我们感谢黄副教授和崔老师对本文的严格指导和支持。该研究得到以下研究：1.国家自然科学基金面上项目（NSFC），82160938，“TGF-β1介导的ERK和Smad信号通路交互效应探索饼状分离艾灸对肾间质纤维化的机制研究”;2.云南省卫生健康委员会“2020年度中医药高水平后备人才”孵化项目（云维中医药发展〔2021〕1号）;3. 云南省科技厅-中药应用基础研究联合专项资金（No.2019FF002-022， No.202101AZ70001-286， No.202301AZ070001-030）;4.云南省针灸推拿防治脑病创新团队（2023）.

该计划遵循云南中医药大学动物护理指南，并已获得云南中医药大学动物伦理委员会的批准。实验动物福利和伦理审查编号为R-062021G098。
1、艾灸实验平台装置（图1）
注：我们根据大鼠前后肢之间的距离及其行为和生理特性，设计和构建了大鼠艾灸实验平台装置。
<ol><li>艾灸器的尺寸<ol><li>使用防打板材料和木板制作设备;整体尺寸为 52 cm L x 23.5 cm W x 64 cm H，设备底部面积为 1222 cm2。 注意：在这个高度的平台上进行艾灸手术，老鼠可以保持冷静。</li>
	</ol></li>
	<li>艾灸器主机架<ol><li>用防打板材料制作整个设备的底座：底板为52厘米长×23.5厘米宽，表面为白色。确保底部在设备重量中占很大比例，以稳定整个设备。</li>
		<li>用实木制作设备的高架平台，高 60 厘米。 注意： 这是整个仪器的主框架，它稳定整个设备并支撑大鼠处于站立位置。</li>
		<li>在高架平台上方放置五个放置槽，尺寸为 10 cm L x 6 cm W x 10 cm H.使用间隔板（10 cm L x 1.3 cm W x 10 cm H）将放置槽分开，对应于站立位置大鼠前肢和后肢之间的距离。</li>
	</ol></li>
	<li>艾灸器的组成部分<ol><li>将两条不锈钢条固定在放置槽的顶部。每个固定条是一根直角等边钢筋，长 40 厘米 x 宽 1.4 厘米（两侧宽度相同），有 15 个孔。固定条上的三个孔对应于每个插槽，用于固定梅花手柄螺钉和移动软管。相邻孔之间的距离为 7 厘米。</li>
		<li>将可调节梅花手柄螺丝放在304不锈钢制成的隔热网两端，并连接到固定隔热网和固定条。</li>
		<li>将每个放置槽的上端与由镀锌板框架包围的可升降隔热网络连接。将此框架连接到两端的梅花手柄螺丝上。在隔热网中间放置304不锈钢圆形网材料，防止灰烬落下烫伤老鼠，固定老鼠，便于灸。</li>
		<li>用不锈钢固定条固定不锈钢活动软管（34厘米长）的底端固定，上端用管夹连接板固定。 注意： 活动软管便于在悬挂艾灸操作过程中调整艾灸棒的位置。</li>
		<li>使用铝合金制成的管夹连接不锈钢活动软管和艾灸夹。</li>
		<li>将不锈钢艾条夹连接到管夹连接板上，同时固定8-10条艾条进行艾灸。 注意：活动软管、管夹连接板和艾灸夹可用于大鼠悬浮艾灸操作。</li>
	</ol></li>
</ol>2.建立慢性肾功能衰竭大鼠模型
<ol><li>驯化24只无特定病原体（SPF）SD雄性大鼠（3-4周龄，体重180-200g）1周。保持进料温度在18-24°C;相对湿度在40-70%;以及 12 小时的光照和 12 小时的黑暗循环。每个笼子里饲养六只动物，并免费提供水和公共饲料。</li>
	<li>建立CRF大鼠模型。根据随机数表，按照本方案，选取6只大鼠作为对照组，其余18只作为模型组。对照组（n = 6）用蒸馏水（10 mL∙kg-1∙day-1）和正常饲料处理，模型组（n = 18）用腺嘌呤混悬液（150 mg kg-1∙day-1）和高蛋白饲料处理21 d。</li>
	<li>每天观察和记录体重、精神状态和饮食情况。21天结束后，确保所有动物在眼眶采血前禁食12小时。</li>
	<li>收集眼眶血样。<ol><li>用异氟醚短暂麻醉大鼠。用非惯用手抓住大鼠的背部，用拇指和食指对称握住大鼠的头和脸，完全露出大鼠的一侧下眦，并保持这个姿势。</li>
		<li>用惯用手在拇指和食指之间握住3厘米长的熔点毛细血管，慢慢扭转毛细血管，以45°进入大鼠的内眦静脉丛，直到血液慢慢从管中流出。</li>
		<li>将 150-200 μL 血样收集在 0.5 mL 采血抗凝管中，轻轻拉出毛细血管，并用棉花按压眼眶。</li>
	</ol></li>
	<li>快速彻底地摇晃抗凝剂管。使用自动生化分析仪测量血浆血清肌酐 （Scr） 和血尿素氮 （BUN）。 注：Scr和BUN的增加表明CRF模型的成功建立。</li>
	<li>将大鼠分为四组： A： 6只对照大鼠，不治疗;18只模型大鼠分为B组 ： （模型组：不用药或艾灸）、 C组（ 氯沙坦钾组：灌胃针12 G给药2.33 mg∙kg-1∙day-1 ）和 D组： （草药-蛋糕分离-艾灸组：草药-蛋糕-分离-艾灸应用于双侧 神书 [B23]和双侧 蕴书 [B20]）。</li>
</ol>3.治疗阶段
注意： A 组和 B 组不接受任何治疗，而 C 组和 D 组将每天治疗一次，持续 24 天，治疗 12 天后休息两天。
<ol><li>治疗前的准备工作<ol><li>准备干黄芪300克，干党参300克，干熟地黄200克，烤短角淫羊藿100克，山茱萸100克 。混合这些粉末。</li>
