Abstract
Purpose :
A common hallmark of Multiple Sclerosis (MS) is the thinning of retinal layers that contain retinal microglia. Microglia are the immune cells of the central nervous system which carry out their dynamic age- and disease-related neuromodulatory functions by dynamically shifting between five distinctive morphotypes (ramified, hyper-ramified, activated, rod and amoeboid). Here, we sought to develop automated algorithms to identify differences in the total and distinct microglia morphotype densities between experimental MS induced by cuprizone (CPZ) treatment, and untreated controls.
Methods :
The MS-modelled male C57 mice were given CPZ (0.2%w/w) mixed with chow for 4 months (m) until their sacrifice at 6m or 28m. Age-matched controls (Ctrl) were fed standard chow. Post-sacrifice, mouse eyes were enucleated, the retinas dissected, immunostained, whole-mounted and then imaged. Automated cell counting and morphotyping methods including supervised machine learning algorithms (SVM-C) were developed to quantify densitometric differences. Non-parametric statistical analyses were performed on the collected data (n: 6mCPZ=7; 6mCtrl=15; 28mCPZ=10; 28mCtrl=7).
Results :
The SVM-C produced performance accuracies of 87.9-100% for each morphotype with AUC >0.99. Experimental groups did not significantly affect the total retinal microglia densities. However, there were more ramified (p<0.05) and amoeboid (ns) cells in old diseased (28mCPZ) retinas compared to young diseased (6mCPZ) and old healthy (28mCtrl) retinas. Also, there were significantly more rod microglia (p<0.01) in young animals (6mCtrl and 6mCPZ) compared to old animals (28mCtrl and 28mCPZ). There were no significant differences between the densities of young animal groups whilst there were between the old, with significantly less activated, hyper-ramified and rod cells (p<0.05) in 28mCPZ.
Conclusions :
The SVM-C worked with a high accuracy and discriminative power. No differences in the total densities suggest that age- and CPZ-induced morphotype density differences might occur as a result of morphological changes rather than proliferation. The current study did not investigate the functional associations, although ramified and amoeboid morphotype trends may suggest increases in debris detection and phagocytosis with the increase of age and CPZ-induced experimental MS.
This is a 2021 ARVO Annual Meeting abstract.