技术领域technical field
本发明涉及神经损伤技术领域，尤其涉及一种大型动物的肺迷走神经损伤的方法。The invention relates to the technical field of nerve injury, in particular to a method for injuring the lung vagus nerve of a large animal.
背景技术Background technique
对于慢性阻塞性肺疾病(COPD)、哮喘等肺部疾病，或者与肺有关的疾病，如：肺气肿，慢性支气管炎等。目前主要的治疗方式是药物治疗，或外科手术治疗(如使用外科手术切除肺神经，使神经的功能全部或部分无效，可以改善哮喘或肺气肿)。慢性阻塞性肺病(COPD)和哮喘的特征在于持续或复发的气流阻塞，这可能与这肺神经传出纤维的过度刺激有关。有研究表明，损伤肺部神经，可以抑制乙酰胆碱释放，抑制平滑肌收缩，进而改善哮喘、COPD症状，改善肺的功能。肺迷走神经损伤方法的研究，有助于研究肺部多种相关疾病的发病机制，有助于相关肺功能的研究，有助于神经修复相关研究。For lung diseases such as chronic obstructive pulmonary disease (COPD), asthma, or diseases related to the lungs, such as: emphysema, chronic bronchitis, etc. At present, the main treatment method is drug therapy, or surgical treatment (such as using surgical operation to remove the pulmonary nerve, so that the function of the nerve is completely or partially invalid, and asthma or emphysema can be improved). Chronic obstructive pulmonary disease (COPD) and asthma are characterized by persistent or recurrent airflow obstruction, which may be related to overstimulation of efferent fibers of this pulmonary nerve. Studies have shown that damage to the lung nerves can inhibit the release of acetylcholine and smooth muscle contraction, thereby improving the symptoms of asthma and COPD, and improving lung function. The research on the method of pulmonary vagus nerve injury is helpful to the study of the pathogenesis of various related diseases of the lung, the study of related lung function, and the study of nerve repair.
针对肺迷走神经损伤的方法，现有的方法主要有：1.外科手术方式；2.局部热烧灼方式。外科手术方式：根据解剖位置，利用外科切开，找准并切除/损伤神经组织。该方法需要逐步分离解剖，找准并分辨神经组织，操作难度大，要求操作者具有专业的经验及背景，耗费时间长。局部热烧灼方式：手术切开组织或气管镜镜下贴合气道内壁进行，使用高温热烧灼方式，使神经组织失活。高温对气道组织损失较大，容易应发气道狭窄、穿孔等并发症，甚至影响动物生存，安全风险高。The existing methods for pulmonary vagus nerve injury mainly include: 1. surgical operation; 2. local thermal ablation. Surgical operation method: According to the anatomical location, surgical incision is used to locate and excise/damage the nerve tissue. This method needs to be separated and dissected step by step, and the nerve tissue is identified and distinguished. The operation is difficult, requires the operator to have professional experience and background, and takes a long time. Local thermal cautery method: Surgical incision of the tissue or fitting the inner wall of the airway under a bronchoscope, using high-temperature thermal cauterization to inactivate the nerve tissue. High temperature will cause great damage to airway tissue, and it is easy to cause complications such as airway stenosis and perforation, and even affect the survival of animals, with high safety risks.
冷冻消融，是利用对局部组织的冷冻，可控地破坏组织的治疗方法。冷冻消融在气道使用时，与热消融相比，冷冻不易导致软骨的损伤，很少发生气道软化、塌陷，防止继发动力性狭窄；冷冻治疗较少引起冰冻部位与周围组织的黏连；可控深度的冷冻治疗对邻近的大血管和气管不易造成严重伤害，不易造成穿孔出血，且利于组织修复；冷冻消融不促进肉芽组织增生，不容易产生瘢痕组织。冷冻消融的特点及优势，使得其更适合用于气道组织，更适合用于肺神经消融、损伤，因此冷冻消融是用于肺神经损伤建模是一种合适的方法。Cryoablation is a treatment that destroys tissue in a controlled manner by freezing local tissue. When cryoablation is used in the airway, compared with thermal ablation, freezing is not easy to cause cartilage damage, and airway softening and collapse rarely occur, preventing secondary dynamic stenosis; cryotherapy is less likely to cause adhesion between the frozen part and surrounding tissues ; The cryotherapy with controllable depth is not easy to cause serious damage to the adjacent large blood vessels and trachea, it is not easy to cause perforation and bleeding, and it is conducive to tissue repair; cryoablation does not promote the proliferation of granulation tissue, and is not easy to produce scar tissue. The characteristics and advantages of cryoablation make it more suitable for airway tissue and pulmonary nerve ablation and injury. Therefore, cryoablation is a suitable method for modeling pulmonary nerve injury.
