支持这项工作是由歌德的法兰克福大学医院和德国研究基金会授予对UC

1。眼科窦静脉注射的Ad-2D6 <ol><li>在无菌条件下，淡化的AD-2D6病毒原液浓度为5×10 9 PFU / ml和在RPMI保持在冰上的Ad-2D6病毒解决方案。注射两个剂量5×10 8 PFU（静脉注射和腹腔）已经证明，导致自身免疫性肝炎小鼠的FVB应变的最可靠的结果。 </li><li> 4％异氟醚麻醉室麻醉鼠标。捏的后腿，以确保动物是麻醉。 </li><li>按住鼠标由颈后，收紧松弛的皮肤用大拇指和中指的头。这样，眼睛凸出轻微的眼睛附近的皮肤牵引。 </li><li>将针头垂直入内眦​​（最接近动物的鼻子，眼皮的交界处），缓慢注入100μLAD-2D6病毒解决方案。重要的是不要施加太大的压力，避免损坏的血管和气管。 </li><li>慢慢地退出针头，以避免溢出和关闭眼皮。 </li><li>行之有效的注射病毒解决方案，如果不泄漏槽的鼻子和眼睛幻灯片回到其初始位置。 </li><li>后立即静脉注射，腹腔注射的第二个100μL的Ad-2D6病毒解决方案的标准执行。 </li></ol>另外，静脉注射，可以执行经尾静脉。然而，通过眼科窦注入更容易执行和病毒解决方案的损失降到最低。它已被证明可以互换使用这两种技术和两条路线是同样有效的10。 感染小鼠的不利影响和苦难和痛苦的迹象明显，如食欲不振，体重减轻，失去流动性，未能新郎，被毛粗糙，监察，弯腰驼背外观，以及舔，咬，抓伤，或晃动某一特定地区（即注射部位）。虽然老鼠肝脏CYP2D6的模型显示一个巨大的损害，老鼠似乎健康无痛苦，疼痛或压力的迹象。然而，严重的肝功能衰竭，如血清转氨酶指标确定定期或实验的一部分。 &gt; 1000 U / L，血清转氨酶水平应该只出现作为病毒感染的直接结果，但不长期。如果血清转氨酶水平长期高于1000 U / L（严重限制）被处死小鼠。 2。老鼠的眼睛出血<ol><li> 4％异氟醚麻醉室麻醉鼠标。捏的后腿，以确保动物是麻醉。 </li><li>按住鼠标由颈后，收紧松弛的皮肤用大拇指和中指的头。这样，眼睛凸出轻微毗邻的眼睛的皮肤牵引。 </li><li>毛细管放置在低或内眼角的眼睛，轻轻的小费，但坚定地沿着眼球下滑眼科静脉窦。静脉毛细血管破裂，导致出血填补眶腔。 </li><li>稍微退出毛细管释放的尖端，使累积的血管绘制。 </li><li>当足够的血液收集管被撤回及眼睑关闭。出血停止后，撤出管和静脉复合物后，正常眼压重建。 </li><li>在毛细管血液转移到Microtainer管和培养冰上30分钟。 </li><li> Microtainer管在4°C离心2分钟在7500 XG </li><li>上清对应血清被转移到一个新的管，并保存在-80°C。 </li><li>可用于血清，以确定用ELISA抗体滴度。籼稻职权范围的肝损害，如使用罗氏诊断试纸（德国曼海姆）和Reflotron加分析仪（血清谷丙转氨酶（ALT / GPT），谷草转氨酶（AST / GOT），可以评估肝酶罗氏诊断，曼海姆，德国）。 </li></ol>另外，采血可以通过经尾静脉或颞浅静脉11。 3。小鼠肝脏灌注 注：这一步很重要，以避免孤立的肝细胞通过血液淋巴细胞的污染<ol><li>安乐死CO 2的致死剂量，其次是颈椎脱位鼠标。 </li><li>装载动物，其背部的发泡板铺设。使用23政针针四肢。 </li><li>滋润和鼠标消毒，用70％乙醇</li><li>约1厘米，离后腿，使切口，通过皮肤和肌肉层。减少动物的两个工作膜片两侧。 </li><li>当隔膜到达皮肤向后翻转。 </li><li>削减隔膜，从撕裂，然后切下腔静脉。 </li><li>移动侧的胃和肠子，露出门静脉。 </li><li>在门静脉插入一个27 G的针头连接到5毫升注射器，充满冷PBS。让PBS的慢慢洗肝脏的血液。 </li><li>肝脏解剖抓住膜片上，并从体内切割膜片离。 </li><li>肝脏转移到培养皿取出膜片，胆囊和其他组织仍连接到肝脏</li><li>转移肝脏上的冰管50毫升或10毫升PBS在华侨城化合物嵌入。 </li></ol> 4。肝细胞的分离<ol><li>准备以下的股票解决方案： <ul><li> 胶原酶的股票 （100毫克/毫升）列印PBS。 （小分装在-20°C和存储）。 </li><li>在PBS的DNA酶的股票 （25毫克/毫升）。 （小分装在-20°C和存储）。 </li><li> 胶原酶缓冲区 ：PBS含0.2毫克/毫升胶原酶，0.02毫克/毫升DNA酶和5％胎牛血清，1肝，混合9.5毫升冰冷的PBS，20μL胶原酶（100毫克/毫升的股票），8μLDNA酶（ 25毫克/毫升的股票），500μL热灭活FCS。 </li><li> 35％的Percoll混合175毫升的Percoll，20毫升10×PBS股票; 305毫升PBS。 </li><li> 的RPMI 完整的RPMI含有10％热灭活FCS，100μ/ ml青霉素，100μg/ mL链霉素，2毫米L-谷氨酰胺。 </li></ul></li><li>灌注肝（见第3节）转移至10毫升新鲜的PBS在培养皿上冰，切成小块，用剪刀。 </li><li>通过用玻璃杵或杵从2毫升注射器转移到70微米的细胞过滤和挤压肝脏帆船。小心加入10毫升冷胶原酶缓冲区和按通过过滤器。收集悬浮液通过过滤器和过滤器的2倍以上。 </li><li>转让暂停50毫升新鲜管上冰。处理下一肝。 </li><li>在37°C孵育60分钟的肝细胞悬液，轻轻混匀，每15分钟。 </li><li>在4°C离心3分钟，30 XG </li><li>将上清转移到一个新的管，留下5毫米以上的颗粒流体</li><li>在4°C离心10分钟在650 XG </li><li>弃上清，留​​3毫米以上的颗粒</li><li>重悬浮颗粒在20毫升的Percoll缓冲区</li><li> 4°C离心20分钟在600 XG </li><li>由弹管，弃掉上清，重悬沉淀。 </li><li>用PBS洗涤沉淀1个。 </li><li>洗的RPMI 完整颗粒1× </li><li>重悬浮颗粒在3毫升的RPMI 完成，并指望在1:10稀释的细胞。 </li><li> 完成 〜10 7细胞/毫升的RPMI重悬细胞和管转移到冰。 </li></ol&gt; 5。胞内细胞因子染色（ICCS的） 5.1刺激： <ol><li>准备〜10 7细胞/ ml肝细胞在RPMI 完整的描述。板100μL（10 6细胞）/到一个单位的96孔板，这是组织培养处理。 </li><li>加入50μL的RPMI 完成包含为2μg/ml布雷菲德菌素A，然后加入50μL的RPMI 完整含有2微克/毫升刺激CYP2D6的肽（即免疫的CD4抗原CYP2D6的41-60 PGLGNLLHVD FQNTPYCFDQ 12的免疫CD8抗原CYP2D6的193-212 RRFEYDDPRF LRLLDLAQEG 12）。混合移液。 </li><li>孵育5小时，在37°C（最佳刺激时间，但过夜培养工程）。 </li></ol> 5.2。染色： <ol><li>准备以下的股票解决方案： <ul type="disc"><li> 流式细胞仪缓冲液 ：PBS含1％小牛小号erum（FCS）。 </li><li> 固定/通透缓冲：流式细胞仪缓冲区含有0.1％的皂甙和4％多聚甲醛</li><li> 流式细胞仪/皂素的洗涤缓冲液 ：流式细胞仪缓冲液含0.1％皂素</li><li> 流式细胞仪/ PFA流式细胞仪的缓冲区 ：缓冲区含有1％多聚甲醛（煤灰） </li></ul></li><li>转入V型底酶标板（96孔）细胞和旋转460 XG为3分钟，在4°C放弃中型和涡板</li><li>在3分钟的460 XG加入150μL流式细胞仪缓冲和离心机在4°C。丢弃介质和涡板。重复洗第一步。 </li><li>块，如果必要的表面FCR（当使用二次抗体）1微克/毫升αCD16/32鸡尾酒流式细胞仪缓冲（FCR块）为15分钟，4°C和150μL染色缓冲液洗2×（460 XG 2分钟4℃）。 </li><li>表面分子：即反CD8a-FITC单克隆抗体染色10微克/毫升，在50μL流式细胞仪为30分钟，在4°C黑暗环境中的缓冲区。 </li><li>加入100μL流式细胞仪在460 XG 3分钟，在4°C的缓冲区和自旋</li><li>洗涤细胞2×150μL流式细胞仪缓冲区（460 XG; 3分钟; 4℃）。涡盘。 </li><li>固定/通透细胞100μL固定/通透性在室温下10分钟的缓冲。 </li><li>在4°C的自旋为7分钟460 XG请注意，重要的是7分钟，延长离心时间以来的固定/ permeabilzation的步骤，改变细胞的整体密度。涡盘。 </li><li>洗2×150μL流式细胞仪/皂素的洗涤缓冲液（460 XG; 7分钟; 4℃）。涡盘。 </li><li>染色细胞内分子：即反IFNγ的聚乙烯单抗10微克/毫升在50μL流式细胞仪/ 30分钟的皂素的洗涤缓冲液在4°C。 </li><li>加入7分钟在460 XG 100μL流式细胞仪/皂素的洗涤缓冲液和自旋在4°C。 </li><li>洗涤细胞2×150μL流式细胞仪/皂素的洗涤缓冲液（460 XG; 7分钟; 4℃）。涡盘。 </li><li>洗细胞1流式细胞仪缓冲区（460 XG; 7分钟; 4℃）。涡盘。 </l我&gt; <li>重悬细胞，直到在200μL流式细胞仪/煤灰缓冲，转移到管，并储存在4°C黑暗环境中，通过流式细胞仪的数据采集。我们通常使用FACSCanto II或FACSCalibur（BD公司，德国海德堡）。 </li></ol> 6。免疫组化 6.1。 H＆E染色，一般肝脏病理的评估： <ol><li>收获肝沉浸，TEK组织华侨城在一次性的基础模具及干冰速冻。 </li><li>削减7微米的肝组织切片，使用徕卡低温恒温器在-17°C和安装在Superfrost加上显微镜幻灯片的组织切片。商店幻灯片在-20°C，直到进一步的使用。 </li><li>冰冻切片在预冷的乙醇固定在-20°C 15分钟10分钟，让干燥。 </li><li>蒸馏水洗2×2分钟在室温（以下步骤6.1.5 - 6.1.13，在室温下进行）。 </li><li>迈尔的苏木解决方案中的染色8分钟。 </li><li> 10分钟用温水（30℃）自来水。改变几次水。 </li><li>在蒸馏水冲洗。 </li><li> 95％的乙醇冲洗20秒。 </li><li>在染液曙红G / Y解决方案，为40秒。 </li><li>脱水2×95％乙醇5分钟，每孵化。 </li><li> 5分钟，每孵化100％乙醇脱水2×。 </li><li>清除二甲苯2×5分钟每结算。 </li><li>安装在ROTI-Histokitt。 </li></ol> 6.2。 I型胶原沉积的免疫组化： <ol><li>收获肝，沉浸在组织-Tek的OCT。在一次性的基础模具及干冰速冻。 </li><li>削减7微米的肝组织切片，使用徕卡低温恒温器在-17°C和安装在Superfrost加上显微镜幻灯片的组织切片。商店幻灯片在-20°C，直到进一步的使用。 </li><li>在15分钟-20℃冷乙醇固定冰冻切片10分钟，让干燥。 </li><li>在PBS清洗，在室温下2分钟2个。 </li> &lt;LI&gt;在PBS孵育含0.3％H 2 O 2和0.1％，钠，叠氮化物在室温为10分钟。 </li><li>在PBS清洗，在室温下2分钟2个。 </li><li>座与亲和素阻断根据制造商的指引套件。 </li><li>阻止10％FCS的PBS（FCS的/ PBS）在室温下30分钟。 </li><li>主要的抗体是一只兔子的抗小鼠I型胶原抗体，其中10％的FCS / PBS稀释1:200。孵育抗体在室温下2小时的部分。 </li><li>洗节在PBS在室温下4分钟3 x。 </li><li> 1:500稀释生物素化的抗兔抗体，室温孵育1.5小时部分。 </li><li>洗节在PBS在室温下4分钟3 x。 </li><li>显色反应得到由顺序孵化与抗生物素蛋白过氧化物酶的共轭二氨基联苯胺 - 过氧化氢，根据制造商的指引。 </li><li>染液在迈耶的他5分钟matoxylin解决方案，在PBS洗2×2分钟在室温和安装在Aquatex。 </li></ol> 7。代表结果 AD-2D6诱导肝损伤的两个不同阶段的小鼠的感染。在感染后的第一天，肝脏的病毒感染引起的急性血清转氨酶水平增加，肝细胞死亡6的指标。这第一，急性期发生的独立的hCYP2D6表达和空控制病毒（AD-控制）或病毒表达绿色荧光蛋白（AD-GFP）感染后，也可以观察。相比之下，第二阶段是自身免疫介导的依赖，触发分子hCYP2D6的表达。这种自身免疫性阶段发生在感染后2-4周，并持续数为6个月。 <img alt="图1" src="/files/ftp_upload/3644/3644fig1.jpg" /> &lt;一类p =“jove_content”&gt; 图1。 CYP2D6的模型。野生型的FVB或C57BL / 6小鼠注射Ad-2D6的两个剂量（静脉注射和腹腔）。在利益时，肝脏和血清收集和评估肝功能损害和hCYP2D6特异性抗体和T细胞的形成。 以下功能是野生型FVB或C57BL / 6小鼠在CYP2D6模型AD-2D6感染后发展为持久性的自身免疫性肝炎的特点：第一，肝脏显示一个aberrantly改变形态。增生结节，尤其是肝叶融合，出现散在整个肝脏和一个巨大的囊纤维化是可见的（图2）。 <img alt="图2" src="/files/ftp_upload/3644/3644fig2.jpg" /> 图2。在4周后感染检测典型的AD-2D6引起的肝损害肝脏形态。。请注意，个别肝叶融合（箭头）。增生结节似乎遍布整个肝脏（箭头）。控制不表达hCYP2D6，腺病毒感染对肝脏的形态没有影响。 第二，广泛和持久性细胞浸润出现在城郊门户和实质（图3A）AD-2D6-感染而不是广告控制感染小鼠肝脏的地区。纤维化为主的发展进入实质（图3B）突出一些胶原束在包膜地区。 <img alt="图3" src="/files/ftp_upload/3644/3644fig3.jpg" /> 图3。肝脏组织学答：无论是广告控制在4周后感染AD-2D6 FVB小鼠感染的肝组织切片H＆E染色。需要注意的是显著的单核细胞浸润，是唯一目前在Ad-2D6感染小鼠（单箭头）。三重箭头指示桥接相邻门户大片肝实质细胞浸润。乙：AD-2D6或广告控制在感染后4周肝纤维化的免疫组化的代表性。使用抗I型胶原抗体的胶原蛋白，肝脏部分都被染红了。注意肝包膜下与一些凸出束进入实质（单箭头）和浸润细胞（箭头）的大型集群（三箭）多层胶原处置。两个代表肝部分显示每个条件。大小酒吧：为100μm。 第三个特点是产生高滴度的抗CYP2D6的抗体，其中有一个类似的抗原特异性人类的LKM-1抗体6,13。最后，hCYP2D6的具体CD4和CD8 T细胞的产生，主要是家里的肝脏。如hCYP2D6特异性T细胞可以检测到刺激与immunodomi南特hCYP2D6肽和细胞内细胞因子染色（ICCS）（图4）后续计量IFNγ的表达。 hCYP2D6-特异性CD8 T细胞杀死体内 12 AD-2D6感染的靶细胞。 <img alt="图4" src="/files/ftp_upload/3644/3644fig4.jpg" /> 图4。 hCYP2D6特异性T细胞。流的hCYP2D6特异性T细胞的流式细胞仪分析。 AD-2D6感染4周后，肝细胞已被隔离，并刺激免疫的CD4或CD8 + hCYP2D6抗原过夜。 γ干扰素生成细胞内细胞因子染色检测和流量使用1流式细胞仪粤语II（BD公司，德国海德堡）流式细胞仪分析。的hCYP2D6特异性CD4和CD8细胞的百分比表示。

