Logic-based mechanistic machine learning on high-content images reveals how drugs differentially regulate cardiac fibroblasts

Abstract

Fibroblasts are essential regulators of extracellular matrix deposition following cardiac injury. These cells exhibit highly plastic responses in phenotype during fibrosis in response to environmental stimuli. Here, we test whether and how candidate anti-fibrotic drugs differentially regulate measures of cardiac fibroblast phenotype, which may help identify treatments for cardiac fibrosis. We conducted a high content microscopy screen of human cardiac fibroblasts treated with 13 clinically relevant drugs in the context of TGFβ and/or IL-1β, measuring phenotype across 137 single-cell features. We used the phenotypic data from our high content imaging to train a logic-based mechanistic machine learning model (LogiMML) for fibroblast signaling. The model predicted how pirfenidone and Src inhibitor WH-4-023 reduce F-actin assembly and F-actin stress fiber formation, respectively. Validating the LogiMML model prediction that PI3K partially mediates the effects of Src inhibition, we found that PI3K inhibition reduces F-actin fiber formation and procollagen I production in human cardiac fibroblasts. In this study, we establish a modeling approach combining the strengths of logic-based network models and regularized regression models, apply this approach to predict mechanisms that mediate the differential effects of drugs on fibroblasts, revealing Src inhibition acting via PI3K as a potential therapy for cardiac fibrosis.SignificanceCardiac fibrosis is a dysregulation of the normal wound healing response, resulting in excessive scarring and cardiac dysfunction. As cardiac fibroblasts primarily regulate this process, we explored how candidate anti-fibrotic drugs alter the fibroblast phenotype. We identify a set of 137 phenotypic features that change in response to drug treatments. Using a new computational modeling approach termed logic-based mechanistic machine learning, we predict how pirfenidone and Src inhibition affect the regulation of the phenotypic features F-actin assembly and F-actin stress fiber formation. We also show that inhibition of PI3K reduces F-actin fiber formation and procollagen I production in human cardiac fibroblasts, supporting a role for PI3K as a mechanism by which Src inhibition may suppress fibrosis.

The Microenvironment of the Pathogenesis of Cardiac Hypertrophy

“…Watts et al extended this model with estrogen signaling, predicting that the effects of some drugs may be sex specific [ 334 ], building on previous experimental studies of how estrogen affects cardiac fibroblast signaling and activation [ 335 ]. Others have combined network modeling with an experimental drug screen through machine learning to identify pathways by which drugs regulate new fibroblast phenotypes such as stress fiber organization [ 336 ].…”

Modular Multiwell Viscoelastic Hydrogel Platform for Two- and Three-Dimensional Cell Culture Applications

“…Additionally, there were significant changes in the angular second moment, a metric quantifying uniformity, of the F-actin cytoskeleton in the elastic groups compared to the viscoelastic group. This decrease in the angular second moment is likely indicative of an increase in F-actin organization into stress fibers that form when fibroblasts are mechanically activated . Altogether, these data demonstrate that this 96-well plate hydrogel platform enables the high-throughput acquisition of rich cell-level data describing changes in the cellular phenotype due to mechanical cues.…”

“…This decrease in the angular second moment is likely indicative of an increase in F-actin organization into stress fibers that form when fibroblasts are mechanically activated. 31 Altogether, these data demonstrate that this 96well plate hydrogel platform enables the high-throughput acquisition of rich cell-level data describing changes in the cellular phenotype due to mechanical cues.…”

Modular multiwell viscoelastic hydrogel platform for two- and three-dimensional cell culture applications

“…Additionally, there were significant changes in the angular second moment, a metric quantifying uniformity, of the F-actin cytoskeleton in the elastic groups compared to the viscoelastic group. This decrease in angular second moment is likely indicative of an increase in F-actin organization into stress fibers that form when fibroblasts are mechanically activated 27 . Altogether, these data demonstrate that this 96-well plate hydrogel platform enables the high-throughput acquisition of rich cell-level data describing changes in cellular phenotype due to mechanical cues.…”