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遵循德国法律的动物研究法规以及实验动物保育原则 (国立卫生研究院出版版 8, 2011)。官方许可来自政府动物护理办公室 (Landesamt Natur, Umwelt 和 Verbraucherschutz Nordrhein, Westfalen, 德国)。
1. 动物
<ol><li>使用雌性德国长白猪 (重25-30 公斤) 存放在开放的笼子里。</li>
	<li>每组使用5只动物 (氩和氦)。</li>
	<li>允许动物在实验前至少一周适应环境。快速动物为 24 h 在手术之前以自由地获得水。</li>
</ol>2. 麻醉
<ol><li>Premedicate 的动物肌肉注射氯胺酮 (15 毫克/千克体重 [体重]), 甲苯噻嗪 (10 毫克/千克体重), 和阿托品 (0.1 毫克/千克体重) 10 分钟前诱导麻醉。</li>
	<li>通过将22口径套管放置到耳静脉中, 可建立外周静脉通路。</li>
	<li>静脉注射丙泊酚2毫克/千克体重诱发全麻。</li>
	<li>将动物置于仰卧位, 在颈静脉沟内进行纵向皮肤切口, 长度为2厘米. 通过钝性的皮下组织的制备找到静脉。插入套管, 然后 Seldinger 导线。</li>
	<li>缩回套管并插入14法郎导管在导丝上。退刀导丝。将导管连接到延伸处, 用皮带或缝合固定导管。</li>
	<li>拔掉舌头, 插入直喉镜。使用喉镜的尖端拉下会厌。用声带插入管子。把袖口放在声门下, 充气。</li>
	<li>通风与40% 氧气在20-26 呼吸/分钟和潮汐容量10毫升/公斤保持末端潮汐部分二氧化碳紧张在36和42毫米汞之间。</li>
	<li>用异氟醚保持麻醉, 浓度为 1-1. 5%, 芬太尼浓度为3-4 µg/千克/小时。</li>
	<li>以4毫升/千克/小时的初始速率供应响铃的乳酸溶液, 在剖腹手术后增加8毫升/公斤/小时的恒定输液率。</li>
</ol>3. 手术和血浆凝固
<ol><li>将动物置于仰卧位, 放置在标准手术台上。</li>
	<li>使用标准的外科消毒剂 (2 丙醇45克/100 克, 1 丙醇10克/100g, Biphenyl-2-ol 0.2 克/100 克) 消毒皮肤, 用手术拭子治疗3次。</li>
	<li>用手术刀进行宽中线剖腹手术, 从剑突过程到耻骨, 并安装手术拉钩。</li>
	<li>开关在等离子凝固装置, 打开氩或氦气瓶, 取决于使用的载体气体。将气体流量调整到3升/分. 选择混凝设备输出功率根据需要。 注: 惰性气体、氩气或氦均可用于等离子凝固。凝固作用是可比较的。有关详细信息, 请参阅参考资料7 。</li>
	<li>在左肝叶上进行血浆凝固, 如前所述7。使用钛模 (方形孔径 1 x 1 厘米2) 来规范凝固区。凝聚为 5 s 与探针距离 1 cm。不同电源设置的 coagulations 可以并排执行, coagulations 之间的距离为5毫米 (图 1)。</li>
	<li>为了收割肝脏, 将所有韧带的连接与肝脏分开。分离和分割上十二指肠弯曲以上的肝蒂, 留下门静脉和胆总管的长部分。将静脉的静脉分在肝脏的上方和下方, 然后取出器官。</li>
	<li>在收获肝脏之后, 猪通过静脉注射管理被安乐死0.16 克/千克戊巴比妥。</li>
	<li>对于爆裂压力实验, 切除一半左内侧肝叶与锋利的剪刀。等离子凝固的切割表面 (100W 输出功率) 或密封的切割表面与纤维蛋白密封剂 (图 2)。</li>
</ol>4. 微循环测量
注: 激光多普勒光谱可以通过测量红细胞运动引起的多普勒移位来确定组织中的血流。激光信号与运动红细胞数目有关。激光多普勒技术是临床应用 (如移植医学), 并已验证了多次15。
