Name: establishment of deep hypothermic circulatory arrest in rats
Uploaded: 2022-12-16T12:30:00
Description: Watch this Scientific Journal Video about Establishment of Deep Hypothermic Circulatory Arrest in Rats at JoVE.com

作者感谢张亮在实验过程中帮助收集视频数据。本研究得到了国家自然科学基金（批准号：82070479）和中央高校基本科研业务费（批准号：3332022128）的支持。

该方案经过机构审查，并获得了中国医学科学院阜外医院机构动物护理和使用委员会的批准（FW-2021-0005）。所有实验程序均按照美国国立卫生研究院出版的实验动物护理和使用指南进行。
注意：雄性Sprague-Dawley大鼠（重量：500-600克，年龄：12-14周）保持在标准实验室条件下，自由获取食物和水。将大鼠随机分为两组（n = 6，每组）：DHCA组和常温CPB组（NtCPB组）。
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插管是在大鼠中建立DHCA的最基本程序。插管前，用 0.5 mL 的 2% 利多卡因浸泡动脉将使其更容易插管。插管后，需要通过颈外静脉 用 500IU / kg肝素进行肝素化，以避免微血栓形成17。我们反复发现，这种剂量的肝素可以达到激活凝血时间（ACT）>480秒的目标。复温期是最困难的部分。在我们的实验中，温度从18°C上升到34°C需要60多分钟，而在其他一些实验中，复温期可以在30?...

自1953年以来，深低温循环骤停（DHCA）已用于心脏手术1。DHCA涉及将患者的核心温度降低到严重低温水平（15-22°C），然后全局中断流向身体的血液2。循环停滞可以提供相对不流血的操作场。深度低温会降低新陈代谢，特别是在大脑和心肌中，这是防止缺血的有效方法3。DHCA通常用于复杂先天性心脏病，主动脉弓疾病，甚至带有腔静脉血栓的肾脏或肾上腺肿瘤4，5的手术中。因此，建立DHCA动物模型为临床环境中程序的细化和并发症的预防提供了重要参考。
虽然可以用犬6、兔7和其他动物建立模型，但由于它们的可操作性和低成本，最好使用大鼠。DHCA大鼠模型于2006年由Jungwirth等人首次描述8。结果发现，循环停止的持续时间对神经系统结局有影响。从那时起，DHCA大鼠模型得到了广泛的研究。已经澄清，DHCA可能引发全身炎症反应综合征（SIRS）9。在随后的研究中，药理学家发现，由SIRS诱导的DHCA相关神经炎症可以通过白藜芦醇10和雷公藤内酯11减弱。我们的团队还发现，DHCA相关的神经炎症可以通过抑制冷诱导的RNA结合蛋白12来减弱。在心血管系统中，超氧化物歧化酶对DHCA13期间的缺血/再灌注（I / R）损伤具有心脏保护作用。这些结果扩大了对DHCA相关病理生理过程的理解，并为改善DHCA的结局提供了新的方向。然而，关于DHCA后内毒素血症，氧化应激和自噬的结果尚无定论。DHCA使用与体外循环（CPB）相同的操作技术14，但其管理策略不同，并且不同团队产生DHCA的步骤不同8，9，10，11。本研究旨在提供一种在大鼠中建立DHCA程序的方法。

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	<tbody>
		<tr>
			<td>Heat Exchanger</td>
			<td>Xi&#8217;an Xijing Medical Appliance Co., Ltd</td>
			<td>Animal-M</td>
			<td></td>
		</tr>
		<tr>
			<td>Membrane Oxygenator</td>
			<td>Dongguan Kewei Medical Instrument Co., Ltd.</td>
			<td>Micro-M</td>
			<td></td>
		</tr>
		<tr>
			<td>Monitor</td>
			<td>Chengdu Techman Co., Ltd</td>
			<td>BL-420s</td>
			<td></td>
		</tr>
		<tr>
			<td>Roller Pump</td>
			<td>Changzhou Prefluid Technology Co.,Ltd</td>
			<td>BL100</td>
			<td></td>
		</tr>
		<tr>
			<td>SD Rat</td>
			<td>HFK Bioscience Co.,Ltd.</td>
			<td>/</td>
			<td></td>
		</tr>
		<tr>
			<td>Sevoflurane</td>
			<td>Maruishi Pharmaceutical Co. Ltd</td>
			<td>H20150020</td>
			<td></td>
		</tr>
		<tr>
			<td>Shaver</td>
			<td>Hangzhou Huayuan Pet Products Co.,Ltd.</td>
			<td>/</td>
			<td></td>
		</tr>
		<tr>
			<td>Vaporizer</td>
			<td>SPACECABS</td>
			<td>/</td>
			<td></td>
		</tr>
		<tr>
			<td>Ventilator</td>
			<td>Shanghai Alcott Biotech Co., Ltd</td>
			<td>ALC-V8S</td>
			<td></td>
		</tr>
		<tr>
			<td>Water Tank</td>
			<td>Maquet Critical Care AB</td>
			<td>Jostra HCU20-600</td>
			<td></td>
		</tr>
	</tbody>
</table>

