Name: a rat model of mild intrauterine hypoperfusion with microcoil stenosis
Uploaded: 2018-01-07T14:00:00.000+00:00
Duration: 6 min 19 s
Description: Watch this Scientific Journal Video about A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis at JoVE.com

这项工作得到了2013财政年度在 jsp 和 CNRS, jsp KAKENHI 授予数字 26860858, 和 Narishige 神经科学研究基金会的研究交流计划的支持。我们感谢 Drs 子和京子盐谷进行了有益的讨论。我们感谢玛丽 Furuta, 睦坂, 律希, 和 Dr. Emi 田中的优秀技术援助。

所有的实验都是按照日本吹国家脑和心血管中心实验动物保育和使用委员会批准的协议进行的。
1. 为 MIUH 手术准备下列动物和材料
<ol><li>在妊娠日 17, 即胚胎日 17 (E17), 准备定时妊娠大鼠。水坝的平均身体重量是307.0 ± 40.7 g (平均± SD, n = 9)。</li>
	<li>准备金属弹簧;内径0.24 毫米, 由金涂层钢制成。</li>
	<li>根据材料表准备所有材料。</li>
</ol>2. 准备 MIUH 手术
<ol><li>在37° c 的大鼠安置处加热热垫</li>
	<li>在暖气垫上放一个无菌尿布</li>
	<li>温水浴37° c 温无菌盐水</li>
</ol>3. 进行 MIUH 手术
<ol><li>在无菌状态下进行手术。使用前用70% 乙醇消毒线圈、镊子、针。使用无菌手套, 偶尔用乙醇喷雾消毒。</li>
	<li>把老鼠放在麻醉感应箱里将4% 异氟醚引入箱内 (约5分钟)。</li>
	<li>在感应箱内彻底麻醉动物后, 将老鼠放在加热垫上的无菌尿布上, 放在仰卧位。检查深度的麻醉, 缺乏对脚趾捏的反应。用鼻锥维持 1.5-2.0% 的异氟醚。</li>
	<li>应用脱毛泡沫到腹部从肚脐向下到区域的骨盆拱。几分钟后, 用纸巾擦拭脱毛泡沫。剃须刀可用作脱毛泡沫的替代品。</li>
	<li>将纱布浸泡在碘消毒剂溶液中, 脱毛皮肤区域。</li>
	<li>在手术切口的区域以外放置一个圆形开口覆盖腹部的外科手术。几块纱布可以用来代替褶皱。</li>
	<li>使用外科手术刀, 使切口大约2.5 厘米长在下腹部从耻骨的上部边缘直接地往肚脐。然后, 通过下面的肌肉层进行切口。</li>
	<li>在褶皱的开口周围放置几片无菌纱布, 用温热的盐水浸湿纱布。</li>
	<li>轻轻推腹两侧, 使子宫角部分从切口中脱出。</li>
	<li>用70% 的乙醇消毒外科医生的手。</li>
	<li>在不使用镊子的情况下, 手动从腹腔轻轻拉出整个子宫角。使用手术钳是不可取的, 因为他们可能很容易破坏脆弱的子宫, 胎儿和血管通过压缩他们强烈和不均匀。</li>
	<li>将子宫角放在湿纱布上。</li>
	<li>为其他子宫角执行相同的程序。</li>
	<li>计算胎儿的数量。</li>
	<li>经常使用温盐水, 使子宫在整个手术过程中保持温暖和湿润。<ol>
<li>换尿布时, 它已经吸收了太多的盐水。为了保持稳定, 重复消毒手套与70% 乙醇和仪器的提示与碘。保持消毒手指远离仪器的无菌提示是至关重要的。</li>
	</ol>
</li>
	<li>在显微镜下确定卵巢和子宫动脉主拱廊的下部。 注意: 重要的是要区分主拱廊从分支到胎盘。请注意, 当子宫角翻转时, 手术操作可能更容易。偶尔地, 在拱廊的极近的部分应用线圈是困难的, 因为它位于深在腹腔之内, 并且动脉到胎盘是非常接近的近的部分拱廊。在这种情况下, 一个线圈可以应用到一个部分的拱廊后的第一个分支叉。</li>
	<li>将动脉从与它平行的静脉中分离出来, 并在它们之间制造空间。</li>
	<li>在动脉下面放置一根绳子 (丝缝 4.0, 5 厘米长)。</li>
	<li>用镊子把绳子的两端抬起, 使动脉从脆弱的静脉中分离出来。</li>
	<li>另一方面, 用细钳夹住圈的边缘, 在提升的动脉旁放置一个圈 (图 2. 1)。</li>
	<li>用圈周围的绳子把圈绕到动脉周围来旋转抬起的动脉。在线圈周围旋转动脉比旋转动脉周围的线圈更容易 (图 2. 2)。</li>
	<li>在旋转动脉3或4匝 (线圈有5匝) 后, 改变线圈被保持到另一边缘的位置。抓住缠绕在动脉上的线圈的边缘, 用镊子夹住。改变的网站的字符串升降机的动脉到另一侧的线圈。围绕线圈旋转动脉, 使线圈完全 (5 匝) 包裹动脉 (图 2. 3, 2.4)。</li>
	<li>删除字符串。</li>
	<li>对其他三动脉进行同样的手术。</li>
	<li>注意, 经常使用温盐水到子宫是至关重要的;否则, 胎儿的死亡率会增加, 体温过低可能是神经保护。</li>
	<li>将子宫角轻轻地放回腹腔。用镊子提起腹壁切口的边缘可以使这个过程更容易。</li>
	<li>施蒂希腹肌, 再施蒂希腹皮缝合 (丝缝 4.0)。在每施蒂希后缝合缝线, 即间断的单缝合线。</li>
	<li>在外科手术切口上和周围涂抹碘消毒液浸泡纱布。</li>
	<li>在0.5 毫克/千克体重的皮下施行镇痛昔。</li>
	<li>允许30分钟的恢复在温暖的笼子里。检查动物是否完全清醒, 并在笼子周围移动, 回到家里的笼子。</li>
</ol>

