City Research Online

Exploring neutrophil behaviour in a zebrafish model of inflammation through the generation of novel parameters using MatLab algorithms

Henry, K. M., Reyes-Aldasoro, C. C. & Renshaw, S. A. (2012). Exploring neutrophil behaviour in a zebrafish model of inflammation through the generation of novel parameters using MatLab algorithms. In: http://ebooks.ekonnect.co/ECI%202012%20Final%20Programme/files/assets/basic-html/page235.html. European Congress of Immunology, 05-09-2012 - 08-09-2012, Glasgow, UK.

Abstract

Purpose/Objective: Tracking of immune cells is key to understanding their behaviour during inflammation. Current software available for tracking of immune cells is limited. The aim of this study was to develop a MatLab package of segmentation and tracking algorithms to apply in the tracking of neutrophils, fluorescently labelled in our zebrafish model [1]. The development of algorithms in MatLab allows us to explore parameters not available in other software packages such as directionality of neutrophil movement and neutrophil behaviour in- side and outside of a wound region.

Materials and methods: Tail fin transection was performed on Tg(mpx:GFP) zebrafish (3 dpf) which were imaged on a spinning disk confocal from 1 h post injury (hpi) to 7 hpi. Images were exported from VolocityTM and analysed using MatLab m-files written for the tracking of immune cells. This is a fully automated analysis, after the user defines the initial thresholds based on fluorescent intensity of the images.

Results: Neutrophils from injured embryos had a lower meandering ratio and a greater speed than neutrophils tracked in uninjured embryos (meandering ratio 0.24 ± 0.03 versus 0.42 ± 0.05, P = 0.003; speed 4.03 ± 0.32 versus 1.31 ± 0.21 pixels/frame, P < 0.0001, n = 3), however there was no significant difference in total distance travelled (166.9 ± 17.4 versus 210.1 ± 30.28 pixels, P > 0.05). In injured embryos with a defined wound region, the oriented velocity towards the wound was 0.31 ± 0.24 pixels/frame. Once within the wound region, the oriented velocity of neutrophil tracks was -0.39 ± 0.32 pixels/frame; indicating that while the neutrophils travel at a similar speed, they are now travelling away from the wound. The ‘in wound ratio’ was 0.91 ± 0.04, indicating that once neutrophils enter the wound region they tend to stay, in the timeframe studied. The ‘leave wound ratio’ was 0.37 ± 0.03, a measure of the rate at which neutrophils move away from the site of injury once they have entered the wound region.

Conclusions: Using these algorithms, we can analyse the behaviour of immune cells in a more detailed way. In addition to previously available parameters such as meandering ratio and speed, more complex parameters such as velocity towards or away from a wound region and a measurement of how neutrophils behave while in a wound region are available. Combining this novel tracking technology with established assays in our laboratory will enable the further dissection of neutrophil fate following an inflammatory stimulus.

Publication Type: Conference or Workshop Item (Paper)
Subjects: Q Science > QR Microbiology > QR180 Immunology
Departments: School of Science & Technology > Engineering
School of Science & Technology > Computer Science > giCentre
[thumbnail of Henry_etal_Glasgow.pdf]
Preview
Text - Accepted Version
Download (45kB) | Preview

Export

Add to AnyAdd to TwitterAdd to FacebookAdd to LinkedinAdd to PinterestAdd to Email

Downloads

Downloads per month over past year

View more statistics

Actions (login required)

Admin Login Admin Login