Name: real-time assessment of spinal cord microperfusion in a porcine model of ischemia/reperfusion
Uploaded: 2020-12-10T14:00:00
Description: Watch this Scientific Journal Video about Real-Time Assessment of Spinal Cord Microperfusion in a Porcine Model of Ischemia/Reperfusion at JoVE.com

The authors would like to thank Lena Brix, V.M.D, Institute of Animal Research, Hannover Medical School, as well as Mrs. Jutta Dammann, Facility of Research Animal Care, University Medical Center Hamburg-Eppendorf, Germany, for providing pre- and perioperative animal care and their technical assistance on animal handling. The authors would further like to thank Dr. Daniel Manzoni, Department of Vascular Surgery, H&#244;pital Kirchberg, Luxembourg, for his technical assistance.

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 &#39;Guide for the Care and Use of Laboratory Animals&#39; (NIH publication No. 86-23, revised 2011) as well as FELASA recommendations and experiments were carried out according to the ARRIVE guidelines24,25. This study was an acute trial, and all animals were euthanized at the end of p...

SCI induced by spinal cord ischemia is a major complication of aortic repair with tremendous impact on patient outcome1,2,3,4,10,11,12. Microcirculation-targeted therapies to prevent and treat SCI are most promising. The protocol provides a reproducible method for real-time spinal cord micro...

Spinal cord injury induced by ischemia/reperfusion (SCI) is one of the most devastating complications of aortic repair associated with reduced outcome1,2,3,4. 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 pressure2,5,6,7,8,9. Despite the implementation of these maneuvers, incidence of SCI still ranges between 2% and 31% depending on the complexity of aortic repair10,11,12.
Recently, microcirculation has gained increased attention13,14. Microcirculation is the area of cellular oxygen uptake and metabolic exchange and therefore, plays a critical role in organ function and cellular integrity13. Impaired microcirculatory blood flow is a major determinant of tissue ischemia associated with increased mortality15,16,17,18,19. Impairment of spinal cord microcirculation is associated with reduced neurological function and outcome20,21,22,23. Therefore, optimization of microperfusion for the treatment of SCI is a most promising approach. Persistence of microcirculatory disturbances, despite macrocirculatory optimization, has been described26,27,28,29. This loss of hemodynamic coherence occurs frequently in various conditions including ischemia/reperfusion, emphasizing the need for direct microcirculatory evaluation and microcirculation-targeted therapies26,27,30.
So far, only few studies have used laser-Doppler probes for real-time assessment of spinal cord microcirculatory behavior20,31. Existing studies have often used microsphere injection techniques, which are limited by intermittent use and post-mortem analysis32,33. The number of different measurements using microsphere injection technique is limited by the availability of microspheres with different wavelengths. Moreover, in contrast to Laser-Doppler techniques, real-time assessment of microperfusion is not possible, as post-mortem tissue processing and analysis is needed for this method. Here, we present an experimental protocol for the real-time assessment of spinal cord microcirculation in a porcine large animal model of ischemia/reperfusion.
This study was part of a large animal project combining a randomized study comparing the influence of crystalloids vs. colloids on microcirculation in ischemia/reperfusion as well as an explorative randomized study on the effects of fluids vs. vasopressors on spinal cord microperfusion. Flow probe 2-point calibration as well as pressure-tip catheter calibration has been previously described34. In addition to the reported protocol, fluorescent microspheres were used for the measurement of spinal cord microperfusion, as previously described, using 12 samples of spinal cord tissue for each animal, with samples 1-6 representing the upper spinal cord and 7-12 representing the lower spinal cord35,36. Microsphere injection was performed for each measurement step after the completion of Laser-Doppler recordings and macrohemodynamic evaluation. Histopathological evaluation was performed using the Kleinman-Score as previously described37.

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real-time assessment of spinal cord microperfusion in a porcine model of ischemia/reperfusion

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.

All six animals survived until the completion of the protocol. Animal weight was 48.2 &#177; 2.9 kg; five animals were male, and one animal was female. Spinal cord needle probe insertion as well as spinal cord Flux measurement was feasible in all animals.
Examples of real-time spinal cord microcirculatory recordings in combination with cerebral microcirculatory and macrohemodynamic recordings during aortic cross-clamping for isc...

Watch this Scientific Journal Video about Real-Time Assessment of Spinal Cord Microperfusion in a Porcine Model of Ischemia/Reperfusion at JoVE.com

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

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.

Spinal cord injury is a devastating complication of aortic repair. Despite developments for the prevention and treatment of spinal cord injury, its ...

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