<ol>
	<li>Ohshima, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mild+intrauterine+hypoperfusion+reproduces+neurodevelopmental+disorders+observed+in+prematurity.">Mild intrauterine hypoperfusion reproduces neurodevelopmental disorders observed in prematurity.</a> Sci Rep. 6, 39377 (2016).</li><li>Anderson, P., Doyle, L. W., Victorian Infant Collaborative Study, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurobehavioral+outcomes+of+school-age+children+born+extremely+low+birth+weight+or+very+preterm+in+the+1990s.">Neurobehavioral outcomes of school-age children born extremely low birth weight or very preterm in the 1990s.</a> JAMA. 289 (24), 3264-3272 (2003).</li><li>Levine, T. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+childhood+neurodevelopment+after+intrauterine+growth+restriction:+a+systematic+review.">Early childhood neurodevelopment after intrauterine growth restriction: a systematic review.</a> Pediatrics. 135 (1), 126-141 (2015).</li><li>Sucksdorff, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preterm+Birth+and+Poor+Fetal+Growth+as+Risk+Factors+of+Attention-Deficit/+Hyperactivity+Disorder.">Preterm Birth and Poor Fetal Growth as Risk Factors of Attention-Deficit/ Hyperactivity Disorder.</a> Pediatrics. 136 (3), e599-e608 (2015).</li><li>Volpe, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brain+injury+in+premature+infants:+a+complex+amalgam+of+destructive+and+developmental+disturbances.">Brain injury in premature infants: a complex amalgam of destructive and developmental disturbances.</a> Lancet Neurol. 8 (1), 110-124 (2009).</li><li>Nardozza, L. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fetal+growth+restriction:+current+knowledge.">Fetal growth restriction: current knowledge.</a> Arch Gynecol Obstet. 295 (5), 1061-1077 (2017).</li><li>Chang, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preterm+birth+and+the+role+of+neuroprotection.">Preterm birth and the role of neuroprotection.</a> Bmj. 350, g6661 (2015).</li><li>Coq, J. O., Delcour, M., Massicotte, V. S., Baud, O., Barbe, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prenatal+ischemia+deteriorates+white+matter,+brain+organization,+and+function:+implications+for+prematurity+and+cerebral+palsy.">Prenatal ischemia deteriorates white matter, brain organization, and function: implications for prematurity and cerebral palsy.</a> Dev Med Child Neurol. 58, 7-11 (2016).</li><li>Jantzie, L. L., Corbett, C. J., Firl, D. J., Robinson, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postnatal+Erythropoietin+Mitigates+Impaired+Cerebral+Cortical+Development+Following+Subplate+Loss+from+Prenatal+Hypoxia-Ischemia.">Postnatal Erythropoietin Mitigates Impaired Cerebral Cortical Development Following Subplate Loss from Prenatal Hypoxia-Ischemia.</a> Cereb Cortex. 25 (9), 2683-2695 (2015).</li><li>Kubo, K. I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Association+of+impaired+neuronal+migration+with+cognitive+deficits+in+extremely+preterm+infants.">Association of impaired neuronal migration with cognitive deficits in extremely preterm infants.</a> JCI Insight. 2 (10), (2017).