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Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Genetics

Exercise Test for Evaluation of the Functional Efficacy of the Pig Cardiovascular System

Published: May 12th, 2023

DOI:

10.3791/65233

1A.I. Virtanen Institute, University of Eastern Finland, 2Heart Center and Gene Therapy Unit, Kuopio University Hospital
* These authors contributed equally

The present protocol describes a large animal exercise test model to assess the functional capacity of the cardiovascular system for evaluating the efficiency of new therapies in the preclinical setting. It is comparable to a clinical exercise test.

Despite the progress in treatments, cardiovascular diseases are still one of the biggest causes of mortality and morbidity worldwide. Gene therapy-based therapeutic angiogenesis is a promising approach for treating patients with significant symptoms, despite optimal pharmacological therapy and invasive procedures. However, many promising cardiovascular gene therapy techniques have failed to accomplish expectations in clinical trials. One explanation is a mismatch between preclinical and clinical endpoints used to measure efficacy. In animal models, the emphasis has usually been on easily quantifiable endpoints, such as the number and area of the capillary vessels calculated from histological sections. Apart from mortality and morbidity, endpoints in clinical trials are subjective, such as exercise tolerance and quality of life. However, the preclinical and clinical endpoints likely measure different aspects of the applied therapy. Nevertheless, both types of endpoints are required to develop successful therapeutic approaches. In clinics, the main goal is always to alleviate patients' symptoms and improve their prognosis and quality of life. To achieve better predictive data from preclinical studies, endpoint measurements must be better matched to those in clinical studies. Here, we introduce a protocol for a clinically relevant treadmill exercise test in pigs. This study aims to: (1) provide a reliable exercise test in pigs that can be used to evaluate the safety and functional efficacy of gene therapy and other novel therapies, and (2) better match the endpoints between preclinical and clinical studies.

Chronic cardiovascular diseases are significant causes of mortality and morbidity worldwide1,2. Although current treatments are effective for the majority of patients, many still cannot benefit from the current therapies due to, for example, diffuse chronic disease or comorbidities. In addition, in some patients, cardiac symptoms are not relieved by the available treatments, and their cardiovascular disease progresses despite optimal medical therapy3. Thus, there is a clear need to develop novel treatment options for severe cardiovascular diseases.

During the....

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All experiments are approved by the Animal Experiment Board of the University of Eastern Finland. This protocol describes a clinically relevant treadmill exercise test for pigs to evaluate the safety and efficacy of novel therapies for heart diseases. Female domestic pigs weighing 25-80 kg were used for the present study. The animals were obtained from a commercial source (see Table of Materials).

1. Setting up the running track

  1. Set up the running .......

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One must have experience working with large animals to succeed with this protocol. Researchers need to be able to evaluate whether an animal stops running due to fatigue or lack of motivation. Recording the speed and distance may help to evaluate this, as usually, animals lacking motivation stop running totally, whereas fatigued animals keep running after slowing down the speed (Figure 3). If necessary, the protocol can be repeated the next day if the results seem unreliable.

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This large animal exercise test mimics the test used in clinics, reducing the gap in endpoints between the preclinical studies and clinical trials. It can be applied to evaluate the efficacy of new treatments for severe cardiovascular diseases, such as arteriosclerosis obliterans, heart failure, and ischemic heart diseases. The time points applied in this protocol may vary depending on the tested treatment. This protocol has been standardized based on a long experience of working with large animals and can be used to eva.......

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The author would like to thank Minna Törrönen, Riikka Venäläinen, Heikki Karhunen, and Inkeri Niemi from the National Laboratory Animal Center for their assistance in animal work. This study is supported by Finnish Academy, ERC, and CardioReGenix EU Horizon grant.

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Name Company Catalog Number Comments
Defibrillator Zoll M series TO9K116790 All portable defribrillators will work
Defibrillator pads Philips M3713A All pads work, as long as the pads are compatible with the defibrillator
ECG electrodes Several providers Prefer ECG electrodes designed for exercise tests
Loop recorder Abbott Oy DM3500 Optional for rhythm monitoring
Patient monitor Schiller Argus LCM Plus 7,80,05,935 All portable ecg monitors will work
Pigs Emolandia Oy
Treadmill NordicTrack All treadmills with adjustable incline and speed are suitable for the exercise test.  The treadmill should be as long and wide as possible.
Ultrasound system Philips EPIQ 7 ultrasound
Various building materials Several providers For building fences, ramps and gates according to the Figure 1 and Figure 2
Various treats for the animals

