Name: contrast imaging in mouse embryos using high-frequency ultrasound
Uploaded: 2015-03-04T14:00:00.000+00:00
Duration: 10 min 39 s
Description: Watch this Scientific Journal Video about Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound at JoVE.com

This work was supported by the Terry Fox Program of the National Cancer Institute of Canada.

<ol>
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F.S. Foster acknowledges his role as consultant to VisualSonics Inc.

초음파 조영제가 말기 임신 마우스 배아 및 비선형 대비 이미지로 주입 하였다는 관류 매개 변수를 측정하기 위해 취득 및 바인딩 마이크로 버블을 대상으로 하였다. 배아의 혈관계 내의 미세 기포 성공적인 이미징은 다수의 인자, 제 인 배아 생존에 의존했다. 모든 장치 및 장치가 분사의 시작에서 배아의 자궁 분리에 필요한 시간을 최소화하기 위해 사전에 준비 하였다. 쥐 배아 마취 단일 또는 ...

조영 증강 초음파 영상 시각화하고 혈관 환경의 특성을 미세 기포 조영제를 사용한다. 이러한 에이전트는 미세 순환, 혈관 및 심장 혈관 기능의 비 침습적 평가를 할 수 있습니다. 또한, 기포 표면의 개질이 가능 혈관 질환의 분자 초음파 영상을 혈관 신생, 염증 및 아테롬성 경화증의 임상 1,2- 애플리케이션에서 입증 된 바와 같이, 내피 표적 바이오 마커에 ​​대한 결합 미세 기포가 발생할 수있다. 콘트라스트 향상 초음파 따라서 혈관 질병과 건강 상태에 영향 3-5 복잡하고 다양한 환경을 식별하기 위해 사용될 수있다. 년의 과거 수 년, 마이크로 버블 영상의 유용성에 대한 관심이 다양한 마우스 배아 모델을 확장했다. 포유류의 개발 모델로, 배아 미세 기포 맥관계 내로 도입 생리 향상개발 순환 시스템 (예를 들어, 혈류, 심장 출력) 및 형질 전환 사례 및 심장 질환 6,7의 대상으로 돌연변이 마우스 모델에서의 연구는 심장 혈관 기능을 변경하는 방법 유전 적 요인에 대한 통찰력을 얻을 수 있습니다. 사실, 양적 및 질적 2D는 배아 뇌 혈관 분석 이미 8을 달성했다. 더욱이, 마우스의 배아는 생체 내에서 혈관 마커 타겟팅 결합 미세 기포를 검사하는 우수한 모델로서 제시한다. Bartelle 등. (9)는, 예를 들어, Biotag - 피라 유전자 변형 배아에 바인딩 및 혈관 해부학을 검토하여 대상을 평가하기 위해 태아 심장의 심실에 아비딘 미세 기포를 도입했습니다. 병원에이 기술을 번역에서 중요한 벤치 마크 - 이질 동형 접합 마우스 모델의 생성은 분자 초음파의 양적 특성을 정의하는 것을 목표로 종양 모델 연구를위한 대리로 사용할 수 있습니다. 8-10 통해 노출로 단일 배아 심장 내 주사를 통해 순환 배아에 도입된다. 자궁 내 주사 그러나 난제에 직면하고있다. 이러한 주입 안내, 어머니 표면화 배아 움직임에 대응하는 필요, 어머니 혈역학 생존을 유지하고 (11)에 의한 출혈 합병증 마취 장기적인 효과를 어드레싱 표면화 배아를 포함한다. 따라서, 조사의 목적은 거실 절연 말기 배아 미세 기포 (12) 내로 주입하는 기술을 개발하는 것이었다. 이 옵션은 분사 제어 및 위치, 장애물없이 촬상면의 재현성 간단한 이미지 분석 및 정량의 관점에서 더 많은 자유를 제공한다. 본 연구에서는 FO 살아있는 쥐의 배아에 미세 기포를 주입위한 새로운 절차 개요R 마이크로 버블 운동 동작을 연구하고의 목적은 내인성 내피 세포 표면 마커에 결합 마이크로 버블을 목표로 공부. 비선형 대비 특정 초음파 영상은 피크 향상 (PE), 세척 된 속도와 시간 고립 E17.5 배아에 (TTP)을 피크를 포함하는 기본 관류 매개 변수들로 측정하는 데 사용됩니다. 우리는 또한 활성 혈관 (13)의 부위에서 혈관 내피 세포에서의 고 발현에 endoglin은 임상 적으로 관련된 대상 함수 트랜스 제닉 마우스 모델의 배아 endoglin 손실 분자 초음파의 정량적 특성을 평가하기위한 배아 모델의 타당성을 입증 . endoglin 타겟 (MB E) 쥐 이소 IgG의 2 제어 (MB C) 및 타겟이 불분명 한 (MB U) 마이크로 버블의 접착 이형 endoglin (영어 +/-)과 동형 접합 endoglin (영어 +가 / +) 표현 배아에서 평가된다. 대상 BINDI 분석NG 분자 초음파 endoglin 유전자형을 구별 정량화 초음파 분자 수준 수용체 밀도를 관련시킬 수 있음을 보여준다.

