Validation of Human Skeletal Muscle Tissue Chip Autonomous Platform to Model Age-Related Muscle Wasting in Microgravity

Abstract

Microgravity-induced muscle atrophy experienced by astronauts shares similar physiological changes to muscle wasting experienced by older adults, known as sarcopenia. These shared attributes provide a rationale for investigating microgravity-induced molecular changes in human bioengineered muscle cells that may also mimic the progressive underlying pathophysiology of sarcopenia. Here, we report the results of an experiment that incorporated three-dimensional myobundles derived from muscle biopsies from young and older adults, that were integrated into an autonomous CubeLabâ"¢, and flown to the International Space Station (ISS) aboard SpaceX CRS-21 in December 2020 as part of the NIH/NASA funded Tissue Chips in Space program. Global transcriptomic RNA-Seq analysis comparing the myobundles in space and on the ground revealed downregulation of shared transcripts related to myoblast proliferation and muscle differentiation for those in space. The analysis also revealed differentially expressed gene pathways related to muscle metabolism unique to myobundles derived from the older cohort exposed to the space environment compared to ground controls. Gene classes related to inflammatory pathways were uniquely modulated in flight samples cultured from the younger cohort compared to ground controls. Our muscle tissue chip platform provides a novel approach to studying the cell autonomous effects of microgravity on muscle cell biology that may not be appreciated on the whole organ or organism level and sets the stage for continued data collection from muscle tissue chip experimentation in microgravity. Thus, we also report on the challenges and opportunities for conducting autonomous tissue-on-chip CubeLab ^TM payloads on the ISS.

Figure 1

Experimental design, implementation, and automation of the skeletal muscle MPS payload for the International Space Station. a Human cell banks. Skeletal muscle biopsies were obtained from the vastus lateralis from volunteers (AdventHealth, Orlando). Isolated muscle precursor cultures were enriched for CD56+ (myogenic) cells. b Experimental flight timeline. Tissue chips were seeded with myoblasts pre-differentiated, loaded into the CubeLab^™ and launched to the ISS on SpaceX CRS-21. Crew members installed the PAUL on the EXPRESS rack locker and the on-orbit experiment was initiated after plug-in. 10 days post launch, crew members, moved the payload to cold stowage following experiment termination with RNALater. c On-orbit real-time telemetry. Recording of YA-derived myobundle during electrical stimulation on-orbit and resultingdominant frequency of contraction determined by Fast Fourier Transform of the time series signal of displacement.

Figure 2

Transcriptional responses of muscle myobundles during flight and ground culture. a, Volcano plots. Differential expressed genes between flight and ground controls for YA (left panel) or OS (right panel). Red: significantly up-regulated genes in flight. Blue: significantly down-regulated genes. Significance determined according to log2 fold change with threshold set at ±2 and -log10 p-value ≤0.05. b. Comparative analysis of differential gene expression changes between flight and ground controls of genes involved in muscle myogenesis and contraction YA (orange bars) and OS (blue bars). c. Venn diagrams. Overlapping and distinct DEGs among Flight YA vs GC and Flight OS vs GC datasets. d. Panther classification system of the 59 shared genes shown in the Venn diagram. The horizontal bar chart identifies the four protein classes and relative number of DEGs of each functional class by applying a cut-off threshold of 10 genes. e. f. Significantly enriched KEGG pathways in response to flight either in YA- or OS-derived myobundles relative to their respective ground controls.