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Development of a Simple ImageJ-Based Method for Dynamic Blood Flow Tracking in Zebrafish Embryos and Its Application in Drug Toxicity Evaluation

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Master Program in Nanotechnology, Chung Yuan Christian University, Chung-Li 32023, Taiwan
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Department of Bioscience Technology, Chung Yuan Christian University, Chung-Li 32023, Taiwan
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Department of Chemistry, Chinese Culture University, Taipei 11114, Taiwan
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Guangxi Key Laboratory of Efficacy Study on Chinese Materia Medica, Guangxi University of Chinese Medicine, Nanning 530200, China
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Guangxi Collaborative Innovation Center for Research on Functional Ingredients of Agricultural Residues, Guangxi University of Chinese Medicine, Nanning 530200, China
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Department of Biological Science & Technology, College of Medicine, I-Shou University, Kaohsiung 82445, Taiwan
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Department of Chemistry, Chung Yuan Christian University, Chung-Li 32023, Taiwan
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Authors to whom correspondence should be addressed.
These authors have equally contributed to this paper.
Inventions 2019, 4(4), 65; https://doi.org/10.3390/inventions4040065
Received: 9 September 2019 / Revised: 18 October 2019 / Accepted: 21 October 2019 / Published: 1 November 2019
(This article belongs to the Section Inventions and innovation in Biotechnology and Materials)
This study aimed to develop a simple and cost-effective method to measure blood flow in zebrafish by using an image-based approach. Three days post fertilization (dpf) zebrafish embryos were mounted with methylcellulose and subjected to video recording for tracking blood flow under an inverted microscope equipped with a high-speed CCD camera. In addition, Hoffman lens was used to enhance the blood cell contrast. The red blood cell movement was tracked by using the TrackMate plug-in in the ImageJ image processing program. Moreover, Stack Difference and Time Series Analyzer plug-in were used to detect dynamic pixel changes over time to calculate the blood flow rate. In addition to blood flow velocity and heart rate, the effect of drug treatments on other cardiovascular function parameters, such as stroke volume and cardiac output remains to be explored. Therefore, by using this method, the potential side effects on the cardiovascular performance of ethyl 3-aminobenzoate methanesulfonate (MS222) and 3-isobutyl-1-methylxanthine (IBMX) were evaluated. MS222 is a common anesthetic, while IBMX is a naturally occurring methylxanthine. Compared to normal embryos, MS222- and IBMX-treated embryos had a reduced blood flow velocity by approximately 72% and 58%, respectively. This study showed that MS222 significantly decreased the heart rate, whereas IBMX increased the heart rate. Moreover, it also demonstrated that MS222 treatment reduced 50% of the stroke volume and cardiac output. While IBMX decreased the stroke volume only. The results are in line with previous studies that used expensive instruments and complicated software analysis to assess cardiovascular function. In conclusion, a simple and low-cost method can be used to study blood flow in zebrafish embryos for compound screening. Furthermore, it could provide a precise measurement of clinically relevant cardiac functions, specifically heart rate, stroke volume, and cardiac output. View Full-Text
Keywords: blood flow tracking; ImageJ; zebrafish; IBMX; MS222; cardiovascular function assessment blood flow tracking; ImageJ; zebrafish; IBMX; MS222; cardiovascular function assessment
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MDPI and ACS Style

Santoso, F.; Sampurna, B.P.; Lai, Y.-H.; Liang, S.-T.; Hao, E.; Chen, J.-R.; Hsiao, C.-D. Development of a Simple ImageJ-Based Method for Dynamic Blood Flow Tracking in Zebrafish Embryos and Its Application in Drug Toxicity Evaluation. Inventions 2019, 4, 65.

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