establishment of deep hypothermic circulatory arrest in rats

深度低温循环骤停（DHCA）常规应用于复杂先天性心脏病和主动脉弓疾病的手术。本研究旨在提供一种在大鼠中建立DHCA的方法。为了评估DHCA过程对生命体征的影响，使用没有循环停止的常温体外循环（CPB）大鼠模型作为对照。正如预期的那样，DHCA导致体温和平均动脉血压显着降低。血气分析表明，DHCA增加了乳酸水平，但不影响血液pH值和血红蛋白、血细胞比容、Na+、Cl−、K+和葡萄糖的浓度。此外，与常温CPB大鼠相比，透射电子显微镜结果显示DHCA大鼠海马自噬体轻度增加。

作为对照组，未发生循环骤停的常温CPB（NtCPB）大鼠在整个手术过程中平均动脉血压（MAP）和体温稳定，而DHCA大鼠心脏骤停期间的MAP下降（p <0.01， 图3A）。DHCA大鼠的温度在冷却阶段迅速下降，并在复温阶段逐渐恢复。当将大鼠从DHCA回路断奶时，DHCA大鼠的温度恢复正常（图3B）。
通过血气分析研究了DHCA过程对大鼠的影响。?...

Watch this Scientific Journal Video about Establishment of Deep Hypothermic Circulatory Arrest in Rats at JoVE.com

Establishment of Deep Hypothermic Circulatory Arrest in Rats

该协议提出了大鼠深低温循环停搏的建立，可用于研究全身炎症反应综合征，缺血/再灌注损伤，氧化应激，神经炎症等。

在大鼠中建立深低温循环停滞

Deep hypothermic circulatory arrest (DHCA) is routinely applied during surgeries for complex congenital heart disease and aortic arch disease. The present ...