		<li>将500克 艾蒿 叶放入中药粉碎机（速度34000转/分钟）中5分钟，将磨碎的粉末转移到板上，然后用600目筛筛。经过两轮筛选后，去除粗艾灸，将细艾灸转移到袋子中使用（图2）。</li>
		<li>通过将细艾灸填充到模具中来准备艾灸筒，制成直径为0.5厘米，高度为0.5厘米的小艾灸筒。或者，将艾灸在双手的拇指和食指之间捣乱，形成艾灸锥（图3A）。</li>
		<li>通过将5mL注射器切成四个0.5cm厚的圆形塑料环并将四个环粘合在同一水平面上（在边长为26mm的方形平面的角上， 图3D）来制备草药蛋糕模具。</li>
		<li>通过将粉末和水以1：1的比例混合并将混合物填充到草药蛋糕模具中来制作药饼（图3E）。将药饼留在模具中，存放在干燥的地方。</li>
	</ol></li>
	<li>标记穴位<ol><li>穴位如下： 神术 （B23），第二腰椎两侧，每侧一个穴位; Pishu （B20），两侧第 11 胸椎的肋间，每侧一个穴位（图 4）。 注意：有关穴位的位置，请参阅 《实验动物针灸手册》20。</li>
		<li>在非惯用手上戴上厚橡胶手套，轻轻覆盖大鼠头部，用惯用手定位大鼠背部的穴位，然后用颜色记号笔标记穴位。</li>
	</ol></li>
	<li>放置老鼠<ol><li>戴上厚橡胶手套。用拇指和惯用手的其他四个手指轻轻捏住大鼠背部和颈部的皮肤。用非惯用手的拇指和食指轻轻捏住老鼠的尾巴。</li>
		<li>抬起大鼠，将大鼠的头部对准专用艾灸平台的放置槽，然后慢慢向前移动大鼠。确保老鼠的前爪靠近放置槽的边缘，并且老鼠爬进槽中。 注意：开始前对整个艾灸平台进行消毒。在抓住老鼠之前，轻轻触摸老鼠的背部或与它交谈以使其平静下来。将保温网的高度从低位置（不使用时）调整到最高，方便老鼠进入。</li>
	</ol></li>
	<li>大鼠固定<ol><li>将双手的拇指和食指放在隔热网两端梅花螺丝的螺丝上。拧下梅花螺丝，将隔热网降低到适当位置，靠近老鼠的背部，但不引起压迫，然后拧紧螺丝。</li>
	</ol></li>
	<li>放置艾灸蛋筒和药饼。<ol><li>根据大鼠身上的穴位痕迹，将准备好的装有药饼的模具放在与貂苒（B20）和神书（B23）穴位对应的隔热网上。有四个穴位，每个穴位对应一个药饼。</li>
		<li>将艾灸蛋筒平滑稳定地放在药饼上。</li>
	</ol></li>
	<li>灸<ol><li>用打火机点燃艾灸，开始艾灸，直到艾灸烧完，余热自然散发（湿的药饼干涸）。</li>
		<li>在同一穴位上用新的艾灸筒和药饼替换烧焦的艾灸筒，然后继续灸灸，直到艾条筒烧完，药饼变干。在每个穴位重复此操作总共 5 次。 注意： 在用新的艾灸筒和药饼更换之前，请清洁和消毒可升降的隔热网络。</li>
		<li>将艾灸蛋筒和药饼放入含水的废液罐中，在每个穴位燃烧五个艾灸球筒后熄灭。 注意：每个穴位接受五个艾灸锥;每个艾灸锥烧坏后都可以更换。</li>
	</ol></li>
	<li>将老鼠放回笼子<ol><li>将空的饲养笼放在大鼠面前，靠近艾灸平台放置槽的下端，并保持静止。轻轻拍打老鼠的臀部，使老鼠可以从放置槽移动到喂食笼。老鼠进来后，放下笼子。用惯用手抓住老鼠背部的皮毛，牢牢握住老鼠，然后将其放在地上进一步观察。</li>
		<li>观察大鼠是否有呼吸急促，情绪是否稳定，皮肤表面是否烫伤。如果没有异常，将其放回笼子里。如果烫伤，请立即将烫伤的软膏涂抹在皮肤表面，并将其放入一个笼子中。</li>
	</ol></li>
</ol>4. 采样阶段
<ol><li>治疗24天后，准备大鼠在取样前禁食12小时。</li>
	<li>将异氟烷倒入气体麻醉机中，用2-3%异氟烷麻醉大鼠。2-3分钟后，确保反射消失（用手术镊子捏住尾巴，检查它们是否没有任何反射）和心跳的持续存在，并切换到用1-1.5%异氟醚维持麻醉。</li>
	<li>将针头插入准腹主动脉，从腹主动脉收集 5 mL 血液。让血液静置2小时，在4°C下以2490× g 离心10分钟，并将血清上清液转移到1.5mL微量离心管中进行进一步分析。</li>
	<li>对大鼠进行长时间深度麻醉，在维持麻醉下从腹主动脉抽血。此外，通过颈椎脱位对大鼠实施安乐死。</li>
	<li>用手术剪刀和镊子剥离双侧肾筋膜和脂肪，并用0.9%生理盐水冲洗，从双肾采集组织。称量左右肾，并记录重量和形态。</li>
	<li>通过将左肾切成六个部分并将它们放入含有 4% 多聚甲醛的 50 mL 离心管中进行浸泡、固定和保存，以保护肾组织。用一把刀在纵向切成六块，在横向用两把刀切成六块。将这些碎片放入 6 个 2 mL 冻存管中，并立即放入液氮中;将它们转移到-80°C冰箱中储存。</li>
</ol>5.检测阶段
<ol><li>使用适当大小的正确肾组织进行蛋白质提取、凝胶电泳、膜转移、封闭、一抗孵育、二抗、化学发光、显影、固定和薄膜扫描。</li>
	<li>使用凝胶图像数据分析系统（例如，Image-Pro Plus 6.0）分析条带的灰度值并计算每种蛋白质的含量。</li>
	<li>将左肾组织脱水，清除，包埋在石蜡中，切片（厚度为4μm），用苏木精 - 伊红染色和Masson三色染色，并在200倍放大倍率的光学显微镜下观察肾组织中的病理变化21 。</li>
</ol>