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Autoimmun. , (2011).</li><li>Ando, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Class+I-restricted+cytotoxic+T+lymphocytes+are+directly+cytopathic+for+their+target+cells+in+vivo.">Class I-restricted cytotoxic T lymphocytes are directly cytopathic for their target cells in vivo.</a> J. Immunol. 152, 3245-3253 (1994).</li><li>Zierden, M., Kuhnen, E., Odenthal, M., Dienes, H. 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Immunol. 28, 2395-2406 (1998).</li><li>Voehringer, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Break+of+T+cell+ignorance+to+a+viral+antigen+in+the+liver+induces+hepatitis.">Break of T cell ignorance to a viral antigen in the liver induces hepatitis.</a> J. Immunol. 165, 2415-2422 (2000).</li><li>Derkow, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+priming+of+CD8+and+CD4+T-cells+in+animal+models+of+autoimmune+hepatitis+and+cholangitis.">Differential priming of CD8 and CD4 T-cells in animal models of autoimmune hepatitis and cholangitis.</a> Hepatology. 46, 1155-1165 (2007).</li><li>Lapierre, P., Djilali-Saiah, I., Vitozzi, S., Alvarez, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+murine+model+of+type+2+autoimmune+hepatitis:+Xenoimmunization+with+human+antigens.">A murine model of type 2 autoimmune hepatitis: Xenoimmunization with human antigens.</a> Hepatology. 39, 1066-1074 (2004).</li><li>Djilali-Saiah, I., Lapierre, P., Vittozi, S., Alvarez, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=DNA+vaccination+breaks+tolerance+for+a+neo-self+antigen+in+liver:+a+transgenic+murine+model+of+autoimmune+hepatitis.">DNA vaccination breaks tolerance for a neo-self antigen in liver: a transgenic murine model of autoimmune hepatitis.</a> J. Immunol. 169, 4889-4896 (2002).</li><li>Christen, U., Hintermann, E., Holdener, M., von Herrath, 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=Viral+triggers+for+autoimmunity:+is+the+'glass+of+molecular+mimicry'+half+full+or+half+empty.">Viral triggers for autoimmunity: is the 'glass of molecular mimicry' half full or half empty.</a> J. Autoimmun. 34, 38-44 (2010).</li><li>Krebs, P., Scandella, E., Odermatt, B., Ludewig, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+functional+exhaustion+and+deletion+of+CTL+following+immunization+with+recombinant+adenovirus.">Rapid functional exhaustion and deletion of CTL following immunization with recombinant adenovirus.</a> J. Immunol. 174, 4559-4566 (2005).</li><li>Hintermann, E., Bayer, M., Pfeilschifter, J., Christen, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adenoviral+delivery+of+the+liver+autoantigen+cytochrome+P450+2D6+chronically+activates+hepatic+stellate+cells+and+induces+fibrosis.">Adenoviral delivery of the liver autoantigen cytochrome P450 2D6 chronically activates hepatic stellate cells and induces fibrosis.</a> Manuscript Submitted. , (2011).</li></ol>

