Name: measurement of the pressure-volume curve in mouse lungs
Uploaded: 2015-01-27T15:00:00.000+00:00
Duration: 9 min 49 s
Description: Watch this Scientific Journal Video about Measurement of the Pressure-volume Curve in Mouse Lungs at JoVE.com

This work has been supported by NIH HL-1034.

존스 홉킨스 대학 동물 관리 및 사용위원회는 모든 동물 프로토콜을 승인했다. 1. 장비 , PV 곡선을 측정 할 준비가 설정 한 복합 시스템은 그림 1에 표시됩니다. <ol><li> 볼륨 측정 : <ol><li> 사용자가 신속하게 압력 한계에 도달 한 후 펌프를 역방향 있도록 스위치 시린지 펌프를 이용하여 팽창과 수축의 일정한 속도를 생성한다. 마우스 PV 곡선의 경우, 공기의 3 ㎖에서 설정 한 초기 볼륨 (이전에 인플레이션)를 매우 가볍게 기름을 바른 5 mL 유리 주사기를 사용합니다. 3 ㎖ 거의 모든 마우스 PV 곡선의 볼륨을 측정 할 수있을만큼 충분히 크다. </li><li> 이동 주사기 플런저에 연결된 작은 센서 막대와, 펌프 하우징에 선형 미분 변압기 부착함으로써 펌프에 의해 전달되는 부피를 측정한다. 주 : 경험적 수단은 시스템의 가스 압축을 보정하기 위해 PV 세제곱에 따라 설명한다RVE 기록부. </li></ol></li><li> 압력 측정 : <ol><li> 0-60cm H 2 O (0-1 PSI)의 범위와 표준 저렴한 압력 게이지를 사용합니다. </li></ol></li><li> 측정 기록 : <ol><li> PV 곡선은 XY 기능 (예를 들어, PowerLab의)와 모든 디지털 레코더를 사용하여 기록합니다. 볼륨 보정 신호 및 PV 곡선을 그래프로 위해서는 경폐 압력 (PTP)을 기록하는 또 다른 채널을 녹화하기 위해 하나의 채널을 설정한다. 압력을 측정하는 주 PowerLab의 다리에 연결 전치 증폭기를 사용한다. 00-40 센티미터 H 2 O로부터 압력 채널을 조정하고, 0-3 ml의 부피에서 채널을 조정. </li></ol></li></ol> 가스 압축 2. 수정 주 :이에 중요한 초기 단계이다 압력 증가, 가스 부피가 감소하고, 마우스로 전달되는 공기의 양으로서 따라서 SYR의 변위보다 더욱 적게되기 때문에, 설정디트로이트 배럴. <ol><li> 폐에 태양 광 발전 시스템을 연결하는 꼭지를 닫습니다, 그래서 가스 시스템을 떠날 수 있습니다. 주입을 시작하고 레코더의 보정 볼륨 채널은 약 40cm의 H 2 O에 압력이 증가함에 따라 측정 가능한 변화를 보여준다 경우 관찰 다음 단계에서와 같이 그렇다면, 올바른합니다. <ol><li> 경험적으로 플런저 변위 측정에서 빼서 가스 압축에 대한 올바른 (즉, 보정 볼륨) 인플레이션 압력에 비례하는 용어입니다. 