</li><li>Delcour, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mild+musculoskeletal+and+locomotor+alterations+in+adult+rats+with+white+matter+injury+following+prenatal+ischemia.">Mild musculoskeletal and locomotor alterations in adult rats with white matter injury following prenatal ischemia.</a> Int J Dev Neurosci. 29 (6), 593-607 (2011).</li><li>Gilbert, J. S., Babcock, S. A., Granger, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hypertension+produced+by+reduced+uterine+perfusion+in+pregnant+rats+is+associated+with+increased+soluble+fms-like+tyrosine+kinase-1+expression.">Hypertension produced by reduced uterine perfusion in pregnant rats is associated with increased soluble fms-like tyrosine kinase-1 expression.</a> Hypertension. 50 (6), 1142-1147 (2007).</li><li>Granger, J. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reduced+uterine+perfusion+pressure+(RUPP)+model+for+studying+cardiovascular-renal+dysfunction+in+response+to+placental+ischemia.">Reduced uterine perfusion pressure (RUPP) model for studying cardiovascular-renal dysfunction in response to placental ischemia.</a> Methods Mol Med. 122, 383-392 (2006).</li><li>Mazur, M., Miller, R. H., Robinson, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Postnatal+erythropoietin+treatment+mitigates+neural+cell+loss+after+systemic+prenatal+hypoxic-ischemic+injury.">Postnatal erythropoietin treatment mitigates neural cell loss after systemic prenatal hypoxic-ischemic injury.</a> J Neurosurg Pediatr. 6 (3), 206-221 (2010).</li><li>Olivier, P., Baud, O., Evrard, P., Gressens, P., Verney, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prenatal+ischemia+and+white+matter+damage+in+rats.">Prenatal ischemia and white matter damage in rats.</a> J Neuropathol Exp Neurol. 64 (11), 998-1006 (2005).</li><li>Robinson, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+changes+induced+by+graded+prenatal+systemic+hypoxic-ischemic+insults+in+rats.">Developmental changes induced by graded prenatal systemic hypoxic-ischemic insults in rats.</a> Neurobiol Dis. 18 (3), 568-581 (2005).</li><li>Rice, D., Barone, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Critical+periods+of+vulnerability+for+the+developing+nervous+system:+evidence+from+humans+and+animal+models.">Critical periods of vulnerability for the developing nervous system: evidence from humans and animal models.</a> Environ Health Perspect. 108, 511-533 (2000).</li><li>Basilious, A., Yager, J., Fehlings, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurological+outcomes+of+animal+models+of+uterine+artery+ligation+and+relevance+to+human+intrauterine+growth+restriction:+a+systematic+review.">Neurological outcomes of animal models of uterine artery ligation and relevance to human intrauterine growth restriction: a systematic review.</a> Dev Med Child Neurol. 57 (5), 420-430 (2015).</li><li>Delcour, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuroanatomical,+sensorimotor+and+cognitive+deficits+in+adult+rats+with+white+matter+injury+following+prenatal+ischemia.">Neuroanatomical, sensorimotor and cognitive deficits in adult rats with white matter injury following prenatal ischemia.</a> Brain Pathol. 22 (1), 1-16 (2012).</li></ol>

