Name: isolation of atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction
Uploaded: 2018-07-26T14:00:00
Description: Watch this Scientific Journal Video about Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction at JoVE.com

이 연구는 (심장 혈관 연구, db 위한 독일 센터), DZHK에 의해 지원 되었다 EKFS (Else Kröner Fresenius 재단, F.H.), 고 (독일 교육 및 연구), BMBF BIH Charité 임상 과학자 프로그램 투자에 의해 의해 Charité-Universitätsmedizin 베를린, 베를린 건강 연구소 (F.H.).

모든 실험 (TVA T0060/15 및 T0003 15) 지역 윤리 위원회에 의해 승인 하 고 관리 및 실험 동물 사용 (건강의 국가 학회, 미국)에 대 한 지침과 일치 하 여 수행 했다.
참고: 절차의 단순화 된 순서도 그림 1에 표시 됩니다.
1입니다. prearrangements
<ol><li>표 1에 따라 버퍼를 준비 합니다.</li>
</ol><table border="1" fo:keep-together.within-page="1" ...

<ol>
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여기, 우리는 먼저 메츠 관련 HFpEF 보여 주는 심 방 개장22표시의 쥐 모델에서 단일 셀 Acm의 격리에 대 한 프로토콜을 설명 합니다. 절차는 수술 준비로 점점 어려운 대동맥의 cannulation 과도 한 지방 조직을 만들 수 있는 유일 하 게 도전. 표 2 주소 격리 절차의 가장 일반적인 문제 문제 해결 가이드 제공.
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메츠 당뇨병 mellitus 타입-2와 심혈 관 질환 위험 요인의 클러스터를 설명 하 고 포함 하는 증가 동맥 혈압, dyslipidemia (트리 글리세라이드를 제기 하 고 고밀도 지 단백 콜레스테롤을 낮 췄 다), 금식 증가 포도 당, 그리고 중앙 비만1. 메츠의 전세계 보급 되며 지속적으로 상승2, 25-30%로 추정 된다. HFpEF은 메츠와 자주 관련 된 이기종 임상 증후군. HFpEF 및 그것의 이전 단계 (즉, 고혈압 성 심장 질환) 중 심장 개장은 또한 동반 atria3을 개장 하는. 감소 된 수축 성 기능 및 구조적인 변화는 왼쪽된 아 트리 움의 사망률 증가, 심 방 세 동, 및 새로운 발병 심장 마비4와 연결 되었습니다. 개장 하는 심 방 이온 채널 기능, 캘리포니아2 + 항상성, 심 방 구조, 섬유 아 세포 활성화 및 조직 섬유 증5에 변화에 의해 특징입니다. 왼쪽에 메츠 관련 HFpEF의 기본 병 적인 메커니즘 아직도 제대로 이해 하 고 더 깊이 있는 조사를 필요로 개장 하는 심 방. 동물 모델은 유용한 도구가 될 고 심 방 cardiomyopathies6,7,,89의 분야에서 많은 발전을 입증 했다.
격리 된 단일 셀 cardiomyocytes와 연구 자주 확증 vivo에서 조사 결과 보완 하는 데 사용 됩니다. 절연 및 잠재적인 후속 세포 배양, 신호 경로, 이온 채널 전류, 및 흥분-수축-커플링의 조사에 대 한 수 있습니다. 생리 적인 조건 하에서 cardiomyocytes 증식 하지 않습니다. Atrial natriuretic 요소의 transcriptional 규제 시퀀스와는 유인원 바이러스 40 큰 T 항 원 유전자 변형 쥐에서 사이의 융합 AT-110라는 첫 번째 불멸 하 게 Acm의 창조로 이끌어 냈다. AT-1 셀의 추가 개발 HL-1 셀만 순차적으로 passaged 수 없습니다 하지만 또한 저절로11계약에 상승을 했다. 그러나 그들은,, 구조와 기능 차이 덜 조직 된 열 대권 외의, hyperpolarization-활성화 안쪽으로 현재12myofibrils11, 개발의 높은 발생 등 갓 고립 된 세포에 비해 표시. 쥐와 쥐 모델의 다양 한에서 심 실 cardiomyocytes (VCM)의 절연은 잘 설립된13,14,15,16,,1718 , 19. 일반적으로, 삭제는 Langendorff 장치에 탑재 마음과 retrogradely 끼얹는다는 캘리포니아2 +-collagenases 및 프로 테아 제와 같은 소화 효소를 포함 하는 무료 버퍼. 칼슘은 다음 생리 적 조건에 단계적 방식으로 재. 그러나, Acm의 고립에 전용 프로토콜 사용할 수 있는20,21, 증가 증과 차이 압력 관련 있더라도 개장 하는 심 질환 모델에서 그들의 유용성은 제한 됩니다.
이 문서에서는, 우리는 심 방 개장 (즉, 특히 메츠 관련 HFpEF의 ZFS1 쥐 모델에 대 한)22를 나타내는 동물에서 심 방 단일 셀 cardiomyocytes의 격리에 대 한 프로토콜을 실행 했다. 기존 격리 프로토콜 최적화 하 고 제어 하 고 형태학 상으로 그리고 기능적으로 그대로 cardiomyocytes의 높은 수율으로 이어지는 심장 충 치의 intraluminal 압력 수정 간단 하 고, 주문 품 장치에 의해 보완 했다. 다음 프로토콜 단계별 가이드와 함께 연구원, 주문 품 장비, 솔루션의 목록 뿐만 아니라 포괄적인 문제 해결 가이드에 대 한 자세한 설명을 제공합니다.