<ol><li>开关激光多普勒流量计和分光光度计。使用平面探头。</li>
	<li>对流速和流速进行基线测量。保存或记下值。</li>
	<li>按照3.5 的描述进行凝固。</li>
	<li>将平探头放在凝固部位, 测量流速和流速。再次保存或记下值。</li>
	<li>对凝固设备的所有电源设置重复。</li>
</ol>5. 测温
<ol><li>切换系统 (红外相机、笔记本和红外线温度计), 并让它在执行测量之前至少运行1小时。</li>
	<li>调整焦点和视图框架在红外相机上凝固地点。红外序列可以检测到的空间分辨率为 1024年 x 768 像素, 温度分辨率大于 20 mK。考虑到, 凝血区和周围组织--受传热影响--位于视图中间。 注意: 它应该包含尽可能多的帧像素, 以实现最佳的空间分辨率。</li>
	<li>用红外相机在2分钟的时间内用等离子凝固在肝脏表面记录凝固过程。</li>
	<li>用热像分析软件分析图像序列: 定义感兴趣的区域。 注: 软件计算随时间推移的相应平均温度的过程。</li>
</ol>6. 混凝深度测量
<ol><li>用锋利的剪刀收割左内侧肝叶。</li>
	<li>用1厘米厚的凝固部位进行消费。切成3毫米厚的纵段进行进一步加工。</li>
	<li>用中性10% 缓冲福尔马林固定4摄氏度的组织样本。热石蜡2°c 在熔点和嵌入切片。进程过夜。</li>
	<li>执行苏木精/伊红染色。<ol>
<li>Deparaffinize 和水合组织随后浸泡在2x 二甲苯, 100% 乙醇 (乙醇), 95% 乙醇, 70% 乙醇, 去离子 H2O 为2分钟每。</li>
		<li>用迈耶的苏木精溶液染色组织样品3分钟。</li>
		<li>现在用自来水冲洗5分钟。</li>
		<li>用3分钟的伊红溶液染色组织。</li>
		<li>冲洗2x 乙醇 95%, 然后二甲苯3分钟。安装标准安装介质。</li>
	</ol>
</li>
	<li>切换系统 (显微镜连接的摄像头, 成像软件)。查看所有具有40X 放大倍数的部分。</li>
	<li>在放大40X 的情况下, 取一个物体千分尺的图像。在 "目标" 窗口中按 "重新校准" 按钮。选择手动校准。在100µm 的千分尺图像上画一条线. 在对话框中输入01毫米, 然后按 "确定"。</li>
	<li>在 "视图" > "分析控件" > "注释和测量" 窗口中选择 "长度"。测量从肝脏表面到凝固边缘与老鼠。导出或记下结果。对同一张幻灯片上的另一个位置重复测量。 注: 凝血深度可以很容易地与正常肝组织的区别, 由正常肝脏线和坏死区之间的明显缩小细胞质, 固缩核, 出血区。</li>
	<li>计算两个测量值的平均值。</li>
</ol>7. 爆破压力测量
<ol><li>切换系统 (自动泵, 压力表)。根据步骤3.7 准备肝脏标本。 注: 使用两个并联泵通过3路旋塞阀连接。单泵不能获得1500毫米汞的最大压力。</li>
	<li>用剪刀将门静脉、普通肝动脉和胆管隔离在触觉蒂内。钳门静脉与 overholt 钳和结扎 monofil 缝合4-0。钳共肝动脉 overholt 钳和结扎 monofil 缝合4-0。</li>
	<li>将 Ch-16 导管插入胆总管, 结扎2-0 丝缝合。将导管连接到自动泵, 安装3路旋塞阀与压力表 (图 3)。</li>
	<li>用生理盐水填满灌注注射器。</li>
	<li>启动自动泵, 交货率为99毫升/小时。</li>
	<li>监测肝切口表面和压力表的渗漏和记录爆裂压力。 注: 为便于识别泄漏, 专利蓝可以添加到生理盐水 (2 毫升专利蓝 + 18 毫升生理盐水)。通过注意压力表压力损失时间, 可以更容易地观察爆破压力。</li>
</ol>