作者没有什么可透露的。

两个子宫角的卵巢和子宫动脉的圈狭窄在所有胎盘和胎儿中产生一致和可再生的宫内灌注。灌注的水平可以通过使用不同内径的弹簧进行修改。大鼠幼崽出生于一个大坝上, 其中动脉狭窄已执行与弹簧0.24 毫米内直径显示 IUGR 和早产 (详见参考 1)。幼崽表现出神经和行为改变, 类似于早产儿所观察到的体征和症状。这些神经的改变包括心室扩大, 皮质和胼胝体变薄, 脑重量减少, 但没有明显的细胞丢?...

胎儿宫内生长受限 (IUGR) (也称为胎生长限制) (出生体重 < 10th百分比为孕龄), 早产 (出生在 < 37 周妊娠), 和/或低出生体重 (< 2500 g) 的帐户新生儿的近 10% 2,3。许多这些婴儿存在神经问题, 如脑瘫和发育障碍 (如, 注意力缺陷/多动障碍 (ADHD) 和学习障碍) 3,4,5。这些条件在其病因和结局上有相同和不同之处。IUGR 的病因是多因素, 胎盘不足与宫内灌注被认为是最常见的原因, non-anomalous 胎儿7。早产的病因也多为多因素, 而炎是最常见的原因8。
轻度宫内灌注 (MIUH) 对大脑发育的影响尚不清楚。目前可用的动物模型宫内灌注/缺血主要涉及严重灌注或梯度程度的灌注有或没有再灌注9,10,11。然而, 在临床环境中, MIUH 的病例被认为比涉及这种严重疾病的情况更为频繁。目前可用的 MIUH 模型是一种啮齿类动物模型, 涉及子宫或卵巢动脉的缝合结扎, 以及一种涉及将双侧卵巢动脉和主动脉夹紧的啮齿动物模型, 其有通畅12,13, 14、15、16、17。这些模型的缺点之一是广泛的 inter-fetus 变异性, 从胎儿与深刻的灌注到胎儿与几乎完整的灌注, 取决于胎儿的位置在子宫动脉拱廊和卵巢动脉。这些模型的另一个缺点是他们无法区分出生后每个胎儿的位置;因此, 研究人员不能区分出生后个体小狗所经历的宫内灌注的严重性。
我们已经开发了一个大鼠模型的 MIUH 涉及多动脉狭窄1。环绕在卵巢和子宫动脉周围的直径0.24 毫米的金属微线圈会导致血管狭窄, 但不会阻塞 (图 1)。应用这些弹簧在所有供子宫的动脉的近端, 即双侧子宫和卵巢动脉, 在胚胎日 17 (相当于胚胎周20-25 在人类18) 引起显著但轻度的减少血液流向胎盘和胎儿线圈后血流的减少被应用到所有四个供子宫的动脉在每个胎盘和胎儿中基本上是相同的。胎儿死亡率低于20%。幼崽是通过自发分娩1-2 天前 (胚胎日 21-22) 比正常出生。几乎所有的幼崽出生都显着低出生体重1。灰色和白色物质体积减少, 没有明显的组织损伤1。幼崽出现延迟获得新生儿反应, 肌肉无力, 并改变自发活动1。该模型模仿早产或 IUGR 的儿童的临床症状和体征;儿童出生早产儿表现为灰质和白质体积减少或无白质损伤6, 目前神经发育延迟的里程碑, 并可能出现行为问题, 如 ADHD 的3,5;儿童与 IUGR 表现出最小的神经的变化, 并有增加的风险, 神经发育障碍, 如马达和认知延迟4,7。早产和 IUGR 是不同的条件, 但这两种情况具有基本的机制, 即在足月孕龄前对未成熟的大脑进行侮辱。

<table><tbody><tr><td>Stereomicroscope</td><td></td><td></td><td></td></tr><tr><td>Isoflurane anesthesia machine</td><td></td><td></td><td></td></tr><tr><td>Anesthesia induction box</td><td></td><td></td><td></td></tr><tr><td>Heating pad</td><td></td><td></td><td></td></tr><tr><td>Diaper 30x40 cm</td><td></td><td></td><td></td></tr><tr><td>Depilatory foam or shaver</td><td></td><td></td><td></td></tr><tr><td>Iodine disinfectant solution</td><td></td><td></td><td></td></tr><tr><td>Gauze 10x20 cm&amp;nbsp;</td><td></td><td></td><td></td></tr><tr><td>Surgical drape 45x45 cm with a round opening 5 cm in diameter</td><td></td><td></td><td></td></tr><tr><td>Spray bottle with ethanol for disinfection</td><td></td><td></td><td></td></tr><tr><td>Cotton swab</td><td></td><td></td><td></td></tr><tr><td>Forceps with large blunt tips</td><td></td><td></td><td></td></tr><tr><td>Forceps with angled fine tips</td><td></td><td></td><td></td></tr><tr><td>Scissors</td><td></td><td></td><td></td></tr><tr><td>Surgical scalpel, blade size is 27mm long (no.10, Axel, AS ONE Corporation, Osaka, Japan)</td><td></td><td></td><td></td></tr><tr><td>Surgical suture needle</td><td></td><td></td><td></td></tr><tr><td>Metal microcoils; inner diameter 0.24 mm, made from gold-coated steel (SAMINI Co. Ltd., Shizuoka, Japan)</td><td></td><td></td><td></td></tr><tr><td>Silk suture 4-0</td><td></td><td></td><td></td></tr><tr><td>Sterile saline (0.9% sodium chloride)</td><td></td><td></td><td></td></tr><tr><td>Heating water bath</td><td></td><td></td><td></td></tr><tr><td>Plastic syringes (50ml) and needles (18G)</td><td></td><td></td><td></td></tr></tbody></table>

