ointment to both eyes.Turn on the temperature-controlled water bath to 42 degrees Celsius in order to maintain the body temperature of the mouse. Then, place a tail vein catheter into the mouse's tail vein. Next, position the mouse for the MRI by placing it prone and with a crunched back on an animal bed equipped with a headlock and tooth bar to minimize head motion.Take care not to straighten the cervical spine of the mouse. Connect a contrast agent injection system filled with 0.3 millimole per kilogram Gd-DTPA to the tail vein catheter. Next, place the 4-Channel Phased Array Surface Receiver head coil onto the head of the mouse.Finally, place a breathing pad onto the back of the mouse and check the respiration on the monitoring device. Turn on the positional laser by pressing F2 on the control panel, and move the mouse until the horizontal positional light is in the middle of the smaller part of the coil, which covers the head of the mouse. Turn off the positional laser by pressing F2 again, and then move the mouse into the final position by pressing F3 on the control panel.Begin by performing a localizing scan to make sure that the mouse brain is in the isocenter of the magnet. Qualitatively assess vasogenic edema by using 3D T2-weighted imaging. Select a multi-slice RARE sequence, and adjust the slice position, making sure that the whole brain, from the nose to the cerebellum is covered.After adjusting the saturation slices, start the sequence, and wait 10 minutes and 48 seconds to acquire the images. Next, perform T2 relaxometry to quantitatively assess vasogenic edema by selecting multi-slice, multiple-spin echo sequence. Adjust the slice position, and start the acquisition of the sequence, which takes eight minutes and 53 seconds.To quantitatively assess both vasogenic edema and cytotoxic edema, carry out diffusion-weighted imaging by selecting a spin-echo EPI diffusion sequence. Adjust the slice position and saturation slice. Then start the sequence and wait seven minutes and 56 seconds until the raw images are acquired.Following that, assess microhemorrhages, using 3D T2 star-weighted imaging. Select a flow-compensated FLASH sequence, adjust the slice position, and start the sequence, which runs for 15 minutes and 40 seconds. Afterwards, assess arterial patency using time of flight angiography by selecting a 3D FLASH sequence.Adjust the slice position, start the sequence, and then wait seven minutes and 16 seconds until the images are acquired. Finally, assess the blood-brain barrier disruption. Select a 3D T1-weighted imaging with a FLASH sequence, adjust the slice position, and start the sequence.Wait one minute and 14 seconds until images without contrast are acquired. Then, inject contrast of 0.3 millimole per kilogram and repeat the measurement. An increase in blood-brain barrier disruption and a fluid increase indicate early vasogenic edema in the olfactory bulb that is already present in mild disease, and starts to spread towards the brain stem as the severity of the disease increases.On T2 maps, the severity of vasogenic edema can be quantified by drawing regions of interest into specific anatomical regions. For each region, a T2 relaxation time that increases with increasing edema can be obtained. Furthermore, brain volume indicative of edema starts to significantly increase in moderately sick mice as compared to baseline, and further increases in severely sick mice.Lastly, microvascular pathology, as evidenced by microhemorrhages and an increase of vessel susceptibility contrast, occurs after the first signs of blood-brain barrier disruption in vasogenic edema. These microhemmorhages predominantly occur in the olfactory bulb. If the whole MRI protocol is obtained, all images including positioning can be obtained within one hour, if it is performed properly.The minimal protocol should include a T2 weighted sequence or T2 relaxometry, and a T2 star-weighted sequence in order to judge the presence of vasogenic edema, microhemmorhage load, and microvascular impairment. After watching this video, you should have a good understanding how whole-brain MRI visualizes pathological changes in experimental cerebral malaria, including such changes as brain swelling, which is difficult to detect ex vivo.

Research

JoVE Journal

Medicine

In Vivo Tracking of Edema Development and Microvascular Pathology in a Model of Experimental Cerebral Malaria Using Magnetic Resonance Imaging

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Myocarditis is an inflammation of the myocardium, but only ~10% of those affected show clinical manifestations of the disease. To study the immune events ...

Tracking the Mammary Architectural Features and Detecting Breast Cancer with Magnetic Resonance Diffusion Tensor Imaging

자기 공명 확산 텐서 영상과 유방 아키텍처 특징을 추적 및 탐지 유방암

Breast cancer is the most common cause of cancer among women worldwide. Early detection of breast cancer has a critical role in improving the quality of ...

Simultaneous PET/MRI Imaging During Mouse Cerebral Hypoxia-ischemia

마우스 대뇌 저산소증 - 허혈 동안 동시 PET / MRI 영상

Dynamic changes in tissue water diffusion and glucose metabolism occur during and after hypoxia in cerebral hypoxia-ischemia reflecting a bioenergetics ...

Using Multi-fluorinated Bile Acids and In Vivo Magnetic Resonance Imaging to Measure Bile Acid Transport

Using Multi-fluorinated Bile Acids and In Vivo Magnetic Resonance Imaging to Measure Bile Acid Transport

멀티 플루오르 담즙 산 및 사용<em&gt; 생체</em&gt; 자기 공명 영상은 측정하는 담즙 산 전송

Along with their traditional role as detergents that facilitate fat absorption, emerging literature indicates that bile acids are potent signaling ...

Quantitative Visualization of Leukocyte Infiltrate in a Murine Model of Fulminant Myocarditis by Light Sheet Microscopy

Light Sheet Microscopy에 의한 심근 성 심근염의 Murine 모델에서의 백혈구 침윤의 정량적 시각화

Light-sheet fluorescence microscopy (LSFM), in combination with chemical clearing protocols, has become the gold standard for analyzing fluorescently ...

Cardiac Magnetic Resonance Imaging at 7 Tesla

CMR at an ultra-high field (magnetic field strength B0&#160;&#8805; 7 Tesla) benefits from the signal-to-noise ratio (SNR) advantage inherent at higher ...