Skeletal Muscle Regeneration is Accelerated Following Injection of Time Release Ion Matrix in Injured Mice

Abstract

IntroductionSkeletal muscle injury remains a significant cause of disability with limited treatment options. Here we report accelerated skeletal muscle regeneration following injection of an inorganic biomaterial alone, cobalt oxide time-release ion matrix (CoO-TRIM). MethodsThe tibialis anterior (TA) muscle of adult C57BL/6J mice was injured with 70 {micro}L of barium chloride, with uninjured limbs serving as contralateral controls. Following the acute inflammatory phase (3 days post injury, dpi), mice were randomly separated into three groups: untreated controls, 70 {micro}l sterile saline injection alone (vehicle control) or 70 {micro}l CoO-TRIM (5 {micro}g/{micro}l) and evaluated at 8- and 14 dpi. Results14 dpi, injured TA muscles receiving a single injection of CoO-TRIM exhibited greater recovery of maximal force production (means {+/-} SEM: Healthy; TRIM, 31.8 {+/-} 0.6 N/cm3; Untreated, 31.7 {+/-} 0.8 N/cm3; Saline, 31.6 {+/-} 0.8 N/cm3, Injured; TRIM, 31.8 {+/-} 0.8 N/cm3; Untreated, 26.1 {+/-} 1.1 N/cm3, P= <0.01 vs. TRIM Injured; Saline, 26.0 {+/-} 1.3 N/cm3, P= <0.01 vs. TRIM Injured), increased fiber size (Healthy; TRIM, 2466 {+/-} 128 {micro}m2 vs. Untreated, 2351 {+/-} 131 {micro}m2; vs. Saline, 2460 {+/-} 129 {micro}m2, Injured; TRIM, 2179 {+/-} 63 {micro}m2 vs. Untreated, 1525 {+/-} 52 {micro}m2, P= <0.01; vs. Saline, 1470 {+/-} 99 {micro}m2, P= <0.01), and accelerated muscle regeneration (TRIM, 32.7 {+/-} 6.8 eMyHC+ fibers/mm2 vs. Untreated, 91.8 {+/-} 11.8 eMyHC+ fibers/mm2, P= <0.01; vs. Saline, 94.0 {+/-} 9.6 eMyHC+ fibers/mm2, P= <0.01). Vascular endothelial growth factor was elevated 14 dpi (TRIM, 17.3 {+/-} 2.9 pg/mg vs. Untreated, 10.3 {+/-} 1.1 pg/mg, P= 0.04; Saline, 5.5 {+/-} 1.2 pg/mg, P= <0.01), with increased muscle microvascular area (TRIM, 119.9 {+/-} 4.2 {micro}m2/fiber vs. Untreated, 94.9 {+/-} 4.2 {micro}m2/fiber, P= <0.01; Saline, 91.3 {+/-} 4.0 {micro}m2/fiber, P= <0.01) following CoO-TRIM treatment. There were early increases in inflammatory responses 8 dpi in injured TA muscles receiving CoO-TRIM (IL-6; TRIM, 29.1 {+/-} 8.2 pg/ml vs. Untreated, 12.4 {+/-} 1.5 pg/ml, P= 0.02; Saline, 11.7 {+/-} 0.8 pg/mg, P= 0.01), with early resolution of degenerative inflammatory cytokines and elevated regenerative cytokines 14 dpi (IL-10; SEM: TRIM, 2.4 {+/-} 0.1 pg/ml vs. Untreated, 2.1 {+/-} 0.1 pg/ml, P= 0.25; Saline, 2.0 {+/-} 0.1 pg/ml, P= 0.04). No differences were observed in healthy contralateral limbs following treatment with CoO-TRIM compared to healthy control mice at 8- and 14 dpi. DiscussionThis work suggests CoO-TRIM enhances muscle regeneration following injury, possibly through an immune cell-mediated mechanism. Statement of Clinical RelevanceThis is the first report of an injected, inorganic biomaterial alone to accelerate regeneration of injured skeletal muscle with no observed effects on healthy muscle. These findings suggest enhancements to skeletal muscle regeneration following CoO-TRIM treatment may be immune cell mediated with an increased local inflammatory response and subsequent improvements to muscle microvasculature and myogenic regulatory factors.