  1. Virani, S., et al. Heart disease and stroke statistics-2020 update: A report from the American Heart Association. Circulation. 141 (9), e139 (2020).
  2. Townsend, N., et al. Epidemiology of cardiovascular disease in Europe. Nature Reviews Cardiology. 19 (2), 133-143 (2022).
  3. Knuuti, J., et al. 2019 ESC Guidelines for the diagnosis and management of chronic coronary syndromes: The Task Force for the diagnosis and management of chronic coronary syndromes of the European Society of Cardiology (ESC). European Heart Journal. 41 (3), 407-477 (2020).
  4. Ylä-Herttuala, S., Baker, A. H. Cardiovascular gene therapy: past, present, and future. Molecular Therapy. 25 (5), 1096-1106 (2017).
  5. Hedman, M., et al. Eight-year safety follow-up of coronary artery disease patients after local intracoronary VEGF gene transfer. Gene Therapy. 16 (5), 629-634 (2009).
  6. Rosengart, T. K., et al. Long-term follow-up of a phase 1 trial of angiogenic gene therapy using direct intramyocardial administration of an adenoviral vector expression the VEGF121 cDNA for the treatment of diffuse coronary artery disease. Human Gene Therapy. 24 (2), 203-208 (2013).
  7. Muona, K., Mäkinen, K., Hedman, M., Manninen, H., Ylä-Herttuala, S. 10-year safety follow-up in patients with local VEGF gene transfer to ischemic lower limb. Gene Therapy. 19 (4), 392-395 (2012).
  8. Leikas, A. J., et al. Long-term safety and efficacy of intramyocardial adenovirus-mediated VEGF-DΔNΔC gene therapy eight-year follow-up of phase I KAT301 study. Gene Therapy. 29 (5), 289-293 (2022).
  9. Telukuntla, K. S., Suncion, V. Y., Schulman, U. H., Hare, J. M. The advancing field of cell-based therapy: insights and lessons from clinical trials. Journal of the American Heart Association. 2 (5), e000338 (2013).
  10. Ylä-Herttuala, S., Bridges, C., Katz, M. G., Korpisalo, P. Angiogenic gene therapy in cardiovascular diseases: dream or vision. European Heart Journal. 38 (18), 1365-1371 (2017).
  11. Lähteenvuo, J., Ylä-Herttuala, S. Advances and challenges in cardiovascular gene therapy. Human Gene Therapy. 28 (11), 1024-1032 (2017).
  12. Ross, R., et al. Importance of assessing cardiorespiratory fitness in clinical practice: a case for fitness as a clinical vital sign: a scientific statement from the American Heart Association. Circulation. 134 (24), e653-e699 (2016).
  13. Sietsema, K. E., Stringer, W. W., Sue, D. Y., Ward, S. . Wasserman & Whipp's Principles of Exercise Testing and Interpretation. 6th. , (2021).
  14. Darmadi, M. A., et al. Exercise-induced sustained ventricular tachycardia without structural heart disease: a case report. The American Journal of Case Reports. 21, e928242 (2020).
  15. Casella, G., Pavesi, P. C., Sangiorgio, P., Rubboli, A., Bracchetti, D. Exercise-induced ventricular arrhythmias in patients with healed myocardial infarction. International Journal of Cardiology. 40 (3), 229-235 (1993).
  16. Gimeno, J. R., et al. Exercise-induced ventricular arrhythmias and risk of sudden cardiac death in patients with hypertrophic cardiomyopathy. European Heart Journal. 30 (21), 2599-2605 (2009).
  17. Lelovas, P. P., Kostomitsopoulos, N. G., Xanthos, T. T. A comparative anatomic and physiologic overview of the porcine heart. Journal of the American Association for Laboratory Animal Science. 53 (5), 432-438 (2014).
  18. Korpela, H., et al. AAV2-VEGF-B gene therapy failed to induce angiogenesis in ischemic porcine myocardium due to inflammatory responses. Gene Therapy. 29 (10-11), 643-652 (2022).
  19. Swindle, M. M. . Swine in the Laboratory: Surgery, Anesthesia, Imaging, and Experimental Techniques. 2nd edition. , (2007).
  20. Poole, D. C., et al. Guidelines for animal exercise and training protocols for cardiovascular studies. American Journal of Physiology. Heart and Circulatory Physiology. 318 (5), H1100-H1138 (2020).

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