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作者没有什么可透露的。

肝脏手术的啮齿动物模型建立了很长时间16。然而, 大型动物模型提供了一定的优势: 不需要显微外科设备作为人体标准手术设备, 手术技术可与临床使用相媲美, 标准的临床评价方法可以转移到实验。例如, 标准的临床血液测试可以进行, 而不需要特殊的实验室测试方法 (图 10)。
猪是适当的实验动物为心肺研究作为他们的生理相似人<sup c...

氩离子凝固 (APC) 是腹部手术中广泛使用的仪器, 三年来1、2。通过封堵肝切口表面, 防止术后出血3, 是大鼠肝切除术后继发止血的标准技术。等离子凝固是射频烙的一种特殊形式, 通过电离气体电弧传递电能。提供单电热止血, 这种非接触技术具有防止电极黏附在组织4的优势。电离气体光束被自动定向到最低电阻的区域, 当电阻上升时, 由于干燥到其他尚未干燥的区域, 就会被转走。这产生一个统一的有限深度的凝固5,6。影响凝血效果的因素有活化时间、凝固装置的功率设置以及探针与组织的距离。氦是另一种载体气体, 可用于等离子凝固7。最近的临床研究集中在临床结果, 而不是组织学和功能发现3,8,9, 而实验研究集中在体外调查10或实验在分离的被灌注的器官11。
基础协议允许研究在接近于临床应用的大型动物模型中的血浆凝固作用在猪附近使用标准人的设备: 微循环由激光多普勒流量计无创性评估和分光光度计, 这是一个标准的临床工具为这个征兆12,13。凝固过程中的温度变化用红外线温度计和红外相机进行监测。在采集组织样品后, 测定组织学上的苏木素/伊红染色切片的凝固深度。为与其他二次止血方法进行比较, 进行了爆破压力实验。与以前描述的技术14相比, 这些都是在说明器官上进行的, 以排除血压相关的影响。除了所述的对血浆凝固局部效应的研究外, 还可以在猪模型中进行标准血液检测。