<table><tbody><tr><td colspan="4">&lt;strong&gt;Reagents&lt;/strong&gt;</td></tr><tr><td colspan="4">&lt;strong&gt;Antibodies&lt;/strong&gt; (biotinylated, eBioscience) &amp;#8212; Antibody choice depends on the experiment</td></tr><tr><td>&lt;ul style=&quot;padding:0px;margin:0px&quot;&gt;&lt;li&gt;rat isotype IgG&lt;sub&gt;2&lt;/sub&gt; control&lt;/li&gt;&lt;/ul&gt;</td><td>eBioscience</td><td>13-4321-85</td><td>This antibody/microbubble combination is often required as experimental control&amp;nbsp;</td></tr><tr><td>&lt;ul style=&quot;padding:0px;margin:0px&quot;&gt;&lt;li&gt;biotin anti-mouse CD309&lt;/li&gt;&lt;/ul&gt;</td><td>eBioscience</td><td>13-5821-85</td><td></td></tr><tr><td>Biotinylated rat MJ 7/18 antibody to mouse endoglin</td><td>In house hybridoma</td><td></td><td>Outside antibodies may also be appropriate: we&amp;nbsp; have used eBioscience (13-1051-85 ) in the past</td></tr><tr><td>Distilled water</td><td></td><td></td><td></td></tr><tr><td colspan="4">&lt;strong&gt;Embryo media&lt;/strong&gt;</td></tr><tr><td>&lt;ul style=&quot;padding:0px;margin:0px&quot;&gt;&lt;li&gt;500 ml Dulbecco&amp;rsquo;s Modified Eagle&amp;rsquo;s Medium with high glucose&lt;/li&gt;&lt;/ul&gt;</td><td>Sigma</td><td>D5796</td><td></td></tr><tr><td>&lt;ul style=&quot;padding:0px;margin:0px&quot;&gt;&lt;li&gt;50 ml Fetal Bovine Serum&lt;/li&gt;&lt;/ul&gt;</td><td>ATCC</td><td>30-2020</td><td>lot # 7592456</td></tr><tr><td>&lt;ul style=&quot;padding:0px;margin:0px&quot;&gt;&lt;li&gt;Hepes&lt;/li&gt;&lt;/ul&gt;</td><td>Gibco</td><td>15630</td><td>5 ml, 1 M</td></tr><tr><td>&lt;ul style=&quot;padding:0px;margin:0px&quot;&gt;&lt;li&gt;Penicillin-Streptomycin&lt;/li&gt;&lt;/ul&gt;</td><td>Gibco</td><td>15140-122</td><td>5 ml, 10,000 units Pen., 10,000 &amp;#956;g Strep</td></tr><tr><td>Ethanol, 70%</td><td></td><td></td><td></td></tr><tr><td>Ice</td><td></td><td></td><td></td></tr><tr><td>Paraformaldehyde</td><td>Sigma</td><td>76240</td><td>4%</td></tr><tr><td>Phosphate Buffered Saline [1x]&amp;nbsp;</td><td>Sigma</td><td>D8537</td><td>1x, w/o calcium chloride &amp;amp; magnesium chloride</td></tr><tr><td>Pregnant mouse, CD-1</td><td>Charles River Laboratories Inc.&amp;nbsp;</td><td></td><td></td></tr><tr><td>0.9% sodium chloride (saline)</td><td>Hospira</td><td>0409-7984-11</td><td></td></tr><tr><td>Ultrasound contrast agent, target ready and untargeted</td><td>MicroMarker; VisualSonics Inc.</td><td></td><td></td></tr><tr><td>Ultrasound gel (Aquasonic 100, colourless)</td><td>CSP Medical</td><td>133-1009</td><td></td></tr><tr><td colspan="4">&lt;strong&gt;Equipment&lt;/strong&gt;</td></tr><tr><td>Cell culture plates (4) :&amp;nbsp; 100&amp;nbsp;x&amp;nbsp;20&amp;nbsp;mm</td><td>Fisher Scientific</td><td>08-772-22</td><td></td></tr><tr><td>Cell culture plates (12) : 60&amp;nbsp;x&amp;nbsp;15&amp;nbsp;mm</td><td>Sigma</td><td>D8054</td><td></td></tr><tr><td>Centrifuge</td><td>Sorvall Legend RT centrifuge&amp;nbsp;</td><td></td><td></td></tr><tr><td>Conical tubes, 50 ml BD Falcon</td><td>VWR</td><td>21008-938</td><td></td></tr><tr><td>Diluent</td><td>Beckman Coulter</td><td>Isoton II Diluent, 8448011</td><td></td></tr><tr><td>Dissection scissors (Wagner)</td><td>Fine Science Tools</td><td>Wagner 14068-12</td><td></td></tr><tr><td>Forceps (2), Dumont SS (0.10 x 0.06 mm)</td><td>Fine Science