볼륨 신호 마이너스 계수 배의 압력을 표시 할 PowerLab의 채널 (라고 VC)에이 작업을 수행합니다. </li><li> 방정식의 계수를 결정합니다. 첫째, 초기 추측에 기록 차트를 켜고 펌프를 시작합니다. 팽창 튜브 밀봉되기 때문에, 압력은 0-40 센티미터 H 2 O.에서 상승 채널 (VC)가 제로 보이게 압력 계수 승산기를 조정 이 위 또는 아래로되면, 단순히까지 보정 계수를 조정이 압력 범위에서 플랫 유지됩니다. 주사기의 체적을 시작 같은 3 ㎖가 변경되지 않은 경우이 보정 계수는 항상 동일하다. </li></ol></li></ol> 마우스 3. 실험 테스트 <ol><li> 마우스의 PV 곡선의 측정을위한 절차. 모든 동물 프로토콜은 존스 홉킨스 대학 동물 관리 및 사용위원회에 의해 승인되었습니다. <ol><li> 케타민 (90 ㎎ / ㎏)과 자일 라진 (15 ㎎ / ㎏)와 (나이 6-12 주에 C57BL / 6 마​​우스) 쥐를 마취하고, 반사 운동의 부재로 마취를 확인합니다. 주 : PV 곡선은 10 분 미만에 마취 된 마우스에서 완료되고 단말 절차 수있다. </li><li> 18 G 스텁 바늘 캐 뉼러와 마우스를 Tracheostomize. 다음 스텁 바늘을 삽입 할 수있는 기관, 작은 슬릿을, 기관을 덮는 피부에 작은 절개를 만드는 기관을 배치함으로써이 작업을 수행합니다. 실로 매서 캐 뉼러를 고정합니다. </li><li> 호흡 하나에 마우스를 허용적어도 4 분 00 % 산소. 이 가방이나 명목상 150 호흡 / 분에서 0.2 ml의 호흡량 설정 인공 호흡기와자가 호흡을 통해 할 수있다. </li><li> 기관 캐뉼라를 종료하고 마우스 3-4 분 모든 산소를 흡수 할 수 있습니다. 이 산소 흡수 과정은 동물의 폐 및 (11)의 거의 전체에서 탈기 죽음을 초래한다. ECG 전극 또는 직접 관찰과 심장 박동의 정지를 측정하여 마우스의 죽음을 확인합니다. </li><li> 폐의 탈기가 완료되고 폐 용적이 제로가되면, / 분으로 3 ㎖의 속도로 시린지 펌프에 실내 공기로 폐를 팽창시키기 시작한다. 디지털 레코더에 압접 흔적을 모니터하고 35cm의 H 2 O에 도달 할 때, 펌프를 역방향. </li><li> 압력이기도가 더 부피 감소를 방지 폐포에서 공기를 포집, 붕괴되는 시간에 의해 부정적인 10cm의 H 2 O를, 도달 할 때까지 디플레이션 곡선을 따르십시오. 즉시 t 역그는 붕괴기도가 여는 폐 reinflate 할 수 있도록, 다시 펌프. 이 이종 개방이 2 인플레이션의 초기 부분에서 잡음이 찾고 인플레이션 사지에 의해 일반적으로 알 수있다. </li><li> 압력이 다시 35cm의 H 2 O에 도달하면, 펌프 방향을 반전하고,이 두번째 디플레이션 사지 0cm의 H 2 O에 도달 할 때까지 폐 수축 계속 그런 다음 펌프를 중지합니다. </li><li> 압력 및 흐름과 PV 곡선의 PowerLab의 차트 기록을 볼 수 있습니다. 그런 다음 다른 폐 병리 발생하는 폐 실질 조직의 표현형 변화를 감지하기 위해 PV 곡선을 분석 할 수 있습니다. </li></ol></li></ol>