基于草药蛋糕治疗大鼠肾功能衰竭大鼠的艾灸平台

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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Using+old+meshy+cage+cover+to+make+a+rat+fixating+device+for+acupuncture+study.">Using old meshy cage cover to make a rat fixating device for acupuncture study.</a> Zhen Ci Yan Jiu = Acupuncture Research. 46 (4), 348-350 (2021).</li><li>Zhang, H. X., 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=Introduction+of+a+new+type+of+mouse+forelimb+fixator+and+two+types+of+batch+mice+moxibustion+device.">Introduction of a new type of mouse forelimb fixator and two types of batch mice moxibustion device.</a> Zhen Ci Yan Jiu = Acupuncture Research. 46 (7), 616-619 (2021).</li><li>Chen, W., 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=A+batch+moxibustion+device+in+the+mouse+abdominal+waist.">A batch moxibustion device in the mouse abdominal waist.</a> Journal of Chengdu University of Traditional Chinese Medicine. 45 (04), 110-112 (2022).</li><li>Gao, S. Y., Wu, R. N., Wang, M. M., Zhang, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+device+for+animal+experiment.">A device for animal experiment.</a> Clinical Journal of Chinese Medicine. 10 (23), 139-140 (2018).</li><li>Hu, Y. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Manual of acupuncture for experimental animals (second edition). , (2014).</li><li>Liang, Y. L., 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=Reflections+on+animal+experiments+with+moxibustion.">Reflections on animal experiments with moxibustion.</a> China Journal of Traditional Chinese Medicine and Pharmacy. 31 (11), 4379-4381 (2016).</li><li>Madalina, V. N., Jinyang, Z. -. B., Malgorzata, W., Liliana, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeting+renal+fibrosis:+Mechanisms+and+drug+delivery+systems.">Targeting renal fibrosis: Mechanisms and drug delivery systems.</a> Advanced Drug Delivery Reviews. 129, 295-307 (2018).</li><li>Wu, R. N., 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=Design+of+convenient+animal+experimental+moxibustion+therapy+device.">Design of convenient animal experimental moxibustion therapy device.</a> Shanghai Journal of Acupuncture and Moxibustion. 38 (02), 243-244 (2019).</li><li>Yan, L. G., Zhou, T., Jia, Z. K., Li, J., Wang, C. X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+Panax+pseudo-ginseng-partition+moxibustion+on+cartilage+damage+in+rats+with+traumatic+osteoarthritis+by+regulating+AMPK/SIRT1/NF-?B+signaling+pathway.">Influence of Panax pseudo-ginseng-partition moxibustion on cartilage damage in rats with traumatic osteoarthritis by regulating AMPK/SIRT1/NF-?B signaling pathway.</a> Hebei Medical Journal. 45 (10), 1450-1454 (2019).</li><li>Xu, Q., 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=Effect+of+mild+moxibustion+of+45+&#176;C+at+&amp;#34;Zusanli&amp;#34;+(ST36)+of+different+durations+on+the+inflammatory+factors+in+the+abdominal+aorta+of+hyperlipidemia+rats.">Effect of mild moxibustion of 45 &#176;C at &amp;#34;Zusanli&amp;#34; (ST36) of different durations on the inflammatory factors in the abdominal aorta of hyperlipidemia rats.</a> Zhen Ci Yan Jiu = Acupuncture Research. 48 (9), 923-932 (2023).</li></ol>

提交人声明不存在利益冲突。大鼠艾灸实验平台装置专利号为ZL202320880924.X。

临床上，针灸治疗要求患者保持冷静放松，避免过度愤怒和饥饿，强调“平安”。然而，目前针灸实验的做法仍是以结合动物为前提，这不仅违背了实验的初衷，也不符合临床实践。捆绑动物，使动物受到惊吓和愤怒，会迅速增加实验动物体内的肾上腺素和其他激素，影响实验结果22。在针灸动物的实验操作过程中，必须紧急解决这些问题，以保证实验的准确性，保护动物的福?...