Tools</td><td>11200-33</td><td></td></tr><tr><td>Forceps, splinter</td><td>VWR</td><td>25601-134</td><td></td></tr><tr><td>Glass beaker, 2 L (Griffin Beaker)</td><td>VWR</td><td>89000-216</td><td></td></tr><tr><td>Glass capillaries, 1 x 90&amp;nbsp;mm GD-1 with filament</td><td>Narishige</td><td>GD-1</td><td></td></tr><tr><td>Glass needle puller</td><td>Narishige</td><td>PN-30</td><td></td></tr><tr><td>Gloves</td><td>Ansell</td><td>4002</td><td></td></tr><tr><td>Gross anatomy probe</td><td>Fine Science Tools</td><td>10088-15</td><td></td></tr><tr><td>Hot plate</td><td>VWR</td><td>89090-994</td><td></td></tr><tr><td>Ice bucket</td><td>Cole Parmer</td><td>RK 06274-01</td><td></td></tr><tr><td>Imaging Platform</td><td>VisualSonics Inc.</td><td>Integrated Rail System</td><td></td></tr><tr><td>Light source, fiber-optic</td><td>Fisher Scientific</td><td>12-562-36</td><td>Ideally has adjustable arms</td></tr><tr><td>Luers (12), polypropylene barbed female &amp;frac14;-28 UNF thread</td><td>Cole Parmer</td><td>45500-30</td><td></td></tr><tr><td>Micro-ultrasound system, high-frequency</td><td>VisualSonics Inc.</td><td>Vevo2100</td><td></td></tr><tr><td>Needles, 21 gauge&amp;nbsp; (1&amp;rdquo;)</td><td>VWR</td><td>305165</td><td></td></tr><tr><td>Particle size analyzer</td><td>Beckman Coulter</td><td>Multisizer 3 Coulter Counter</td><td></td></tr><tr><td>Perforated spoon (Moria)</td><td>Fine Science Tools</td><td>MC 17 10373-17</td><td></td></tr><tr><td>Pins (6), black anodized minutien 0.15 mm</td><td>Fine Science Tools</td><td>26002-15</td><td></td></tr><tr><td>Pipettors [2-20 &amp;#956;l, 20-200 &amp;#956;l, 100-1,000 &amp;#956;l]</td><td>Eppendorf</td><td>Research Plus&amp;nbsp; adjustable 3120000038;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp; 3120000054;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp; 3120000062</td><td></td></tr><tr><td>Pipettor tips [2-200&amp;nbsp;&amp;#956;l, 50-1,000&amp;nbsp;&amp;#956;l]</td><td>Eppendorf</td><td>epT.I.P.S.&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp; 22491334;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp;&amp;nbsp; 022491351</td><td></td></tr><tr><td>Scissors</td><td></td><td></td><td></td></tr><tr><td>Sylgard 184 Silicone Elastomer Kit</td><td>Dow Corning</td><td></td><td></td></tr><tr><td>Tubing, Tygon laboratory 1/32&amp;rdquo; x 3/32&amp;rdquo;</td><td>VWR</td><td>63010-007</td><td></td></tr><tr><td>Wooden applicator stick (swab, cotton head)</td><td>VWR</td><td>CA89031-270</td><td></td></tr><tr><td>Surgical microscope 5-8X magnification</td><td>Fisher Scientific</td><td>Steromaster</td><td></td></tr><tr><td>Syringes, 1 ml Normject</td><td>Fisher</td><td>14-817-25</td><td></td></tr><tr><td>Syringes (10), 30 ml</td><td>VWR</td><td>CA64000-041</td><td></td></tr><tr><td>Syringe infusion pump&amp;nbsp;</td><td>Bio-lynx&amp;nbsp;</td><td>NE-1000</td><td></td></tr><tr><td>Thermometer, -20-110&amp;nbsp;&amp;#176;C</td><td>VWR</td><td>89095-598</td><td></td></tr><tr><td>Timer</td><td>VWR</td><td>33501-418</td><td></td></tr><tr><td>Tubes, Eppendorf</td><td>VWR</td><td>20170-577</td><td></td></tr><tr><td>Tube racks (3)</td><td>VWR</td><td>82024-462</td><td></td></tr><tr><td>Ultrasound transducer, 20 MHz</td><td>VisualSonics Inc.</td><td>MS250</td><td></td></tr><tr><td>Vannas-Tubingen, angled up</td><td>Fine Science Tools</td><td>15005-08</td><td></td></tr></tbody></table>

