Real-Time Assessment of Spinal Cord Microperfusion in a Porcine Model of Ischemia/Reperfusion

Summary

Spinal cord microcirculation plays a pivotal role in spinal cord injury. Most methods do not allow real-time assessment of spinal cord microcirculation, which is essential for the development of microcirculation-targeted therapies. Here, we propose a protocol using Laser-Doppler-Flow Needle probes in a large animal model of ischemia/reperfusion.

Abstract

Spinal cord injury is a devastating complication of aortic repair. Despite developments for the prevention and treatment of spinal cord injury, its incidence is still considerably high and therefore, influences patient outcome. Microcirculation plays a key role in tissue perfusion and oxygen supply and is often dissociated from macrohemodynamics. Thus, direct evaluation of spinal cord microcirculation is essential for the development of microcirculation-targeted therapies and the evaluation of existing approaches in regard to spinal cord microcirculation. However, most of the methods do not provide real-time assessment of spinal cord microcirculation. The aim of this study is to describe a standardized protocol for real-time spinal cord microcirculatory evaluation using laser-Doppler needle probes directly inserted in the spinal cord. We used a porcine model of ischemia/reperfusion to induce deterioration of the spinal cord microcirculation. In addition, a fluorescent microsphere injection technique was used. Initially, animals were anesthetized and mechanically ventilated. Thereafter, laser-Doppler needle probe insertion was performed, followed by the placement of cerebrospinal fluid drainage. A median sternotomy was performed for exposure of the descending aorta to perform aortic cross-clamping. Ischemia/reperfusion was induced by supra-celiac aortic cross-clamping for a total of 48 min, followed by reperfusion and hemodynamic stabilization. Laser-Doppler Flux was performed in parallel with macrohemodynamic evaluation. In addition, automated cerebrospinal fluid drainage was used to maintain a stable cerebrospinal pressure. After completion of the protocol, animals were sacrificed, and the spinal cord was harvested for histopathological and microsphere analysis. The protocol reveals the feasibility of spinal cord microperfusion measurements using laser-Doppler probes and shows a marked decrease during ischemia as well as recovery after reperfusion. Results showed comparable behavior to fluorescent microsphere evaluation. In conclusion, this new protocol might provide a useful large animal model for future studies using real-time spinal cord microperfusion assessment in ischemia/reperfusion conditions.

Introduction

Spinal cord injury induced by ischemia/reperfusion (SCI) is one of the most devastating complications of aortic repair associated with reduced outcome^1,2,3,4. Current prevention and treatment options for SCI include the optimization of macrohemodynamic parameters as well as the normalization of cerebrospinal fluid pressure (CSP) to improve spinal cord perfusion pressure^2,5,6,7,8

Protocol

The study was approved by the Governmental Commission on the Care and Use of Animals of the City of Hamburg (Reference-No. 60/17). The animals received care in compliance with the 'Guide for the Care and Use of Laboratory Animals' (NIH publication No. 86-23, revised 2011) as well as FELASA recommendations and experiments were carried out according to the ARRIVE guidelines^24,25. This study was an acute trial, and all animals were euthanized at the end of p.......

Discussion

SCI induced by spinal cord ischemia is a major complication of aortic repair with tremendous impact on patient outcome^{1,2,3,4,10,11,12}. Microcirculation-targeted therapies to prevent and treat SCI are most promising. The protocol provides a reproducible method for real-time spinal cord micro.......

Materials

Name	Company	Catalog Number	Comments
CardioMed Flowmeter	Medistim AS, Oslo, Norway	CM4000	Flowmeter for Flow-Probe Femoral Artery
CardioMed Flow-Probe, 5mm	Medistim AS, Oslo, Norway	PS100051	Flow-Probe Femoral Artery
COnfidence probe,	Transonic Systems Inc., Ithaca, NY, USA	MA16PAU	Flow-Probe Aorta
16 mm liners
DIVA Sevoflurane Vapor	Dräger Medical, Lübeck, Germany		Vapor
Hotline Level 1 Fluid Warmer	Smiths Medical Germany GmbH, Grasbrunn, Germany	HL-90-DE-230	Fluid Warmer
Infinity Delta	Dräger Medical, Lübeck, Germany		Basic Monitoring Hardware
Infinity Hemo	Dräger Medical, Lübeck, Germany		Basic Pressure Monitoring and Pulmonary Thermodilution Hardware
LabChart Pro	ADInstruments Ltd., Oxford, UK	v8.1.16	Synchronic Laser-Doppler, Blood Pressure, ECG and Blood-Flow Aquisition Software
LiquoGuard 7	Möller Medical GmbH, Fulda, Germany		Cerebrospinal Fluid Drainage System
Millar Micro-Tip Pressure Catheter (5F, Single, Curved, 120cm, PU/WD)	ADInstruments Ltd., Oxford, UK	SPR-350	Pressure-Tip Catheter Aorta
moor VMS LDF	moor Instruments, Devon, UK		Designated Laser-Doppler Hardware
moor VMS Research Software	moor Instruments, Devon, UK		Designated Laser-Doppler Software
Perivascular Flow Module	Transonic Systems Inc., Ithaca, NY, USA	TS 420	Flow-Module for Flow-Probe Aorta
PiCCO 2, Science Version	Getinge AB, Göteborg, Sweden	v. 6.0	Blood Pressure and Transcardiopulmonary Monitoring Hard- and Software
PiCCO 5 Fr. 20cm	Getinge AB, Göteborg, Sweden		Thermistor-tipped Arterial Line
PowerLab	ADInstruments Ltd., Oxford, UK	PL 3516	Synchronic Laser-Doppler, Blood Pressure, ECG and Blood-Flow Aquisition Hardware
QuadBridgeAmp	ADInstruments Ltd., Oxford, UK	FE 224	Four Channel Bridge Amplifier for Laser-Doppler and Invasive Blood Pressure Aquisition
Silverline	Spiegelberg, Hamburg, Germany	ELD33.010.02	Cerebrospinal Fluid Drainage
SPSS statistical software package	IBM SPSS Statistics Inc., Armonk, New York, USA	v. 27	Statistical Software
Twinwarm Warming System	Moeck & Moeck GmbH, Hamburg, Germany	12TW921DE	Warming System
Universal II Warming Blanket	Moeck & Moeck GmbH, Hamburg, Germany	906	Warming Blanket
VP 3 Probe, 8mm length (individually manufactured)	moor Instruments, Devon, UK		Laser-Doppler Probe
Zeus	Dräger Medical, Lübeck, Germany		Anesthesia Machine