<table border="0" cellpadding="0" cellspacing="0" width="824">
	<tbody>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Xylazine 20 mg/mL</td>
			<td style="width:184px;">Vetoquinol GmbH</td>
			<td style="width:192px;">Xylapan</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Ketamine 100 mg/mL</td>
			<td style="width:184px;">Ceva GmbH</td>
			<td style="width:192px;">Ceva Ketamine Injection</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Atropine 100 mg / 10 mL</td>
			<td style="width:184px;">Dr. Franz K&#246;hler Chemie GmbH</td>
			<td style="width:192px;">Atropinsulfat K&#246;hler 100mg Amp.</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;">Propofol</td>
			<td style="width:184px;">Fresenius Kabi GmbH</td>
			<td style="width:192px;">Propofol 1% MCT Fresenius</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;">Fentanyl</td>
			<td style="width:184px;">KG Rotexmedica GmbH</td>
			<td style="width:192px;">Fentanyl 0,5mg Rotexmedica</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;">Isoflurane</td>
			<td style="width:184px;">Abbot GmbH</td>
			<td style="width:192px;">Forene 100% (V/V) 250 mL</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;">Ringer&#39;s lactate solution</td>
			<td style="width:184px;">Baxter Deutschland GmbH</td>
			<td style="width:192px;"></td>
			<td style="width:232px;">sodium 131mmol/l, potassium 5 mmol/l, calcium 2 mmol/l, cloride 111 mmol/l, lactate 29 mmol/l</td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Surgical disinfactant</td>
			<td style="width:184px;">Sch&#252;lke &#38; Mayr GmbH</td>
			<td style="width:192px;">Kodan Tinktur forte gef&#228;rbt 1l 104804</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Motorized microscope</td>
			<td style="width:184px;">Nikon Instruments Europe</td>
			<td style="width:192px;">Eclipse TE2000-E</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Microscope camera</td>
			<td style="width:184px;">Nikon Instruments Europe</td>
			<td style="width:192px;">Digitalsight DS-Qi1Mc</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Imaging software</td>
			<td style="width:184px;">Nikon Instruments Europe</td>
			<td style="width:192px;">NIS elements Vers. 4.40</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Plasma coagulator</td>
			<td style="width:184px;">S&#246;ring GmbH</td>
			<td style="width:192px;">CPC-1000</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Argon gas</td>
			<td style="width:184px;">Linde AG</td>
			<td style="width:192px;">Argon 4.8&#160;</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Helium gas</td>
			<td style="width:184px;">Linde AG</td>
			<td style="width:192px;">Helium 4.8</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">O2C</td>
			<td style="width:184px;">LEA Medizintechnik GmbH</td>
			<td style="width:192px;">O2C Version 1212</td>
			<td style="width:232px;">with LF-2 or LF-3 probe</td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;">Infrared thermometer</td>
			<td style="width:184px;">Voltcraft</td>
			<td>VOLTCRAFT IR 260-8S</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Thermographic camera</td>
			<td>InfraTec GmbH</td>
			<td>VarioCAM HD head 820</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Thermographic analysis sofrtware</td>
			<td>InfraTec GmbH</td>
			<td style="width:192px;">IRBIS 3</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Mayer&#39;s Hematoxylin solution</td>
			<td></td>
			<td style="width:192px;">Merck 1.09249</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Eosin solution</td>
			<td style="width:184px;">VWR International GmbH</td>
			<td style="width:192px;">Merck 1.09844</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Rollerpump Masterflex L/S easy Load</td>
			<td style="width:184px;">Cole-Parmer Instrument Company</td>
			<td style="width:192px;">model 7518-10</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Perfusorpump</td>
			<td style="width:184px;">B. Braun Melsungen AG</td>
			<td style="width:192px;">Perfusor secura FT</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Digital pressure meter</td>
			<td style="width:184px;">Greisinger electronic</td>
			<td style="width:192px;">GMH 3161</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Perfusorsyringe, 50 mL</td>
			<td style="width:184px;">B. Braun Melsungen AG</td>
			<td style="width:192px;">REF 8728810 F</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Perfusor line, Type IV Standard, PVC Luer lock</td>
			<td style="width:184px;">B. Braun Melsungen AG</td>
			<td style="width:192px;">REF 8722960</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">3-Way stopcock, Dicofix C35C</td>
			<td style="width:184px;">B. Braun Melsungen AG</td>
			<td style="width:192px;">REF 16494 C</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Silk 2-0. 3 metric</td>
			<td style="width:184px;">Resorba</td>
			<td style="width:192px;">REF H5F</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">Vicryl 4-0 Sutupak</td>
			<td style="width:184px;">Ethicon</td>
			<td>V1224H</td>
			<td style="width:232px;"></td>
		</tr>
		<tr height="47">
			<td height="47" style="height:47px;width:216px;">NaCl 0.9 %</td>
			<td style="width:184px;">B. Braun Melsungen AG</td>
			<td style="width:192px;"></td>
			<td style="width:232px;"></td>
		</tr>
	</tbody>
</table>

assessment of plasma coagulation on liver tissue in a large animal model in vivo

血浆凝固作为一种烙的形式, 用于肝脏手术数十年, 以封闭大肝切除术后, 主要切除后, 以防止出血的后期阶段。血浆凝固对肝脏组织的确切的作用只被检查的很少。在我们的猪模型中, 凝血作用可以接近临床应用。联合激光多普勒流量计和分光光度计文件在8毫米组织深度无创性凝血过程中微循环的变化, 提供了超出主观临床印象的可量化的止血信息。在凝固过程中, 用红外线温度计对凝血部位的温度进行评估, 在凝固期间用红外相机测量气体束温度, 因为器件的上阈值是不可能的。用目标测微仪标定后的苏木精/红/红染色切片, 对凝固深度进行了显微测量, 并给出了功率设定-凝固深度关系的准确信息。在胆管上检查密封效果, 因为血浆凝固无法封堵较大的血管。对说明器官进行了爆裂压力实验, 排除了血压的相关影响。

微循环:通过对血浆凝固后止血的诊断装置, 可以通过微循环的改变来证明。毛细管血流 (显示为任意单位 (AU)) 从基线值减少 142.7, 76.08 au 到 57.78, 49.57 au 在 25 w 设备输出功率, 48.5 @ 7.26 au 在 50 w 和 5.04 @ 1.31 au 在 100 w (图 4)。
温度:用红外相机测量凝固部位的温度 (图 ...