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None of the authors have any financial interests that would be in conflict with the material presented in this paper.

본 논문에서는 간단한 재현 방법은 마우스의 표현형 폐 탄성, 전체 폐 PV 곡선의 고전적인 방법을 측정하는 기술되었다. 이러한 곡선은 폐 계면 활성제의 발견과 폐 안정성을 제공의 중요성에 쓸모 있었다. 여기가 PV 곡선 성인 마우스 폐에서 폐 탄성 관련된 여러 변수를 측정하는 수단을 제공하기에 유용한 방법을 도시한다. 마우스 폐의 병리학 적 변화를 생성하기 위해 일반적으로 사용되는 두 ?...

이제 마우스 폐 질환의 다양한 일차 동물 모델이다. 폐기종 또는 섬유증의 모델에서, 필수적인 표현형 변화는 가장 폐 탄성의 변화를 측정함으로써 평가된다. 탄성을 측정하는 방법은 여러 가지가 있겠지만, 고전적인 방법은 그 총 폐 용량 (TLC)에 잔여 볼륨 (RV)에서 측정 된 전체 압력 - 볼륨 (PV) 곡선. 이 측정은 거의 백년 1-3 거슬러 올라가는 거의 모든 포유류에서 성인 폐에 만들어졌다. 이러한 PV 곡선 태아 폐 개발 4-7 폐 계면 활성제의 기능의 발견 및 이해에 중요한 역할을했다. 폐의 표현형의 측정치로 PV 곡선의 중요성에도 불구하고,이 측정을 수행 할 수있는 표준화 된 방법이 없었다. 그것은 팽창 및 이산 단계와 폐 (각 후 평형에 대한 변수 대기 시간)을 배출하기 또는 펌프를 간단하게 수행되었음을지속적으로 팽창하고 폐를 수축 할 수 있습니다. PV 곡선들은 제로 및 일부 사용자 정의 폐활량 사이의 체적 범위에서 수행되지만 다른 실험실에 의해보고 된 각 압력 볼륨 루프의 지속 시간은 5.6 시간 (2)에 몇 초 (8)로부터 변화, 매우 변수였다. 일부 연구자들은 정적 또는 quasistatic 등이 총 폐 PV 곡선을 참조하지만, 이러한 작은 통찰력을 제공 질적이며, 그들은 여기에 사용되지 않습니다. 또한, PV 곡선 널리이 폐에서의 구조적 변화의 거시적 효과에 관한 유용한 정보를 제공 할 수 있다는 사실에도 불구하고, 마우스에서보고되지 않았다. 몇 가지 문제가 포함 PV 곡선 획득의 가변성에 성공 : 인플레이션과 디플레이션 1) 비율; 2) 인플레이션과 디플레이션에 대한 압력 여행; 3) 수단은 절대 폐 량 측정을 결정한다. 여기에 방법 존재에서, 3 ㎖의 속도 / 분 compromis로 선정되었다대규모 코호트 연구 특히, 측정 비실용적로서 전자, 너무 짧게있는 것은 통풍 너무 느린하지와 관련된 동적 탄성을 반영합니다. C57BL / 6 마우스의 건강한 공칭 전체 폐 용량 1.2 ㎖의 9 정도이기 때문에,이 속도는 전형적으로는 두 개의 완전한 PV는 약 1.5 분에서 수행 될 수 있도록 폐쇄 루프. PV 곡선이보고 된 확장 된 문헌에서 사용 피크 인플레이션 압력은 40 센티미터 H 2 O로 낮은 20에서 다양한 매우 변수왔다 이 변동의 일부는 종에 관련 될 수 있지만, PV 곡선의 상한 압력 한계를 설정하는 기본 목표 총 폐활량 (TLC) 또는 최대 폐 용적 폐 팽창한다. 인간의 TLC는 개인이 만들 수있는 최대한의 자발적인 노력에 의해 정의된다, 그러나 불행하게도이 모든 동물 모델에서 중복 될 수 없다. 따라서, 실험 PV 곡선의 최대 볼륨을 억제하다임의로 연구자가 설정 한 최대 압력에 의해 채굴. 목표는 PV 곡선이 평탄 압력을 설정할 수 있지만, 불행히도 포유류 폐 PV 곡선의 팽창 사지 결코 평탄한. 그래서 대부분의 연구자들은 일반적으로 인플레이션 곡선이 실질적으로 평평하게 시작 압력, 30cm의 H 2 O를 설정 마우스 그러나 PV 곡선은 팽창 사지 이중 혹으로 더 복잡하며,이 팽창 사지 종종 여전히 30cm의 H 2 O (10)에서 급격하게 상승되는 경우, 그래서 30 대 좋은 종점 아니다 PV 곡선. 이러한 이유로, 우리는 우리가 검토 한 모든 균주의 팽창 사지가 평평하게 시작하는 압력이 마우스 PV 곡선의 압력 한계, 같은 35cm의 H 2 O를 사용합니다. PV 곡선 자체가 매우 비선형이기 때문에, PV 루프의 외관은 곡선이 어디서 시작 부피​​에 의존 할 것이다. 일부 상용 팬은 F에서 시작하여, 사용자가 큰 PV 루프를 수행 할 수RC하지만 FRC 볼륨이 미지 인 경우, 이들 변경 단순히 볼륨 개시까지 변화로부터 발생할 수 있기 때문에, 임의의 병리와 같은 PV 곡선의 변화를 해석하는 것은 불가능하고, 폐하지 구조적 변화. 따라서 절대 볼륨 측정하지 않고, PV 곡선을 해석하고, 따라서 작은 유틸리티가 거의 불가능하다. 폐 볼륨을 측정하는 방법에는 여러 가지가 있습니다,하지만, 이들은 종종 복잡하고 특별한 장비가 필요합니다. 여기에 설명 된 간단한 방법으로, PV 곡선은 생체 내 가스 제거 과정을 거친 후 제로 볼륨에서 시작한다. 요약하면, 본 논문에서는 마우스의 폐에서 폐 PV 곡선 측정을 표준화하기위한 간단한 방법을 설명하고, 폐 구조에 연결되어있는이 곡선으로부터 계산 될 수있는 몇 가지 지표를 정의합니다. PV 곡선 따라서 통신과 마우스에서 표현형의 구조적인 변화를 감지 할 수있는 직접 응용 프로그램이있는 폐 기능 검사를 제공합니다폐기종 및 섬유증과 같은 폐 병리에.