艾灸是随着火的应用而出现的。古代医生无意中发现，燃烧身体的某些部位可以缓解不适的症状。经过长期的实践，逐渐选择“艾蒿叶”作为主要艾灸材料1。艾灸是中医的重要组成部分之一，当与中医汤剂和穴位针灸并列时，这三大中医疗法在许多疾病中被广泛使用。
早在黄帝内经（黄帝内经）时期，中国医生就经常使用艾灸来治疗疾病。艾蒿是一种中草药，具有温顺经络、止血、祛寒、止痛的作用。常用于中药的外用治疗，也可用于中药汤剂的治疗。艾灸是一种外置疗法2 ，其中将艾灸或其他药物置于体表的穴位上燃烧，并利用艾灸火的温和热量和药物的效果来治疗疾病3。
艾灸作为常用的治疗方法之一，在临床实践中得到了广泛的应用。随着艾灸疗法的不断拓展，越来越多的研究对艾灸的机制进行了探究，并进行了许多动物实验4,5。
艾灸法可分为直接艾灸和间接艾灸。将艾灸放在人体皮肤上的方法称为直接艾灸。间接艾灸用药物或其他材料将艾灸锥表面和穴位分离。艾灸可以治疗多种疾病，如肠易激综合征6、慢性疲劳综合征7、膝骨关节炎8。
本文主要探讨中药-蛋糕-分离-艾灸的机理。草药-蛋糕分离-艾灸属于间接艾灸法，将艾锥放在药饼上以应用艾灸。目前的研究表明，艾灸锥燃烧产生的红外线和温热效应、艾灸叶焦油的化学成分、穴位刺激等，可以通过增加药物对穴位的渗透来实现疾病的预防和治疗9 。因此，中药-蛋糕-分离-艾灸主要通过艾灸、药物、穴位产生“综合效果”10。
既往研究表明，艾灸是治疗CRF11、12、13、14、15的有效方法，但其潜在机制尚不明确。我们一直致力于CRF的临床和动物研究。目前，我们正在对CRF大鼠进行基于中药蛋糕分离艾灸介导的蛋白激酶RNA样内质网激酶（PERK）内质网激酶信号通路的内质网应激的艾灸实验研究。通过这项动物实验，我们旨在初步解释艾灸治疗慢性肾脏病的潜在机制。在本实验中，大鼠中药-蛋糕分离艾灸的治疗是关键操作。
在前面的实验16中，饲养笼的盖子用于捆绑和固定大鼠。为了保护操作者，在结合之前需要对大鼠进行麻醉。然而，Sprague-Dawley（SD）老鼠很容易害怕和惊吓。结合操作容易引起大鼠烦躁不安。大鼠姿势固定不良、穴位定位偏移、实验时间增加、操作人员受伤等问题时有发生。
此外，如果捆绑太紧，很容易引起大鼠呼吸困难，不利于动物福利。如果结合太松，大鼠容易挣扎和移动身体，导致艾灸锥掉落并灼伤皮肤，从而影响实验结果。根据文献资料，动物实验仍然涉及结合16,17,18，大多数实验装置只能固定，阻碍了艾灸19等其他技术的实施。因此，在保证动物福利和实验规范与效率的同时，实现实验装置中固定和艾灸的一体化，成为研究者的重点领域之一。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.5 mL heparin lithium biochemical tube</td>
			<td>Shijiazhuang Kangweishi Medical Equipment Co., LTD</td>
			<td>220518</td>
			<td>Blood collection</td>
		</tr>
		<tr>
			<td>0.9% normal saline</td>
			<td>Kunming Nanjiang Pharmaceutical Co., LTD</td>
			<td>/</td>
			<td>Rinse</td>
		</tr>
		<tr>
			<td>1.5 mL Eppendorf tube</td>
			<td>Beijing Lanjieke Biotechnology Co., LTD</td>
			<td>12100024</td>
			<td>Subpackage</td>
		</tr>
		<tr>
			<td>2 mL cryopreservation tube</td>
			<td>Baroque Biotechnology Co., LTD</td>
			<td>88-0151</td>
			<td>Packaged kidney&#160;</td>
		</tr>
		<tr>
			<td>4 &#176;C refrigerator</td>
			<td>Haier Group Co. LTD</td>
			<td>BCD-236WDGL</td>
			<td>Store</td>
		</tr>
		<tr>
			<td>5 mL syringe</td>
			<td>Sichuan New century Medical polymer products Co., LTD</td>
			<td>20211115</td>
			<td>Gavage&#160;</td>
		</tr>
		<tr>
			<td>&#160;50 mL centrifuge tube</td>
			<td>Beijing Langeke Biotechnology Co., LTD</td>
			<td>CT-002-50A</td>
			<td>Packaged kidney&#160;</td>
		</tr>
		<tr>
			<td>12 G gavage needle</td>
			<td>Beijing Zhongke Hengtian Technology Co., Ltd.</td>
			<td>12#55mm</td>
			<td>Gavage&#160;</td>
		</tr>
		<tr>
			<td>96-well plates</td>
			<td>Beijing Lanjieke Biotechnology Co., LTD</td>
			<td>11510</td>
			<td>Determination of protein concentration</td>
		</tr>
		<tr>
			<td>&#160;-80 &#176;C freezer</td>
			<td>Thermo Fisher Scientific</td>
			<td>991</td>
			<td>Store</td>
		</tr>
		<tr>
			<td>Adenine</td>
			<td>Beijing Boaotuo Technology Co., LTD</td>
			<td>A0607</td>
			<td>Molding drug</td>
		</tr>
		<tr>
			<td>BCA developer</td>
			<td>Biyuntian Biological Engineering Co., LTD</td>
			<td>P0010</td>
			<td>Determination of protein concentration</td>
		</tr>
		<tr>
			<td>BCL-2 protein antibody reagent</td>
			<td>Abcam Shanghai Trading Co., LTD</td>
			<td>ab196495</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>Caspase-3 protein antibody reagent</td>
			<td>Abcam Shanghai Trading Co., LTD</td>
			<td>ab184787</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>Chemiluminescence imaging system</td>
			<td>Shanghai Qinxiang Scientific Instrument Co., LTD</td>
			<td>100240073</td>
			<td>Protein banding imaging</td>
		</tr>
		<tr>
			<td>Chinese medicine powder</td>
			<td>The Second Affiliated Hospital of Yunnan University of Chinese Medicine</td>
			<td>/</td>
			<td>Cure</td>
		</tr>
		<tr>
			<td>Direct-load Color Prestained Marker</td>
			<td>Beijing Kangrunchengye Biotechnology Co., LTD (GenStar)</td>
			<td>M221</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>Disposable blood collection needle</td>
			<td>Liuyang Sanli Medical science and technology development Co., LTD</td>
			<td>YZB/ Country 0411-2010</td>
			<td>Blood sampling</td>
		</tr>
		<tr>
			<td>Disposable human venous blood sample collection container</td>
			<td>Liuyang Sanli Medical science and technology development Co., LTD</td>
			<td>20210901</td>
			<td>Blood sampling</td>
		</tr>
		<tr>
			<td>DK-30 Automatic snow ice maker</td>
			<td>Henan Brothers instrument equipment Co., LTD</td>
			<td>SHDX0023</td>
			<td>Ice-making</td>
		</tr>
		<tr>
			<td>ECL luminescent substrate kit</td>
			<td>Beijing Lanjieke Biotechnology Co., LTD</td>
			<td>BL520B</td>
			<td>Convert latent images in exposed film into visible images</td>
		</tr>
		<tr>
			<td>electronic scales</td>
			<td>Shanghai Ran Hao Electronics Co., LTD</td>
			<td>JCS-31002C</td>
			<td>The body weight and kidney weight of rats were measured</td>
		</tr>
		<tr>
			<td>Freezing microtome&#160;</td>
			<td>Thermo Fisher Scientific</td>
			<td>HM525 NX</td>
			<td>Microtome</td>
		</tr>
		<tr>
			<td>fully automatic biochemical analyser</td>
			<td>Tianjin Micro nano core Technology Co., LTD</td>
			<td>PV2112001Z02410305</td>
			<td>Renal function test</td>