Watch this Scientific Journal Video about Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo at JoVE.com

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

在这里, 我们提出了一个实验性的评估血浆凝固在体内的肝脏组织的协议。在猪模型中, 通过激光多普勒检查微循环, 用红外测温仪和红外相机对凝血深度进行组织学测量, 凝血部位温度, 并通过爆裂压力记录管道密封效果。实验。

大动物模型肝组织血浆凝固的评价

Plasma coagulation as a form of electrocautery is used in liver surgery for decades to seal the large liver cut surface after major hepatectomy to prevent ...

assessment-plasma-coagulation-on-liver-tissue-large-animal-model

Research

JoVE Journal

Medicine

Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

8.8K 次观看.  University of Bonn. 在这里, 我们提出了一个实验性的评估血浆凝固在体内的肝脏组织的协议。在猪模型中, 通过激光多普勒检查微循环, 用红外测温仪和红外相机对凝血深度进行组织学测量, 凝血部位温度, 并通过爆裂压力记录管道密封效果。实验。

视频: Assessment of Plasma Coagulation on Liver Tissue in a Large Animal Model In Vivo

An Isolated Working Heart System for Large Animal Models

Since its introduction in the late 19th century, the Langendorff isolated heart perfusion apparatus, and the subsequent development of the working heart model, have been invaluable tools for studying cardiovascular function and disease1-15. Although the Langendorff heart preparation can be used for any mammalian heart, most studies involving this apparatus use small animal models (e.g., mouse, rat, and rabbit) due to the increased complexity of systems for larger mammals1,3,11. One major difficulty is ensuring a constant coronary perfusion pressure over a range of different heart sizes &#8211; a key component of any experiment utilizing this device1,11. By replacing the classic hydrostatic afterload column with a centrifugal pump, the Langendorff working heart apparatus described below allows for easy adjustment and tight regulation of perfusion pressures, meaning the same set-up can be used for various species or heart sizes. Furthermore, this configuration can also seamlessly switch between constant pressure or constant flow during reperfusion, depending on the user&#8217;s preferences. The open nature of this setup, despite making temperature regulation more difficult than other designs, allows for easy collection of effluent and ventricular pressure-volume data.

Most studies involving the Langendorff apparatus use small animal models due to the increased complexity of systems for larger mammals. We describe a Langendorff system for large animal models that allows for use across a range of species, including humans, and relatively easy data acquisition.

一个孤立的工作心脏系统的大型动物模型

Since its introduction in the late 19th century, the Langendorff isolated heart perfusion apparatus, and the subsequent development of the working heart ...

科研

医学

Depletion of Mouse Cells from Human Tumor Xenografts Significantly Improves Downstream Analysis of Target Cells

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic patient tumors in different assays1. Human tumor xenografts represent the gold standard with respect to tumor biology, drug discovery, and metastasis research 2-4. Tumor xenografts can be derived from different types of material like tumor cell lines, tumor tissue from primary patient tumors4 or serially transplanted tumors. When propagated in vivo, xenografted tissue is infiltrated and vascularized by cells of mouse origin. Multiple factors such as the tumor entity, the origin of xenografted material, growth rate and region of transplantation influence the composition and the amount of mouse cells present in tumor xenografts. However, even when these factors are kept constant, the degree of mouse cell contamination is highly variable.
Contaminating mouse cells significantly impair downstream analyses of human tumor xenografts. As mouse fibroblasts show high plating efficacies and proliferation rates, they tend to overgrow cultures of human tumor cells, especially slowly proliferating subpopulations. Mouse cell derived DNA, mRNA, and protein components can bias downstream gene expression analysis, next-generation sequencing, as well as proteome analysis 5. To overcome these limitations, we have developed a fast and easy method to isolate untouched human tumor cells from xenografted tumor tissue. This procedure is based on the comprehensive depletion of cells of mouse origin by combining automated tissue dissociation with the benchtop tissue dissociator and magnetic cell sorting. Here, we demonstrate that human target cells can be can be obtained with purities higher than 96% within less than 20 min independent of the tumor type.

Human tumor xenografts are vascularized and infiltrated by cells of mouse origin during the growth phase in vivo. To circumvent the bias caused by these contaminating cells during downstream analysis, we developed a method allowing for comprehensive depletion of all mouse cells by magnetic cell sorting.