作者没有透露。

在以往的研究，实验性肝炎经常报道只是短暂的，许多自身免疫性肝病的电流模式依赖于一个相当复杂的疾病诱导协议（评论见3,5）。例如，一些车型使用转基因小鼠表达特定的靶抗原，继转移目标抗原特异性的，大多是TCR转基因T细胞14,15。往往是必要的额外，与livertropic病毒，细菌或寄生虫感染诱发疾病16-18。另外，为靶抗原，包括CYP2D6的19，编码质粒DNA疫苗用于诱发肝炎。然而，额外的疫苗与质粒编码促炎细胞因子，如IL-12，20。不幸的是，有少数例外，15,20型肝炎仅是短暂的。 CYP2D6的模型6,21使用一个简单的方法TO诱导例AIH-2 7,8的主要自身抗原，表达hCYP2D6，腺病毒感染的小鼠自身免疫性肝炎。 CYP2D6的交付由腺病毒的结构，保证了肝脏的直接定位以及局部炎症，促进宽容破裂。这是重要的，但是要注意，病毒滴度以及给药途径是这种模式的关键。一方面，AD-2D6是一种复制缺陷病毒，因此，需要有足够高的病毒滴度肝脏内产生的抗原的关键。另一方面，滴度过高，可能会导致致命的急性肝功能衰竭。此外，它已被证明以前高滴度腺病毒的小鼠感染导致功能衰竭的免疫反应22。在我们的模型中，我们使用了静脉注射和腹腔感染的组合以获得慢性细胞浸润以及分机ensive纤维化。我们已经看到，单靠静脉注射感染仍然造成大量细胞浸润，但没有纤维化，表明由于腹腔注射，腹腔炎症，包膜地区可能会在胶原蛋白的不断的包膜积累（欣特曼和克里森，准备手稿）参与。然而，重要的是要注意观察肝纤维化是抗原特异性的，因为我们没有检测感染后，等于AD-GFP或Ad-23控制病毒滴度与肝星状细胞活化及胶原沉积。 CYP2D6的模型反映了人类AIH的1,2的许多方面，如慢性肝炎特点是持续性细胞浸润和肝纤维化6。此外，存在高滴度的抗体具有相似的抗原特异性和传播13例AIH-2患者中发现的LKM-1抗体的抗CYP2D6的指示前SENCE的一种常见的慢性自身免疫性反应。 CYP2D6的是，在2 AIH的主要抗原，但它不是由更频繁的1型AIH的确诊患者的认可。此外，病人患上其他肝病的病因与自身免疫性疾病，如原发性胆汁性肝硬化（PBC），原发性硬化性胆管炎（PSC），生成的标志抗体，显示特异性明显的肝自身抗原和肝脏不同的病理特点，围绕对小（中国人民银行）或大（PSC）的胆管。然而，CYP2D6的模型分析慢性肝脏炎症过程中涉及发生在自身免疫性肝病，确定关键的球员，推动肝细胞自身免疫性破坏，并评估可能治愈疾病的治疗干预的免疫病理机制提供了机会。

