Name: an in vivo assessment of blood-brain barrier disruption in a rat model of ischemic stroke
Uploaded: 2018-03-11T14:00:00.000+00:00
Duration: 12 min 19 s
Description: Watch this Scientific Journal Video about An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke at JoVE.com

저자는 재정 지원에 대 한 아르다 빌 대학 의료 과학 (아르다 빌, 이란)의 연구에 대 한 부 장관에 게 감사 (No를 부여: 9607).

모든 절차는 동물 연구를 실시 위한 아르다 빌 대학교 의료 과학 연구 위원회의 지침에 따라 수행한 (윤리적 ID 번호: 적외선 ARUMS입니다. REC.1394.08).이 시각된 연구에서 사용 하는 성인 남성 Sprague-Dawley 쥐 (300-350 g) 목초지 연구소 (테헤란, 이란)에서 얻은.
1. 마 취와 Flowmetry
<ol><li>4 %isoflurane 사용 하 여 마 취를 유도 하 고 수술 하는 동안 질소 산화물 (70 %v / v)와 산소 (30 %v / v)의 혼합물에 isoflurane (1-1.5%)으로 유지 합니다.</li>
	<li>피드 백 제어 난방 담요에 발생 하기 쉬운 자세에서 anaesthetized 동물을 놓고이 난방 장치에 연결 된 프로브를 직장에 의하여 37±0.5 ° C에서 체온을 유지 합니다.</li>
	<li>건조를 방지 하기 위해 양쪽 눈에 안과 연 고의 작은 금액을 적용 합니다.</li>
	<li>전기 면도기와 두개골의 왼쪽에 외과 영역을 면도. 거 즈 패드와 함께 betadine 솔루션을 사용 하 여 피부를 소독. 린스이 지역 메 마른 패드 70% 에탄올을 포함 하 고 3 주기17두 단계를 반복 합니다.</li>
	<li>진통, 대 수술 지역17에 0.5 %bupivacaine 0.2 mL을 피하 주사.</li>
	<li>(왼쪽된 눈의 측면 법에서의 왼쪽된 귀에 확장 되는) 두개골에 1cm 긴 피부 절 개를 하 고 두개골 노출 왼쪽된 temporalis 근육 해 부. 그런 다음 만들 작은 버 구멍 5mm 측면 하 고 1mm 도플러 유량 계 연필 프로브 팁의 용이성 배치 bregma18 후부.</li>
	<li>경향이 부정사의 위치에서 쥐 바뀌고 레이저 연필 조사 지역 대뇌 혈 류 (rCBF)을 모니터링 하는 데 이전 드릴된 두개골 블라인드 구멍에 넣어.</li>
	<li>각 동물의 초기 rCBF를 기록 하 고 100%로 이것을 고려 하십시오. 특히, 중간 대뇌 동맥 폐색 (MCAO) rCBF의 80%에 감소는 검색된19,20때마다 시작 됩니다.</li>
</ol>2입니다. 초점 Cerebra 허 혈의 유도
<ol><li>목 영역을 면도 하 고 betadine 솔루션 및 70% 에탄올으로 피부를 소독. 진통17수술 사이트에서 피하 0.5 %bupivacaine 0.2 mL를 주사.</li>
	<li>좌 일반 경 동맥 (LCCA) 액세스할 수 목의 복 부 표면에 2 cm 긴 외과 절 개를 확인 합니다. 외부 경 동맥 (ECA) 및 내부 경 동맥 (ICA)의 분기점에 액세스 하려면 인접 근 막과 미주 신경에서 LCCA를 격리 합니다.</li>
	<li>LCCA 또는 5-0 비단 봉합 사를 고용 하는 ECA 영구적으로 선 하 고 ICA pterygopalatine 동맥의 수준에 무료 부.</li>
	<li>느슨하게 LCCA, 주위 5-0 비단 봉합의 넥타이 배치 후 일시적으로 사라지는 혈관 마이크로 클립 클램프.</li>
	<li>이전 배치 느슨한 넥타이 이전 LCCA에 작은 절 개를 하 게 다음 ICA의 luminal 공간에 4-0 실리콘 코팅 나일론 봉합을 삽입 하 고 나일론 봉합 확보 및 혈액 누수 방지 LCCA 주위 봉합 조입니다.</li>
	<li>ICA에서 혈관 마이크로 클립을 제거 합니다. 그런 다음, MCA 근원의 폐색을 나타내는 rCBF에 표시 된 감소를 관찰까지 실리콘 코팅된 intraluminal 필 라 멘 트를 사전. 허 혈 성 기간 (90 분)의 끝에, intraluminal 봉합21철수 여 reperfusion를 시작 합니다.</li>
</ol>3. 경 정 맥 Cannulation 및 에반스 블루 (EB) 주입
<ol><li>1 cm 세로 절 개를 목에는 중간의 왼쪽을 만들고 퉁 명 스럽게 왼쪽된 경 정 맥 (LGV)의 외부 분기 액세스 멀리 표면 근을 해 부. 다음, 영구적으로 5-0 비단 봉합으로 LGV의 두개골 끝 선. 느슨하게 주위 정 맥, 두 관계를 놓고 일시적으로 혈관 마이크로 클램프와 LGV의 심장 끝을 클램프.</li>
	<li>두 봉합 사이 LGV에 작은 절 개를 하 게 그리고 LGV의 luminal 공간으로 가득 heparinized 혈 청 카 테 터 삽입 및 사전 그것은 약 10 m m. 나중, 조여 정 맥 카 테 터를 확보 하 고 출혈 방지 주위 합자. 적은 양의 정 맥 붕괴를 피하기 위해 혈 청을 주입.</li>
	<li>Reperfusion 기간 시작 되는 시점에서 5 분 동안 EB 염료 (식 염 수에 2 %EB 솔루션의 1 mL/kg) 주사 천천히. 그 후,는 정 0.5 mL 정상 식 염 수를 주입 하 여 세척 하 고 주입 정 맥12,22에서 철수.</li>
	<li>마지막으로, 목과 머리의 절 개를 봉합 하 고 intraperitoneally buprenorphine 0.05 mg/kg을 주사 (IP) 수술 후 진통17reperfusion 기간 동안 모든 6-8 h 주사를 반복.</li>
	<li>복구 케이지17수 화를 제공 하기 위해 미리 데워 진된 saline(IP)의 5 mL를 주사.</li>
	<li>장소 특별 한 복구에 동물 온도 및 공기 조화 제어 하 고 마 취에서 동물의 복구를 모니터링 합니다.</li>
</ol>4. 혈액 뇌 장벽 침투성의 평가
<ol><li>Reperfusion의 24 h 후 깊이 thiopental 나트륨과 동물 anesthetize. 그런 다음, 흉 골 아래에 작은 구멍을 만들어 흉 강을 엽니다.</li>
	<li>오른쪽 아 트리 움에 작은 개통을 확인 하 고 미리 따뜻하게 0.9% 식 염 수 (37 ˚C) 안마당에서 정상적인 염 분 출구23무색 될 때까지 순환에서 EB를 씻어 15 분 동안 좌 심 실을 통해 110 mmHg의 압력의 250 mL를 주입.</li>
	<li>즉시 두개골에서 뇌를 제거 하 고 뇌 매트릭스에 배치. 후 각 전구 및 소 뇌를 해 부하 고 중간 선 따라 왼쪽 (Lesioned)에서 뇌의 오른쪽 반구를 분리 후 깨끗 한 면도날을 사용 하 여.</li>
	<li>각 반구의 무게 하 고 별도로 버퍼링 하는 인산 염의 2.5 mL에 균질 trichloroacetic 산 (60%)의 2.5 mL 소용돌이 기계를 사용 하 여 2 분이 혼합.</li>
	<li>1,322 x g에서 30 분 동안 뇌 샘플 원심 고 샘플 10 분 동안 4 ℃ 냉장고에 진정을 허용 합니다.</li>
	<li>610 nm23에 분 광 광도 계에 의해 EB 흡 광도 추정 하는 supernatants를 사용 합니다.</li>
	<li>EB 농도 표준 곡선 및 표현 결과 대 한 뇌 조직의 µ g/g으로 계산 합니다.</li>
</ol>5입니다. 생산 EB 표준 곡선의
<ol><li>EB 버퍼링 하는 인산 염의 5 ml에서의 µ g 1-10의 농도와 EB의 10 샘플 솔루션을 준비 합니다.</li>
	<li>610에 각 샘플의 흡 광도 값 nm는 분 광 광도 계를 사용 하 여.</li>
	<li>Y 축 (그림 1)에 스프레드시트와 X 축에 현재 EB 농도 흡 광도 값을 사용 하 여 분산형 차트를 확인 합니다.</li>
	<li>곡선의 방정식에 대 한 선형 추세선을 정의 합니다. 그런 다음 샘플의 EB 농도를 사용 합니다.</li>
	<li>뇌 조직 무게의 µ g/g으로 EB 넘쳐 흐름의 최종 결과 보고 합니다.</li>
</ol>6. 가짜 작업
<ol><li>가짜 운영 그룹 (포함 절 개 목 지역에 EB 주입)에 쥐와 같은 수술 과정을 수행 하지만 MCAO는 제외.</li>
</ol>

