Cell Shape Characteristics of Human Skeletal Muscle Cells as a Predictor of Myogenic Competency: A New Paradigm Towards Precision Cell Therapy for Incontinence

Desprez C1, Danovi D2, Knowles C3, Day R1

Research Type

Pure and Applied Science / Translational

Abstract Category

Continence Care Products / Devices / Technologies

Abstract 515
The Best of the Rest in Science
Scientific Podium Short Oral Session 33
Saturday 10th September 2022
12:37 - 12:45
Hall G1
Anal Incontinence Basic Science Pre-Clinical testing Stem Cells / Tissue Engineering
1. Centre for Precision Healthcare, UCL Division of Medicine, University College London, Gower Street, London, United Kingdom, 2. Centre for Stem Cells and Regenerative Medicine, King's College London, London, United Kingdom, 3. National Bowel Research Centre and GI Physiology Unit, Blizard Institute, Centre for Neuroscience, Surgery & Trauma, Queen Mary University of London, London, United Kingdom

Charlotte Desprez



Hypothesis / aims of study
Skeletal muscle derived cells (SMDC) hold tremendous potential for replenishing dysfunctional muscle associated with incontinence. Autologous SMDC-based therapy for faecal incontinence (FI) is reported to be safe, but the level of efficacy reported in previous studies suggests that heterogeneity in the proposed therapy may contribute to inconsistent clinical outcomes. 

Improved methods for patient stratification to identify patients who are (un)likely to respond well to regenerative therapies are advocated to ensure the potential benefits of cell-based regenerative medicine can be fully exploited. Predicting the therapeutic potency of SMDC in vitro prior to implantation will facilitate development of successful therapeutics in regenerative medicine and reduce implementation costs. 

Here, we report on the development of a novel SMDC profiling tool to examine populations of cells in vitro derived from different donors. The primary aim of the study was to evaluate the applicability of multi-parametric imaging-based phenotypic characterization to distinguish myogenic potency of SMDC. Heterogeneity in the formation of myotubes from different donors was correlated with cell shape descriptors.
Study design, materials and methods
Commercially available human SMDC (Cook MyoSite) consisting of non-differentiated primary human muscle cells were derived from 14 individual human donors. Image acquisition of SMDC growth was performed using a CytoSMART Lux2 microscope. Images were acquired every 5 minutes over a period of 72 hours. CellProfiler software was used to create an image analysis pipeline for analysis of different cell shape characteristics during 2 hours of tracking at 12- and 24-hours post-culture. The Konstanz Information Miner (KNIME; open-source data analytics) software was used to calculate the mean and SD of the characteristic for all the objects identified in one image. These data were then used for statistical correlation with the fusion index calculated from myotube formation assays established in the same cultures after 5 days of subsequent incubation in differentiation medium. The fusion index was calculated as the ratio of nuclei number in myocytes with 2 or more nuclei divided by the total number of nuclei. Correlation studies used the Spearman test with r < -0.5 or > +0.5 indicating correlation.
Several early cell shape characteristics were found to negatively correlate with the fusion index. Images collected at 12 hours after initiating culture revealed that 5 cell shape characteristics negatively correlated with the fusion index, including the total area occupied by cells (r=-0.815, p=0.001), area shape (r=-0.631; p=0.028), bounding box area (r=-0.576; p=0.049), minimum ferret diameter (r=-0.589; p=0.044) and minor axis length (r=-0.614; p=0.034).  Images collected at 24 hours after initiating revealed 8 cell shape characteristics that negatively correlated with the fusion index. These included: total area occupied by cells (r=-0.686; p=0.007), area shape (r=-0.709; p=0.005), bounding box area (r=-0.563; p=0.036), compactness (r=-0.534; p=0.049), equivalent diameter (r=-0.576; p=0.031), minimum ferret diameter (r=-0.620; p=0.018), minor axis length (r=-0.590; p=0.026) and perimeter (r=-0.654; p=0.011). There was a high correlation between all these characteristics at 24 hours of imaging after initiating culture.
Interpretation of results
Results indicate that monitoring of cell shape during the early stages of bioprocessing using real-time imaging could be used to predict cellular competency necessary for differentiation and myofibre formation in vivo.
Concluding message
Further studies could establish whether selection of either patients or cell populations on this basis have a higher probability of yielding better outcomes in cell-based therapy for incontinence.
Funding Confidence in Collaboration Award, London Advanced Therapies, a Research England - Connecting Capabilities Fund initiative AMELIE consortium via the European Union's Horizon 2020 Research and Innovation Programme under grant agreement No 874807. National Institute of Health Research (NIHR) for the University College London Hospitals Biomedical Research Centre UCL Grand Challenges and the Higher Education, Research and Innovation Department at the French Embassy in the UK Clinical Trial No Subjects None

Continence 2S2 (2022) 100466
DOI: 10.1016/j.cont.2022.100466

02/07/2024 20:52:14