contrast imaging in mouse embryos using high-frequency ultrasound

Ultrasound contrast-enhanced imaging can convey essential quantitative information regarding tissue vascularity and perfusion and, in targeted applications, facilitate the detection and measure of vascular biomarkers at the molecular level. Within the mouse embryo, this noninvasive technique may be used to uncover basic mechanisms underlying vascular development in the early mouse circulatory system and in genetic models of cardiovascular disease. The mouse embryo also presents as an excellent model for studying the adhesion of microbubbles to angiogenic targets (including vascular endothelial growth factor receptor 2 (VEGFR2) or &#945;v&#946;3) and for assessing the quantitative nature of molecular ultrasound. We therefore developed a method to introduce ultrasound contrast agents into the vasculature of living, isolated embryos. This allows freedom in terms of injection control and positioning, reproducibility of the imaging plane without obstruction and motion, and simplified image analysis and quantification. Late gestational stage (embryonic day (E)16.6 and E17.5) murine embryos were isolated from the uterus, gently exteriorized from the yolk sac and microbubble contrast agents were injected into veins accessible on the chorionic surface of the placental disc. Nonlinear contrast ultrasound imaging was then employed to collect a number of basic perfusion parameters (peak enhancement, wash-in rate and time to peak) and quantify targeted microbubble binding in an endoglin mouse model. We show the successful circulation of microbubbles within living embryos and the utility of this approach in characterizing embryonic vasculature and microbubble behavior.

전 자궁 마우스 배아에 초음파 조영제의 주입은 주입 및 관련 초음파 영상의 과정을 통해 자궁과 생존의 유지 보수에서 생활, 늦은 임신 단계 배아의 성공적인 분리에 따라 달라집니다. 배아가 표면화하고도 1에 도시 된 바와 같이, 위치되면 배아 혈관에 조영제 주입 조심 가능하다. E17.5 마우스 태아의 전형적인 B 모드 초음파 이미지는도 2a에 도시된다. 미세 기포 ?...

Watch this Scientific Journal Video about Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound at JoVE.com

Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound

여기서 우리는 생활에 초음파 미세 기포 조영제, 고립 늦은 임신 단계 쥐의 배아를 주입하는 프로토콜을 제시한다. 이 방법은 관류의 매개 변수 및 조영 증강 고주파 초음파 영상을 이용한 혈관 내 배아 분자 마커의 연구를 가능하게한다.

고주파 초음파를 사용하여 마우스 태아에 대비 이미징

Ultrasound contrast-enhanced imaging can convey essential quantitative information regarding tissue vascularity and perfusion and, in targeted ...

contrast-imaging-in-mouse-embryos-using-high-frequency-ultrasound

Universidad San Sebastian

Research

JoVE Journal

Developmental Biology

15.3K 조회수.  University of Toronto.  여기서 우리는 생활에 초음파 미세 기포 조영제, 고립 늦은 임신 단계 쥐의 배아를 주입하는 프로토콜을 제시한다. 이 방법은 관류의 매개 변수 및 조영 증강 고주파 초음파 영상을 이용한 혈관 내 배아 분자 마커의 연구를 가능하게한다. 