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A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Candesartan+attenuates+ischemic+brain+edema+and+protects+the+blood-brain+barrier+integrity+from+ischemia/reperfusion+injury+in+rats.">Candesartan attenuates ischemic brain edema and protects the blood-brain barrier integrity from ischemia/reperfusion injury in rats.</a> Iran Biomed J. 18 (4), 232-238 (2014).</li><li>Kaya, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+magnesium+sulfate+on+blood-brain+barrier+disruption+caused+by+intracarotid+injection+of+hyperosmolar+mannitol+in+rats.">The effects of magnesium sulfate on blood-brain barrier disruption caused by intracarotid injection of hyperosmolar mannitol in rats.</a> Life sci. 76 (2), 201-212 (2004).</li><li>Sch&#246;ller, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+microvascular+basal+lamina+damage+and+blood-brain+barrier+dysfunction+following+subarachnoid+hemorrhage+in+rats.">Characterization of microvascular basal lamina damage and blood-brain barrier dysfunction following subarachnoid hemorrhage in rats.</a> Brain Res. 1142, 237-246 (2007).</li><li>Bodsch, W., Hossmann, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=125I-Antibody+Autoradiography+and+Peptide+Fragments+of+Albumin+in+Cerebral+Edema.">125I-Antibody Autoradiography and Peptide Fragments of Albumin in Cerebral Edema.</a> J Neurochem. 41 (1), 239-243 (1983).</li><li>Sandoval, K. E., Witt, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blood-brain+barrier+tight+junction+permeability+and+ischemic+stroke.">Blood-brain barrier tight junction permeability and ischemic stroke.</a> Neurobiol Dis. 32 (2), 200-219 (2008).</li><li>Zhu, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Baicalin+reduces+the+permeability+of+the+blood-brain+barrier+during+hypoxia+in+vitro+by+increasing+the+expression+of+tight+junction+proteins+in+brain+microvascular+endothelial+cells.">Baicalin reduces the permeability of the blood-brain barrier during hypoxia in vitro by increasing the expression of tight junction proteins in brain microvascular endothelial cells.</a> J Ethnopharmacol. 141 (2), 714-720 (2012).</li><li>Kucuk, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+losartan+on+the+blood-brain+barrier+permeability+in+long-term+nitric+oxide+blockade-induced+hypertensive+rats.">Effects of losartan on the blood-brain barrier permeability in long-term nitric oxide blockade-induced hypertensive rats.</a> Life Sci. 71 (8), 937-946 (2002).</li><li>Uyama, O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+evaluation+of+vascular+permeability+in+the+gerbil+brain+after+transient+ischemia+using+Evans+blue+fluorescence.">Quantitative evaluation of vascular permeability in the gerbil brain after transient ischemia using Evans blue fluorescence.</a> J Cereb Blood Flow Metab. 8 (2), 282-284 (1988).</li><li>Kleinig, T. J., Vink, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Suppression+of+inflammation+in+ischemic+and+hemorrhagic+stroke:+therapeutic+options.">Suppression of inflammation in ischemic and hemorrhagic stroke: therapeutic options.</a> Curr Opin Neurol. 22 (3), 294-301 (2009).</li><li>Del Zoppo, G. J., Mabuchi, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cerebral+microvessel+responses+to+focal+ischemia.">Cerebral microvessel responses to focal ischemia.</a> J Cereb Blood Flow Metab. 23 (8), 879-894 (2003).</li><li>Fluri, F., Schuhmann, M. K., Kleinschnitz, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+models+of+ischemic+stroke+and+their+application+in+clinical+research.">Animal models of ischemic stroke and their application in clinical research.</a> Drug Des Devel Ther. 9, 3445-3454 (2015).</li><li>Noor, R., Wang, C. X., Shuaib, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+hyperthermia+on+infarct+volume+in+focal+embolic+model+of+cerebral+ischemia+in+rats.">Effects of hyperthermia on infarct volume in focal embolic model of cerebral ischemia in rats.</a> Neurosci Lett. 349 (2), 130-132 (2003).</li><li>Shin, H. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mild+induced+hypertension+improves+blood+flow+and+oxygen+metabolism+in+transient+focal+cerebral+ischemia.">Mild induced hypertension improves blood flow and oxygen metabolism in transient focal cerebral ischemia.</a> Stroke. 39 (5), 1548-1555 (2008).</li><li>Bottiger, B. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+cerebral+ischemia+due+to+cardiocirculatory+arrest+in+mice+causes+neuronal+degeneration+and+early+induction+of+transcription+factor+genes+in+the+hippocampus.">Global cerebral ischemia due to cardiocirculatory arrest in mice causes neuronal degeneration and early induction of transcription factor genes in the hippocampus.</a> Brain Res Mol Brain Res. 65 (2), 135-142 (1999).</li></ol>