从人类肿瘤异种移植的小鼠细胞的消耗显着改善靶细胞下游分析

The use of in vitro cell line models for cancer research has been a useful tool. However, it has been shown that these models fail to reliably mimic ...

Assessment of Child Anthropometry in a Large Epidemiologic Study

A high proportion of children have overweight and obesity in the United States and other countries. Accurate assessment of anthropometry is essential to understand health effects of child growth and adiposity. Gold standard methods of measuring adiposity, such as dual X-ray absorptiometry (DXA), may not be feasible in large field studies. Research staff can, however, complete anthropometric measurements, such as body circumferences and skinfold measurements, using inexpensive portable equipment. In this protocol we detail how to obtain manual anthropometric measurements from children, including standing and sitting height, weight, waist circumference, hip circumference, mid-upper arm circumference, triceps skinfold thickness, and subscapular skinfold thickness, and procedures to assess the quality of these measurements. To demonstrate accuracy of these measurements, among 1,110 school-aged children in the pre-birth cohort Project Viva we calculated Spearman correlation coefficients comparing manual anthropometric measurements with a gold standard measure of body fat, DXA fat mass1. To address reliability, we evaluate intra-rater technical error of measurement at a quality control session conducted on adult female volunteers.

In epidemiologic studies of children, well-trained research staff can accurately and precisely assess weight, height, sitting height, skinfold thicknesses, and body circumferences.

大型流行病学研究儿童人体测量评估

A high proportion of children have overweight and obesity in the United States and other countries. Accurate assessment of anthropometry is essential to ...

Surgical Angiogenesis in Porcine Tibial Allotransplantation: A New Large Animal Bone Vascularized Composite Allotransplantation Model

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic options have significant risk of failure and substantial morbidity.
Use of bone vascularized composite allotransplantation (VCA) would offer both a close match of resected bone size and shape and the healing and remodeling potential of living bone. At present, life-long drug immunosuppression (IS) is required. Organ toxicity, opportunistic infection and neoplasm risks are of concern to treat such non-lethal indications.
We have previously demonstrated that bone and joint VCA viability may be maintained in rats and rabbits without the need of long-term-immunosuppression by implantation of recipient derived vessels within the VCA. It generates an autogenous, neoangiogenic circulation with measurable flow and active bone remodeling, requiring only 2 weeks of IS. As small animals differ from man substantially in anatomy, bone physiology and immunology, we have developed a porcine bone VCA model to evaluate this technique before clinical application is undertaken. Miniature swine are currently widely used for allotransplantation research, given their immunologic, anatomic, physiologic and size similarities to man. Here, we describe a new porcine orthotopic tibial bone VCA model to test the role of autogenous surgical angiogenesis to maintain VCA viability.
The model reconstructs segmental tibial bone defects using size- and shape-matched allogeneic tibial bone segments, transplanted across a major swine leukocyte antigen (SLA) mismatch in Yucatan miniature swine. Nutrient vessel repair and implantation of recipient derived autogenous vessels into the medullary canal of allogeneic tibial bone segments is performed in combination with simultaneous short-term IS. This permits a neoangiogenic autogenous circulation to develop from the implanted tissue, maintaining flow through the allogeneic nutrient vessels for a short time. Once established, the new autogenous circulation maintains bone viability following cessation of drug therapy and subsequent nutrient vessel thrombosis.

Currently any kind of vascularized composite allotransplantation depends on long-term-immunosuppression, difficult to support for non-life-critical indications. We present a new porcine tibial VCA model that can be used to study bone VCA and demonstrate the use of surgical angiogenesis to maintain bone viability without the need of long-term immune-modulation.

猪胫骨移植术后血管生成的动物模型

Segmental bone loss resulting from trauma, infection malignancy and congenital anomaly remains a major reconstructive challenge. Current therapeutic ...

Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to pulmonary hypertension. For a better understanding and treatment of this disease, precise hemodynamic monitoring of left and right ventricular parameters is important. For this reason, it is essential to establish experimental pig models of cardiac hemodynamics and measurements for research purpose. 
This article shows the induction of ARDS by using oleic acid (OA) and consequent right ventricular dysfunction, as well as the instrumentation of the pigs and the data acquisition process that is needed to assess hemodynamic parameters. To achieve right ventricular dysfunction, we used oleic acid (OA) to cause ARDS and accompanied this with pulmonary artery hypertension (PAH). With this model of PAH and consecutive right ventricular dysfunction, many hemodynamic parameters can be measured, and right ventricular volume load can be detected. 
All vital parameters, including respiratory rate (RR), heart rate (HR) and body temperature were recorded throughout the whole experiment. Hemodynamic parameters including femoral artery pressure (FAP), aortic pressure (AP), right ventricular pressure (peak systolic, end systolic and end diastolic right ventricular pressure), central venous pressure (CVP), pulmonary artery pressure (PAP) and left arterial pressure (LAP) were measured as well as perfusion parameters including ascending aortic flow (AAF) and pulmonary artery flow (PAF). Hemodynamic measurements were performed using transcardiopulmonary thermodilution to provide cardiac output (CO). Furthermore, the PiCCO2 system (Pulse Contour Cardiac Output System 2) was used to receive parameters such as stroke volume variance (SVV), pulse pressure variance (PPV), as well as extravascular lung water (EVLW) and global end-diastolic volume (GEDV). Our monitoring procedure is suitable for detecting right ventricular dysfunction and monitoring hemodynamic findings before and after volume administration.

We present a protocol of creating right ventricular dysfunction in a pig model by inducing ARDS. We demonstrate invasive monitoring of left and right ventricular cardiac output using flow probes around the aorta and the pulmonary artery, as well as blood pressure measurements in the aorta and pulmonary artery.

ards 大型动物模型主动脉和肺动脉血流动力学的侵袭血流动力学监测

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to ...

A Small Animal Model of Ex Vivo Normothermic Liver Perfusion

There is a significant shortage of liver allografts available for transplantation, and in response the donor criteria have been expanded. As a result, normothermic ex vivo liver perfusion (NEVLP) has been introduced as a method to evaluate and modify organ function. NEVLP has many advantages in comparison to hypothermic and subnormothermic perfusion including reduced preservation injury, restoration of normal organ function under physiologic conditions, assessment of organ performance, and as a platform for organ repair, remodeling, and modification. Both murine and porcine NEVLP models have been described. We demonstrate a rat model of NEVLP and use this model to show one of its important applications &#8212; the use of a therapeutic molecule added to liver perfusate. Catalase is an endogenous reactive oxygen species (ROS) scavenger and has been demonstrated to decrease ischemia-reperfusion in the eye, brain, and lung. Pegylation has been shown to target catalase to the endothelium. Here, we added pegylated-catalase (PEG-CAT) to the base perfusate and demonstrated its ability to mitigate liver preservation injury. An advantage of our rodent NEVLP model is that it is inexpensive in comparison to larger animal models. A limitation of this study is that it does not currently include post-perfusion liver transplantation. Therefore, prediction of the function of the organ post-transplantation cannot be made with certainty. However, the rat liver transplant model is well established and certainly could be used in conjunction with this model. In conclusion, we have demonstrated an inexpensive, simple, easily replicable NEVLP model using rats. Applications of this model can include testing novel perfusates and perfusate additives, testing software designed for organ evaluation, and experiments designed to repair organs.

A Small Animal Model of Ex Vivo Normothermic Liver Perfusion

There is a significant liver donor shortage, and criteria for liver donors have been expanded. Normothermic ex vivo liver perfusion (NEVLP) has been developed to evaluate and modify organ function. This study demonstrates a rat model of NEVLP and tests the ability of pegylated-catalase, to mitigate liver preservation injury.

体外常温肝灌注的小动物模型

There is a significant shortage of liver allografts available for transplantation, and in response the donor criteria have been expanded. As a result, ...

Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, we have translated a vascular injury model from a small animal into a large animal (canine), with slight modifications that can be used for pre-clinical screening of prophylactic and thrombolytic agents. In addition to the surgical methods, the modified protocol describes the step-by-step methods to assess carotid artery canalization by angiography, detailed instructions to process both the brain and carotid artery for histological analysis to verify carotid canalization and cerebral hemorrhage, and specific parameters to complete an assessment of downstream thromboembolic events by utilizing magnetic resonance imaging (MRI). In addition, specific procedural changes from the previously well-established small animal model necessary to translate into a large animal (canine) vascular injury are discussed.