비디오: Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound

Ultrasound-Guided Microinjection into the Mouse Forebrain In Utero at E9.5

In utero survival surgery in mice permits the molecular manipulation of gene expression during development. However, because the uterine wall is opaque during early embryogenesis, the ability to target specific parts of the embryo for microinjection is greatly limited. Fortunately, high-frequency ultrasound imaging permits the generation of images that can be used in real time to guide a microinjection needle into the embryonic region of interest. Here we describe the use of such imaging to guide the injection of retroviral vectors into the ventricular system of the mouse forebrain at embryonic day (E) 9.5. This method uses a laparotomy to permit access to the uterine horns, and a specially designed plate that permits host embryos to be bathed in saline while they are imaged and injected. Successful surgeries often result in most or all of the injected embryos surviving to any subsequent time point of interest (embryonically or postnatally). The principles described here can be used with slight modifications to perform injections into the amnionic fluid of E8.5 embryos (thereby permitting infection along the anterior posterior extent of the neural tube, which has not yet closed), or into the ventricular system of the brain at E10.5/11.5. Furthermore, at mid-neurogenic ages (~E13.5), ultrasound imaging can be used direct injection into specific brain regions for viral infection or cell transplantation. The use of ultrasound imaging to guide in utero injections in mice is a very powerful technique that permits the molecular and cellular manipulation of mouse embryos in ways that would otherwise be exceptionally difficult if not impossible.

Ultrasound-Guided Microinjection into the Mouse Forebrain In Utero at E9.5

In utero survival surgery in mice permits the molecular manipulation of gene expression during development. Here we describe the use of high-frequency ultrasound imaging to guide the injection of retroviral vectors into the mouse brain at embryonic day (E) 9.5.

마우스 Forebrain에 초음파 유도 Microinjection Utero에서 E9.5에서

In utero survival surgery in mice permits the molecular manipulation of gene expression during development.  However, because the uterine wall is opaque ...

연구

신경과학

Murine Fetal Echocardiography

Transgenic mice displaying abnormalities in cardiac development and function represent a powerful tool for the understanding the molecular mechanisms underlying both normal cardiovascular function and the pathophysiological basis of human cardiovascular disease. Fetal and perinatal death is a common feature when studying genetic alterations affecting cardiac development 1-3. In order to study the role of genetic or pharmacologic alterations in the early development of cardiac function, ultrasound imaging of the live fetus has become an important tool for early recognition of abnormalities and longitudinal follow-up. Noninvasive ultrasound imaging is an ideal method for detecting and studying congenital malformations and the impact on cardiac function prior to death 4. It allows early recognition of abnormalities in the living fetus and the progression of disease can be followed in utero with longitudinal studies 5,6. Until recently, imaging of fetal mouse hearts frequently involved invasive methods. The fetus had to be sacrificed to perform magnetic resonance microscopy and electron microscopy or surgically delivered for transillumination microscopy. An application of high-frequency probes with conventional 2-D and pulsed-wave Doppler imaging has been shown to provide measurements of cardiac contraction and heart rates during embryonic development with databases of normal developmental changes now available 6-10. M-mode imaging further provides important functional data, although, the proper imaging planes are often difficult to obtain. High-frequency ultrasound imaging of the fetus has improved 2-D resolution and can provide excellent information on the early development of cardiac structures 11.

Fetal and perinatal death is a common feature when studying genetic alterations affecting cardiac development. High-frequency ultrasound imaging has improved 2-D resolution and can provide excellent information on early cardiac development and is an ideal method to detect the impact on cardiac structure and function prior to death.

손쥐 태아 Echocardiography

Transgenic mice displaying abnormalities in  cardiac development and function represent a powerful tool for the understanding  the molecular mechanisms ...

의학

A Novel Use of Three-dimensional High-frequency Ultrasonography for Early Pregnancy Characterization in the Mouse

High-frequency ultrasonography (HFUS) is a common method to non-invasively monitor the real-time development of the human fetus in utero. The mouse is routinely used as an in vivo model to study embryo implantation and pregnancy progression. Unfortunately, such murine studies require pregnancy interruption to enable follow-up phenotypic analysis. To address this issue, we used three-dimensional (3-D) reconstruction of HFUS imaging data for early detection and characterization of murine embryo implantation sites and their individual developmental progression in utero. Combining HFUS imaging with 3-D reconstruction and modeling, we were able to accurately quantify embryo implantation site number as well as monitor developmental progression in pregnant C57BL6J/129S mice from 5.5 days post coitus (d.p.c.) through to 9.5 d.p.c. with the use of a transducer. Measurements included: number, location, and volume of implantation sites as well as inter-implantation site spacing; embryo viability was assessed by cardiac activity monitoring. In the immediate post-implantation period (5.5 to 8.5 d.p.c.), 3-D reconstruction of the gravid uterus in both mesh and solid overlay format enabled visual representation of the developing pregnancies within each uterine horn. As genetically engineered mice continue to be used to characterize female reproductive phenotypes derived from uterine dysfunction, this method offers a new approach to detect, quantify, and characterize early implantation events in vivo. This novel use of 3-D HFUS imaging demonstrates the ability to successfully detect, visualize, and characterize embryo-implantation sites during early murine pregnancy in a non-invasive manner. The technology offers a significant improvement over current methods, which rely on the interruption of pregnancies for gross tissue and histopathologic characterization. Here we use a video and text format to describe how to successfully perform ultrasounds of early murine pregnancy to generate reliable and reproducible data with reconstruction of the uterine form in mesh and solid 3-D images.