저자는 공개 없다.

지금까지, autoradiography 방사성 추적기24,25, 면역 형광 검사 현미경 검사 법26,27, EB 넘쳐 흐름 기술은20, 의 검출 등 다양 한 방법 23 혈액-뇌 장벽 손상을 평가 하는 데 사용 되었습니다. EB 염료 강력 하 게 혈 청 알 부 민에 바인딩할 수 이며 혈관 누설을 감지 하 고 측정 BBB 붕괴11,,2829는 추적 프로그램으로 사용 됩니다. 매우 허용 하 고 안정적인 방법으로 EB 넘쳐 흐름 기술 허 혈 성 뇌졸중을 포함 한 다른 뇌 부상에 의해 영향을 받는 BBB의 무결성에 직접 견적을 제공 합니다.
BBB의 vivo에서 평가 연구원을 허 혈 유발 vasogenic 뇌 부 종의 가능한 병 태 생리 기계 장치를 공부 하 고 새로운 치료 개입을 찾을 수 있습니다. 이 모델 특별 한 시설을 필요로 하지 않습니다 및 실험 (80% 이상)13,20의 높은 성공률을 신뢰할 수 있는 결과 얻을 수 있습니다. 뇌 조직에 직접 액세스,이 모델 BBB 무결성의 매우 정확한 평가 가능 하 게 하지만 장기 연구로 제한 됩니다.
허 혈 성 뇌졸중으로 인 한 BBB에 병 적인 변화 3 단계에서 개발: (시간), 급성 아 급성 (일 시간), 만성 (일 개월)30,31. 물론, 초기 치료 내정간섭 급성 병 적인 단계에서 귀중 한 보호 효과 생산. EB 복용량과 주입의 시간 포인트는 허 혈 성 모욕 다음 BBB의 동적 특성으로 인해 신뢰할 수 있는 결과 얻기 위한 두 가지 중요 한 매개 변수. 따라서, reperfusion 기간의 시작 후 적절 한 복용량 (1 mg/kg의 식 염 수에 2 %EB 솔루션)을 사용 하 여 정 맥 정 맥을 통해 천천히 EB 염료의 주입 중요 한 요소 이며 뇌졸중의 초기 단계에서 병 태 생리 변화 연구를 수 .
여러 가지 실험 방법은 허 혈 성 뇌졸중 연구 도입 되었습니다. 이 실험 모델에서 우리 intraluminal 필 라 멘 트 메서드를 조건 인간의 선21,32와 비슷한 MCAO 사용. 이 기술은 간단 하 고 믿을 수 있다; 그러나, 실행 기술의 성능 향상의 정확성 수 몇 가지 기술 포인트를 고려 해야 합니다. 체온 유지 되어야 한다 생리 적 범위 내에서 수술 하는 동안 혈압과 혈액 가스 모니터링된33,,3435이어야 하는 동안. 레이저 도플러 유량 계와 rCBF의 기록 하 고 적합 한 준비를 사용 하 여 일정 한 실리콘 코팅 필 라 멘 트 수 MCAO 성공률 증가 뿐만 아니라 또한 사망 율을 줄이기 위해.