但经过文献和相关的专利检索，在现有技术中，并未发现任何采用冷冻消融进行肺迷走神经损伤的方法。However, after searching the literature and related patents, in the prior art, no method of using cryoablation for pulmonary vagus nerve injury has been found.
发明内容Contents of the invention
为了克服现有技术中所存在的缺点与不足，本发明提供一种大型动物的肺迷走神经损伤的方法，其为采用冷冻消融方法构建一种创伤小、操作简单、重复性好的肺迷走神经损伤的新方法。In order to overcome the shortcomings and deficiencies in the prior art, the present invention provides a method for lung vagus nerve injury in large animals, which uses cryoablation to construct a lung vagus nerve injury method with small trauma, simple operation, and good repeatability. new method.
为实现上述目的，本发明采用如下技术方案：To achieve the above object, the present invention adopts the following technical solutions:
本发明的第一个目的是提供一种大型动物的肺迷走神经损伤的方法，其包括如下步骤：在离体的肺组织上，放入内窥镜，在内窥镜监视下，放入球囊冷冻消融导管，推送球囊冷冻消融导管到达指定部位，扩张球囊，确认冷冻消融位置，开启冷冻消融程序，对指定位置进行冷冻消融；其中，所述冷冻消融位置为分布有肺迷走神经的气管和/或支气管外壁。The first object of the present invention is to provide a method for pulmonary vagus nerve injury in large animals, which includes the following steps: putting an endoscope on the isolated lung tissue, and putting a balloon under the monitoring of the endoscope The cryoablation catheter pushes the balloon cryoablation catheter to the designated site, expands the balloon, confirms the cryoablation position, starts the cryoablation program, and performs cryoablation on the designated position; wherein, the cryoablation position is the trachea and / or the outer wall of the bronchi.
为了进一步优化上述大型动物的肺迷走神经损伤的方法，本发明采取的技术措施还包括：In order to further optimize the method for the injury of the pulmonary vagus nerve of the above-mentioned large animals, the technical measures taken by the present invention also include:
进一步地，所述冷冻消融位置为气管和/或支气管整个管壁的圆周方向。Further, the cryoablation position is in the circumferential direction of the entire tube wall of the trachea and/or bronchi.
进一步地，所述大型动物的肺迷走神经损伤的方法还包括步骤：冷冻消融程序完成后，开启复温程序，待复温完成后，将球囊轻轻从冷冻部位抽出。Further, the method for pulmonary vagus nerve injury in large animals further includes the step of: after the cryoablation procedure is completed, the rewarming procedure is started, and after the rewarming is completed, the balloon is gently withdrawn from the frozen part.
进一步地，所述球囊扩张后有效直径为6-18mm，球囊冷冻时表面温度低于-100℃，冷冻消融时间为10-120秒。Further, the effective diameter of the balloon after expansion is 6-18mm, the surface temperature of the balloon is lower than -100°C when frozen, and the cryoablation time is 10-120 seconds.
进一步地，所述球囊冷冻消融导管的参数为：球囊直径为15mm，球囊冷冻时表面温度为-140℃，冷冻消融时间为60秒。Further, the parameters of the balloon cryoablation catheter are as follows: the diameter of the balloon is 15 mm, the surface temperature of the balloon is -140°C when frozen, and the cryoablation time is 60 seconds.
进一步地，所述内窥镜为支气管镜。Further, the endoscope is a bronchoscope.