a rat model of mild intrauterine hypoperfusion with microcoil stenosis

宫内灌注/缺血是宫内/胎儿生长受限、早产和低出生体重的主要原因之一。这一现象的大多数研究都是在严重的宫内缺血模型中进行的, 或者是子宫灌注的梯度度模型。未对均匀轻度宫内灌注 (MIUH) 模型进行研究。两个模型已用于研究 MIUH: 一个模型的基础上, 两侧的动脉拱廊形成的子宫和卵巢动脉, 和一个瞬态模型的基础上修剪双侧卵巢动脉和主动脉有通畅。这两个啮齿动物模型的 MIUH 有一定的局限性, 例如, 不是所有的胎儿都受到 MIUH, 取决于他们在子宫角的位置。在我们的 MIUH 模型中, 所有胎儿的宫内灌注。MIUH 是通过轻度狭窄的所有四动脉喂养子宫, 即, 双侧子宫和卵巢动脉。
动脉狭窄是由金属弹簧包裹在喂养动脉周围。产生动脉狭窄与弹簧允许我们控制, 优化和繁殖减少血流量, 很少 inter-animal 变异性和低死亡率, 从而使准确的评价。当使用内径为0.24 毫米的弹簧时, 胎盘和胎儿的血流轻度减少 (约30% 来自胎盘的 pre-stenosis 水平)。我们的 MIUH 模型的后代清楚地显示了神经、神经和行为测试结果的长期变化。

在将弹簧应用于所有供子宫的动脉, 即双侧子宫和卵巢动脉后, 所有胎儿都受到了类似的灌注。内直径为0.24 毫米的弹簧的应用导致了这些动脉的轻度狭窄, 从而导致胎盘和胎儿的血流轻度减少 (图 3; 大约30% 从胎盘的 pre-stenosis 水平, 看见参考1的详细信息)。从大坝出生的幼崽演示 IUGR (表 1)。
?...

Watch this Scientific Journal Video about A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis at JoVE.com

A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis

轻度宫内灌注是由动脉狭窄, 金属弹簧包裹周围的子宫和卵巢动脉在大鼠胚胎日17。这一程序产生了产前灌注和宫内生长的限制。

轻度宫内灌注圈狭窄的大鼠模型

Intrauterine hypoperfusion/ischemia is one of the major causes of intrauterine/fetal growth restriction, preterm birth, and low birth weight. Most studies ...

a-rat-model-of-mild-intrauterine-hypoperfusion-with-microcoil-stenosis

Research

JoVE Journal

Neuroscience

10.2K 次观看.  National Cerebral and Cardiovascular Center. 轻度宫内灌注是由动脉狭窄, 金属弹簧包裹周围的子宫和卵巢动脉在大鼠胚胎日17。这一程序产生了产前灌注和宫内生长的限制。

视频: A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis

The Hypoxic Ischemic Encephalopathy Model of Perinatal Ischemia

Hypoxic-Ischemic Encephalopathy (HIE) is the consequence of systemic asphyxia occurring at birth. Twenty five percent of neonates with HIE develop severe and permanent neuropsychological sequelae, including mental retardation, cerebral palsy, and epilepsy. The outcomes of HIE are devastating and permanent, making it critical to identify and develop therapeutic strategies to reduce brain injury in newborns with HIE. To that end, the neonatal rat model for hypoxic-ischemic brain injury has been developed to model this human condition. The HIE model was first validated by Vannucci et al 1 and has since been extensively used to identify mechanisms of brain injury resulting from perinatal hypoxia-ischemia 2 and to test potential therapeutic interventions 3,4. The HIE model is a two step process and involves the ligation of the left common carotid artery followed by exposure to a hypoxic environment. Cerebral blood flow (CBF) in the hemisphere ipsilateral to the ligated carotid artery does not decrease because of the collateral blood flow via the circle of Willis; however with lower oxygen tension, the CBF in the ipsilateral hemisphere decreases significantly and results in unilateral ischemic injury. The use of 2,3,5-triphenyltetrazolium chloride (TTC) to stain and identify ischemic brain tissue was originally developed for adult models of rodent cerebral ischemia 5, and is used to evaluate the extent of cerebral infarctin at early time points up to 72 hours after the ischemic event 6. In this video, we demonstrate the hypoxic-ischemic injury model in postnatal rat brain and the evaluation of the infarct size using TTC staining.

The postnatal rat model for hypoxic-ischemic brain injury is a well-established model of human neonatal hypoxic ischemic encephalopathy (HIE). In this article, we describe the model of HIE in post-natal rat pups.

围产期缺血缺氧缺血性脑病模型

Hypoxic-Ischemic Encephalopathy (HIE) is the consequence of systemic asphyxia occurring at birth. Twenty five percent of neonates with HIE develop severe ...