<table border="1"><tbody><tr><td> 设备名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td> 23G针</td><td> BD公司</td><td> 300800 </td><td></td></tr><tr><td> 27½Ğ针</td><td>厂商VWR </td><td> 612-0151 </td><td></td></tr><tr><td> 30½Ğ针</td><td> BD公司</td><td> 304000 </td><td></td></tr><tr><td>谷丙转氨酶试纸（谷丙转氨酶/ ALT） </td><td>罗氏诊断</td><td> 10 745 138 202 </td><td></td></tr><tr><td>谷草转氨酶试纸（GOT /助攻） </td><td>罗氏诊断</td><td> 10 745 120 202 </td><td></td></tr><tr><td>麻醉组Univentor 400 </td><td> <a href="http://www.agnthos.se" target="_blank" >AgnTho的</a> </td><td></td><td></td></tr><tr><td>相应的模具，一次性37x24x10毫米</td><td>厂商VWR </td><td> 720-0208 </td><td></td></tr><tr><td>细胞过滤70毫米</td><td>厂商VWR </td><td> 734-0003 </td><td></td></tr><tr><td>低温恒温器</td><td>徕卡</td><td> CM1850紫外线</td><td></td></tr><tr><td>肝素化毛细管管</td><td>费舍尔</td><td> 3123987 </td><td></td></tr><tr><td>显微镜载玻片Superfrost加</td><td>门泽尔格拉泽</td><td> J1800AMN​​Z </td><td></td></tr><tr><td> microtainer不锈钢管</td><td> BD公司</td><td> 365951 </td><td></td></tr><tr><td>酶标板，V型底部</td><td>厂商VWR </td><td> 391-1924 </td><td></td></tr><tr><td> reflotron加</td><td>罗氏诊断</td><td></td><td> determiniation的血清AST / ALT </td></tr><tr><td colspan="4"></td></tr><tr><td> 姓名of的试剂</td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td>兔抗鼠抗I型胶原抗体， </td><td> Chemicon公司</td><td> AB765P </td><td></td></tr><tr><td>生物素标记的羊抗兔IgG </td><td>载体（AXXORA） </td><td>的VC-BA-1000-MC15 </td><td></td></tr><tr><td> FITC标记的抗鼠CD8a抗体</td><td>南方生物技术</td><td> 1550至1502年</td><td></td></tr><tr><td> PE标记的抗小鼠IFNγ的抗体</td><td> BD公司</td><td> 554412 </td><td></td></tr><tr><td> PE标记的抗小鼠CD16/CD32抗体（FCR块） </td><td> BD公司</td><td> 553141 </td><td></td></tr><tr><td> aquatex </td><td>厂商VWR </td><td> 1.08562.0050 </td><td></td></tr><tr><td>抗生物素蛋白/生物素阻断试剂盒</td><td矢量（AXXORA） </td><td>的VC-SP-2001-KI01 </td><td></td></tr><tr><td>布雷菲德菌素A </td><td>西格玛</td><td> B6542 </td><td></td></tr><tr><td>胶原酶</td><td>西格玛</td><td> C5138-100MG </td><td></td></tr><tr><td>民建联基板套件3&#39;3&#39;diaminobenzidine </td><td>载体（AXXORA） </td><td> VC - SK-4100-KI01 </td><td></td></tr><tr><td> DNA酶</td><td>西格玛</td><td> DN-25 </td><td></td></tr><tr><td>精英农行试剂</td><td>载体（AXXORA） </td><td> VC - PK-7100-L050 </td><td></td></tr><tr><td>曙红G / Y解决方案</td><td>罗斯</td><td> X883.1 </td><td></td></tr><tr><td>小牛血清（FCS） </td><td> biochrom公司</td><td>小号0115 </td><td></td></tr><tr><td>异氟醚</td><td> forene </td><td> B506 </td><td></td></tr><tr><td>迈尔的苏木解决方案</td><td> AppliChem </td><td> A4840，1000 </td><td></td></tr><tr><td>华侨城复合</td><td>欧洲樱花Finetek </td><td> 4583 </td><td></td></tr><tr><td>甲醛PBS缓冲的10％ </td><td>罗斯</td><td> A146.3 </td><td></td></tr><tr><td> Percoll密度</td><td>阿默舍姆</td><td> 17-0891-01 </td><td></td></tr><tr><td> ROTI-Histokit </td><td>罗斯</td><td> 6638.1 </td><td></td></tr><tr><td>的RPMI </td><td> Invitrogen公司</td><td> 61870 </td><td></td></tr><tr><td>从quillaja皂皂甙的树皮</td><td>西格玛</td><td> S7900 </td><td></td></tr></tbody></table>