<table><tbody><tr><td>Syringe pump</td><td>Harvard Apparatus</td><td>55-2226</td><td>Infuse/withdraw syringe pump</td></tr><tr><td>Pump 22 reversing switch</td><td>Harvard Apparatus</td><td>552217</td><td>Included with pump</td></tr><tr><td>Linear displacement transformer</td><td>Trans-Tek, Inc.</td><td>0244-0000</td><td></td></tr><tr><td>5 ml glass syringe</td><td>Becton Dickenson</td><td></td><td>Several other possible vendors</td></tr><tr><td>Digital recorder</td><td>ADInstruments</td><td>PL3504</td><td>Several other possible vendors</td></tr><tr><td>Bridge amp signal conditioner</td><td>ADInstruments</td><td>FE221</td><td></td></tr><tr><td>Gas tank, 100% oxygen</td><td>Airgas, Inc</td><td></td><td>Any supplier or hospital source will work</td></tr><tr><td>Pressure transducer: 0 - 1 psi&amp;nbsp;mV output</td><td>Omega Engineering</td><td>PX-137</td><td>Range &amp;asymp; 0 - 60 cm H&lt;sub&gt;2&lt;/sub&gt;O</td></tr></tbody></table>

measurement of the pressure-volume curve in mouse lungs

최근 수십 년 동안 마우스가 폐 질환의 다양한 차 동물 모델이되고있다. 폐기종 또는 섬유증의 모델에서, 필수적인 표현형 변화는 가장 폐 탄성 변화의 측정에 의해 평가된다. 마우스에서 가장 이러한 병리를 특정 기본 메커니즘을 이해하기 위해서는 병리 현상을 반영 할 수있는 기능을 측정하는 데 필수적이다. 탄성을 측정하는 방법에는 여러 가지가 있지만, 전형적인 방법은 전체 폐 용적의 압력 (PV)는 폐 곡선 볼륨의 전체 범위에 걸쳐 수행한다는 것이다. 이 측정은 거의 백년 거슬러 거의 모든 포유 동물 종으로부터 성인 폐에서 이루어진 것으로, 이러한 PV 곡선은 태아 폐 개발 폐 계면 활성제의 기능의 발견 및 이해에 중요한 역할을했다. 불행히도, 이러한 전체 PV 곡선들이 널리 macrosc에 유용한 정보를 제공 할 수 있다는 사실에도 불구하고, 마우스에서보고되지 않은폐의 구조적인 변화의 OPIC 효과. 폐 용적의 단지 부분적인 변화를 측정 PV 곡선 때때로보고되어 있지만, 절대 체적의 측정치없이 전체 PV 곡선의 비선형 특성을 해석하는 이러한 부분들 매우 어렵게 만든다. 본 연구에서는 총 PV 곡선을 측정하는 표준화 된 방법을 설명합니다. 우리는 다음 두 가지 일반적인 폐 병변, 폐기종 및 섬유증 마우스 폐 구조의 변화를 감지하는이 곡선의 능력을 시험 하였다. 결과는 이러한 병리와 예상 구조적 변화와 일치하는 여러 변수에 큰 변화를 보였다. 마우스에서 폐 PV 곡선이 측정은 시간이 지남에 따라서, 병태 생리 학적 변화 및 ​​치료 방법의 효과 전위의 진행을 모니터링하는 간단한 수단을 제공한다.

PV 곡선에 대한 절차는 전용 제어 건강한 쥐 비디오에서 입증되었지만, 우리는 두 가지 일반적인 병리, 폐기종 및 섬유증 마우스에서 기능적이고 병리 변화를 검출하기 위해 PV 곡선의 능력을 조사 하였다. 이러한 전통적인 모델의 세부 사항은 다른 곳에서 12, 13 설명했다. 아주 간단히, 3 % 이소 플루 란 마취 후 폐기종은 기관지에 주입 3 또는 6 U 돼지의 췌장 엘라 스타 제에 의해 발생 된 3 ?...

Watch this Scientific Journal Video about Measurement of the Pressure-volume Curve in Mouse Lungs at JoVE.com

Measurement of the Pressure-volume Curve in Mouse Lungs

Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung&#8217;s structural changes with developing pathology.