Vasogenic 뇌 부 종 혈액-뇌 장벽 (BBB) 중단으로 인해 허 혈 성 뇌졸중의 중요 한 합병증 및 뇌졸중 환자1,2에서 생존 율의 주요 결정 요인이 남아 있다. 혈액-뇌 장벽 (BBB)는 뇌 모 세관 내 피 세포 (BCECs)에 의해 형성 된 뚜렷한 혈관 구성 요소 (예: BCECs, pericytes, astroglial, 및 신경 세포3중 꽉 접합)의 구성, 제공 하는 중앙 신경 조직 (CNS) 말 초 혈액 순환4,5사이의 특수 하 고 역동적인 인터페이스. 국 소 빈 혈 reperfusion 상해 등 모욕 BBB의 기능적 무결성을 방해 하 고 궁극적으로 트리거 대뇌 염증 및 기본 뇌 손상 뇌 실질에 순환 하는 백혈구의 침투를 후속으로 이어질 수 있습니다. 6 , 7. 뇌졸중의 발생에 따라 BBB의 부전의 정확한 검출을 위한 동물 모델 필요. 이러한 모델은 새로운 신경 보호 전략을 소개 하 고 기본 병 태 생리 기계 장치 공부에 대 한 매우 중요. 체 외에서 세포 문화 기반 모델 BBB의 높은 개발 하 고 되었습니다 BBB physiopathology8,,910의 분자 연구에 사용. 그럼에도 불구 하 고, vivo에서 동물 모델, 인간의 임상 조건에 비슷한 BBB의 허 혈 성 손상, 생산은 또한 매우 보람 이와. 에반스 블루 (EB)의 넘쳐 흐름의 정량적 검출은 잘 허용 하 고 민감한 기술 평가 BBB 무결성 및 신경 퇴행 성 질환, 허 혈 성 뇌졸중11, 를 포함 하 여 기능에 대 한 사용 되었습니다. 12 , 13 , 14.이 방법은 비용, 재현성, 및 완전히 적용 가능한 모든 실험 실험실에서. 그것의 구현 방사성 트레이 서15 또는 자기 공명 영상 (MRI)16다른 방법에 대 한 필수 구성 요소를 같은 고급 장비를 필요 하지 않습니다. 이 문서에서는, BBB 평가 EB 넘쳐 흐름을 사용 하 여 허 혈 성 뇌졸중의 쥐 모델에서의 기본적인 기술적 프로세스 포괄적으로 설명 합니다.

<table><tbody><tr><td>Isoflurane</td><td>Piramal</td><td>AWN 34041100</td><td>20 - 25 &amp;deg;C</td></tr><tr><td>2,3,5-Triphenyltetrazolium chloride (TTC)</td><td>Molekula</td><td>31216368</td><td>4 years</td></tr><tr><td>Sprague&amp;ndash;Dawley rats&amp;nbsp;</td><td>Pasture Institute (Tehran, Iran)</td><td></td><td>300-350g</td></tr><tr><td>Evans Blue&amp;nbsp;</td><td>Sigma-Aldrich&amp;nbsp;</td><td>314-13-6</td><td></td></tr><tr><td>Trichloroacetic acid&amp;nbsp;</td><td>Sigma-Aldrich&amp;nbsp;</td><td>76-03-9</td><td>2 years</td></tr><tr><td>Bupivacaine HCl (0.5%)</td><td>Delpharm Tours</td><td></td><td>below&amp;nbsp; 25 &amp;deg;C</td></tr><tr><td>Bupernorphine</td><td>Exir (Iran)</td><td></td><td></td></tr><tr><td>Sodium Carbonate</td><td>Sigma-Aldrich&amp;nbsp;</td><td>497-19-8</td><td></td></tr><tr><td>Sodium chloride&amp;nbsp;</td><td>Sigma-Aldrich&amp;nbsp;</td><td>7647-14-5</td><td></td></tr><tr><td>Di- Sodium hydrogen phosphate</td><td>EMD Millipore&amp;nbsp;</td><td>231-448-7</td><td></td></tr><tr><td>Potassium chloride</td><td>Sigma-Aldrich&amp;nbsp;</td><td>&amp;nbsp;7447-40-7</td><td></td></tr><tr><td>Ethanol&amp;nbsp;</td><td>Sigma-Aldrich&amp;nbsp;</td><td>64-17-5</td><td></td></tr><tr><td>silicone(Xantopren)</td><td>Heraeus</td><td>EN ISO 4823</td><td></td></tr><tr><td>Activator universal plus</td><td>Heraeus</td><td>66037445</td><td></td></tr><tr><td>Micro-Dissecting forceps</td><td>Stoelting</td><td>52100-41</td><td></td></tr><tr><td>Spring Scisors</td><td>Stoelting</td><td>52130-00</td><td></td></tr><tr><td>Operating&amp;nbsp; Scissors</td><td>Roboz</td><td>52140-70</td><td></td></tr><tr><td>Brain matrix&amp;nbsp;</td><td>Stoelting</td><td>51390</td><td></td></tr><tr><td>Anesthesia Machine for Small Animals |</td><td>&amp;nbsp;Kent Scientific</td><td>SS-01</td><td></td></tr><tr><td>Power Lab system</td><td>AD Instruments</td><td>ML880</td><td></td></tr><tr><td>Laser Doppler flowmeter</td><td>AD Instruments</td><td>ML191</td><td></td></tr><tr><td>Heating feed back system</td><td>Harvard Appratus</td><td>72-7560</td><td></td></tr><tr><td>Vascular micro clamp</td><td>FineScience Tools</td><td>18055-03</td><td></td></tr><tr><td>Silk 5-0 suture thread</td><td>Ethicon</td><td>682G</td><td></td></tr><tr><td>Ethilon 4-0 suture thread&amp;nbsp;</td><td>Ethicon</td><td>EH6740G</td><td></td></tr></tbody></table>

an in vivo assessment of blood-brain barrier disruption in a rat model of ischemic stroke

허 혈 성 뇌졸중 vasogenic 대뇌 부 종 및 후속 기본 뇌 손상, 혈액-뇌 장벽 (BBB)의 파괴를 통해 중재 된다 이끌어 낸다. 유발 허 혈 성 뇌졸중 쥐 설립 되었고 BBB의 기능 무결성을 조사 하 비보에 모델로 사용. 허 혈 성 상해 두뇌 샘플에서 에반스 블루 (EB)의 spectrophotometric 감지 소설 치료 modalities의 개발과 연구에 대 한 신뢰할 수 있는 정당성을 제공할 수 있습니다. 이 방법은 재현성 결과 생성 하 고 특별 한 장비를 위한 필요 없이 어떤 실험실에 적용 됩니다. 여기, 선물이 쥐에서 허 혈 성 뇌졸중의 유도 따라 EB의 넘쳐 흐름의 검출에 시각 및 기술 지침.