科研

生物学

Fetal Echocardiography and Pulsed-wave Doppler Ultrasound in a Rabbit Model of Intrauterine Growth Restriction

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ultrasound and fetal echocardiography. Increasingly, these imaging modalities are being employed clinically to examine cardiac function and assess wellbeing in utero, thereby guiding timing of birth decisions. Here, we used a rabbit model of IUGR that allows analysis of cardiac function in a clinically relevant way. Using isoflurane induced anesthesia, IUGR is surgically created at gestational age day 25 by performing a laparotomy, exposing the bicornuate uterus and then ligating 40-50% of uteroplacental vessels supplying each gestational sac in a single uterine horn. The other horn in the rabbit bicornuate uterus serves as internal control fetuses. Then, after recovery at gestational age day 30 (full term), the same rabbit undergoes examination of fetal cardiac function. Anesthesia is induced with ketamine and xylazine intramuscularly, then maintained by a continuous intravenous infusion of ketamine and xylazine to minimize iatrogenic effects on fetal cardiac function. A repeat laparotomy is performed to expose each gestational sac and a microultrasound examination (VisualSonics VEVO 2100) of fetal cardiac function is performed. Placental insufficiency is evident by a raised pulsatility index or an absent or reversed end diastolic flow of the umbilical artery Doppler waveform. The ductus venosus and middle cerebral artery Doppler is then examined. Fetal echocardiography is performed by recording B mode, M mode and flow velocity waveforms in lateral and apical views. Offline calculations determine standard M-mode cardiac variables, tricuspid and mitral annular plane systolic excursion, speckle tracking and strain analysis, modified myocardial performance index and vascular flow velocity waveforms of interest. This small animal model of IUGR therefore affords examination of in utero cardiac function that is consistent with current clinical practice and is therefore useful in a translational research setting.

We describe examination of fetal cardiac function with contemporary functional fetal echocardiography and fetoplacental Doppler ultrasound using the VisualSonics VEVO 2100 microultrasound in a surgically induced model of intrauterine fetal growth restriction in a rabbit.

在家兔模型宫内生长受限胎儿超声心动图和多普勒超声脉冲波

Fetal intrauterine growth restriction (IUGR) results in abnormal cardiac function that is apparent antenatally due to advances in fetoplacental Doppler ...

医学

Transuterine Fetal Tracheal Occlusion Model in Mice

Fetal tracheal occlusion (TO), an established treatment modality, promotes fetal lung growth and survival in severe congenital diaphragmatic hernia (CDH). Following TO, retention of the secreted epithelial fluid increases luminal pressure and induces lung growth. Various animal models have been defined to understand the pathophysiology of CDH and TO. All have their own advantages and disadvantages such as the difficulty of the technique, the size of the animal, cost, high mortality rates, and the availability of genetic tools. Herein, a novel transuterine model of murine fetal TO is described. Pregnant mice were anesthetized, and the uterus exposed via a midline laparotomy. The trachea of selected fetuses were ligated with a single transuterine suture placed behind the trachea, one carotid artery, and one jugular vein. The dam was closed and allowed to recover. Fetuses were collected just before parturition. Lung to body weight ratio in TO fetuses was higher than that in control fetuses. This model provides researchers with a new tool to study the impact of both TO and increased luminal pressure on lung development.

Various animal models of congenital diaphragmatic hernia and fetal tracheal occlusion present advantages and disadvantages regarding ethical issues, cost, surgical difficulty, size, survival rates, and availability of genetic tools. This model provides a new tool to study the impact of both tracheal occlusion and increased luminal pressure on lung development.

小鼠的转宫胎儿气管闭塞模型

Fetal tracheal occlusion (TO), an established treatment modality, promotes fetal lung growth and survival in severe congenital diaphragmatic hernia (CDH). ...

发育生物学

Stenosis of the Inferior Vena Cava: A Murine Model of Deep Vein Thrombosis

Deep vein thrombosis (DVT) and its devastating complication, pulmonary embolism, are a severe health problem with high mortality. Mechanisms of thrombus formation in veins remain obscure. Lack of mobility (e.g., after surgery or long-haul flights) is one of the main factors leading to DVT. The pathophysiological consequence of the lack of mobility is blood flow stagnation in venous valves. Here, a model is described that mimics such flow disturbance as a thrombosis-driving factor. In this model, partial flow restriction (stenosis) in the inferior vena cava (IVC) is created. Closure of about 90% of the IVC lumen for 48 h results in development of thrombi structurally similar to those in humans. The similarities are: i) most of the thrombus volume is red, i.e., consists of red blood cells and fibrin, ii) presence of a white part (lines of Zahn), iii) non-denuded endothelial monolayer, iv) elevated plasma D-Dimer levels, and v) possibility to prevent thrombosis by low molecular weight heparin. Limitations include variable size of thrombi and the fact that a certain percentage of wild-type mice (0 - 35%) may not produce a thrombus. In addition to visual observation and measurement, thrombi may be visualized by non-invasive technologies, such as ultrasonography, which allows for monitoring the dynamics of thrombus development. At shorter time points (1 - 6 h), intravital microscopy may be applied to directly observe events (e.g., recruitment of cells to the vessel wall) preceding thrombus formation. Use of this method by several teams around the world has made it possible to uncover basic mechanisms of DVT initiation and identify potential targets that might be beneficial for its prevention.