the cyp2d6 animal model: how to induce autoimmune hepatitis in mice

自身免疫性肝炎是一种罕见但危及生命的自身免疫性疾病的病因不明，1,2肝脏。在过去的许多尝试已作出生成一个动物模型，反映了人类疾病的特点3-5。然而，在各种型号的感应疾病是相当复杂的，往往肝炎只是短暂的3-5。因此，我们已经开发出一种简单的小鼠模型，在2型自身免疫性肝炎（AIH的-2），即hCYP2D6，使用人类的主要自身抗原作为触发6。 1型肝肾微粒体抗体（LKM-1）抗体承认hCYP2D6例AIH-2 7,8的标志。 hCYP2D6交付是由腺病毒结构（AD-2D6），确保触发肝抗原直接传送到野生型FVB或C57BL / 6小鼠。因此，随之而来的局部炎症产生的自身免疫性疾病的后续发展肥田9。一个CAD-2D6 ombination静脉注射和腹腔注射是最有效的途径，诱发一种持久的自身免疫性损害肝脏（1节）。在这里，我们提供了一个如何在CYP2D6模型诱导自身免疫性肝病和肝功能损害的不同方面如何可以评估的详细协议。首先，表明肝细胞的破坏，如转氨酶以及hCYP2D6抗体的滴度的血清标记确定采样的血液retroorbitaly（第2节）。其次，在hCYP2D6特异性T细胞反应的特点是通过收集脾淋巴细胞和肝脏。为了获得纯的肝细胞，肝脏灌注通过门静脉（第3）胶原蛋白和消化，通过Percoll梯度（第4）纯化，用PBS。分析与刺激hCYP2D6肽和鉴定流式细胞仪检测细胞IFNγ的生产（第5条）的hCYP2D6特异性T细胞的频率。第三，由肝部分（第6条）免疫组化细胞浸润和纤维化。这样的分析方案，以证明该模型的长期性疾病开始后要进行多次。特点是数量级的频率和hCYP2D6特异性T和/或B细胞的活性和肝损伤和肝纤维化程度的免疫反应，被评价为后续可能的治疗，以防止，延缓或废除autodestructive评估肝脏的过程。

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The CYP2D6 Animal Model: How to Induce Autoimmune Hepatitis in Mice

小鼠与表达人类的主要自身抗原的腺病毒感染的细胞色素P450 2D6（hCYP2D6）血清持久的特点是广泛的肝炎，肝纤维化和生成一种CYP2D6的形式在介导的自身免疫性肝病患者患有2型自身免疫性肝炎的结果确认特异性免疫反应。

CYP2D6动物模型：如何诱导小鼠自身免疫性肝炎

Autoimmune hepatitis is a rare but life threatening autoimmune disease of the liver of unknown etiology1,2. In the past many attempts have been made to ...

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18.9K 次观看.  Goethe University Hospital Frankfurt. 小鼠与表达人类的主要自身抗原的腺病毒感染的细胞色素P450 2D6（hCYP2D6）血清持久的特点是广泛的肝炎，肝纤维化和生成一种CYP2D6的形式在介导的自身免疫性肝病患者患有2型自身免疫性肝炎的结果确认特异性免疫反应。

视频: The CYP2D6 Animal Model: How to Induce Autoimmune Hepatitis in Mice

Pressure Controlled Ventilation to Induce Acute Lung Injury in Mice

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

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

压力控制通气诱导小鼠急性肺损伤

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

科研

医学

Biomarkers in an Animal Model for Revealing Neural, Hematologic, and Behavioral Correlates of PTSD

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for objective diagnosis but also for the evaluation of therapeutic efficacy and resilience to trauma. Ongoing research is directed at identifying molecular biomarkers for PTSD, including traumatic stress induced proteins, transcriptomes, genomic variances and genetic modulators, using biologic samples from subjects' blood, saliva, urine, and postmortem brain tissues. However, the correlation of these biomarker molecules in peripheral or postmortem samples to altered brain functions associated with psychiatric symptoms in PTSD remains unresolved. Here, we present an animal model of PTSD in which both peripheral blood and central brain biomarkers, as well as behavioral phenotype, can be collected and measured, thus providing the needed correlation of the central biomarkers of PTSD, which are mechanistic and pathognomonic but cannot be collected from people, with the peripheral biomarkers and behavioral phenotypes, which can.
Our animal model of PTSD employs restraint and tail shocks repeated for three continuous days - the inescapable tail-shock model (ITS) in rats. This ITS model mimics the pathophysiology of PTSD 17, 7, 4, 10. We and others have verified that the ITS model induces behavioral and neurobiological alterations similar to those found in PTSD subjects 17, 7, 10, 9. Specifically, these stressed rats exhibit (1) a delayed and exaggerated startle response appearing several days after stressor cessation, which given the compressed time scale of the rat's life compared to a humans, corresponds to the one to three months delay of symptoms in PTSD patients (DSM-IV-TR PTSD Criterian D/E 13), (2) enhanced plasma corticosterone (CORT) for several days, indicating compromise of the hypothalamopituitary axis (HPA), and (3) retarded body weight gain after stressor cessation, indicating dysfunction of metabolic regulation.
The experimental paradigms employed for this model are: (1) a learned helplessness paradigm in the rat assayed by measurement of acoustic startle response (ASR) and a charting of body mass; (2) microdissection of the rat brain into regions and nuclei; (3) enzyme-linked immunosorbent assay (ELISA) for blood levels of CORT; (4) a gene expression microarray plus related bioinformatics tools 18. This microarray, dubbed rMNChip, focuses on mitochondrial and mitochondria-related nuclear genes in the rat so as to specifically address the neuronal bioenergetics hypothesized to be involved in PTSD.