마우스 폐의 압력 볼륨 곡선의 측정

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential ...

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18.6K 조회수.  Johns Hopkins University. Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung’s structural changes with developing pathology.

비디오: Measurement of the Pressure-volume Curve in Mouse Lungs

Pressure Controlled Ventilation to Induce Acute Lung Injury in Mice

Murine models are extensively used to investigate acute injuries of different organs systems (1-34). Acute lung injury (ALI), which occurs with prolonged mechanical ventilation, contributes to morbidity and mortality of critical illness, and studies on novel genetic or pharmacological targets are areas of intense investigation (1-3, 5, 8, 26, 30, 33-36). ALI is defined by the acute onset of the disease, which leads to non-cardiac pulmonary edema and subsequent impairment of pulmonary gas exchange (36). We have developed a murine model of ALI by using a pressure-controlled ventilation to induce ventilator-induced lung injury (2). For this purpose, C57BL/6 mice are anesthetized and a tracheotomy is performed followed by induction of ALI via mechanical ventilation. Mice are ventilated in a pressure-controlled setting with an inspiratory peak pressure of 45 mbar over 1 - 3 hours. As outcome parameters, pulmonary edema (wet-to-dry ratio), bronchoalveolar fluid albumin content, bronchoalveolar fluid and pulmonary tissue myeloperoxidase content and pulmonary gas exchange are assessed (2). Using this technique we could show that it sufficiently induces acute lung inflammation and can distinguish between different treatment groups or genotypes (1-3, 5). Therefore this technique may be helpful for researchers who pursue molecular mechanisms involved in ALI using a genetic approach in mice with gene-targeted deletion.

A murine model for ventilator induced lung injury is an important tool to study an acute lung injury in vivo. Here, we report an easy applicable in situ model for acute lung injury using high-pressure mechanical ventilation to induce acute failure of the lung.

마우스의 급성 폐 상해를 유발하는 압력 제어 환기

Murine models are extensively used to investigate acute injuries of different organs systems (1-34). Acute lung injury (ALI), which occurs with prolonged ...

연구

의학

Right Ventricular Systolic Pressure Measurements in Combination with Harvest of Lung and Immune Tissue Samples in Mice

The function of the right heart is to pump blood through the lungs, thus linking right heart physiology and pulmonary vascular physiology. Inflammation is a common modifier of heart and lung function, by elaborating cellular infiltration, production of cytokines and growth factors, and by initiating remodeling processes 1.
Compared to the left ventricle, the right ventricle is a low-pressure pump that operates in a relatively narrow zone of pressure changes. Increased pulmonary artery pressures are associated with increased pressure in the lung vascular bed and pulmonary hypertension 2. Pulmonary hypertension is often associated with inflammatory lung diseases, for example chronic obstructive pulmonary disease, or autoimmune diseases 3. Because pulmonary hypertension confers a bad prognosis for quality of life and life expectancy, much research is directed towards understanding the mechanisms that might be targets for pharmaceutical intervention 4. The main challenge for the development of effective management tools for pulmonary hypertension remains the complexity of the simultaneous understanding of molecular and cellular changes in the right heart, the lungs and the immune system.
Here, we present a procedural workflow for the rapid and precise measurement of pressure changes in the right heart of mice and the simultaneous harvest of samples from heart, lungs and immune tissues. The method is based on the direct catheterization of the right ventricle via the jugular vein in close-chested mice, first developed in the late 1990s as surrogate measure of pressures in the pulmonary artery5-13. The organized team-approach facilitates a very rapid right heart catheterization technique. This makes it possible to perform the measurements in mice that spontaneously breathe room air. The organization of the work-flow in distinct work-areas reduces time delay and opens the possibility to simultaneously perform physiology experiments and harvest immune, heart and lung tissues.
The procedural workflow outlined here can be adapted for a wide variety of laboratory settings and study designs, from small, targeted experiments, to large drug screening assays. The simultaneous acquisition of cardiac physiology data that can be expanded to include echocardiography5,14-17 and harvest of heart, lung and immune tissues reduces the number of animals needed to obtain data that move the scientific knowledge basis forward. The procedural workflow presented here also provides an ideal basis for gaining knowledge of the networks that link immune, lung and heart function. The same principles outlined here can be adapted to study other or additional organs as needed.

A specific and rapid protocol to simultaneously investigate right heart function, lung inflammation, and the immune response is described as a learning tool. Video and figures describe physiology and microdissection techniques in an organized team-approach that is adaptable to be used for small to large sized studies.