Here, we present the modifications necessary to a well characterized and commonly used small animal ferric chloride-induced (FeCl3) carotid artery injury model for use in a large animal vascular injury model. The resulting model can be utilized for pre-clinical trial assessment of both prophylactic and thrombolytic pharmacological and mechanical interventions.

氯化铁诱导的犬颈动脉血栓形成: 一种大的血管损伤动物模型

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

用线 Myography 测定小鼠肠系膜动脉的等轴收缩力

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

In Vitro Model of Coronary Angiogenesis

Here, we describe an in vitro culture assay to study coronary angiogenesis. Coronary vessels feed the heart muscle and are of clinical importance. Defects in these vessels represent severe health risks such as in atherosclerosis, which can lead to myocardial infarctions and heart failures in patients. Consequently, coronary artery disease is one of the leading causes of death worldwide. Despite its clinical importance, relatively little progress has been made on how to regenerate damaged coronary arteries. Nevertheless, recent progress has been made in understanding the cellular origin and differentiation pathways of coronary vessel development. The advent of tools and technologies that allow researchers to fluorescently label progenitor cells, follow their fate, and visualize progenies in vivo have been instrumental in understanding coronary vessel development. In vivo studies are valuable, but have limitations in terms of speed, accessibility, and flexibility in experimental design. Alternatively, accurate in vitro models of coronary angiogenesis can circumvent these limitations and allow researchers to interrogate important biological questions with speed and flexibility. The lack of appropriate in vitro model systems may have hindered the progress in understanding the cellular and molecular mechanisms of coronary vessel growth. Here, we describe an in vitro culture system to grow coronary vessels from the sinus venosus (SV) and endocardium (Endo), the two progenitor tissues from which many of the coronary vessels arise. We also confirmed that the cultures accurately recapitulate some of the known in vivo mechanisms. For instance, we show that the angiogenic sprouts in culture from SV downregulate COUP-TFII expression similar to what is observed in vivo. In addition, we show that VEGF-A, a well-known angiogenic factor in vivo, robustly stimulates angiogenesis from both the SV and Endo cultures. Collectively, we have devised an accurate in vitro culture model to study coronary angiogenesis.

In vitro models of coronary angiogenesis can be utilized for the discovery of the cellular and molecular mechanisms of coronary angiogenesis. In vitro explant cultures of sinus venosus and endocardium tissues show robust growth in response to VEGF-A and display a similar pattern of COUP-TFII expression as in vivo.

冠状血管生成体模型

Here, we describe an in vitro culture assay to study coronary angiogenesis. Coronary vessels feed the heart muscle and are of clinical importance. Defects ...

Using a Chemical Biopsy for Graft Quality Assessment

Kidney transplantation is a life-saving treatment for a large number of people with end-stage renal dysfunction worldwide. The procedure is associated with an increased survival rate and greater quality of patient&#8217;s life when compared to conventional dialysis. Regrettably, transplantology suffers from a lack of reliable methods for organ quality assessment. Standard diagnostic techniques are limited to macroscopic appearance inspection or invasive tissue biopsy, which do not provide comprehensive information about the graft. The proposed protocol aims to introduce solid phase microextraction (SPME) as an ideal analytical method for comprehensive metabolomics and lipidomic analysis of all low molecular compounds present in kidneys allocated for transplantation. The small size of the SPME probe enables performance of a chemical biopsy, which enables extraction of metabolites directly from the organ without any tissue collection. The minimum invasiveness of the method permits execution of multiple analyses over time: directly after organ harvesting, during its preservation, and immediately after revascularization at the recipient&#8217;s body. It is hypothesized that the combination of this novel sampling method with a high-resolution mass spectrometer will allow for discrimination of a set of characteristic compounds that could serve as biological markers of graft quality and indicators of possible development of organ dysfunction.

The protocol presents utilization of the chemical biopsy approach followed by comprehensive metabolomic and lipidomic analysis for quality assessment of kidney grafts allocated for transplantation.

使用化学活检进行移植质量评估

Kidney transplantation is a life-saving treatment for a large number of people with end-stage renal dysfunction worldwide. The procedure is associated ...