Largo Plazo La cateterización de la linfa intestinal Tronco y colección de linfa en cerdos neonatales
Summary
We present a surgical procedure to catheterize the intestinal lymph trunk in neonatal pigs to collect large quantities of lipid metabolism components from efferent lymph.
Abstract
Catheterization of the intestinal lymph trunk in neonatal pigs is a technique allowing for the long-term collection of large quantities of intestinal (central) efferent lymph. Importantly, the collection of central lymph from the intestine enables researchers to study both the mechanisms and lipid constitutes associated with lipid metabolism, intestinal inflammation and cancer metastasis, as well as cells involved in immune function and immunosurveillance. A ventral mid-line surgical approach permits excellent surgical exposure to the cranial abdomen and relatively easy access to the intestinal lymph trunk vessel that lies near the pancreas and the right ventral segment of the portal vein underneath the visceral aspect of the right liver lobe. The vessel is meticulously dissected and released from the surrounding fascia and then dilated with sutures allowing for insertion and subsequent securing of the catheter into the vessel. The catheter is exteriorized and approximately 1 L/24 hr of lymph is collected over a 7 day period. While this technique enables the collection of large quantities of central lymph over an extended period of time, the success depends on careful surgical dissection, tissue handling and close attention to proper surgical technique. This is particularly important with surgeries in young animals as the lymph vessels can easily tear, potentially leading to surgical and experimental failure. The video demonstrates an excellent surgical technique for the collection of intestinal lymph.
Introduction
El sistema linfático es un área poco estudiada de la fisiología. Los modelos preclínicos de cateterismo linfático se producen en diferentes especies animales 1-8 y son utilizados por las industrias farmacéuticas e instituciones de investigación para investigar los mecanismos implicados en lípidos y metabolismo de los fármacos 8-12 13-15, 16 metástasis del cáncer con el tratamiento experimental de 17, 18 y la función inmune -26. Este estudio explora el uso de cateterismo intestinal linfático tronco en un modelo de cerdo doméstico para medir los componentes del metabolismo de las lipoproteínas. metabolismo de las lipoproteínas está implicada en la producción y secreción de quilomicrones, así como cambios en los lípidos asociados y proteína total. Estas consideraciones son importantes, ya que hay grandes diferencias en el metabolismo de los lípidos entre los modelos de roedores y humanos utilizados habitualmente y, como tal, empleando modelos porcinos para recoger la linfa intestinal podría proporcionar información comparable para el estudio de lípidos mímetabo- en las personas 27-31.
Varias técnicas quirúrgicas se utilizan para recoger la linfa intestinal en grandes especies animales: un enfoque hombro craneal (es decir, torácica cateterización del conducto) 5, una aproximación lateral flanco superior 32-34, y una línea media ventral o enfoque paramedian 22,35. Este video describe en detalle el procedimiento quirúrgico en los cerdos utilizando un enfoque quirúrgico línea media ventral para la cateterización del tronco linfático intestinal. Cuidadosas permisos técnica quirúrgica este método de cateterismo linfático para recoger grandes cantidades de la linfa y de sus componentes durante períodos prolongados de tiempo.
Esta técnica abre una gran variedad de aplicaciones en muchas disciplinas que examinan diversas funciones fisiológicas. Las aplicaciones podrían incluir, pero no se limitan a, las lipoproteínas de todo el cuerpo y el metabolismo de los lípidos, la inmunovigilancia, génesis tumoral y la metástasis, la función intestinal, y la DESARROLLOent y la progresión de la enfermedad inflamatoria intestinal.
Protocol
Representative Results
Discussion
Recogida linfático intestinal es un excelente método para investigar los mecanismos que participan en los lípidos 8-12 y 13-15 de drogas metabolismo, metástasis de cáncer de 16,17, el tráfico de células y la función inmune 18-26, en varios modelos animales experimentales. De hecho, la capacidad para cosechar grandes cantidades de cualquiera periférica (aferente) y central (eferente y los grandes vasos del tronco) linfático durante un período prolongado ha sido espec…
Disclosures
The authors have nothing to disclose.
Acknowledgements
The work was supported in part by funding from Alberta Livestock and Meat Agency and Natural Science and Research Council Discovery grant to S. D. Proctor.
Materials
| Miller laryngoscope blade | Welch Allyn | 68044 | 182 mm length |
| Surgivet advisor: Vital signs monitor | Surgivet | V9203 | |
| Rectal temperature probe | Surgivet | V3417 | |
| Mono-polar electrosurgery generator | Valley Lab | ||
| Metzenbaum scissors | Fine Science | 14518-18 | |
| Tuffier retractor | Stevens | 162-11-676 | |
| Mosquito forceps | Stevens | 162-7-10 | |
| Kelly forceps-curved (14cm) | Stevens | 162-7-38 | |
| Allis tissue forceps | Stevens | 162-7-38 | |
| Forceps dressing-eye (10.2cm) | Stevens | 162-18-780 | |
| Forceps dressing-Adison (12.1cm) | Stevens | 162-17-2510 | |
| Needle Drivers | Stevens | 162-V98-42 | |
| Iris scissors | Fine science | 14058-11 | |
| Circulating water pump | Jorvet | J-783X | |
| Maxitherm-Vinyl blanket | Jorvet | J-784C | |
| Q tip applicators | Fisher Scientific | 22-037-960 | |
| Catheterization tubing (4.06 OD X 2.31 ID) | Braintree Scientific Inc. | MRE-160 | Micro-Renethane implantation tubing |
| 2-0 silk suture | Ethicon | LA556 | |
| 2-0 polyglactin suture | Ethicon | J443H | 2-0 vicryl |
| Large animal jacket | Lomir Biomedical Inc. | SSJ2YC | |
| Polypropylene wash bottles | Fisher Scientific | 03-409-22C | 500 ml |
| Penicillin-Streptomycin | Sigma Aldrich | D4333 | |
| EDTA | Sigma Aldrich | 60-00-4 | |
| Amphotericin B | Sigma Aldrich | A2411 | |
| Azaperone | Elanco Animal Health | Stresnil | |
| Dexmedetomidine hydrochloride | Zoetis | 6295 | Dexdomitor |
| Isoflurane | Abbott Animal Health | 05260-5 | IsoFlo |
| Ketamine hydrochloride | Zoetis | 2626 | Ketaset |
| Bupenorphine hydrochloride | Champion Alstoe Animal Health | DIN:02347510 | |
| 6 mm Endotracheal tube | Jorvet | J-165d | |
| 10% Lidocaine spray | AstraZeneca | DIN:02003767 | |
| 4 % Chlorhexidine surgical scrub | Partnar Animal Health | PCH-011 | Diluted: 2.0% solution |
| 3M Surgical steri- drape | 3M Health Care | 1040 | |
| SDS page gel | Invitrogen | EA0375BOX | 3-8 % tris acetate |
| Polyvinylidene fluoride membrane | Millipore | IPVH00010 | 0.45 μm pore size |
| ApoB antibody | EMD Millipore | AB742 | 1:4000 dilution |
| Donkey anti-goat IgG-HRP | Santa Cruz Biotechnology | Sc-2304 | |
| ECL Prime Western Blotting Reagent | GE Healthcare LifeSciences | RPN2232 | |
| Triglyceride Kit | Wako Pure Chemicals | 998-40391/994-40491 | |
| Total Cholesterol Kit | Wako Pure Chemicals | 439-17501 | |
| Total Protein | Pierce | 23225 | Bicinchoninic Acid Assay |
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