폐 마우스의 면역 조직 샘플의 수확과 함께 오른쪽 심실 수축기 압력 측정

The function of the right heart is to pump blood through the lungs, thus linking right heart physiology and pulmonary vascular physiology. Inflammation is ...

면역학 및 감염병학

A Standardized Method for Measuring Internal Lung Surface Area via Mouse Pneumonectomy and Prosthesis Implantation

Pulmonary morphology, physiology, and respiratory functions change in both physiological and pathological conditions. Internal lung surface area (ISA), representing the gas-exchange capacity of the lung, is a critical criterion to assess respiratory function. However, observer bias can significantly influence measured values for lung morphological parameters. The protocol that we describe here minimizes variations during measurements of two morphological parameters used for ISA calculation: internal lung volume (ILV) and mean linear intercept (MLI). Using ISA as a morphometric and functional parameter to determine the outcome of alveolar regeneration in both pneumonectomy (PNX) and prosthesis implantation mouse models, we found that the increased ISA following PNX treatment was significantly blocked by implantation of a prosthesis into the thoracic cavity1. The ability to accurately quantify ISA is not only expected to improve the reliability and reproducibility of lung function studies in injured-induced alveolar regeneration models, but also to promote mechanistic discoveries of multiple pulmonary diseases.

Internal lung surface area (ISA) is a critical criterion for assessing lung morphology and physiology in lung diseases and injury-induced alveolar regeneration. We describe here a standardized method that can minimize the measurement bias for ISA in both lung pneumonectomy and prosthesis implantation mouse models.

마우스 폐 전 절제술 및 보철물 삽입을 통한 내시경 표면 측정을위한 표준화 된 방법

Pulmonary morphology, physiology, and respiratory functions change in both physiological and pathological conditions. Internal lung surface area (ISA), ...

발생학

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

마우스 폐 삽관 법의 간단한 방법

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Videomorphometric Analysis of Hypoxic Pulmonary Vasoconstriction of Intra-pulmonary Arteries Using Murine Precision Cut Lung Slices

Acute alveolar hypoxia causes pulmonary vasoconstriction (HPV) - also known as von Euler-Liljestrand mechanism - which serves to match lung perfusion to ventilation. Up to now, the underlying mechanisms are not fully understood. The major vascular segment contributing to HPV is the intra-acinar artery. This vessel section is responsible for the blood supply of an individual acinus, which is defined as the portion of lung distal to a terminal bronchiole. Intra-acinar arteries are mostly located in that part of the lung that cannot be selectively reached by a number of commonly used techniques such as measurement of the pulmonary artery pressure in isolated perfused lungs or force recordings from dissected proximal pulmonary artery segments1,2. The analysis of subpleural vessels by real-time confocal laser scanning luminescence microscopy is limited to vessels with up to 50 &#181;m in diameter3.
We provide a technique to study HPV of murine intra-pulmonary arteries in the range of 20-100 &#181;m inner diameters. It is based on the videomorphometric analysis of cross-sectioned arteries in precision cut lung slices (PCLS). This method allows the quantitative measurement of vasoreactivity of small intra-acinar arteries with inner diameter between 20-40 &#181;m which are located at gussets of alveolar septa next to alveolar ducts and of larger pre-acinar arteries with inner diameters between 40-100 &#181;m which run adjacent to bronchi and bronchioles. In contrast to real-time imaging of subpleural vessels in anesthetized and ventilated mice, videomorphometric analysis of PCLS occurs under conditions free of shear stress. In our experimental model both arterial segments exhibit a monophasic HPV when exposed to medium gassed with 1% O2 and the response fades after 30-40 min at hypoxia.

Hypoxic pulmonary vasoconstriction (HPV) is an important physiological phenomenon by which at alveolar hypoxia lung perfusion is matched to ventilation. The major vascular segment contributing to HPV is the intra-acinar artery. Here, we describe our protocol for the analysis of HPV of murine pulmonary vessels with diameters of 20-100 &mu;m.

Murine 정밀 절단 폐 슬라이스를 사용하여 폐 내 동맥의 저산소 폐 혈관 수축의 비디오 형태 측정 분석

Acute alveolar hypoxia causes pulmonary vasoconstriction (HPV) - also known as von Euler-Liljestrand mechanism - which serves to match lung perfusion to ...

