Development of Functional, Contractile 3‐D Human Muscle Constructs for Studies of the Effects of Microgravity on Muscle Mass and Function.

Major demographic changes are affecting modern societies, leading to rapidly increasing numbers of older people with relatively poor health and quality of life. The mechanisms underlying age‐related loss of muscle have not been fully evaluated, but we have previously demonstrated that muscle from ageing animals and humans show attenuated adaptations to exercise that compromises their ability to maintain muscle mass and function. We have evaluated key mechanisms underlying adaptations of skeletal muscle to exercise and demonstrated that the exercising muscle of young or adult animals and humans generates reactive oxygen species (ROS) that stimulate activation of specific transcription factors leading to increased generation of a number of protective and cytoprotective proteins. Such responses do not occur in muscle from older animals and man in response to exercise. Astronauts and animals exposed to microgravity also lose skeletal muscle mass and their muscles are also relatively unresponsive to aerobic or resistance training in microgravity. Within a call from the UK Space Agency for an upcoming national mission on board the International Space Station (ISS), we will utilise in vitro studies of 3‐D skeletal muscle constructs to determine whether an analogous failure of muscle adaptations to contractile activity occurs in muscle fibres exposed to microgravity as that which occurs in older people on earth. The aim of the current study was to develop appropriate 3‐D constructs which remained viable and contractile following field electrical stimulation in specifically designed, sealed hardware for space flight for up to 7 days. Initial experiments examined a variety of immortalised and primary murine myocytes which were found unsuitable due to lack of excitability and high oxygen consumption. Subsequent experiments have utilised immortalised human myocytes obtained from Dr. Vincent Mouly (Paris, France) which have been fabricated on 3D printed scaffolds encapsulated in a fibrin hydrogel solution (4×10 6 cells/construct) and polymerised at 37°C. Once matured the constructs were checked for contractility and the medium was replaced with Leibrovitz‐L15 medium (supplemented with 2 mg/mL 6‐aminocaproic acid) to minimise oxygen consumption while retaining viability. These constructs retain viability and contractility for up to 7 days and display significant adaptations to short periods of contractile activity in normal gravity. Specific hardware to maintain the constructs and permit electrical stimulation of contractions has been designed by Kayser Space Ltd., (Harwell, UK), and the experiment will fly to the ISS in 2021. Support or Funding Information Supported by the UK Space Agency