We describe here stenosis in the inferior vena cava as a murine model of deep vein thrombosis. This model recapitulates blood flow restriction, one of the major triggers of venous thrombosis in humans.

下腔静脉狭窄: 深静脉血栓形成的小鼠模型

Deep vein thrombosis (DVT) and its devastating complication, pulmonary embolism, are a severe health problem with high mortality. Mechanisms of thrombus ...

免疫与感染

Transient Middle Cerebral Artery Occlusion Model of Neonatal Stroke in P10 Rats

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature and mature brain. Stroke occurs commonly in the perinatal period in humans, and transient ischemia-reperfusion is the most common form of stroke in neonates. The reperfusion phase is a critical component of injury progression, which occurs over a period of days to weeks, and of the endogenous response to injury. This postnatal day 10 (p10) rat model of transient middle cerebral artery occlusion (tMCAO) creates a unilateral, non-hemorrhagic focal ischemia-reperfusion injury that can be utilized to study the mechanisms of focal injury and repair in the full-term-equivalent brain. The injury pattern that is produced by tMCAO is consistent and highly reproducible and can be confirmed with MRI or histological analyses. The severity of injury can be manipulated through changes in occlusion time and other methods that will be discussed.

Neonatal stroke is a significant cause of early brain injury requiring a translational model with consistent focal injury patterns and high reproducibility in order to enable study. This study describes the detailed surgical procedure for creating a non-hemorrhagic, unilateral focal ischemia-reperfusion injury in full-term-equivalent rodents.

在P10大鼠新生儿中风的短暂大脑中动脉栓塞模型

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature ...

Murine Model of Central Venous Stenosis using Aortocaval Fistula with an Outflow Stenosis

Central venous stenosis is an important entity contributing to arteriovenous fistula (AVF) failure. A murine AVF model was modified to create a partial ligation of the inferior vena cava (IVC) in the outflow of the fistula, mimicking central venous stenosis. Technical aspects of this model are introduced. The aorta and IVC are exposed, following an abdominal incision. The infra-renal aorta and IVC are dissected for proximal clamping, and the distal aorta is exposed for puncture. The IVC at the midpoint between the left renal vein and the aortic bifurcation is carefully dissected to place an 8-0 suture beneath the IVC. After clamping the aorta and IVC, an AVF is created by puncturing the infra-renal aorta through both walls into the IVC with a 25 G needle, followed by ligating a 22 G intra-venous (IV) catheter and IVC together. The catheter is then removed, creating a reproducible venous stenosis without occlusion. The aorta and IVC are unclamped after confirming primary hemostasis. This novel model of central vein stenosis is easy to perform, reproducible, and will facilitate studies on AVF failure.

An aortocaval fistula was created by puncturing the murine infra-renal aorta through both walls into the inferior vena cava and was followed by creation of a stenosis in its outflow via partial ligation of the inferior vena cava. This reproducible model can be used to study central venous stenosis.

中央静脉狭窄病的Murine模型,使用具有流出狭窄病的动脉静脉瘘管病

Central venous stenosis is an important entity contributing to arteriovenous fistula (AVF) failure. A murine AVF model was modified to create a partial ...

Real-Time Assessment of Spinal Cord Microperfusion in a Porcine Model of Ischemia/Reperfusion

Spinal cord injury is a devastating complication of aortic repair. Despite developments for the prevention and treatment of spinal cord injury, its incidence is still considerably high and therefore, influences patient outcome. Microcirculation plays a key role in tissue perfusion and oxygen supply and is often dissociated from macrohemodynamics. Thus, direct evaluation of spinal cord microcirculation is essential for the development of microcirculation-targeted therapies and the evaluation of existing approaches in regard to spinal cord microcirculation. However, most of the methods do not provide real-time assessment of spinal cord microcirculation. The aim of this study is to describe a standardized protocol for real-time spinal cord microcirculatory evaluation using laser-Doppler needle probes directly inserted in the spinal cord. We used a porcine model of ischemia/reperfusion to induce deterioration of the spinal cord microcirculation. In addition, a fluorescent microsphere injection technique was used. Initially, animals were anesthetized and mechanically ventilated. Thereafter, laser-Doppler needle probe insertion was performed, followed by the placement of cerebrospinal fluid drainage. A median sternotomy was performed for exposure of the descending aorta to perform aortic cross-clamping. Ischemia/reperfusion was induced by supra-celiac aortic cross-clamping for a total of 48 min, followed by reperfusion and hemodynamic stabilization. Laser-Doppler Flux was performed in parallel with macrohemodynamic evaluation. In addition, automated cerebrospinal fluid drainage was used to maintain a stable cerebrospinal pressure. After completion of the protocol, animals were sacrificed, and the spinal cord was harvested for histopathological and microsphere analysis. The protocol reveals the feasibility of spinal cord microperfusion measurements using laser-Doppler probes and shows a marked decrease during ischemia as well as recovery after reperfusion. Results showed comparable behavior to fluorescent microsphere evaluation. In conclusion, this new protocol might provide a useful large animal model for future studies using real-time spinal cord microperfusion assessment in ischemia/reperfusion conditions.

