Contractile force assessment methods for in vitro skeletal muscle tissues

Abstract

Over the last few years, there has been growing interest in measuring the contractile force (CF) of engineered muscle tissues to evaluate their functionality. However, there are still no standards available for selecting the most suitable experimental platform, measuring system, culture protocol, or stimulation patterns. Consequently, the high variability of published data hinders any comparison between different studies. We have identified that cantilever deflection, post deflection, and force transducers are the most commonly used configurations for CF assessment in 2D and 3D models. Additionally, we have discussed the most relevant emerging technologies that would greatly complement CF evaluation with intracellular and localized analysis. This review provides a comprehensive analysis of the most significant advances in CF evaluation and its critical parameters. In order to compare contractile performance across experimental platforms, we have used the specific force (sF, kN/m2), CF normalized to the calculated cross-sectional area (CSA). However, this parameter presents a high variability throughout the different studies, which indicates the need to identify additional parameters and complementary analysis suitable for proper comparison. We propose that future contractility studies in skeletal muscle constructs report detailed information about construct size, contractile area, maturity level, sarcomere length, and, ideally, the tetanus-to-twitch ratio. These studies will hopefully shed light on the relative impact of these variables on muscle force performance of engineered muscle constructs. Prospective advances in muscle tissue engineering, particularly in muscle disease models, will require a joint effort to develop standardized methodologies for assessing CF of engineered muscle tissues.

Adaptive exhaustion during prolonged intermittent hypoxia causes dysregulated skeletal muscle protein homeostasis

“…Our C2C12 cell line is a robust in vitro model of skeletal muscle which we then validated with hiPSC‐derived myotubes. While hiPSC myotubes have been utilized in the literature to provide in vitro human translational relevance (Guo et al., 2020), and demonstrate contractile force similar to C2C12 and primary muscle cultures in humans and mice (Vesga‐Castro et al., 2022), future studies are needed to further characterize how well these cell lines compare in terms of molecular and functional characteristics.…”

Engineering of human myotubes toward a mature metabolic and contractile phenotype

“…We are aware that high levels of functionality can also be achieved and observed with 3D culture tissue engineering approaches also giving way to even more elaborate readouts [69–71]. However, it was our goal to develop an “easy to use” 2D protocol which can be implemented in many laboratories, driven by metabolic or diabetes related research questions, where monolayer myotube culture is readily available while establishing complicated tissue engineered 3D solutions is no valid option.…”

“…Thus, our data show and give suggestions why IGF1-guided human myotube differentiation overcomes the previously inadequate capability of established human myotube models with our newly described "easy to use" differentiation protocol. We are aware that high levels of functionality can also be achieved and observed with 3D culture tissue engineering approaches also giving way to even more elaborate readouts [69][70][71]. However, it was our goal to develop an "easy to use" 2D protocol which can be implemented in many laboratories, driven by metabolic or diabetes related research questions, where monolayer myotube culture is readily available while establishing complicated tissue engineered 3D solutions is no valid option.…”

l-Anserine Increases Muscle Differentiation and Muscle Contractility in Human Skeletal Muscle Cells

“…In vitro studies using skeletal muscle cells are useful for directly evaluating the effects of functional materials on skeletal muscle, because unlike in in vivo studies, the effects of complex organ–organ interactions do not need to be considered. Although rodent-derived skeletal muscle cells have been widely used in in vitro studies, it is preferable to use human-derived skeletal muscle cells for evaluating functional materials for food ingredients and drugs, because cells from different species may respond differently to the materials. , Skeletal muscle tissues constructed using human skeletal muscle cells can be employed as a tool for evaluating functional materials in vitro. , In particular, the contractile force, which can be measured noninvasively, is considered to be a useful index for tissue characterization, because it reflects the complex interactions of multiple second messenger pathways and structural proteins. , We previously developed microdevices to measure the contractile force of engineered skeletal muscle tissues. − More recently, we developed a 96-well contractile force-measuring system with engineered human skeletal muscle tissues and were able to screen for antiatrophic peptides using this system …”

“…13,14 In particular, the contractile force, which can be measured noninvasively, is considered to be a useful index for tissue characterization, because it reflects the complex interactions of multiple second messenger pathways and structural proteins. 15,16 We previously developed microdevices to measure the contractile force of engineered skeletal muscle tissues. 17−19 More recently, we developed a 96-well contractile force-measuring system with engineered human skeletal muscle tissues and were able to screen for antiatrophic peptides using this system.…”