进一步地，所述指定部位为左主支气管和右主支气管。更进一步地，所述大型动物的肺迷走神经损伤的方法还包括步骤：当左主支气管和右主支气管中的一个完成消融损伤后，开启复温程序，待复温完成后，将球囊轻轻从冷冻部位抽出；以同样的步骤的进行左主支气管和右主支气管中的另一个的冷冻消融步骤，待完成消融损伤后，开启复温程序，待复温完成后，将球囊轻轻从冷冻部位抽出。Further, the designated sites are the left main bronchus and the right main bronchus. Furthermore, the method for the injury of the pulmonary vagus nerve in large animals further includes the step of: when one of the left main bronchus and the right main bronchus has completed the ablation injury, start the rewarming program, and after the rewarming is completed, the balloon is gently Pull out from the frozen part; carry out the cryoablation step of the left main bronchus and the right main bronchus in the same steps, after the ablation injury is completed, start the rewarming procedure, and after the rewarming is completed, gently remove the balloon from the The frozen part is drawn out.
进一步地，所述球囊冷冻消融导管的制冷原理为液氮制冷原理。Further, the refrigeration principle of the balloon cryoablation catheter is the principle of liquid nitrogen refrigeration.
进一步地，所述确认冷冻消融位置在内窥镜直视下或在X光机下显示。Further, the confirmed cryoablation position is displayed under direct vision of an endoscope or under an X-ray machine.
进一步地，所述离体的肺组织来源于羊、猪、牛等大型动物；更进一步地，所述离体的肺组织来源于羊。Further, the isolated lung tissue is derived from large animals such as sheep, pigs, and cattle; further, the isolated lung tissue is derived from sheep.
本发明的第二个目的是提供一种由任一上述方法制备的肺迷走神经损伤模型。The second object of the present invention is to provide a model of pulmonary vagus nerve injury prepared by any of the above methods.
本领域技术人员可理解的是上述肺迷走神经损伤的方法也同样直接适用于实验动物，实验动物需进行麻醉后进行相应的损伤步骤；若指定部位为左、右主支气管，其可在同一次操作中均进行冷冻消融，也可以间隔一段时间分开进行冷冻消融，上述间隔时间为2～4周。Those skilled in the art can understand that the above method of lung vagus nerve injury is also directly applicable to experimental animals, and the experimental animals need to be anesthetized and then undergo corresponding injury steps; if the designated parts are the left and right main bronchi, they can be operated at the same time The cryoablation can be performed in both cases, and the cryoablation can also be performed separately at intervals, and the above-mentioned interval is 2 to 4 weeks.
本发明采用上述技术方案，具有如下技术效果：The present invention adopts above-mentioned technical scheme, has following technical effect:
本发明采用冷冻消融原理，利用动物气道，以非常小的创伤完成对神经组织的损伤，构建了一种大型动物离体肺组织的肺迷走神经损伤模型。本发明所采用的冷冻消融方法，利用动物自然通道，构建一种创伤小、安全性高、操作简单、重复性好的大型动物的肺迷走神经损伤的方法。The invention adopts the principle of cryoablation, utilizes the airway of animals, completes the damage to the nerve tissue with very small trauma, and constructs a model of pulmonary vagus nerve injury of the isolated lung tissue of large animals. The cryoablation method adopted in the present invention utilizes the natural channel of animals to construct a method for lung vagus nerve injury in large animals with less trauma, high safety, simple operation and good repeatability.
附图说明Description of drawings
图1为本发明一实施例中在支气管镜下，羊右主支气管放置球囊冷冻消融导管图片；Fig. 1 is a picture of a balloon cryoablation catheter placed in the right main bronchus of a sheep under a bronchoscope in an embodiment of the present invention;
图2为本发明一实施例中羊右主支气管的冷冻消融图片；Fig. 2 is a cryoablation picture of the right main bronchus of a sheep in an embodiment of the present invention;
图3A～图3B为本发明一实施例中羊一侧主支气管不同时间冷冻消融免疫组化病理结果图；其中，图3A为40s即刻免疫组化图，图3B为60S即刻免疫组化图，图3C为90S即刻免疫组化图，图3D为120S即刻免疫组化图；Figures 3A to 3B are the immunohistochemical pathological results of cryoablation of the main bronchus of sheep at different times in one embodiment of the present invention; among them, Figure 3A is the immediate immunohistochemical picture at 40s, and Figure 3B is the immediate immunohistochemical picture at 60s, Figure 3C is the immediate immunohistochemical image of 90S, and Figure 3D is the immediate immunohistochemical image of 120S;
图4A～图4B为本发明一实施例中冷冻消融处理后羊肺部的解剖图；其中，图4A为肺气管/支气管解剖分离图，图4B为分离气管/支气管剖开图。4A-4B are anatomical diagrams of sheep lungs after cryoablation in an embodiment of the present invention; among them, FIG. 4A is an anatomical separation diagram of lung trachea/bronchus, and FIG. 4B is a dissection diagram of separated trachea/bronchus.
