​Harmonization timeline was generated by using Biorender.

In preclinical research, the same protocol can be carried out in multiple different ways, introducing bias and confounding variables. In order to overcome these common issues, SPRINT launched the Harmonization Trial.

The SPRINT Harmonization Trial aimed to utilize the unique expertise of each laboratory by integrating them into single, unified standard operating procedures (SOPs). To start off, representatives from each site were sent to learn specialized MCA stroke induction and behavior testing. These included multiple workshops, training videos, site visits, and specialized instructions organized by the SPRINT coordinating center. From then, each lab ran a pilot experiment with both sham and MCA occluded mice, and the data was then collected and analyzed centrally. The necessary adjustments were then made, turning SPRINT into the polished, collaborative network it is today. 

Below are a few short descriptions of some of the factors that were adjusted during the harmonization process, and links to our unified SOPs.  

 

Stroke induction 

:Before the trial began, one surgeon from each lab was sent for a two-day training workshop by the Plesnila group in Munich, to learn the PURE MCAo model. Surgeons then returned to their respective laboratories and practiced the technique, until successfully achieving stroke volumes with a 15% standard deviation of one another. Upon completion, surgeons received a certificate of competency. 

Link to SOP:

 

Behavior tests and equipment 

The SPRINT Network included tests such as the Irregular Ladder Rung, Barnes Maze, Adhesive Tape Removal, Cylinder Test, and the personalized SPAN score to measure behavior outcomes. All testing equipment was built centrally by the Charité workshop to ensure uniformity. Several training videos were made for conduction of the tests, lighting conditions, as well as setup. Finally, visits were made by the Coordinating Center to each laboratory for in-person demonstrations of the performance and recording of the tests. 

Link to SOP:

 

Magnetic Resonance Imaging (MRI)

To confirm surgery success and access lesion volumes, a specific imaging protocol was development by the EPIC3R Neuroimaging group at the Charité and the Aswendt Laboratory in Frankfurt. MRI Phantoms were sent to each laboratory and accessed using the SPRINT MRI protocol. Each MRI scanner then had its settings adjusted to achieve the highest accuracy and clearest images. 

Link to SOP:

 

Data collection 

Each site received both paper and digital data collection forms (using the REDCap software) that were filled out at each step of the trial. This included information on animal handling, behavior tests and scoring, in addition to any comments along the way. All behavior videos and MRI scans were uploaded centrally to the Charité Virtual Research Environment and analyzed blinded using automated kinematic analysis algorithms and manual scoring.  

Link to SOP:

 

Biological Sample Collection

Samples were collected at several timepoints from mice during the harmonization trial and at the end of the experiment. Tissue, stool, and blood were collected and analyzed centrally following pre-specified extraction and analysis protocols. 

Link to SOP:

 

Statistical Analysis, Legal Criteria, and Systematic Review

Power calculations and variability between sites was centrally calculated by the DECIDE, BIH group and Statistics Core at the Charite. This helped to ensure the greatest possible efficacy in fine-tuning the collaboration between laboratory sites. All legal amendments and applications were advised under the Responsible PrecliniX group in order to ensure the best possible translation and clinical relevance with the 3R principles in mind. Finally, systematic reviews and meta analyses were carried out under the McCann group at the BIH, Charité. This aided in establishing robust, high-quality measures that were implemented into the Harmonization Trial. 

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