가짜 운영 쥐의 좌 반구와 우 반구에서 EB 수준에 중요 한 차이가 없었다 (1.06 ± 0.1 µ g/g과 1.1 ± 0.09 µ g/g, 각각). 그림 2A 2B, 일시적인 허 혈의 유도에서 같이 (90 분 허 혈 / 24 h reperfusion) EB 수준에 상당한 차이 발생 (10.41 ± 0.84 µ g/g, p < 0.001) 각각에 비해 허 혈 성 쥐의 왼쪽된 반구에 운영 하는 가짜 쥐에 반구입니다. 샨 다, 이러한 결과 나타냅니다 정상적인 조건 하에서 EB 쉽게 뇌 실질에 BBB를 교차할 수 없습니다 대뇌 허 혈 성 모욕 (그림 2A와 2B) BBB의 향상 된 침투성 통해 EB의 넘쳐 흐름을 유도.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/57156/57156fig1.jpg"> 그림 1 : 표준 곡선 흡 광도 값에서 EB 농도 결정 하는 데 사용 됩니다. <a href="/files/ftp_upload/57156/57156fig1large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭 하십시오.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/57156/57156fig2.jpg"> 그림 2 : EB 넘쳐 흐름 허 혈 성 뇌졸중 후 24 시간에 의해 BBB 장애의 평가. 가짜 운영 및 허 혈 성 동물 (A)에서 두뇌의 사진. 뇌 조직 (파란색) 넘쳐 흐름 EB의 강도 lesioned에서 BBB 붕괴의 범위에서 발생합니다. EB 농도 (lesioned) 왼쪽에서 준비 하는 샘플에 가짜 운영 및 허 혈 성 동물 (B) 두뇌의 반구를 오른쪽 (n = 6, *p< 0.001 p가짜 그룹에서 왼쪽된 반구에 비해 < 0.001에 비해 동측 반구의 같은 그룹)입니다. <a href="/files/ftp_upload/57156/57156fig2large.jpg" target="_blank">이 그림의 더 큰 버전을 보려면 여기를 클릭 하십시오.</a>

Watch this Scientific Journal Video about An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke at JoVE.com

An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

허 혈 성 뇌졸중의 쥐 모델에서 뇌혈관 장애의 비보에 평가

이 절차의 전반적인 목표는 비보에 허 혈 성 뇌졸중의 쥐 모델에서 뇌혈관 장애의 평가 대 한 높은 재현 기법을 제공 하는입니다.

Ischemic stroke leads to vasogenic cerebral edema and subsequent primary brain injury, which is mediated through destruction of the blood-brain barrier ...

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Nanyang Technological University

Research

JoVE Journal

Neuroscience

An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

11.3K 조회수.  Ardabil University of Medical Sciences. 이 절차의 전반적인 목표는 비보에 허 혈 성 뇌졸중의 쥐 모델에서 뇌혈관 장애의 평가 대 한 높은 재현 기법을 제공 하는입니다.

비디오: An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

Focal Cerebral Ischemia Model by Endovascular Suture Occlusion of the Middle Cerebral Artery in the Rat

Stroke is the leading cause of disability and the third leading cause of death in adults worldwide1. In human stroke, there exists a highly variable clinical state; in the development of animal models of focal ischemia, however, achieving reproducibility of experimentally induced infarct volume is essential. The rat is a widely used animal model for stroke due to its relatively low animal husbandry costs and to the similarity of its cranial circulation to that of humans2,3. In humans, the middle cerebral artery (MCA) is most commonly affected in stroke syndromes and multiple methods of MCA occlusion (MCAO) have been described to mimic this clinical syndrome in animal models. Because recanalization commonly occurs following an acute stroke in the human, reperfusion after a period of occlusion has been included in many of these models. In this video, we demonstrate the transient endovascular suture MCAO model in the spontaneously hypertensive rat (SHR). A filament with a silicon tip coating is placed intraluminally at the MCA origin for 60 minutes, followed by reperfusion. Note that the optimal occlusion period may vary in other rat strains, such as Wistar or Sprague-Dawley. Several behavioral indicators of stroke in the rat are shown. Focal ischemia is confirmed using T2-weighted magnetic resonance images and by staining brain sections with 2,3,5-triphenyltetrazolium chloride (TTC) 24 hours after MCAO.

Surgical induction of ischemic brain damage in the rat is a widely used model for stroke research. Here we demonstrate the induction of focal cerebral ischemia by occlusion of the middle cerebral artery. Visualization of the resulting infarct by histological staining and magnetic resonance imaging is also shown.

쥐 중간 대뇌 동맥의 폐색의 Endovascular 봉합사에 의해 초점 대뇌 국소 빈혈 모델

Stroke is the leading cause of disability and the third leading cause of death in adults worldwide1. In human stroke, there exists a highly variable ...

연구

신경과학

Rat Model of Blood-brain Barrier Disruption to Allow Targeted Neurovascular Therapeutics

Endothelial cells with tight junctions along with the basement membrane and astrocyte end feet surround cerebral blood vessels to form the blood-brain barrier1. The barrier selectively excludes molecules from crossing between the blood and the brain based upon their size and charge. This function can impede the delivery of therapeutics for neurological disorders. A number of chemotherapeutic drugs, for example, will not effectively cross the blood-brain barrier to reach tumor cells2. Thus, improving the delivery of drugs across the blood-brain barrier is an area of interest.
The most prevalent methods for enhancing the delivery of drugs to the brain are direct cerebral infusion and blood-brain barrier disruption3. Direct intracerebral infusion guarantees that therapies reach the brain; however, this method has a limited ability to disperse the drug4. Blood-brain barrier disruption (BBBD) allows drugs to flow directly from the circulatory system into the brain and thus more effectively reach dispersed tumor cells. Three methods of barrier disruption include osmotic barrier disruption, pharmacological barrier disruption, and focused ultrasound with microbubbles. Osmotic disruption, pioneered by Neuwelt, uses a hypertonic solution of 25% mannitol that dehydrates the cells of the blood-brain barrier causing them to shrink and disrupt their tight junctions. Barrier disruption can also be accomplished pharmacologically with vasoactive compounds such as histamine5 and bradykinin6. This method, however, is selective primarily for the brain-tumor barrier7. Additionally, RMP-7, an analog of the peptide bradykinin, was found to be inferior when compared head-to-head with osmotic BBBD with 25% mannitol8. Another method, focused ultrasound (FUS) in conjunction with microbubble ultrasound contrast agents, has also been shown to reversibly open the blood-brain barrier9. In comparison to FUS, though, 25% mannitol has a longer history of safety in human patients that makes it a proven tool for translational research10-12.
In order to accomplish BBBD, mannitol must be delivered at a high rate directly into the brain's arterial circulation. In humans, an endovascular catheter is guided to the brain where rapid, direct flow can be accomplished. This protocol models human BBBD as closely as possible. Following a cut-down to the bifurcation of the common carotid artery, a catheter is inserted retrograde into the ECA and used to deliver mannitol directly into the internal carotid artery (ICA) circulation. Propofol and N2O anesthesia are used for their ability to maximize the effectiveness of barrier disruption13. If executed properly, this procedure has the ability to safely, effectively, and reversibly open the blood-brain barrier and improve the delivery of drugs that do not ordinarily reach the brain 8,13,14.