Mice are widely used to study gestational biology. However, pregnancy termination is required for such studies which precludes longitudinal investigations and necessitates the use of large numbers of animals. Therefore, we describe a non-invasive technique of high-frequency ultrasonography for early detection and monitoring of post-implantation events in the pregnant mouse.

마우스에서 초기 임신 특성에 대 한 3 차원 높은-주파수 초음파의 새로운 사용

High-frequency ultrasonography (HFUS) is a common method to non-invasively monitor the real-time development of the human fetus in utero. The mouse is ...

발생학

Fetal Mouse Cardiovascular Imaging Using a High-frequency Ultrasound (30/45MHZ) System

Congenital heart defects (CHDs) are the most common cause of childhood morbidity and early mortality. Prenatal detection&#160;of the underlying molecular mechanisms of CHDs is&#160;crucial for inventing new preventive and therapeutic strategies. Mutant mouse models are powerful tools to discover new&#160;mechanisms&#160;and environmental stress modifiers that drive cardiac development and their potential&#160;alteration in CHDs. However, efforts to establish the causality of these putative contributors have been limited to histological&#160;and molecular studies&#160;in non-survival animal experiments, in which&#160;monitoring the key physiological and hemodynamic parameters is&#160;often absent. Live imaging technology has become an essential tool to establish the&#160;etiology of CHDs. In particular,&#160;ultrasound imaging can be used prenatally without surgically exposing the fetuses,&#160;allowing maintaining&#160;their baseline physiology while monitoring the impact of environmental stress&#160;on the hemodynamic and structural aspects of cardiac chamber development. Herein, we use the High-Frequency Ultrasound (30/45)&#160;system to examine the cardiovascular system in fetal mice at E18.5 in utero at the baseline and in response to prenatal hypoxia exposure. We demonstrate the feasibility of the system to measure cardiac chamber size, morphology, ventricular function, fetal heart rate, and umbilical artery flow indices, and their alterations in fetal mice exposed to systemic chronic hypoxia in utero in real time.

Fetal Mouse Cardiovascular Imaging Using a High-frequency Ultrasound 30/45MHZ System

High-frequency ultrasound imaging of the fetal mouse has improved imaging resolution and can provide precise non-invasive characterization of cardiac development and structural defects. The protocol outlined herein is designed to perform real-time fetal mice echocardiography in vivo.

태아 마우스 심장 혈관 이미징 높은 주파수 초음파 (30/45 m h Z) 시스템을 사용 하 여

Congenital heart defects (CHDs) are the most common cause of childhood morbidity and early mortality. Prenatal detection&#160;of the underlying molecular ...

In utero Measurement of Heart Rate in Mouse by Noninvasive M-mode Echocardiography

Congenital heart disease (CHD) is the most frequent noninfectious cause of death at birth. The incidence of CHD ranges from 4 to 50/1,000 births (Disease and injury regional estimates, World Health Organization, 2004). Surgeries that often compromise the quality of life are required to correct heart defects, reminding us of the importance of finding the causes of CHD. Mutant mouse models and live imaging technology have become essential tools to study the etiology of this disease. Although advanced methods allow live imaging of abnormal hearts in embryos, the physiological and hemodynamic states of the latter are often compromised due to surgical and/or lengthy procedures. Noninvasive ultrasound imaging, however, can be used without surgically exposing the embryos, thereby maintaining their physiology. Herein, we use simple M-mode ultrasound to assess heart rates of embryos at E18.5 in utero. The detection of abnormal heart rates is indeed a good indicator of dysfunction of the heart and thus constitutes a first step in the identification of developmental defects that may lead to heart failure.