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			<td height="20" style="height:20px;width:281px;">ZSF-1 Obese rat</td>
			<td style="width:292px;">Charles River Laboratories, Inc.</td>
			<td style="width:136px;"></td>
			<td style="width:227px;">21 weeks old</td>
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			<td height="20" style="height:20px;width:281px;">Fine Iris Scissors</td>
			<td style="width:292px;">Fine Science Tools GmbH</td>
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</table>

isolation of atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction

이 문서에서는, 우리는 변화 증후군 (메츠)의 쥐 모델에서 단일 셀 심 방 cardiomyocytes (Acm)의 분리에 대 한 최적화, Langendorff 기반 절차 설명-관련 보존된 방출 분수 (HFpEF)와 함께 심장 마비. 메츠 관련 HFpEF의 보급은 상승 하 고, 그리고 심 cardiomyopathies 개장 하는 심 방 및 심 방 세 동와 관련 된 관련성이 높은 임상으로 심 방 개장 사망률의 독립적인 예측 인자. 격리 된 단일 셀 cardiomyocytes와 연구 자주 확증 vivo에서 조사 결과 보완 하는 데 사용 됩니다. 순환 선박 rarefication 및 조직 간 질 성 섬유 증이이 질환의 동물 모델에서 Acm의 성공적인 단일 셀 격리에 대 한 잠재적으로 제한 요소를 포즈.
우리가 실질적으로 형태학 상으로 그리고 기능적으로 그대로 Acm의 수확량을 증가 수동으로 격리 절차 동안 심장 구멍의 intraluminal 압력을 조절 하는 장치를 사용 하 여이 문제를 해결 했습니다. 인수 세포 세포 배양 및 기능성 칼슘 이미징 (즉, 흥분-수축-커플링) 등 다양 한 실험의 다양 한에서 사용할 수 있습니다.
우리 연구원 단계별 프로토콜, 최적화 된 솔루션, 필요한 장비 및 포괄적인 문제 해결 가이드를 준비 하는 철저 한 지침의 목록 제공 한다. 절차의 초기 구현을 오히려 어려울 수도 있습니다, 그러나 성공적인 적응 메츠 관련 광범위 한 실험에 대 한 HFpEF의 쥐 모델에서 최신의 ACM 격리를 수행 하는 독자 수 있게 됩니다.

나이의 21 주에 가능한 Acm (6.1 단계에 설명 된 대로 추정), 칼슘 다시 적응 (단계 5.4-5.7), 후의 60-90% (그림 4A)이 메서드에서 ZSF-1 비만 쥐에서 분리 수 있습니다. 쥐, Acm 다르고 VCMs24,25에 비해 더 많은 이기종 표현 형 특징입니다. 그림 4B 와 보존된 막 sarcomere 구조를 기능적으로 통합 셀의 ...

Watch this Scientific Journal Video about Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction at JoVE.com

Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction

심 방 Cardiomyocytes 보존된 방출 분수와 함께 대사 증후군 관련 심장 마비의 쥐 모델에서의 격리

여기, 우리가 보존된 방출 분수와 함께 대사 증후군 관련 심장 마비의 쥐 모델에서 단일 셀 심 방 cardiomyocytes의 격리에 대 한 최적화, Langendorff 기반 절차를 설명합니다. 심장 구멍의 intraluminal 압력의 수동 조절 기능 그대로 myocytes 흥분-수축-결합 연구에 대 한 적합 한를 구현 됩니다.

In this article, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes (ACMs) from a rat model of ...

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