		</tr>
		<tr>
			<td>Gas anesthesia system</td>
			<td>Zhongshi Technology Co., Ltd</td>
			<td>ZS-MV-1</td>
			<td>Anesthesia</td>
		</tr>
		<tr>
			<td>Glycine</td>
			<td>Beijing Soleibao Technology Co., Ltd.</td>
			<td>Cat#G8200</td>
			<td>Running buffer, Western transfer buffer</td>
		</tr>
		<tr>
			<td>HE staining reagent</td>
			<td>Beijing Soleibao Technology Co., Ltd.</td>
			<td>G1120&#160;</td>
			<td>Staining</td>
		</tr>
		<tr>
			<td>High-protein feed</td>
			<td>Jiangsu Synergetics Biological Engineering Co., LTD</td>
			<td>XT19008</td>
			<td>Feeding</td>
		</tr>
		<tr>
			<td>Intelligent laboratory ultra-pure water machine</td>
			<td>Chongqing huachuag water treatment engineering co.,LTD</td>
			<td>15L</td>
			<td>Filtration</td>
		</tr>
		<tr>
			<td>Internal reference &#946;-actin</td>
			<td>Jiangsu Qinke Biological Research Co., LTD</td>
			<td>AF7018</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Jiangsu Zhongmu Beikang Pharmaceutical Co., Ltd</td>
			<td>20220701</td>
			<td>Anesthesia</td>
		</tr>
		<tr>
			<td>liquid nitrogen</td>
			<td>Kunming Botai dry ice sales Co., LTD</td>
			<td>/</td>
			<td>Renal tissue freezing</td>
		</tr>
		<tr>
			<td>Losartan potassium tablets</td>
			<td>Merck Sharp &#38; Dohme (China) Investment Co., LTD</td>
			<td>J20180054</td>
			<td>Cure</td>
		</tr>
		<tr>
			<td>Low temperature high speed tissue grinder</td>
			<td>Wuhan Servicebio technology CO.,LTD</td>
			<td>SKZ3F20200191</td>
			<td>Tissue grinding</td>
		</tr>
		<tr>
			<td>Maason staining kit</td>
			<td>Beijing Soleibao Technology Co., Ltd.</td>
			<td>G1340</td>
			<td>Staining</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Guangdong Guanghua Sci-Tech Co., Ltd</td>
			<td>20220519</td>
			<td>Western transfer buffer</td>
		</tr>
		<tr>
			<td>Mini-PROTEAN Tetra</td>
			<td>Bole Life Medical Products (Shanghai) Co., Ltd</td>
			<td>552BR 233193</td>
			<td>Electrophoresis</td>
		</tr>
		<tr>
			<td>Moxa</td>
			<td>The Second Affiliated Hospital of Yunnan University of Chinese Medicine</td>
			<td>/</td>
			<td>Cure</td>
		</tr>
		<tr>
			<td>Multiskan Spectrum Microplate Spectrophotometer</td>
			<td>TECAN</td>
			<td>Spark</td>
			<td>The absorbance and concentration of tissue protein were detected</td>
		</tr>
		<tr>
			<td>operating scissors</td>
			<td>Zhangjiagang city Jinfeng deer knife cutting factory</td>
			<td>201118</td>
			<td>Cut the skin</td>
		</tr>
		<tr>
			<td>Paraformaldehyde</td>
			<td>Beijing Lanjieke Biotechnology Co., LTD</td>
			<td>BL539A</td>
			<td>Fixed tissue</td>
		</tr>
		<tr>
			<td>PERK protein antibody reagent</td>
			<td>Shanghai Pumai Biotechnology Co., LTD (CST China Agent)</td>
			<td>3192</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>Positive fluorescence microscope</td>
			<td>Carl Zeiss Suzhou Co.,Ltd.</td>
			<td>5446000456</td>
			<td>Capture picture</td>
		</tr>
		<tr>
			<td>Prefabricated glue</td>
			<td>Derun Cheng (Beijing) Biological Co., LTD (ACE China Agent)</td>
			<td>B230418</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>ProteinFind Goat Anti-Rabbit LgG(H+L), HRP Conjugate</td>
			<td>Shanghai Dinran electronic Technology Co., LTD (TRAN)</td>
			<td>HS101</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>pulverizer</td>
			<td>Hefei Rongzhida small household appliances Co., LTD</td>
			<td>RS-FS1401</td>
			<td>Crushed Chinese medicine</td>
		</tr>
		<tr>
			<td>PVDF membrane</td>
			<td>MerckMillipore Corporation&#160;</td>
			<td>IPVH00010&#160;</td>
			<td>Western Blot</td>
		</tr>
		<tr>
			<td>Refrigerated centrifuge</td>
			<td>Hettich Precision Technology (Zhuhai) Co., LTD</td>
			<td>MIKRO 220R</td>
			<td>Centrifuge</td>
		</tr>
		<tr>
			<td>RIPA lysate</td>
			<td>Beijing Solaibao Technology Co., LTD (Solarbio)</td>
			<td>R0010</td>
			<td>Lytic tissue</td>
		</tr>
		<tr>
			<td>Sampling capillary</td>
			<td>Jiangsu Kangjianhua Medical Supplies Co., LTD</td>
			<td>20160157</td>
			<td>Blood from inner canthus</td>
		</tr>
		<tr>
			<td>Scald ointment (Zilian ointment)</td>
			<td>Yunnan Provincial Hospital of Traditional Chinese Medicine</td>
			<td>/</td>
			<td>Used for the treatment of scalding from moxibustion</td>
		</tr>
		<tr>
			<td>Skim milk powder</td>
			<td>Guangzhou Saiguo Biotechnology Co., LTD (BIOFROXX)</td>
			<td>1172GR500</td>
			<td>Western blocking buffer</td>
		</tr>
		<tr>
			<td>surgical blade</td>
			<td>Shanghai Medical Equipment Co., LTD</td>
			<td>JOBO10</td>
			<td>Incision kidney tissue</td>
		</tr>
		<tr>
			<td>surgical knife handle</td>
			<td>Shanghai Medical Equipment Co., LTD</td>
			<td>J11010</td>
			<td>Mounting blade</td>
		</tr>
		<tr>
			<td>TS-200 Orbital shaker</td>
			<td>Haimen Qilin Bell Manufacturing Co., Ltd.</td>
			<td>TS-8S</td>
			<td>Gel fixation</td>
		</tr>
		<tr>
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using herbal-cake-separated moxibustion for the treatment of rats with chronic renal failure