具体实施方式Detailed ways
本发明涉及一种大型动物的肺迷走神经损伤的方法及其构建的肺迷走神经损伤模型，该方法包括如下步骤：在离体的肺组织上，放入内窥镜，在内窥镜监视下，放入球囊冷冻消融导管，推送球囊冷冻消融导管到达指定部位，扩张球囊，确认冷冻消融位置，开启冷冻消融程序，对指定位置进行冷冻消融；其中，所述冷冻消融位置为分布有肺迷走神经的气管和/或支气管外壁。The present invention relates to a method of lung vagus nerve injury in large animals and a model of lung vagus nerve injury constructed thereof. The method comprises the following steps: placing an endoscope on the isolated lung tissue, and placing Insert the balloon cryoablation catheter, push the balloon cryoablation catheter to the designated site, expand the balloon, confirm the cryoablation position, start the cryoablation program, and perform cryoablation on the designated position; outer wall of the trachea and/or bronchi.
下面结合附图和实施例，对本发明的具体实施方式作进一步描述。以下实施例仅用于更加清楚地说明本发明的技术方案，而不能以此来限制本发明的保护范围。The specific implementation manners of the present invention will be further described below in conjunction with the drawings and examples. The following examples are only used to illustrate the technical solution of the present invention more clearly, but not to limit the protection scope of the present invention.
下述实施例中所涉及的球囊有效直径、球囊有效长度、冷冻消融时间为多次实验获得的基本参数。下述实施例中实验所选择的离体肺组织为30-40Kg的羊的肺组织。可理解的是，当选取其他大型动物，例如猪或牛的离体肺组织时，上述参数需进行适应性的调整。The effective diameter of the balloon, the effective length of the balloon, and the cryoablation time involved in the following examples are basic parameters obtained from multiple experiments. The isolated lung tissue selected for the experiments in the following examples is 30-40Kg sheep lung tissue. It is understandable that when the isolated lung tissue of other large animals, such as pigs or cattle, is selected, the above parameters need to be adjusted adaptively.
实施例1Example 1
本实施例为一较佳的一种肺迷走神经损伤的方法，包括如下步骤：The present embodiment is a preferred method for pulmonary vagus nerve injury, comprising the following steps:
选择新鲜的离体肺组织(羊)，在离体的肺组织上，插入支气管镜与球囊冷冻消融导管，在支气管镜监视下，将球囊冷冻消融导管推送至羊一侧主支气管，到达位置后，扩张球囊，确认位置无误后，开启冷冻消融程序，计时，到达预定时间时，冷冻程序停止；其中球囊扩张后有效直径为15mm，球囊冷冻时表面温度为-140℃，冷冻消融时间为60秒。图1所示为，支气管镜下，羊右主支气管放置球囊冷冻消融导管图片。图2所示为，羊右主支气管冷冻消融图片。Select fresh isolated lung tissue (sheep), insert a bronchoscope and balloon cryoablation catheter on the isolated lung tissue, and push the balloon cryoablation catheter to the main bronchus of the sheep side under the monitoring of the bronchoscope, and reach After the location, expand the balloon. After confirming that the location is correct, start the cryoablation program, count the time, and stop the freezing program when the scheduled time is reached; the effective diameter of the balloon after expansion is 15mm, and the surface temperature of the balloon is -140°C when frozen The ablation time was 60 seconds. Figure 1 shows, under the bronchoscopic view, the balloon cryoablation catheter placed in the right main bronchus of sheep. Figure 2 shows the images of cryoablation of the right main bronchi of sheep.