在大鼠中建立深度低温循环停滞。DHCA深度低温循环停滞，常规应用于复杂先天性心脏病和主动脉弓疾病的手术。本研究旨在提供一种在大鼠中建立DHCA的方法。该方案接受了机构审查，并获得了中国医学科学院阜外医院机构动物护理和使用委员会的批准。准备工作。确保CPB回路包含由变形滴管，辊泵，热交换器，膜氧合器，连接管和连接回路的水箱改编而成的储液器，并用1毫升肝素钠和1毫升盐水使12毫升羟基淀粉与40毫升灌注溶液一起，辊泵轻轻旋转。二、麻醉和插管。将管子与呼吸机和侧面参数连接。放一个电加热毯和右边，用胶带面向右边。将眼药膏涂抹在眼睛上以防止干燥。用剃须刀剃除右腹股沟区域、右颈椎区域和尾巴上的头发。然后用醛和酒精对皮肤进行消毒。切开皮肤和左腹股沟区域，解剖肌肉和组织软骨，露出左股静脉和动脉。小心地分开动脉。插管 22 规静脉导管进入左股动脉。结扎动脉和丝绸导管。使用含有肝素的生理盐水冲洗套管以避免凝血。将导管与按压传感器连接以监测血压。切开尾部的皮肤，然后用手术刀切开尾动脉的浅表部分，露出位于手术视野中间的尾动脉。用 22 号静脉导管插管尾动脉。用丝绸结扎动脉和导管。使用含有肝素的生理盐水冲洗导管以避免凝血。切开右颈静脉的皮肤，然后分离肌肉和组织以暴露静脉。将 16 号自制多龄静脉导管插入右颈外静脉，并小心地将其放入右下腔静脉或右心房。给予肝素钠，其中右外静脉用湿球覆盖每个空心区域，以避免污染。三、将回路与尾动脉导管连接，保持泵每分钟一到两毫升的低速率。然后将储液器与右颈外静脉中的导管连接。确保储液罐中的血液水平始终约为一厘米。打开水箱，先将水温设置为37摄氏度。血压稳定后，轻轻增加流量泵至每分钟80至100毫升。四、降温。将室温设置为 20 摄氏度左右。将冰块放入这种一次性手套中，然后将它们放在右侧头部和侧面。根据大米的直肠温度实时建议水箱的温度。根据血气分析结果适当改变呼吸机的相关参数。五、深部低温循环停滞。然后直肠温度下降到15至20摄氏度。更换冰袋并丢弃手套，以确保在循环停滞期间维持深度体温过低。停止滚筒泵。保持储液器与环境接触，并将血液从颈外静脉缓慢排出到储液器。注意血压改革。没有血压和心跳后，停止引流并保持储液器关闭。关闭呼吸机。六、预热再灌注。摘下所有一次性手套，将室温升高至25摄氏度。恢复记忆氧间通气，同时保持静脉引流管夹紧。打开滚筒泵，确保储液罐中的血液慢慢回到正确的身体。在储液罐中的血位保持一厘米后打开呼吸机。使用引流管将血液从右心房缓慢引流到储液器。打开加热灯、加热垫和水箱。将水箱温度设置为25摄氏度。然后及时将其调整到左侧温度。直肠温度恢复到34摄氏度后取下加热灯。七、断奶CPB。缓慢而逐渐地降低泵的滚筒流量，并调整静脉属的速度，使流量降低到每分钟一毫升。保持储液罐与环境接触。将剩余的血液注入回路中，流速为每分钟一毫升。停止记忆氧合和滚筒泵。经过一段时间的机械通气和深度麻醉后安乐死。具有代表性的结果。作为对照组，常温CPB、NtCPB、大鼠。无循环骤停显示稳定的平均动脉血压MAP，整个过程中体温正常。虽然DHCA大鼠的MAP在心脏骤停期间下降。DHCA大鼠的温度在冷却阶段快速下降，在旋转阶段逐渐恢复。DHCA断奶电路使DHCA大鼠体温恢复正常。以上数据表明，我们的DHCA程序模仿了诊所患者的实际温度和MAP变化。通过血气分析研究了DHCA处理器和上升的影响。全血与初级溶液接触后，两组血红蛋白浓度均高于每分升6克。从DHCA表面断奶时，由于CPB回路中的剩余血液重新注入右侧，因此每分升增加至9克。血细胞比容HCT显示出与HB相似的倾向。DHCA再灌注后乳酸快速增加，这在DHCA组中更为明显。DHCA手术后pH值下降，这很可能是乳酸积累的结果。在整个实验过程中，钠、氯化物、钾离子和葡萄糖的浓度在每10个点上没有表现出显着差异。结果表明，DHCA仅诱导乳酸升高，而不影响血pH值和血红蛋白、血细胞比容、钠离子、氯离子、钾离子和葡萄糖的浓度。为了评估DHCA对神经元自噬的影响，我们使用透射电显微镜，令人惊讶地发现并增加了DHCA大鼠海马体中的自噬体数量。以上数据表明，我们的DHCA模型模仿了诊所患者的实际温度和MAP变化。该模型可用于研究全身炎症反应综合征、神经炎症等。

establishment-of-deep-hypothermic-circulatory-arrest-in-rats

Research

JoVE Journal

Immunology and Infection

Orthotopic Hind-Limb Transplantation in Rats

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel therapeutic entity in reconstructive surgery. However, long term high-dose multi-drug immunosuppressive therapy is required to ensure graft survival, bearing the risk of serious side effects which halters broader application. Further progression in this field may depend on better understanding of basic immunology and ischemia reperfusion injury in composite tissue grafts.
To date, orthotopic hind limb transplantation in rats has been the preferred rodent model for reconstructive transplantation (RT), however, it is an extremely demanding procedure that requires extraordinary microsurgical skills for reattachment of vasculature, bones, muscles and nerves.
We have introduced the vascular cuff anastomosis technique to this model, providing a rapid and reliable approach to rat hind limb transplantation. This technique simplifies and shortens the surgical procedure and enables surgeons with basic microsurgical experience to successfully perform the operation with high survival and low complication rates. The technique seems to be well suited for immunological as well as ischemia reperfusion injury (IRI) studies.

Here we describe the orthotopic rat hind-limb transplantation procedure, which seems to be the gold standard in vivo model for composite tissue allotransplantation research.