在临床实践中，受试者在艾灸过程中，需要保持情绪稳定、姿势舒适，避免出现艾灸灰落下、皮肤烫伤、疗效不佳等问题。这样的问题在大鼠艾灸实验中也存在。为了模拟临床实践，有必要在动物实验中引入一种实验仪器，即固定大鼠和灸治疗同步的艾灸装置，可以使实验大鼠在非麻醉下安静舒适地接受艾灸治疗。
课题组设计了大鼠艾灸实验平台。该设备由一块木板构成，木板带有支撑底板、多个固定组件和分区组件。该装置可实现大鼠艾灸的操作模式，无需捆绑，避免麻醉烫伤，同时暴露背部多个穴位。该操作可以避免对大鼠和操作人员的身心伤害，提高了研究效率，进一步推动了艾灸动物实验的发展和研究。
该装置结构简单，易于操作和推广，全面创新设计，可重复使用，适用于以艾灸为主的大鼠实验。本文主要介绍了大鼠艾灸实验平台装置的结构、使用该装置的实验大鼠中草药-蛋糕分离-艾灸的基本过程，并介绍了慢性肾功能衰竭（CRF）大鼠模型的建立和具有代表性的实验结果。

肾脏形态学 从 图5可以看出，与其他组相比， D 组（草食-蛋糕分离-艾灸组）肾脏的大小、颜色和质地与正常组更接近。模型肾形态和氯沙坦钾基团均表现为不同程度的白色纤维化。
肾脏重量 图6 显示，A组的左肾重量与B组的左肾重量有显著差异，表明CRF模型是成功的。 D 组?...

Watch this Scientific Journal Video about Using Herbal-Cake-Separated Moxibustion for the Treatment of Rats with Chronic Renal Failure at JoVE.com

作者聚焦：了解草药蛋糕分离艾灸对肾病大鼠的影响

艾灸实验平台可以不受限制地固定大鼠，大鼠在清醒状态下可以进行多穴位艾灸，保证了大鼠的福利，保护了操作人员的安全，保证了治疗效果，提高了实验的效率。

采用中药蛋糕分离艾灸治疗慢性肾功能衰竭大鼠

In the process of moxibustion in clinical practice, subjects need to be in a stable mood and comfortable posture to avoid problems such as moxa ash ...

我们研究了中草药蛋糕分离艾灸对慢性肾功能衰竭大鼠的影响。本实验试图回答中草药-蛋糕分离艾灸的机理。该协议的优点是大鼠在非捆绑和非麻醉条件下进行破除，使动物实验接近临床实践。艾灸平台的应用不仅保护了大鼠的动物福利，而且在节省操作人员的时间和精力方面减少了艾灸操作过程，在一定程度上促进了动物艾灸实验的发展。未来，我们的实验室将进一步扩大该平台的使用范围，探索实验大鼠的针灸效果。使用大鼠后，在相同条件下，对大鼠用该装置和其他艾灸方法的艾灸效果进行比较研究。

using-herbal-cake-separated-moxibustion-for-treatment-rats-with

Research

JoVE Journal

Medicine

Using Herbal-Cake-Separated Moxibustion for the Treatment of Rats with Chronic Renal Failure

671 次观看.  Yunnan University of Chinese Medicine. 我们研究了中草药蛋糕分离艾灸对慢性肾功能衰竭大鼠的影响。本实验试图回答中草药-蛋糕分离艾灸的机理。该协议的优点是大鼠在非捆绑和非麻醉条件下进行破除，使动物实验接近临床实践。艾灸平台的应用不仅保护了大鼠的动物福利，而且在节省操作人员的时间和精力方面减少了艾灸操作过程，在一定程度上促进了动物艾灸实验的发展。未来，我们的实验室将进一...