Blood-brain barrier disruption aids the delivery of certain drugs to the brain. Mannitol delivered intra-arterially shrinks cells surrounding blood vessels in order to physically disrupt the barrier.

혈액 - 뇌 장벽 장애의 쥐 모델 Neurovascular 치료학 타겟이 분명한 허용하려면

Endothelial cells with tight junctions along with the basement membrane and astrocyte end feet surround cerebral blood vessels to form the blood-brain ...

의학

Setting-up an In Vitro Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport

The blood brain barrier (BBB) specifically regulates molecular and cellular flux between the blood and the nervous tissue. Our aim was to develop and characterize a highly reproducible rat syngeneic in vitro model of the BBB using co-cultures of primary rat brain endothelial cells (RBEC) and astrocytes to study receptors involved in transcytosis across the endothelial cell monolayer. Astrocytes were isolated by mechanical dissection following trypsin digestion and were frozen for later co-culture. RBEC were isolated from 5-week-old rat cortices. The brains were cleaned of meninges and white matter, and mechanically dissociated following enzymatic digestion. Thereafter, the tissue homogenate was centrifuged in bovine serum albumin to separate vessel fragments from nervous tissue. The vessel fragments underwent a second enzymatic digestion to free endothelial cells from their extracellular matrix. The remaining contaminating cells such as pericytes were further eliminated by plating the microvessel fragments in puromycin-containing medium. They were then passaged onto filters for co-culture with astrocytes grown on the bottom of the wells. RBEC expressed high levels of tight junction (TJ) proteins such as occludin, claudin-5 and ZO-1 with a typical localization at the cell borders. The transendothelial electrical resistance (TEER) of brain endothelial monolayers, indicating the tightness of TJs reached 300 ohm&#183;cm2 on average. The endothelial permeability coefficients (Pe) for lucifer yellow (LY) was highly reproducible with an average of 0.26 &#177;&#160;0.11 x 10-3 cm/min. Brain endothelial cells organized in monolayers expressed the efflux transporter P-glycoprotein (P-gp), showed a polarized transport of rhodamine 123, a ligand for P-gp, and showed specific transport of transferrin-Cy3 and DiILDL across the endothelial cell monolayer. In conclusion, we provide a protocol for setting up an in vitro BBB model that is highly reproducible due to the quality assurance methods, and that is suitable for research on BBB transporters and receptors.

Setting-up an In Vitro Model of Rat Blood-brain Barrier BBB: A Focus on BBB Impermeability and Receptor-mediated Transport

The aim of the present study was to validate the reproducibility of an in vitro BBB model involving a rat syngeneic co-culture of endothelial cells and astrocytes. The endothelial cell monolayer presented high TEER and low LY permeability. Expression of specific TJ proteins, functional responses to inflammation and functionality of transporters and receptors were assessed.

설정 - 업<em&gt; 체외</em쥐 뇌 혈관 벽의&gt; 모델 (BBB)​​ : BBB 불 침투성 및 수용체 매개 운송에 초점

The blood brain barrier (BBB) specifically regulates molecular and cellular flux between the blood and the nervous tissue. Our aim was to develop and ...

A Triple Primary Cell Culture Model of the Human Blood-Brain Barrier for Studying Ischemic Stroke In Vitro

Ischemic stroke is a major cause of death and disability worldwide with limited therapeutic options. The neuropathology of ischemic stroke is characterized by an interruption in blood supply to the brain leading to cell death and cognitive dysfunction. During and after ischemic stroke, blood-brain barrier (BBB) dysfunction facilitates injury progression and contributes to poor patient recovery. Current BBB models primarily include endothelial monocultures and double co-cultures with either astrocytes or pericytes.
Such models lack the ability to fully imitate a dynamic brain microenvironment, which is essential for cell-to-cell communication. Additionally, commonly used BBB models often contain immortalized human endothelial cells or animal-derived (rodent, porcine, or bovine) cell cultures that pose translational limitations. This paper describes a novel well-insert-based BBB model containing only primary human cells (brain microvascular endothelial cells, astrocytes, and brain vascular pericytes) enabling the investigation of ischemic brain injury in vitro. 
The effects of oxygen-glucose deprivation (OGD) on barrier integrity were assessed by passive permeability, transendothelial electrical resistance (TEER) measurements,and direct visualization of hypoxic cells. The presented protocol offers a distinct advantage inmimicking the intercellular environment of the BBB in vivo, serving as a more realistic in vitro BBB model for developing new therapeutic strategies in the setting of ischemic brain injury.

A Triple Primary Cell Culture Model of the Human Blood-Brain Barrier for Studying Ischemic Stroke In Vitro

Here, we describe the method for establishing a triple cell culture model of the blood-brain barrier based on primary human brain microvascular endothelial cells, astrocytes, and pericytes. This multicellular model is suitable for studies of neurovascular unit dysfunction during ischemic stroke in vitro or for the screening of drug candidates.

시험관 내 허혈성 뇌졸중 연구를 위한 인간 혈액-뇌 장벽의 삼중 일차 세포 배양 모델

Ischemic stroke is a major cause of death and disability worldwide with limited therapeutic options. The neuropathology of ischemic stroke is ...