We describe a rat model of post traumatic stress disorder (PTSD) that reveals the persistent alterations in neuroendocrine function and the delayed long-term, exaggerated fear response, characteristic of PTSD patients. The animal model and methods described here are useful for correlating biomarkers in brain nuclei, which are mechanistic but cannot be measured in patients, with biomarkers in peripheral white blood cells, which can.

在动物模型揭示神经，血液，创伤后应激障碍和行为相关因素的生物标志物

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for ...

Protocol for Long Duration Whole Body Hyperthermia in Mice

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body temperature that is observed during fever. The use of hyperthermia as an adjuvant has emerged as a promising procedure for tumor regression in the field of cancer biology. For this purpose, the most important requirement is to have reliable and uniform heating protocols. We have developed a protocol for hyperthermia (whole body) in mice. In this protocol, animals are exposed to cycles of hyperthermia for 90 min followed by a rest period of 15 min. During this period mice have easy access to food and water. High body temperature spikes in the mice during first few hyperthermia exposure cycles are prevented by immobilizing the animal. Additionally, normal saline is administered in first few cycles to minimize the effects of dehydration. This protocol can simulate fever like conditions in mice up to 12-24 hr. We have used 8-12 weeks old BALB/Cj female mice to demonstrate the protocol.

This paper describes a protocol for whole body hyperthermia in mice that can stimulate fever like conditions up to 12-24 hr.

持续时间较长的小鼠全身热疗协议

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body ...

Use of Animal Model of Sepsis to Evaluate Novel Herbal Therapies

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several techniques, including infusion of exogenous bacterial toxin (endotoxemia) or bacteria (bacteremia), as well as surgical perforation of the cecum by cecal ligation and puncture (CLP)1-3. CLP allows bacteria spillage and fecal contamination of the peritoneal cavity, mimicking the human clinical disease of perforated appendicitis or diverticulitis. The severity of sepsis, as reflected by the eventual mortality rates, can be controlled surgically by varying the size of the needle used for cecal puncture2. In animals, CLP induces similar, biphasic hemodynamic cardiovascular, metabolic, and immunological responses as observed during the clinical course of human sepsis3. Thus, the CLP model is considered as one of the most clinically relevant models for experimental sepsis1-3.
Various animal models have been used to elucidate the intricate mechanisms underlying the pathogenesis of experimental sepsis. The lethal consequence of sepsis is attributable partly to an excessive accumulation of early cytokines (such as TNF, IL-1 and IFN-&gamma;)4-6 and late proinflammatory mediators (e.g., HMGB1)7. Compared with early proinflammatory cytokines, late-acting mediators have a wider therapeutic window for clinical applications. For instance, delayed administration of HMGB1-neutralizing antibodies beginning 24 hours after CLP, still rescued mice from lethality8,9, establishing HMGB1 as a late mediator of lethal sepsis. The discovery of HMGB1 as a late-acting mediator has initiated a new field of investigation for the development of sepsis therapies using Traditional Chinese Herbal Medicine. In this paper, we describe a procedure of CLP-induced sepsis, and its usage in screening herbal medicine for HMGB1-targeting therapies.

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, and can be simulated by a surgical technique termed cecal ligation and puncture (CLP). Here we describe a method to use CLP-induced animal model to screen medicinal herbs for therapeutic agents.

脓毒症动物模型，用于评估新型草药疗法

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several ...

Visualization and Analysis of Blood Flow and Oxygen Consumption in Hepatic Microcirculation: Application to an Acute Hepatitis Model

There is a considerable discrepancy between oxygen supply and demand in the liver because hepatic oxygen consumption is relatively high but about 70% of the hepatic blood supply is poorly oxygenated portal vein blood derived from the gastrointestinal tract and spleen. Oxygen is delivered to hepatocytes by blood flowing from a terminal branch of the portal vein to a central venule via sinusoids, and this makes an oxygen gradient in hepatic lobules. The oxygen gradient is an important physical parameter that involves the expression of enzymes upstream and downstream in hepatic microcirculation, but the lack of techniques for measuring oxygen consumption in the hepatic microcirculation has delayed the elucidation of mechanisms relating to oxygen metabolism in liver. We therefore used FITC-labeled erythrocytes to visualize the hepatic microcirculation and used laser-assisted phosphorimetry to measure the partial pressure of oxygen in the microvessels there. Noncontact and continuous optical measurement can quantify blood flow velocities, vessel diameters, and oxygen gradients related to oxygen consumption in the liver. In an acute hepatitis model we made by administering acetaminophen to mice we observed increased oxygen pressure in both portal and central venules but a decreased oxygen gradient in the sinusoids, indicating that hepatocyte necrosis in the pericentral zone could shift the oxygen pressure up and affect enzyme expression in the periportal zone. In conclusion, our optical methods for measuring hepatic hemodynamics and oxygen consumption can reveal mechanisms related to hepatic disease.

An optical system was developed to visualize hepatic microcirculation with FITC-labeled erythrocytes and to measure the partial pressure of oxygen in the microvessels with laser-assisted phosphorimetry. This method can be used to investigate physiological and pathological mechanisms by analyzing microvascular structure, diameter, blood flow velocity, and oxygen tension.

可视化和肝脏微循环血流量和耗氧量分析：急性肝炎模型中的应用

There is a considerable discrepancy between oxygen supply and demand in the liver because hepatic oxygen consumption is relatively high but about 70% of ...