Automated Measurement of Pulmonary Emphysema and Small Airway Remodeling in Cigarette Smoke-exposed Mice

COPD is projected to be the third most common cause of mortality world-wide by 2020(1). Animal models of COPD are used to identify molecules that contribute to the disease process and to test the efficacy of novel therapies for COPD. Researchers use a number of models of COPD employing different species including rodents, guinea-pigs, rabbits, and dogs(2). However, the most widely-used model is that in which mice are exposed to cigarette smoke. Mice are an especially useful species in which to model COPD because their genome can readily be manipulated to generate animals that are either deficient in, or over-express individual proteins. Studies of gene-targeted mice that have been exposed to cigarette smoke have provided valuable information about the contributions of individual molecules to different lung pathologies in COPD(3-5). Most studies have focused on pathways involved in emphysema development which contributes to the airflow obstruction that is characteristic of COPD. However, small airway fibrosis also contributes significantly to airflow obstruction in human COPD patients(6), but much less is known about the pathogenesis of this lesion in smoke-exposed animals. To address this knowledge gap, this protocol quantifies both emphysema development and small airway fibrosis in smoke-exposed mice. This protocol exposes mice to CS using a whole-body exposure technique, then measures respiratory mechanics in the mice, inflates the lungs of mice to a standard pressure, and fixes the lungs in formalin. The researcher then stains the lung sections with either Gill&#8217;s stain to measure the mean alveolar chord length (as a readout of emphysema severity) or Masson&#8217;s trichrome stain to measure deposition of extracellular matrix (ECM) proteins around small airways (as a readout of small airway fibrosis). Studies of the effects of molecular pathways on both of these lung pathologies will lead to a better understanding of the pathogenesis of COPD.

The goal of this protocol is to provide automated methods to quantify chronic lung pathologies in a murine model of COPD. The protocol includes exposing mice to cigarette smoke (CS), measuring pulmonary function, inflating the lungs, and using morphometry methods to measure emphysema and small airway remodeling in mice.

담배 연기에 노출 된 마우스의 폐 폐기종 및 작은기도 리모델링 자동 측정

COPD is projected to be the third most common cause of mortality world-wide by 2020(1).  Animal models of COPD are used to identify molecules that ...

Characterization of the Isolated, Ventilated, and Instrumented Mouse Lung Perfused with Pulsatile Flow

The isolated, ventilated and instrumented mouse lung preparation allows steady and pulsatile pulmonary vascular pressure-flow relationships to be measured with independent control over pulmonary arterial flow rate, flow rate waveform, airway pressure and left atrial pressure. Pulmonary vascular resistance is calculated based on multi-point, steady pressure-flow curves; pulmonary vascular impedance is calculated from pulsatile pressure-flow curves obtained at a range of frequencies. As now recognized clinically, impedance is a superior measure of right ventricular afterload than resistance because it includes the effects of vascular compliance, which are not negligible, especially in the pulmonary circulation. Three important metrics of impedance - the zero hertz impedance Z0, the characteristic impedance ZC, and the index of wave reflection RW - provide insight into distal arterial cross-sectional area available for flow, proximal arterial stiffness and the upstream-downstream impedance mismatch, respectively. All results obtained in isolated, ventilated and perfused lungs are independent of sympathetic nervous system tone, volume status and the effects of anesthesia. We have used this technique to quantify the impact of pulmonary emboli and chronic hypoxia on resistance and impedance, and to differentiate between sites of action (i.e., proximal vs. distal) of vasoactive agents and disease using the pressure dependency of ZC. Furthermore, when these techniques are used with the lungs of genetically engineered strains of mice, the effects of molecular-level defects on pulmonary vascular structure and function can be determined.

The following protocol outlines the process of isolating, ventilating and instrumenting mouse lungs to measure steady or pulsatile pulmonary vascular pressure-flow relationships in order to quantify the effects of blood flow, airflow, airway changes and vascular changes on right ventricular afterload.

절연, 환기 및기구 마우스 폐의 특성화는 타악기 흐름 Perfused

The isolated, ventilated and instrumented mouse lung preparation allows steady and pulsatile pulmonary vascular pressure-flow relationships to be measured ...