冷冻消融程序完成后，开启复温程序，待复温完成后，将球囊轻轻从冷冻部位抽出。注意，须轻柔移动导管，防止冷冻消融部位发生损伤。当一侧主支气管完成消融损伤后，进行另一侧主支气管的消融，冷冻消融方法同上一次相同。After the cryoablation procedure is completed, start the rewarming procedure. After the rewarming is completed, gently withdraw the balloon from the frozen site. Note that the catheter must be moved gently to prevent damage to the cryoablation site. After the ablation injury of one main bronchus was completed, the other main bronchus was ablated, and the cryoablation method was the same as the previous one.
冷冻消融以其特有的优势，在气道应用广泛。本实施例是通过球囊与气道壁接触，利用球囊内部的超低温，通过接触部分将冷量传递至目标组织，利用低温可控的将目标组织灭活。液氮原理的冷冻消融使用越来越多，液氮深低温(-196℃)，易获得，成本低，使用安全，这些特点促进液氮使用技术快速发展。如图2所示，冷冻消融过程中，气道壁与球囊接触的部分，受低温的影响，形成较明显的冷冻消融损伤区域。Cryoablation is widely used in the airway due to its unique advantages. In this embodiment, the balloon is in contact with the airway wall, and the ultra-low temperature inside the balloon is used to transmit the cold energy to the target tissue through the contact part, and the target tissue is controlled to be inactivated by the low temperature. The cryoablation based on the principle of liquid nitrogen is being used more and more. The cryogenic temperature of liquid nitrogen (-196°C) is easy to obtain, low in cost, and safe to use. These characteristics promote the rapid development of liquid nitrogen use technology. As shown in Figure 2, during the cryoablation process, the part of the airway wall in contact with the balloon is affected by the low temperature, forming a more obvious cryoablation injury area.
实施例2Example 2
本实施例阐述了实施例1中冷冻消融各工艺参数的筛选过程。This example illustrates the screening process of various process parameters of cryoablation in Example 1.
本实施例中，冷冻消融的时间为关键参数，其直接影响冷冻消融的效果。冷冻消融的时间长时，有效损伤深度大、损伤范围大，对应的，冷冻消融时间短，有效损伤深度和范围小，可能达不到预期的效果。所以，在保证有效损伤神经的情况下，尽量减少冷冻消融时间，即减少非必要的损伤。肺迷走神经损伤过程中，需根据选择的消融部位，选择合理的冷冻时间。本实施例中，选择羊肺组织的左、右主支气管进行消融，损伤该部位迷走神经组织。In this embodiment, the time of cryoablation is a key parameter, which directly affects the effect of cryoablation. When the cryoablation time is long, the effective damage depth is large and the damage area is large. Correspondingly, the cryoablation time is short, the effective damage depth and range are small, and the expected effect may not be achieved. Therefore, in the case of ensuring effective nerve damage, the cryoablation time should be minimized, that is, unnecessary damage should be reduced. In the process of pulmonary vagus nerve injury, it is necessary to choose a reasonable freezing time according to the selected ablation site. In this embodiment, the left and right main bronchi of sheep lung tissue are selected for ablation, and the vagus nerve tissue in this part is damaged.
30-40Kg的羊的左右主支气管的管壁厚度约为3-5mm，根据经验，选取40S、60S、90S、120S，完成冷冻消融程序后，解剖取病理查看结果。如图3A-3D所示，羊一侧主支气管不同时间梯度冷冻消融免疫组化病理结果图，其中，图3A为40s即刻免疫组化图，图3B为60S即刻免疫组化图，图3C为90S即刻免疫组化图，图3D为120S即刻免疫组化图。根据病理结果，40S冷冻时，神经损伤不够彻底，60S、90s、120S冷冻时，均达到有效损伤神经组织的目的。综合考虑冷冻消融对肺部气道的损伤，选取60S作为优选的肺部神经损伤的参数。The wall thickness of the left and right main bronchi of a 30-40Kg sheep is about 3-5mm. Based on experience, select 40S, 60S, 90S, and 120S. As shown in Figures 3A-3D, the immunohistochemical results of the main bronchus on one side of the sheep at different time gradients were frozen and ablated. Immediate immunohistochemistry of 90S, and Figure 3D is immediate immunohistochemistry of 120S. According to the pathological results, when frozen at 40S, the nerve damage was not complete enough, and when frozen at 60S, 90S, and 120S, the purpose of effectively damaging nerve tissue was achieved. Taking comprehensive consideration of the damage to the lung airway caused by cryoablation, 60S was selected as the optimal parameter for lung nerve injury.