생화학

Quantification of Neurovascular Protection Following Repetitive Hypoxic Preconditioning and Transient Middle Cerebral Artery Occlusion in Mice

Experimental animal models of stroke are invaluable tools for understanding stroke pathology and developing more effective treatment strategies. A 2 week protocol for repetitive hypoxic preconditioning (RHP) induces long-term protection against central nervous system (CNS) injury in a mouse model of focal ischemic stroke. RHP consists of 9 stochastic exposures to hypoxia that vary in both duration (2 or 4 hr) and intensity (8% and 11% O2). RHP reduces infarct volumes, blood-brain barrier (BBB) disruption, and the post-stroke inflammatory response for weeks following the last exposure to hypoxia, suggesting a long-term induction of an endogenous CNS-protective phenotype. The methodology for the dual quantification of infarct volume and BBB disruption is effective in assessing neurovascular protection in mice with RHP or other putative neuroprotectants. Adult male Swiss Webster mice were preconditioned by RHP or duration-equivalent exposures to 21% O2 (i.e. room air). A 60 min transient middle cerebral artery occlusion (tMCAo) was induced 2 weeks following the last hypoxic exposure. Both the occlusion and reperfusion were confirmed by transcranial laser Doppler flowmetry. Twenty-two hr after reperfusion, Evans Blue (EB) was intravenously administered through a tail vein injection. 2 hr later, animals were sacrificed by isoflurane overdose and brain sections were stained with 2,3,5- triphenyltetrazolium chloride (TTC). Infarcts volumes were then quantified. Next, EB was extracted from the tissue over 48 hr to determine BBB disruption after tMCAo. In summary, RHP is a simple protocol that can be replicated, with minimal cost, to induce long-term endogenous neurovascular protection from stroke injury in mice, with the translational potential for other CNS-based and systemic pro-inflammatory disease states.

This protocol describes repetitive hypoxic preconditioning, or brief exposures to systemic hypoxia that reduce infarct volumes and blood-brain barrier disruption following transient middle cerebral artery occlusion in mice. It also details dual quantification of infarct volume and blood-brain barrier disruption after stroke to assess the efficacy of neurovascular protection.

마우스의 반복 저산소 전처리 및 과도 중간 대뇌 동맥 폐쇄 후 신경 혈관 보호의 정량화

Experimental animal models of stroke are invaluable tools for understanding stroke pathology and developing more effective treatment strategies. A 2 week ...

Assessment of Blood-brain Barrier Permeability by Intravenous Infusion of FITC-labeled Albumin in a Mouse Model of Neurodegenerative Disease

Disruption of blood-brain barrier (BBB) integrity is a common feature for different neurological and neurodegenerative diseases. Although the interplay between perturbed BBB homeostasis and the pathogenesis of brain disorders needs further investigation, the development and validation of a reliable procedure to accurately detect BBB alterations may be crucial and represent a useful tool for potentially predicting disease progression and developing targeted therapeutic strategies.
Here, we present an easy and efficient procedure for evaluating BBB leakage in a neurodegenerative condition like that occurring in a preclinical mouse model of Huntington disease, in which defects in the permeability of BBB are clearly detectable precociously in the disease. Specifically, the high molecular weight fluorescein isothiocyanate labelled (FITC)-albumin, which is able to cross the BBB only when the latter is impaired, is acutely infused into a mouse jugular vein and its distribution in the vascular or parenchymal districts is then determined by fluorescence microscopy.
Accumulation of green fluorescent-albumin in the brain parenchyma functions as an index of aberrant BBB permeability and, when quantitated by using Image J processing software, is reported as Green Fluorescence Intensity.

In this study, we present an easy and efficient procedure for evaluating the disruption of the blood-brain barrier under neurodegenerative conditions. To achieve our objective, we infused high molecular weight fluorescein isothiocyanate labelled (FITC)-albumin into mouse jugular vein and evaluated its leakage into the brain parenchyma by fluorescence microscopy.

FITC 표시 된 알 부 민 신경 질환의 마우스 모델에서의 정 맥 주입에 의해 혈액-뇌 장벽 침투성의 평가

Disruption of blood-brain barrier (BBB) integrity is a common feature for different neurological and neurodegenerative diseases. Although the interplay ...

An In Vivo Blood-brain Barrier Permeability Assay in Mice Using Fluorescently Labeled Tracers

Blood-brain barrier (BBB) is a specialized barrier that protects the brain microenvironment from toxins and pathogens in the circulation and maintains brain homeostasis. The principal sites of the barrier are endothelial cells of the brain capillaries whose barrier function results from tight intercellular junctions and efflux transporters expressed on the plasma membrane. This function is regulated by pericytes and astrocytes that together form the neurovascular unit (NVU). Several neurological diseases such as stroke, Alzheimer&#39;s disease (AD), brain tumors are associated with an impaired BBB function. Assessment of the BBB permeability is therefore crucial in evaluating the severity of the neurological disease and the success of the treatment strategies employed.
We present here a simple yet robust permeability assay that have been successfully applied to several mouse models both, genetic and experimental. The method is highly quantitative and objective in comparison to the tracer fluorescence analysis by microscopy that is commonly applied. In this method, mice are injected intraperitoneally with a mix of aqueous inert fluorescent tracers followed by anesthetizing the mice. Cardiac perfusion of the animals is performed prior to harvesting brain, kidneys or other organs. Organs are homogenized and centrifuged followed by fluorescence measurement from the supernatant. Blood drawn from the cardiac puncture just before perfusion serves for normalization purpose to the vascular compartment. The tissue fluorescence is normalized to the wet weight and serum fluorescence to obtain a quantitative tracer permeability index. For additional confirmation, the contralateral hemi-brain preserved for immunohistochemistry can be utilized for tracer fluorescence visualization purposes.

An In Vivo Blood-brain Barrier Permeability Assay in Mice Using Fluorescently Labeled Tracers

Here we present a mouse brain vascular permeability assay using intraperitoneal injection of fluorescent tracers followed by perfusion that is applicable to animal models of blood-brain barrier dysfunction. One hemi-brain is used for assessing permeability quantitatively and the other for tracer visualization/immunostaining. The procedure takes 5 - 6 h for 10 mice.