原位大鼠后肢移植

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel ...

科研

免疫与感染

Diagnosis of Ecto- and Endoparasites in Laboratory Rats and Mice

Internal and external parasites remain a significant concern in laboratory rodent facilities, and many research facilities harbor some parasitized animals. Before embarking on an examination of animals for parasites, two things should be considered. One: what use will be made of the information collected, and two: which test is the most appropriate. Knowing that animals are parasitized may be something that the facility accepts, but there is often a need to treat animals and then to determine the efficacy of treatment. Parasites may be detected in animals through various techniques, including samples taken from live or euthanized animals. Historically, the tests with the greatest diagnostic sensitivity required euthanasia of the animal, although PCR has allowed high-sensitivity testing for several types of parasite. This article demonstrates procedures for the detection of endo- and ectoparasites in mice and rats. The same procedures are applicable to other rodents, although the species of parasites found will differ.

This article describes various procedures for screening rats and mice to detect endo- or ectoparasitism. Several diagnostic assays will be demonstrated, both those suitable for use on live animals and those used after euthanasia of the animal. Photographs to aid in identification of rat and mouse parasites will be included.

Ecto和实验室大鼠和小鼠体内寄生虫的诊断

Internal and external parasites remain a significant concern in laboratory rodent facilities, and many research facilities harbor some parasitized ...

Establishment of Epstein-Barr Virus Growth-transformed Lymphoblastoid Cell Lines

Infection of B cells with Epstein-Barr virus (EBV) leads to proliferation and subsequent immortalization, resulting in establishment of lymphoblastoid cell lines (LCL) in vitro. Since LCL are latently infected with EBV, they provide a model system to investigate EBV latency and virus-driven B cell proliferation and tumorigenesis1. LCL have been used to present antigens in a variety of immunologic assays2, 3. In addition, LCL can be used to generate human monoclonal antibodies4, 5 and provide a potentially unlimited source when access to primary biologic materials is limited6, 7.
A variety of methods have been described to generate LCL. Earlier methods have included the use of mitogens such as phytohemagglutinin, lipopolysaccharide8, and pokeweed mitogen9 to increase the efficiency of EBV-mediated immortalization. More recently, others have used immunosuppressive agents such as cyclosporin A to inhibit T cell-mediated killing of infected B cells7, 10-12.
The considerable length of time from EBV infection to establishment of cell lines drives the requirement for quicker and more reliable methods for EBV-driven B cell growth transformation. Using a combination of high titer EBV and an immunosuppressive agent, we are able to consistently infect, transform, and generate LCL from B cells in peripheral blood. This method uses a small amount of peripheral blood mononuclear cells that are infected in vitroclusters of cells can be demonstrated. The presence of CD23 with EBV in the presence of FK506, a T cell immunosuppressant. Traditionally, outgrowth of proliferating B cells is monitored by visualization of microscopic clusters of cells about a week after infection with EBV. Clumps of LCL can be seen by the naked eye after several weeks. We describe an assay to determine early if EBV-mediated growth transformation is successful even before microscopic clusters of cells can be demonstrated. The presence of CD23hiCD58+ cells observed as early as three days post-infection indicates a successful outcome.

We describe a method for generating transformed B cell lines using Epstein-Barr virus. We also illustrate a novel assay that can identify B cells destined to undergo transformation as early as three days after infection.

Epstein - Barr病毒生长转化的淋巴细胞系的建立

Infection of B cells with Epstein-Barr virus (EBV) leads to proliferation and subsequent immortalization, resulting in establishment of lymphoblastoid ...

Establishment of an In vitro System to Study Intracellular Behavior of Candida glabrata in Human THP-1 Macrophages

A cell culture model system, if a close mimic of host environmental conditions, can serve as an inexpensive, reproducible and easily manipulatable alternative to animal model systems for the study of a specific step of microbial pathogen infection. A human monocytic cell line THP-1 which, upon phorbol ester treatment, is differentiated into macrophages, has previously been used to study virulence strategies of many intracellular pathogens including Mycobacterium tuberculosis. Here, we discuss a protocol to enact an in vitro cell culture model system using THP-1 macrophages to delineate the interaction of an opportunistic human yeast pathogen Candida glabrata with host phagocytic cells. This model system is simple, fast, amenable to high-throughput mutant screens, and requires no sophisticated equipment. A typical THP-1 macrophage infection experiment takes approximately 24 hr with an additional 24-48 hr to allow recovered intracellular yeast to grow on rich medium for colony forming unit-based viability analysis. Like other in vitro model systems, a possible limitation of this approach is difficulty in extrapolating the results obtained to a highly complex immune cell circuitry existing in the human host. However, despite this, the current protocol is very useful to elucidate the strategies that a fungal pathogen may employ to evade/counteract antimicrobial response and survive, adapt, and proliferate in the nutrient-poor environment of host immune cells.