생물학

Evaluation of Respiratory System Mechanics in Mice using the Forced Oscillation Technique

The forced oscillation technique (FOT) is a powerful, integrative and translational tool permitting the experimental assessment of lung function in mice in a comprehensive, detailed, precise and reproducible manner. It provides measurements of respiratory system mechanics through the analysis of pressure and volume signals acquired in reaction to predefined, small amplitude, oscillatory airflow waveforms, which are typically applied at the subject's airway opening. The present protocol details the steps required to adequately execute forced oscillation measurements in mice using a computer-controlled piston ventilator (flexiVent; SCIREQ Inc, Montreal, Qc, Canada). The description is divided into four parts: preparatory steps, mechanical ventilation, lung function measurements, and data analysis. It also includes details of how to assess airway responsiveness to inhaled methacholine in anesthetized mice, a common application of this technique which also extends to other outcomes and various lung pathologies. Measurements obtained in na&iuml;ve mice as well as from an oxidative-stress driven model of airway damage are presented to illustrate how this tool can contribute to a better characterization and understanding of studied physiological changes or disease models as well as to applications in new research areas.

The present protocol provides a detailed step-by-step description of the procedures required to execute measurements of respiratory system mechanics as well as the assessment of airway responsiveness to inhaled methacholine in mice using the forced oscillation technique (flexiVent; SCIREQ Inc, Montreal, Qc, Canada).

강제 진동 기술을 사용하여 마우스의 호흡기 시스템 역학의 평가

The  forced oscillation technique (FOT) is a powerful, integrative and translational  tool permitting the experimental assessment of lung function in mice ...

Lung Fixation under Constant Pressure for Evaluation of Emphysema in Mice

Emphysema is a significant feature of chronic obstructive pulmonary disease (COPD). Studies involving an emphysematous mouse model require optimal lung fixation to produce reliable histological specimens of the lung. Due to the nature of the lung&rsquo;s structural composition, which consists largely of air and tissue, there is a risk that it collapses or deflates during the fixation process. Various lung fixation methods exist, each of which has its own advantages and disadvantages. The lung fixation method presented here utilizes constant pressure to enable optimal tissue evaluation for studies using an emphysematous mouse lung model. The main advantage is that it can fix many lungs with the same condition at one time. Lung specimens are obtained from chronic cigarette smoke-exposed mice. Lung fixation is performed using specialized equipment that enables the production of constant pressure. This constant pressure maintains the lung in a reasonably inflated state. Thus, this method generates a histological specimen of the lung that is suitable to evaluate cigarette smoke-induced mild emphysema.

Presented here is a useful protocol for lung fixation that creates a stable condition for histological evaluate of lung specimens from a mouse model of emphysema. The main advantage of this model is that it can fix many lungs with the same constant pressure without lung collapse or deflation.

마우스에 있는 기종의 평가를 위한 일정한 압력의 밑에 폐 고정

Emphysema is a significant feature of chronic obstructive pulmonary disease (COPD). Studies involving an emphysematous mouse model require optimal lung ...

Measuring Respiratory Function in Mice Using Unrestrained Whole-body Plethysmography

Respiratory dysfunction is one of the leading causes of morbidity and mortality in the world and the rates of mortality continue to rise. Quantitative assessment of lung function in rodent models is an important tool in the development of future therapies. Commonly used techniques for assessing respiratory function including invasive plethysmography and forced oscillation. While these techniques provide valuable information, data collection can be fraught with artefacts and experimental variability due to the need for anesthesia and/or invasive instrumentation of the animal. In contrast, unrestrained whole-body plethysmography (UWBP) offers a precise, non-invasive, quantitative way by which to analyze respiratory parameters. This technique avoids the use of anesthesia and restraints, which is common to traditional plethysmography techniques. This video will demonstrate the UWBP procedure including the equipment set up, calibration and lung function recording. It will explain how to analyze the collected data, as well as identify experimental outliers and artefacts that results from animal movement. The respiratory parameters obtained using this technique include tidal volume, minute volume, inspiratory duty cycle, inspiratory flow rate and the ratio of inspiration time to expiration time. UWBP does not rely on specialized skills and is inexpensive to perform. A key feature of UWBP, and most appealing to potential users, is the ability to perform repeated measures of lung function on the same animal.

The assessment of respiratory physiology has traditionally relied upon techniques, which require restraint or sedation of the animal. Unrestrained whole-body plethysmography, however, provides precise, non-invasive, quantitative analysis of respiratory physiology in animal models. In addition, the technique allows repeated respiratory assessment of mice allowing for longitudinal studies.

무제한 전신 혈량 측정법을 사용하여 마우스의 호흡 기능을 측정

Respiratory dysfunction is one of the leading causes of morbidity and mortality in the world and the rates of mortality continue to rise. Quantitative ...