In utero Measurement of Heart Rate in Mouse by Noninvasive M-mode Echocardiography

Ultra-high frequency ultrasound is a powerful live imaging tool to examine cardiac abnormalities in small animals. Its noninvasiveness allows maintaining the physiologic state of embryos. Herein, we show the use of M-mode ultrasound to measure the heart rates of embryos at E18.5 in utero.

비 침습 M-모드 초음파 심장 검진 법에 의한 마우스의 심장 박동의 자궁 측정

Congenital heart disease (CHD) is the most frequent noninfectious cause of death at birth. The incidence of CHD ranges from 4 to 50/1,000 births (Disease ...

High Frequency Ultrasound for the Analysis of Fetal and Placental Development In Vivo

Ultrasound imaging is a widespread method used to detect organ anomalies and tumors in human and animal tissues. The method is non-invasive, harmless, and painless, and the application is easy, fast, and can be done anywhere, even with mobile devices. During pregnancy, ultrasound imaging is standardly used to closely monitor fetal development. The technique is important to assess intrauterine growth restriction (IUGR), a pregnancy complication with short- and long-term health consequences for both the mother and fetus. Understanding the process of IUGR is indispensable for developing effective therapeutic strategies. 
The ultrasound system used in this manuscript is an ultrasound device produced for the analysis of small animals and can be used in various research fields, including pregnancy research. Here we describe the usage of the system for in vivo analysis of fetuses from natural killer (NK) cell/mast cell (MC)-deficient mothers that give birth to growth-restricted pups. The protocol includes preparation of the system, handling of the mice before and during measurements, and the usage of the B-mode, color doppler mode, and pulse-wave doppler mode. Fetal size, placental size, and blood supply to the fetus were analyzed. We found reduced implantation sizes and smaller placentas in NK/MC-deficient mice from mid-gestation onwards. In addition, MC/NK-deficiency was associated with absent and reversed end diastolic flow in the fetal Arteria umbilicalis(UmA) and an elevated resistance index. The methods described in the protocol can easily be used for related and non-related research topics.

High Frequency Ultrasound for the Analysis of Fetal and Placental Development In Vivo

Here we describe the technique of high frequency ultrasound for in vivo analysis of fetuses in mice. This method allows the follow-up of fetuses and the analysis of placental parameters as well as maternal and fetal blood flow throughout pregnancy.

비보에 태아 및 태 반 개발의 분석에 대 한 높은 주파수 초음파

Ultrasound imaging is a widespread method used to detect organ anomalies and tumors in human and animal tissues. The method is non-invasive, harmless, and ...

Non-Invasive Ultrasound Assessment of Endometrial Cancer Progression in Pax8-Directed Deletion of the Tumor Suppressors Arid1a and Pten in Mice

Uterine cancers can be studied in mice due to the ease of handling and genetic manipulation in these models. However, these studies are often limited to assessing pathology post-mortem in animals euthanized at multiple time points in different cohorts, which increases the number of mice needed for a study. Imaging mice in longitudinal studies can track the progression of disease in individual animals, reducing the number of mice needed. Advances in ultrasound technology have allowed for the detection of micrometer-level changes in tissues. Ultrasound has been used to study follicle maturation in ovaries and xenograft growth but has not been applied to morphological changes in the mouse uterus. This protocol examines the juxtaposition of pathology with in vivo imaging comparisons in an induced endometrial cancer mouse model. The features observed by ultrasound were consistent with the degree of change seen by gross pathology and histology. Ultrasound was found to be highly predictive of the observed pathology, supporting the incorporation of ultrasonography into longitudinal studies of uterine diseases such as cancer in mice.

Non-Invasive Ultrasound Assessment of Endometrial Cancer Progression in Pax8-Directed Deletion of the Tumor Suppressors Arid1a and Pten in Mice

This protocol describes a method for monitoring the progression of morphological changes over time in the uterus in an inducible mouse model of endometrial cancer using ultrasound imaging with correlation to gross and histological changes.

마우스에서 종양 억제인자 Arid1a 및 Pten 의 Pax8 지시 결실에서 자궁내막암 진행의 비침습적 초음파 평가

Uterine cancers can be studied in mice due to the ease of handling and genetic manipulation in these models. However, these studies are often limited to ...

