Automated Image Analysis Pipeline Development to Monitor Disease Progression in Muscular Dystrophy Using CellProfiler

Abstract

Muscular dystrophies are inherited disorders that are characterized by progressive muscle degeneration. These disorders are caused by mutation in the genes encoding structural elements within the muscle, which leads to increased vulnerability to mechanical stress and sarcolemma damage. Although myofibers have the capacity to regenerate, the newly formed myofibers still harbor the genetic mutation, which induces continuous cycles of muscle fiber death and regeneration. This repeated cycling is accompanied by an inflammatory response which eventually provokes excessive fibrotic deposition. The histopathological changes in skeletal muscle tissue are central to the disease pathogenesis. Analysis of muscle histopathology is the gold standard for monitoring muscle health and disease progression. However, manual, or semi-manual quantification methods are not only immensely tedious but can be subjective. Here, we present four image analysis pipelines built in CellProfiler which enable users without a background in computer vision or programming to quantitatively analyze biological images. These image analysis pipelines are designed to quantify skeletal muscle histopathological staining for membrane damage, the abundance and size distribution of regenerating muscle fibers, inflammation via quantification of CD68 + M1 macrophages, and collagen deposition. Additionally, we discuss methods to address common errors associated with the quantification of microscopy images. These automated tools can not only improve workflow efficiency but can provide a better understanding of the histopathological progression of muscular dystrophy.

Exploring lipin1 as a promising therapeutic target for the treatment of Duchenne muscular dystrophy

“…Quantification analysis of Picrosirius red and EBD staining were used pipelines generated in our lab [ 30 ]. For immunostaining and Western blot, statistical analysis was performed with one-way analysis of variance (ANOVA) followed by Bonferroni’s multiple comparison test to determine significant changes between groups using GraphPad Prism 9 (GraphPad Software, version 9.4.0).…”