Establishment of an In vitro System to Study Intracellular Behavior of Candida glabrata in Human THP-1 Macrophages

The current article outlines a protocol to establish an in vitro cell culture model system to study the interaction of a facultative intracellular human fungal pathogen Candida glabrata with human macrophages which will be a useful tool to advance our knowledge of fungal virulence mechanisms.

建立一个<em&gt;在体外</em&gt;系统来研究细胞内的行为<em&gt;光滑念珠菌</em&gt;在人THP-1巨噬细胞

A cell culture model system, if a close mimic of host environmental conditions, can serve as an inexpensive, reproducible and easily manipulatable ...

Complete Thymectomy in Adult Rats with Non-invasive Endotracheal Intubation

Thymectomy in neonatal rodents is an established and reliable procedure for immunological studies. However, in adult rats, complications of hemorrhage and pneumothorax from pleural disruption can result in a significant mortality rate. This protocol is a simple method of rat thymectomy that utilizes a mini-sternotomy and endotracheal intubation. Intubation is accomplished with a non-invasive and easily reproducible method and allows for positive pressure ventilation to prevent pneumothorax and a controlled airway that allows sufficient time for careful thymus dissection to minimize pleural disruption. A 1.5 cm sternal incision decreases contact with mediastinal vessels and pleura, while still providing full visualization of the thymus. Following exposure of the mediastinum, the thymus is removed by blunt dissection under magnification. The pleural space is then sealed by suture closure of the pre-tracheal muscles followed by the application of surgical glue. The thorax is then closed by suture closure of the sternum, followed by suture closure of the skin. All thymectomies were complete as evidenced by immunohistochemical (IHC) staining of mediastinal tissue, and absence of na&#239;ve T-cells by flow cytometry, and the procedure had a 96% survival rate. This method is suitable when complete thymectomy with minimal complications is desired for further immunological studies in athymic adult rats.

Rodent thymectomy is a valuable technique in immunological research. Here, a protocol for complete thymectomy in adult rats using a mini-sternotomy along with non-invasive intubation and positive pressure ventilation to minimize perioperative morbidity and mortality is described.

完整的胸腺切除术的成年大鼠非侵入气管插管

Thymectomy in neonatal rodents is an established and reliable procedure for immunological studies. However, in adult rats, complications of hemorrhage and ...

Prevention of Heat Stress Adverse Effects in Rats by Bacillus subtilis Strain

This study was designed to evaluate the protective effect of the Bacillus subtilis strain against complications related to heat stress. Thirty-two Sprague-Dawley rats were used in this study. Animals were orally treated twice a day for two days with B. subtilis BSB3 strain or PBS. The next day after the last treatment, each group was divided and two experimental groups (one treated with PBS and one treated with B. subtilis) were placed at 45 oC for 25 min. Two control groups stayed for 25 min at room temperature. All rats were euthanized and different parameters were analyzed in all groups. Adverse effects of heat stress are registered by the decrease of villi height and total mucosal thickness in the intestinal epithelium; translocation of bacteria from the lumen; increased vesiculation of erythrocytes and elevation of the lipopolysaccharides (LPS) level in the blood. The protective efficacy of treatment is evaluated by prevention of these side effects. The protocol was set up for the oral treatment of rats with bacteria for prevention of heat stress complications, but this protocol can be modified and used for other routes of administration and for analysis of different compounds.

Prevention of Heat Stress Adverse Effects in Rats by Bacillus subtilis Strain

We described a protocol for prevention of heat stress effects in rats by oral pre-treatment with beneficial bacteria. This protocol can be modified and used for various routes of administration and for analysis of different compounds.

热火预防应激在大鼠不利影响<em&gt;枯草芽孢杆菌</em&gt;应变

This study was designed to evaluate the protective effect of the Bacillus subtilis strain against complications related to heat stress. Thirty-two ...

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 ...