Spinal cord microcirculation plays a pivotal role in spinal cord injury. Most methods do not allow real-time assessment of spinal cord microcirculation, which is essential for the development of microcirculation-targeted therapies. Here, we propose a protocol using Laser-Doppler-Flow Needle probes in a large animal model of ischemia/reperfusion.

猪缺血/再灌注模型中脊髓微灌注的实时评估

Spinal cord injury is a devastating complication of aortic repair. Despite developments for the prevention and treatment of spinal cord injury, its ...

Myocardial Infarction by Percutaneous Embolization Coil Deployment in a Swine Model

Myocardial infarction (MI) is the leading cause of mortality worldwide. Despite the use of evidence-based treatments, including coronary revascularization and cardiovascular drugs, a significant proportion of patients develop pathological left-ventricular remodeling and progressive heart failure following MI. Therefore, new therapeutic options, such as cellular and gene therapies, among others, have been developed to repair and regenerate injured myocardium. In this context, animal models of MI are crucial in exploring the safety and efficacy of these experimental therapies before clinical translation. Large animal models such as swine are preferred over smaller ones due to the high similarity of swine and human hearts in terms of coronary artery anatomy, cardiac kinetics, and the post-MI healing process. Here, we aimed to describe an MI model in pig by permanent coil deployment. Briefly, it comprises a percutaneous selective coronary artery cannulation through retrograde femoral access. Following coronary angiography, the coil is deployed at the target branch under fluoroscopic guidance. Finally, complete occlusion is confirmed by repeated coronary angiography. This approach is feasible, highly reproducible, and emulates the pathogenesis of human non-revascularized MI, avoiding the traditional open-chest surgery and the subsequent postoperative inflammation. Depending on the time of follow-up, the technique is suitable for acute, sub-acute, or chronic MI models.

Myocardial infarction (MI) animal models that emulate the natural process of the disease in humans are crucial to understanding pathophysiological mechanisms and testing the safety and efficacy of new emergent therapies. Here, we describe an MI swine model created by deploying a percutaneous embolization coil.

猪模型中经皮栓塞线圈部署的心肌梗死

Myocardial infarction (MI) is the leading cause of mortality worldwide. Despite the use of evidence-based treatments, including coronary revascularization ...

Early Pathological and Magnetic Resonance Detection of Cerebral Injury Using a Rat Model of Neonatal Hypoxic Ischemic Encephalopathy

Perinatal hypoxic-ischemic encephalopathy (HIE) is an acute disease that may afflict newborns, resulting in variable long- and short-term neurodevelopmental outcomes. Early diagnosis is critical to identifying infants who may benefit from intervention; however, early diagnosis relies heavily on clinical criteria. No molecular or radiological tests have shown promise in detecting early cerebral injury. Studies have shown that magnetic resonance imaging (MRI) can show changes in both blood flow/ischemia and metabolic disruption. However, they have all been used to evaluate the secondary phase of the disease (&gt;12 h) after the onset of the injury. Early diagnosis is critical to rapidly starting therapeutic hypothermia in eligible infants, which is currently recommended to be initiated within 6 h of birth. The rat model of hypoxic-ischemic injury was developed in 1981 and has been validated and used extensively to study changes in brain perfusion, cerebral injury markers, and morphology. However, it has primarily been used as a "late model", evaluating injury several days after the initial ischemic insult. The model has been known to have poor sensitivity in evaluating reliable and reproducible early cerebral changes. The objective of this study was to develop a reliable model to study early gross morphological and radiological markers of HIE using pathological staining and cerebral magnetic resonance imaging/magnetic resonance spectroscopy.

<meta charset="utf8"></head><body>使用新生儿缺氧缺血性脑病大鼠模型进行脑损伤的早期病理和磁共振检测

The present protocol describes a rodent model of newborn hypoxic-ischemic injury for identifying early changes in cerebral tissue by gross morphology and magnetic resonance imaging. This has benefits over existing models, which can be used to study late injury but do not allow the evaluation of reproducible early changes.