JoVE Journal > Cancer Research
Summary
Abstract
Introduction
Protocol
Results
Discussion
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Acknowledgements
Materials
References
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小鼠模型中上颈神经节切除术的手术技术

Published: December 02, 2022
doi:
10.3791/64527
, , , ,
1Department of Surgery,Memorial Sloan Kettering Cancer Center, 2Department of Otolaryngology-Head and Neck Surgery, Beijing Tongren Hospital,Capital Medical University, 3David M. Rubenstein Center for Pancreatic Cancer Research,Memorial Sloan Kettering Cancer Center

Summary

本协议描述了通过识别和切除颈上神经节来消融肾上腺素能神经支配的小鼠模型。

Abstract

越来越多的证据表明，交感神经系统在癌症进展中起着重要作用。肾上腺素能神经支配调节唾液腺分泌、昼夜节律、黄斑变性、免疫功能和心脏生理学。小鼠外科交感神经切除术是一种研究肾上腺素能神经支配作用的方法，允许完全、单侧肾上腺素能消融，同时避免重复药物干预和相关副作用。然而，由于颈上神经节的尺寸较小，小鼠的手术交感神经切除术在技术上具有挑战性。本研究描述了一种手术技术，用于可靠地识别和切除颈上神经节以消融交感神经系统。通过使用转基因小鼠对荧光交感神经节进行成像、鉴定切除后的霍纳综合征、对切除的神经节中的肾上腺素能标志物进行染色以及观察交感神经切除术后靶器官中肾上腺素能免疫荧光减弱，验证了神经节的成功鉴定和去除。该模型能够对癌症进展以及交感神经系统调节的其他生理过程进行未来研究。

Introduction

多项研究报道，肿瘤微环境中的神经在支持肿瘤进展方面起着积极作用。肾上腺素能交感神经的消融已被证明会损害体内前列腺癌和胃癌的肿瘤发展和播散1,2,3，而肾上腺素能受体的药理学阻断抑制头颈癌的肿瘤生长4。交感神经受累也见于胰腺癌、宫颈癌和基底细胞癌进展 5,6,7。

在交感神经系统中，颈上神经节（SCG）是交感神经干中唯一支配头部的神经节。SCG调节各种生理功能，如唾液分泌和昼夜节律，并直接支配颈部淋巴结8,9,10。SCG 还与病理过程有关，例如黄斑变性11 和主动脉夹层的进展 12。此外，据报道，SCG切除会加重缺血再灌注引起的急性肾损伤13，并改变大鼠的肠道微生物群14。

在小鼠模型中完全消融SCG将代表一种有价值的实验技术，使癌症和自主神经系统研究成为可能。虽然许多研究已利用药物学肾上腺素能受体阻断作为肾上腺素能消融术15,16,17,18,19,20，但手术切除允许完全，单侧肾上腺素能消融，同时避免需要重复的药物干预和相关副作用21,22,23。

SCG的手术切除已在大鼠24中描述，大多数研究颈上神经节切除术（SCGx）效果的报告都采用了大鼠模型。与大鼠模型相比，由于SCG的尺寸小，SCGx在小鼠中的技术更具挑战性。然而，小鼠相对更容易处理，更具成本效益，并且更适合基因操作。Garcia等人是最早在小鼠中报告SCGx的人之一，发现它会影响胰岛素释放25。最近，Ziegler等人根据已发表的大鼠24,26技术描述了小鼠中的SCGx。本文和其他文章描述了一种方法，其中首先识别和解剖颈总动脉（CCA），随后从CCA 21,22,27,28的分叉中取出SCG。在本文中，描述了一种侵入性较小且更安全的小鼠技术，该技术避免了CCA的解剖，从而最大限度地减少了该过程最严重的并发症 – CCA损伤引起的出血。

Protocol

这里描述的动物程序得到了纪念斯隆凯特琳癌症中心的机构动物护理和使用委员会的批准。这里使用了八周龄的雄性和雌性NSG小鼠。这些动物是从商业来源获得的（见 材料表）。器械经过消毒，手术工作表面消毒，动物皮肤表面消毒，外科医生在整个手术过程中戴无菌手套。 1.小鼠的准备和术前设置 在手术前一天，在诱导室（3.75英寸宽x 9英?…

Representative Results

该协议描述了小鼠模型中SCG的手术切除。 图 2 显示了解剖标志，包括 CCA、颈前静脉和 SCG。通过夹层（图2A），可以看到右颈前静脉沿着气管的侧缘流动。由于它位于颈前静脉更深的地方，因此左 CCA 及其分叉到颈内动脉 （ICA） 和颈外动脉 （ECA） 仅在静脉外侧隐约可见。在 NSG 中检查此问题时。B6-P0TdTomato转基因小鼠（一种P0-Cre TdTomato小鼠，其中雪旺?…

Discussion

该协议描述了用于SCG输入的手术单侧消融的小鼠模型。该技术允许研究肾上腺素能神经支配在各种环境中的影响。此外，切除的交感神经节也可以在3D基质胶培养物中生长，用于 体外 实验30。

涉及SCGx的研究大多在大鼠中进行，因为它们较大的解剖结构允许更容易的解剖可视化和解剖。虽然Ziegler等人之前已经描述了小鼠中的SCGx26，并…

Disclosures

The authors have nothing to disclose.

Acknowledgements

Q. W.得到了NIH T32CA009685的支持。R. J. W. 得到了 NIH R01CA219534 的支持。纪念斯隆凯特琳癌症中心的核心设施得到了NIH P30CA008748的支持。

Materials

Anti-Tyrosine Hydroxylase Antibody EMD Millipore AB152
Artificial Tears Lubricant Ophthalmic Ointment Akorn 59399-162-35
Curity 2 x 2 Inch Gauze Sponge 8 Ply, Sterile Covidien 1806
Derf Needle Holder Thomas Scientific 1177K00
Dissecting Microscope
Dumont #5/45 Forceps Fine Science Tools 11251-35
Dumont #7b Forceps Fine Science Tools 11270-20
ETHILON Nylon Suture Ethicon 698H
Fine Scissors – ToughCut Fine Science Tools 14058-09
Hypoallergenic Surgical Tape 3M Blenderm 70200419342
Induction Chamber, 2 Liter VetEquip 941444
Isoflurane Baxter 1001936060
Nair Church & Dwight Co., Inc 40002957 chemical hair removing agent
NORADRENALINE RESEARCH ELISA Labor Diagnostika Nord (Rocky Mountain Diagnostics) BA E-5200
NSG Mouse Jackson Laboratory JAX:005557
Povidone-Iodine Swabstick PDI S41350
Webcol Alcohol Preps Covidien 5110
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Tags

Surgical TechniqueSuperior Cervical GanglionectomyMurine ModelSympathetic Nervous SystemCancer ProgressionSurgical SympathectomyAdrenergic AblationCarotid Artery DissectionAnatomical LandmarkAnesthesiaSkin IncisionSubmandibular Salivary GlandsDigastric MuscleOmohyoid MuscleDeep Cervical FasciaSuperior Cervical Ganglion

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Wang, Q., Chen, C., Xu, H., Deborde, S., Wong, R. J. Surgical Technique for Superior Cervical Ganglionectomy in a Murine Model. J. Vis. Exp. (190), e64527, doi:10.3791/64527 (2022).
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