Deep Vein Thrombosis Induced by Stasis in Mice Monitored by High Frequency Ultrasonography

Venous thrombosis is a common condition affecting 1 - 2% of the population, with an annual incidence of 1 in 500. Venous thrombosis can lead to death through pulmonary embolism or results in the post-thrombotic syndrome, characterized by chronic leg pain, swelling, and ulceration, or in chronic pulmonary hypertension resulting in significant chronic respiratory compromise. This is the most common cardiovascular disease after myocardial infarction and ischemic stroke and is a clinical challenge for all medical disciplines, as it can complicate the course of other disorders such as cancer, systemic disease, surgery, and major trauma.
Experimental models are necessary to study these mechanisms. The stasis model induces consistent thrombus size and a quantifiable amount of thrombus. However, it is necessary to systematically ligate side branches of the inferior vena cava to avoid variability in thrombus sizes and any erroneous data interpretation. We have developed a non-invasive technique to measure thrombus size using ultrasonography. Using this technique, we can assess thrombus development and resolution over time in the same animal. This approach limits the number of mice required for quantification of venous thrombosis consistent with the principle of replacement, reduction, and refinement of animals in research. We have demonstrated that thrombus weight and histological analysis of thrombus size correlate with measurement obtained with ultrasonography. Therefore, the current study describes how to induce deep vein thrombosis in mice using the inferior vena cava stasis model and how to monitor it using high frequency ultrasound.

The present protocol describes the steps to obtain venous thrombosis using a stasis model. In addition, we are using a non-invasive method to measure thrombus formation and resolution over time.

高频超声监测小鼠瘀血致深静脉血栓形成

Venous thrombosis is a common condition affecting 1 - 2% of the population, with an annual incidence of 1 in 500. Venous thrombosis can lead to death ...

Deep Dermal Injection As a Model of Candida albicans Skin Infection for Histological Analyses

The skin is an extremely extended organ of the body and, due to this large surface, it is continuously exposed to microorganisms. Skin damage can easily lead to infections in the dermis which can, in turn, result in the dissemination of pathogens into the bloodstream. Understanding how the immune system fights infections at the very early stage and how the host can eliminate the pathogens is an important step to set the base for future therapeutic interventions. Here we describe a model of Candida albicans infection that can visualize the processes that occur early during an infection, including when the pathogen has passed the epithelial barrier, as well as the immune response elicited by the C. albicans invasion. We used this infection model to perform histological analyses that show the immune cells that infiltrate the skin as well as the presence and localization of the pathogen. Samples collected after the infection can be processed for RNA extraction.

Deep Dermal Injection As a Model of Candida albicans Skin Infection for Histological Analyses

Here we describe a protocol that allows histological and molecular analysis of skin samples after Candida albicans intradermal injection. This protocol maintains the structural integrity of the skin and allows for the localization of tissue-resident or newly recruited immune cells as well as the pathogen distribution.

深部真皮注射液作为白色念珠菌皮肤感染模型的组织学分析

The skin is an extremely extended organ of the body and, due to this large surface, it is continuously exposed to microorganisms. Skin damage can easily ...

Establishment of Viral Infection and Analysis of Host-Virus Interaction in Drosophila Melanogaster

Virus spreading is a major cause of epidemic diseases. Thus, understanding the interaction between the virus and the host is very important to extend our knowledge of prevention and treatment of viral infection. The fruit fly Drosophila melanogaster has proven to be one of the most efficient and productive model organisms to screen for antiviral factors and investigate virus-host interaction, due to powerful genetic tools and highly conserved innate immune signaling pathways. The procedure described here demonstrates a nano-injection method to establish viral infection and induce systemic antiviral responses in adult flies. The precise control of the viral injection dose in this method enables high experimental reproducibility. Protocols described in this study include the preparation of flies and the virus, the injection method, survival rate analysis, the virus load measurement, and an antiviral pathway assessment. The influence effects of viral infection by the flies' background were mentioned here. This infection method is easy to perform and quantitatively repeatable; it can be applied to screen for host/viral factors involved in virus-host interaction and to dissect the crosstalk between innate immune signaling and other biological pathways in response to viral infection.

Establishment of Viral Infection and Analysis of Host-Virus Interaction in Drosophila Melanogaster

This protocol describes how to establish viral infection in vivo in Drosophila melanogaster using the nano-injection method and basic techniques to analyze virus-host interaction.

果蝇病毒感染的建立及宿主病毒相互作用分析

Virus spreading is a major cause of epidemic diseases. Thus, understanding the interaction between the virus and the host is very important to extend our ...