Vivo에서 혈액-뇌 장벽 침투성 분석 결과 사용 하 여 마우스에 붙일 추적기 표시

Blood-brain barrier (BBB) is a specialized barrier that protects the brain microenvironment from toxins and pathogens in the circulation and maintains ...

Measuring Post-Stroke Cerebral Edema, Infarct Zone and Blood-Brain Barrier Breakdown in a Single Set of Rodent Brain Samples

One of the most common causes of morbidity and mortality worldwide is ischemic stroke. Historically, an animal model used to stimulate ischemic stroke involves middle cerebral artery occlusion (MCAO). Infarct zone, brain edema and blood-brain barrier (BBB) breakdown are measured as parameters that reflect the extent of brain injury after MCAO. A significant limitation to this method is that these measurements are normally obtained in different rat brain samples, leading to ethical and financial burdens due to the large number of rats that need to be euthanized for an appropriate sample size. Here we present a method to accurately assess brain injury following MCAO by measuring infarct zone, brain edema and BBB permeability in the same set of rat brains. This novel technique provides a more efficient way to evaluate the pathophysiology of stroke.

This protocol describes a novel technique of measuring the three most important parameters of ischemic brain injury on the same set of rodent brain samples. Using only one brain sample is highly advantageous in terms of ethical and economic costs.

설치류 뇌 샘플의 단일 세트에서 뇌졸중 후 뇌 부종, 경색 영역 및 혈액 뇌 장벽 고장 측정

One of the most common causes of morbidity and mortality worldwide is ischemic stroke. Historically, an animal model used to stimulate ischemic stroke ...

Real-Time Monitoring and Modulation of Blood Pressure in a Rabbit Model of Ischemic Stroke

Control of blood pressure, in terms of both absolute values and its variability, affects outcomes in ischemic stroke patients. However, it remains challenging to identify the mechanisms that lead to poor outcomes or evaluate measures by which these effects can be mitigated because of the prohibitive limitations inherent to human data. In such cases, animal models can be utilized to conduct rigorous and reproducible evaluations of diseases. Here we report refinement of a previously described model of ischemic stroke in rabbits that is augmented with continuous blood pressure recording to assess the impacts of modulation on blood pressure. Under general anesthesia, femoral arteries are exposed through surgical cutdowns to place arterial sheaths bilaterally. Under fluoroscopic visualization and roadmap guidance, a microcatheter is advanced into an artery of the posterior circulation of the brain. An angiogram is performed by injecting the contralateral vertebral artery to confirm occlusion of the target artery. With the occlusive catheter remaining in position for a fixed duration, blood pressure is continuously recorded to allow for tight titration of blood pressure manipulations, whether through mechanical or pharmacological means. At the completion of the occlusion interval, the microcatheter is removed, and the animal is maintained under general anesthesia for a prescribed length of reperfusion. For acute studies, the animal is then euthanized and decapitated. The brain is harvested and processed to measure the infarct volume under light microscopy and further assessed with various histopathological stains or spatial transcriptomic analysis. This protocol provides a reproducible model that can be utilized for more thorough preclinical studies on the effects of blood pressure parameters during ischemic stroke. It also facilitates effective preclinical evaluation of novel neuroprotective interventions that might improve care for ischemic stroke patients.

Continuous arterial blood pressure recording allows the investigation of impacts of various hemodynamic parameters. This report demonstrates the application of continuous arterial blood pressure monitoring in a large animal model of ischemic stroke for determination of stroke pathophysiology, impact of different hemodynamic factors, and the assessment of novel treatment approaches.

허혈성 뇌졸중의 토끼 모델에서 혈압의 실시간 모니터링 및 조절

Control of blood pressure, in terms of both absolute values and its variability, affects outcomes in ischemic stroke patients. However, it remains ...

<meta charset="utf8"></head><body>쥐 MCAO 뇌졸중 모델의 세포 사멸을 연구하기 위한 면역형광 염색

Evaluating Cell Death Signaling by Immunofluorescence in a Rat Model of Ischemic Stroke

Stroke is a leading cause of death and disability worldwide. Most cases of stroke are ischemic and result from the occlusion of the middle cerebral artery (MCA). Current pharmacological approaches for the treatment of ischemic stroke are limited; therefore, novel therapies providing effective neuroprotection against ischemic injury following stroke are urgently needed. In the brain tissue following ischemic stroke, the area of the ischemic penumbra is salvageable but at risk of progressing to irreversible damage. The penumbral area surrounding the infarcted core is a conceptual target for neuroprotection. Since the blood flow is partially maintained in the penumbral area, neuronal and non-neuronal cells in this area transiently survive after stroke, and these cells that are still viable may be rescued by appropriate medical interventions. Understanding the pathophysiology of the penumbra is important in the development of neuroprotective therapies because the cell death pathways activated in the ischemic penumbra may indicate therapeutic targets, such as RUNX1 and cathepsins. These protein targets functioning as mediators of programmed cell death can be further exploited in translational research. With moderate size and similarity to the human brain, the in vivo rat model of MCA occlusion (MCAO) mimics the human ischemic stroke and offers an applicable tool to investigate the penumbral pathology, examine the cell death signaling, and evaluate the effects of potential targets in the context of MCAO. Here, we describe how to induce MCAO in rats and how to perform immunofluorescence staining for the detection of cell death signaling in the rat brain following MCAO.

<meta charset="utf8"></head><body>허혈성 뇌졸중의 쥐 모델에서 면역형광에 의한 세포 사멸 신호 평가

Rat in vivo models are indispensable tools to investigate the pathological mechanisms and therapeutic targets for ischemic stroke. This work will describe performing immunofluorescence staining in infarcted brain slices following middle cerebral artery occlusion (MCAO) in rats.

허혈성 뇌졸중의 쥐 모델에서 면역형광에 의한 세포 사멸 신호 평가

Stroke is a leading cause of death and disability worldwide. Most cases of stroke are ischemic and result from the occlusion of the middle cerebral artery ...