Implanting Glass Spinal Cord Windows in Adult Mice with Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as multiple sclerosis. Here we present a low-cost and reproducible glass window implantation protocol that is suitable for intravital microscopy and studying the dynamics of spinal cord cytoarchitecture with subcellular resolution in live adult mice with EAE. Briefly, we surgically expose the vertebrae T12-L2 and construct a chamber around the exposed vertebrae using a combination of cyanoacrylate and dental cement. A laminectomy is performed from T13 to L1, and a thin layer of transparent silicone elastomer is applied to the dorsal surface of the exposed spinal cord. A modified glass cover slip is implanted over the exposed spinal cord taking care that the glass does not directly contact the spinal cord. To reduce the infiltration of inflammatory cells between the window and spinal cord, anti-inflammatory treatment is administered every 2 days (as recommended by ethics committee) for the first 10 days after implantation. EAE is induced only 2-3 weeks after the cessation of anti-inflammatory treatment.
Using this approach we successfully induced EAE in 87% of animals with implanted windows and, using Thy1-CFP-23 mice (blue axons in dorsal spinal cord), quantified axonal loss throughout EAE progression. Taken together, this protocol may be useful for studying the recruitment of various cell populations as well as their interaction dynamics, with subcellular resolution and for extended periods of time. This intravital imaging modality represents a valuable tool for developing therapeutic strategies to treat autoimmune demyelinating diseases such as EAE.

We describe a method for implanting and maintaining glass windows over the exposed spinal cords of adult mice for studying experimental autoimmune encephalomyelitis. These windows provide chronic optical access to the spinal cord for monitoring cell dynamics at the subcellular level in live animals using two-photon microscopy.

玻璃植入脊髓的Windows在成年小鼠实验性自身免疫性脑脊髓炎

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as ...

A Magnetic Microbead Occlusion Model to Induce Ocular Hypertension-Dependent Glaucoma in Mice

The use of rodent models of glaucoma has been essential to understand the molecular mechanisms that underlie the pathophysiology of this multifactorial neurodegenerative disease. With the advent of numerous transgenic mouse lines, there is increasing interest in inducible murine models of ocular hypertension. Here, we present an occlusion model of glaucoma based on the injection of magnetic microbeads into the anterior chamber of the eye using a modified microneedle with a facetted bevel. The magnetic microbeads are attracted to the iridocorneal angle using a handheld magnet to block the drainage of aqueous humour from the anterior chamber. This disruption in aqueous dynamics results in a steady elevation of intraocular pressure, which subsequently leads to the loss of retinal ganglion cells, as observed in human glaucoma patients. The microbead occlusion model presented in this manuscript is simple compared to other inducible models of glaucoma and also highly effective and reproducible. Importantly, the modifications presented here minimize common issues that often arise in occlusion models. First, the use of a bevelled glass microneedle prevents backflow of microbeads and ensures that minimal damage occurs to the cornea during the injection, thus reducing injury-related effects. Second, the use of magnetic microbeads ensures the ability to attract most beads to the iridocorneal angle, effectively reducing the number of beads floating in the anterior chamber avoiding contact with other structures (e.g., iris, lens). Lastly, the use of a handheld magnet allows flexibility when handling the small mouse eye to efficiently direct the magnetic microbeads and ensure that there is little reflux of the microbeads from the eye when the microneedle is withdrawn. In summary, the microbead occlusion mouse model presented here is a powerful investigative tool to study neurodegenerative changes that occur during the onset and progression of glaucoma.

Here, we present a protocol to induce ocular hypertension in the murine eye that results in the loss of retinal ganglion cells as observed in glaucoma. Magnetic microbeads are injected into the anterior chamber and attracted to the iridocorneal angle using a magnet to block the outflow of aqueous humour.

A磁条微珠缺血模型，以诱导小鼠高眼压，青光眼相关

The use of rodent models of glaucoma has been essential to understand the molecular mechanisms that underlie the pathophysiology of this multifactorial ...

Balloon-based Injury to Induce Myointimal Hyperplasia in the Mouse Abdominal Aorta

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine balloon denudation model, which is comparable with established vessel injury models in large animals. The aorta denudation model with balloon catheters mimics the clinical setting and leads to comparable pathobiological and physiological changes. Briefly, after performing a horizontal incision in the aorta abdominalis, a balloon catheter will be inserted into the vessel, inflated, and introduced retrogradely. Inflation of the balloon will lead to intima injury and overdistension of the vessel. After removing the catheter, the aortic incision will be closed with single stiches. The model shown in this article is reproducible, easy to perform, and can be established quickly and reliably. It is especially suitable for evaluating expensive experimental therapeutic agents, which can be applied in an economical fashion. By using different knockout-mouse strains, the impact of different genes on MH development can be assessed.

This article demonstrates a murine model to study the development of myointimal hyperplasia (MH) after aortic balloon injury.

球囊损伤诱发小鼠腹主动脉 Myointimal 增生的研究

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine ...

A Closed-chest Model to Induce Transverse Aortic Constriction in Mice

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to perform a partial thoracotomy to visualize the transverse aortic arch. However, the surgical trauma caused by the thoracotomy in open-chest models changes the respiratory physiology as the ribs are dissected and left unattached after chest closure. To prevent this, we established a minimally invasive, closed chest approach via lateral thoracotomy. Herein we approach the aortic arch via the 2nd intercostal space without entering the chest cavities, leaving the mouse with a less traumatic injury to recover from. We perform this operation using standard laboratory settings for open chest TAC procedures with equal survival rates. Apart from maintaining physiological breathing patterns due to the closed chest approach, the mice seem to benefit by showing rapid recovery, as the less invasive technique appears to facilitate a fast healing process and to reduce immune response after trauma.

Here, we present a protocol of Transverse Aortic constriction (TAC) via a lateral thoracotomy. This technique is a minimally invasive, closed chest surgical procedure aiming to simulate pressure overload and heart failure in mice utilizing standard TAC laboratory settings.

诱导小鼠横向主动脉狭窄的闭合胸模型

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to ...

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 ...