암 연구학

The Preparation of Chicken Ex Ovo Embryos and Chorioallantoic Membrane Vessels as In Vivo Model for Contrast-Enhanced Ultrasound Imaging and Microbubble-Mediated Drug Delivery Studies

The chicken embryo and the blood-vessel rich chorioallantoic membrane (CAM) is a valuable in vivo model to investigate biomedical processes, new ultrasound pulsing schemes, or novel transducers for contrast-enhanced ultrasound imaging and microbubble-mediated drug delivery. The reasons for this are the accessibility of the embryo and vessel network of the CAM as well as the low costs of the model. An important step to get access to the embryo and CAM vessels is to take the egg content out of the eggshell. In this protocol, three methods for taking the content out of the eggshell between day 5 and&#160;8 of incubation are described thus allowing the embryos to develop inside the eggshell up to these days. The described methods only require simple tools and equipment and yield a higher survival success rate of 90% for 5-day, 75% for 6-day, 50% for 7-day, and 60% for 8-day old incubated eggs in comparison to ex ovo cultured embryos (~50%). The protocol also describes how to inject cavitation nuclei, such as microbubbles, into the CAM vascular system, how to separate the membrane containing the embryo and CAM from the rest of the egg content for optically transparent studies, and how to use the chicken embryo and CAM in a variety of short-term ultrasound experiments. The in vivo&#160;chicken embryo and CAM model is extremely relevant to investigate novel imaging protocols, ultrasound contrast agents, and ultrasound pulsing schemes for contrast-enhanced ultrasound imaging, and to unravel the mechanisms of ultrasound-mediated drug delivery.

This protocol describes three methods on how to obtain and use 5 to 8-day old chicken embryos and their chorioallantoic membrane (CAM) as an in vivo&#160;model to study contrast-enhanced ultrasound imaging and microbubble-mediated drug delivery.

조영제 강화 초음파 영상 및 마이크로버블 매개 약물 전달 연구를 위한 생체 내 모델로서의 닭 Ex Ovo 배아 및 융모막 막 혈관의 준비

The chicken embryo and the blood-vessel rich chorioallantoic membrane (CAM) is a valuable in vivo model to investigate biomedical processes, new ...

생체공학

High-frequency Ultrasound Imaging of the Abdominal Aorta

Source: Amelia R. Adelsperger, Evan H. Phillips, and <a href="https://www-jove-com.bdigitaluss.remotexs.co/author/Craig+J._Goergen">Craig J. Goergen</a>, Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana
High-frequency ultrasound systems are used to acquire high resolution images. Here, the use of a state-of-the-art system will be demonstrated to image the morphology and hemodynamics of small pulsatile arteries and veins found in mice and rats. Ultrasound is a relatively inexpensive, portable, and versatile method for the noninvasive assessment of vessels in humans as well as large and small animals. These are several key advantages that ultraound offers compared to other techniques, such as computed tomography (CT), magnetic resonance imaging (MRI), and near-infrared fluorescence tomography (NIRF). CT requires ionizing radiation and MRI can be prohibitively expensive and even impractical in some scenarios. NIRF, on the other hand, is limited by the penetration depth of light required to excite the fluorescent contrast agents.
Ultrasound has limitations in terms of imaging depth; however, this may be overcome by sacrificing resolution and using a lower frequency transducer. Abdominal gas and excess body weight can severely diminish image quality. In the first case, the propagation of sound waves is limited, while in the latter case, they are attenuated by overlying tissues, such as fat and connective tissue. As a result, no contrast or faint contrast may be observed. Finally, ultrasound is a highly user-dependent technique, requiring the sonographer to be familiar with anatomy and to be able to work around issues, such as the appearance of imaging artifacts or acoustic interference.

High-frequency Ultrasound Imaging and Image Acquisition

복부 대동맥의 고주파 초음파 영상

Source: Amelia R. Adelsperger, Evan H. Phillips, and Craig J. Goergen, Weldon School of Biomedical Engineering, Purdue University, West Lafayette, 
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JoVE Science Education

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의공학

Injection of Microbubbles into Isolated Mouse Embryos: A Technique to Deliver Microbubble Contrast Agents into the Vasculature of Living Murine Embryos

Source: Denbeigh, J. M. et al. <a href="https://www-jove-com.bdigitaluss.remotexs.co/video/52520" target="_blank">Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound</a>. J. Vis. Exp. (2015)
This video demonstrates a procedure to inject microbubble contrast agents into an isolated late gestation stage mouse embryo. The microbubble contrast agent-based ultrasound imaging helps visualize and characterize the vascular environment of the embryo.

고립된 쥐 배아에 microbubbles의 주입: 살아있는 쥐 배아의 혈관 구조로 microbubble 조영제를 전달하는 기술

Source: Denbeigh, J. M. et al. Contrast Imaging in Mouse Embryos Using High-frequency Ultrasound. J. Vis. Exp. (2015)
This video demonstrates a procedure ...