视频: 作者聚焦：了解草药蛋糕分离艾灸对肾病大鼠的影响

Acute and Chronic Tactile Sensory Testing after Spinal Cord Injury in Rats

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory testing in rat SCI models is needed. However, until recently, no single testing approach had been validated for SCI so that standardized methods have not been implemented across labs. Additionally, available testing methods could not be implemented acutely or when severe motor impairments existed, preventing studies of the development of SCI-induced allodynia3. Here we present two validated sensory testing methods using von Frey Hair (VFH) monofilaments which quantify changes in tactile sensory thresholds after SCI4-5. One test is the well-established Up-Down test which demonstrates high sensitivity and specificity across different SCI severities when tested chronically5. The other test is a newly-developed dorsal VFH test that can be applied acutely after SCI when allodynia develops, prior to motor recovery4-5. Each VFH monofilament applies a calibrated force when touched to the skin of the hind paw until it bends. In the up-down method, alternating VFHs of higher or lower forces are used on the plantar L5 dermatome to delineate flexor withdrawal thresholds. Successively higher forces are applied until withdrawal occurs then lower force VFHs are used until withdrawal ceases. The tactile threshold reflects the force required to elicit withdrawal in 50% of the stimuli. For the new test, each VFH is applied to the dorsal L5 dermatome of the paw while the rat is supported by the examiner. The VFH stimulation occurs in ascending order of force until at least 2 of 3 applications at a given force produces paw withdrawal. Tactile sensory threshold is the lowest force to elicit withdrawal 66% of the time. Acclimation, testing and scoring procedures are described. Aberrant trials that require a retest and typical trials are defined. Animal use was approved by Ohio State University Animal Care and Use Committee.

We describe two tactile sensory testing methods for acute or chronic periods of spinal cord injury in rats. These validated procedures can detect the development and maintenance of allodynia-like sensations.

在大鼠脊髓损伤后的急性和慢性触觉测试

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory ...

科研

医学

Assessment of Vascular Function in Patients With Chronic Kidney Disease

Patients with chronic kidney disease (CKD) have significantly increased risk of cardiovascular disease (CVD) compared to the general population, and this is only partially explained by traditional CVD risk factors. Vascular dysfunction is an important non-traditional risk factor, characterized by vascular endothelial dysfunction (most commonly assessed as impaired endothelium-dependent dilation [EDD]) and stiffening of the large elastic arteries. While various techniques exist to assess EDD and large elastic artery stiffness, the most commonly used are brachial artery flow-mediated dilation (FMDBA) and aortic pulse-wave velocity (aPWV), respectively. Both of these noninvasive measures of vascular dysfunction are independent predictors of future cardiovascular events in patients with and without kidney disease. Patients with CKD demonstrate both impaired FMDBA, and increased aPWV. While the exact mechanisms by which vascular dysfunction develops in CKD are incompletely understood, increased oxidative stress and a subsequent reduction in nitric oxide (NO) bioavailability are important contributors. Cellular changes in oxidative stress can be assessed by collecting vascular endothelial cells from the antecubital vein and measuring protein expression of markers of oxidative stress using immunofluorescence. We provide here a discussion of these methods to measure FMDBA, aPWV, and vascular endothelial cell protein expression.

The degree of vascular dysfunction and contributing physiological mechanisms can be assessed in patients with chronic kidney disease by measuring brachial artery flow-mediated dilation, aortic pulse-wave velocity, and vascular endothelial cell protein expression.

血管功能在慢性肾脏疾病的评估

Patients with chronic kidney disease (CKD) have significantly increased risk of cardiovascular disease (CVD) compared to the general population, and this ...

Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is estimated to contribute to 19- 25% of all clinical cases of acute kidney injury (AKI) in critically ill patients. AKI detection has historically relied on metrics such as serum creatinine (sCr) or blood urea nitrogen (BUN) which are demonstrably inadequate in full assessment of nephrotoxicity in the early phase of renal dysfunction. Currently, there is no robust diagnostic method to accurately detect hemodynamic alteration in the early phase of AKI while such alterations might actually precede the rise in serum biomarker levels. Such early detection can help clinicians make an accurate diagnosis and help in in decision making for therapeutic strategy. Rats were treated with Cisplatin to induce AKI. Nephrotoxicity was assessed for six days using high-frequency sonography, sCr measurement and upon histopathology of kidney. Hemodynamic evaluation using 2D and Color-Doppler images were used to serially study nephrotoxicity in rats, using the sonography. Our data showed successful drug-induced kidney injury in adult rats by histological examination. Color-Doppler based sonographic assessment of AKI indicated that resistive-index (RI) and pulsatile-index (PI) were increased in the treatment group; and peak-systolic velocity (mm/s), end-diastolic velocity (mm/s) and velocity-time integral (VTI, mm) were decreased in renal arteries in the same group. Importantly, these hemodynamic changes evaluated by sonography preceded the rise of sCr levels. Sonography-based indices such as RI or PI can thus be useful predictive markers of declining renal function in rodents. From our sonography-based observations in the kidneys of rats that underwent AKI, we showed that these noninvasive hemodynamic measurements may consider as an accurate, sensitive and robust method in detecting early stage kidney dysfunction. This study also underscores the importance of ethical issues associated with animal use in research.