由于肺迷走神经分布于气管/支气管外壁，需要对气管/支气管整个管壁圆周方向进行冷冻消融，以达到最佳损伤效果。进而，根据消融选择的部位，须选择合适冷冻球囊尺寸、温度等参数。在本实施例中，由于选择羊的肺组织，左右主支气管，选择球囊直径15mm，冷冻球囊表面温度-140℃。Since the pulmonary vagus nerve is distributed on the outer wall of the trachea/bronchus, it is necessary to perform cryoablation on the entire wall of the trachea/bronchus in order to achieve the best damage effect. Furthermore, according to the site selected for ablation, appropriate parameters such as the size and temperature of the cryoballoon must be selected. In this embodiment, since sheep lung tissue and left and right main bronchi are selected, the diameter of the balloon is 15mm, and the surface temperature of the frozen balloon is -140°C.
实施例3Example 3
本实施例为对实施例1完成冷冻消融的离体肺组织的解剖病理分析。This example is an anatomical and pathological analysis of the isolated lung tissue that underwent cryoablation in Example 1.
待冷冻消融程序完成后，解剖分离肺组织，取出冷冻消融部位，取得病理并做病理切分分析。After the cryoablation procedure was completed, the lung tissue was dissected and separated, and the cryoablation site was taken out for pathological analysis.
为查验冷冻消融损伤神经情况，对完成冷冻消融程序的羊肺组织进行解剖，取冷冻部位组织，用于查看神经损伤的病理结果，如图4A-4B所示，羊肺部解剖图，其中，图4A为肺气管/支气管解剖分离图，图4B为分离气管/支气管剖开图。由上述分析结果可知，已成功完成了对肺迷走神经的损伤。In order to check the nerve damage caused by cryoablation, the sheep lung tissue that has completed the cryoablation procedure was dissected, and the tissue from the frozen part was taken to view the pathological results of the nerve injury, as shown in Figure 4A-4B, the sheep lung anatomy diagram, in which, Figure 4A is an anatomical separation diagram of the pulmonary trachea/bronchi, and Figure 4B is a dissection diagram of the separated trachea/bronchi. From the above analysis results, it can be seen that the injury to the pulmonary vagus nerve has been successfully completed.
由上述实施例可知，本发明采用冷冻消融来损伤肺迷走神经，需根据选择的消融部位，选择合适的球囊大小、冷冻球囊温度、冷冻消融时间，以达到最佳的实验效果，其可建立以一种重复性好，损伤小，操作简单，稳定可靠的肺神经损伤的新方法。It can be seen from the above examples that the present invention uses cryoablation to damage the pulmonary vagus nerve. It is necessary to select the appropriate balloon size, cryoballoon temperature, and cryoablation time according to the selected ablation site to achieve the best experimental effect. It can be established A new method for pulmonary nerve injury with good repeatability, less damage, simple operation, stable and reliable.
以上对本发明的具体实施例进行了详细描述，但其只作为范例，本发明并不限制于以上描述的具体实施例。对于本领域技术人员而言，任何对本发明进行的等同修改和替代也都在本发明的范畴之中。因此，在不脱离本发明的精神和范围下所作的均等变换和修改，都应涵盖在本发明的范围内。The specific embodiments of the present invention have been described in detail above, but they are only examples, and the present invention is not limited to the specific embodiments described above. For those skilled in the art, any equivalent modifications and substitutions to the present invention are also within the scope of the present invention. Therefore, equivalent changes and modifications made without departing from the spirit and scope of the present invention shall fall within the scope of the present invention.
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910713802.XACN110393603A (en) | 2019-08-02 | 2019-08-02 | Method for treating pulmonary vagus nerve injury of large animals |
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910713802.XACN110393603A (en) | 2019-08-02 | 2019-08-02 | Method for treating pulmonary vagus nerve injury of large animals |
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| CN110393603Atrue CN110393603A (en) | 2019-11-01 |
| Application Number | Title | Priority Date | Filing Date |
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| CN201910713802.XAPendingCN110393603A (en) | 2019-08-02 | 2019-08-02 | Method for treating pulmonary vagus nerve injury of large animals |
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