Early stage hemodynamic dysfunction is critical to the development of kidney disease. Yet, detection methodologies are limited. Recent advances in sonography provide a noninvasive, accurate option for early detection of kidney injury. This study outlines a step-by-step, sonographic methodology for detecting kidney dysfunction using a drug-induced nephrotoxicity rat model.

大鼠使用超声技术药物引起的肾血流动力学障碍的早期检测

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is ...

温针灸缓解慢性阻塞性肺病腹胀

A Preliminary Study on Warm Acupuncture and Moxibustion for Treating Chronic Obstructive Pulmonary Disease with Abdominal Distension

Most patients with COPD have a combination of abdominal distension, which has been shown to adversely affect pulmonary symptoms, frequency of acute exacerbations, and quality of life in patients with COPD. Warm acupuncture and moxibustion have been shown to be effective in relieving symptoms in patients with COPD combined with abdominal distention. Warm acupuncture and moxibustion are highly effective, easy to perform, and inexpensive forms of traditional Chinese medicine treatments. The standardized practice of warm acupuncture and moxibustion is very important for the treatment of COPD combined with abdominal distension. The specific steps include selecting the appropriate acupoints for needling through syndrome differentiation treatment and selecting moxa sticks of appropriate length for moxibustion for about 30 min after the De-qi. The course of treatment lasts for one week. The following indicators are specifically assessed: the score of the COPD Assessment Test (CAT) and the abdominal distension visual analog scale (VAS). This article will clearly illustrate how to standardize the manipulation of warm acupuncture and moxibustion to relieve COPD combined with abdominal distention.

作者聚焦:慢性阻塞性肺疾病腹胀的有效中医治疗方法 

Here, we present a protocol to properly manipulate warm acupuncture and moxibustion for treating chronic obstructive pulmonary disease (COPD) with abdominal distension.

温针灸治疗慢性阻塞性肺疾病伴腹胀的初步研究

Most patients with COPD have a combination of abdominal distension, which has been shown to adversely affect pulmonary symptoms, frequency of acute ...

使用改进的 Videman 方法在兔子中建立 KOA 模型

Application of Acupotomy in a Knee Osteoarthritis Model in Rabbit

Knee osteoarthritis (KOA) is one of the most frequently encountered diseases in the orthopedic department, which seriously reduces the quality of life of people with KOA. Among several pathogenic factors, the biomechanical imbalance of the knee joint is one of the main causes of KOA. Acupotomology believes that restoring the mechanical balance of the knee joint is the key to treating KOA. Clinical studies have shown that acupotomy can effectively reduce pain and improve knee mobility by reducing adhesion, contracture of soft tissues, and stress concentration points in muscles and tendons around the knee joint. 
In this protocol, we used the modified Videman method to establish a KOA model by immobilizing the left hindlimb in a straight position. We have outlined the method of operation and the precautions related to acupotomy in detail and evaluated the efficacy of acupotomy in conjunction with the theory of "Modulating Muscles and Tendons to Treat Bone Disorders" through the detection of the mechanical properties of quadriceps femoris and tendon, as well as cartilage mechanics and morphology. The results show that acupotomy has a protective effect on cartilage by adjusting the mechanical properties of the soft tissues around the knee joint, improving the cartilage stress environment, and delaying cartilage degeneration.

作者聚焦:研究针尖切开术治疗膝骨关节炎的作用机制

In this protocol, a knee osteoarthritis model was prepared using the modified Videman method, and the operation procedures and precautions of acupotomy are detailed. The effectiveness of acupotomy has been demonstrated by testing the mechanical properties of quadriceps femoris and tendon and the mechanical and morphological properties of cartilage.

针灸术在兔膝骨关节炎模型中的应用

Knee osteoarthritis (KOA) is one of the most frequently encountered diseases in the orthopedic department, which seriously reduces the quality of life of ...

<meta charset="utf8"></head><body>岭南火针疗法治疗大鼠骨质疏松症

Operation Procedure and Precautions of Lingnan Fire-Needle Therapy in Osteoporosis Model Rats

Compared to filiform needle therapy, fire-needle therapy has both the stimulation of needles and the warming effect of heat, making it have unexpected effects on some chronic diseases and incurable diseases. Osteoporosis (OP) has a high incidence in postmenopausal women and middle-aged and elderly men, and the treatment cycle is long. According to Traditional Chinese Medicine (TCM), Lingnan fire-needle therapy has shown potential in treating osteoporosis. However, there is still a long way to go before it can be widely used. This article focuses on the application of Lingnan fire-needle therapy in the intervention of OP in rats. It covers the selection of needle tools, acupuncture point selection, positioning of rats' bodies, and fixation methods. We also outline the steps and precautions to be taken during and after needling with fire needles. The experiment was done with three groups: a normal group, a model group, and a fire-needle group, each containing 10 rats. The rats in the fire-needle group were treated with fire-needle intervention for six sessions. After the intervention period, we collected femoral specimens and performed micro-CT scans. The results suggest that fire needling can enhance bone morphology and mineral density in OP rats. This information can serve as a methodological basis for conducting basic research on fire-needle therapy.

<meta charset="utf8"></head><body>作者聚焦：探索岭大火针疗法对骨质疏松症干预的影响

Here, we present a protocol for the application of Lingnan Fire-Needle therapy in the intervention of osteoporosis model rats.

岭南火针疗法治疗骨质疏松模式大鼠的操作流程及注意事项

Compared to filiform needle therapy, fire-needle therapy has both the stimulation of needles and the warming effect of heat, making it have unexpected ...