Early fracture callus displays smooth muscle‐like viscoelastic properties ex vivo: Implications for fracture healing

Abstract

Cells of early, fibrous callus in bone fractures possess much alpha smooth muscle actin. This callus contracts and relaxes; however, active and passive components of its force production have yet to be defined. We aimed to establish whether passive viscoelastic properties of early soft fracture callus are smooth muscle-like in nature. Under anesthesia one rib was fractured in rats and calluses removed 7 days later for analysis. Urinary bladder detrusor muscle and Achilles tendon were also resected and analyzed. Force production in these tissues was measured using a force transducer when preparations were immersed in calcium-free Krebs-Henseleit solution (pH 7.4, 228C). Viscoelastic responses were measured in each preparation in response to 50 mN increases and decreases in force after achieving basal tissue tension by preconditioning. Callus, bladder, and tendon all displayed varying, reproducible degrees of stress relaxation (SR) and reverse stress relaxation (RSR) (n ¼ 7 for all groups). Hysteresis was observed in callus, with the first SR response significantly larger than that produced in subsequent stretches (p < 0.05). Callus SR responses were greater than tendon (p < 0.001) but less than bladder (p < 0.001). Callus RSR responses were greater than tendon (p < 0.001), but no significant difference was seen between RSR of callus and bladder. We concluded that early, soft callus displayed significant SR and RSR phenomena similar to smooth muscle tissue, and SR and RSR may be important in maintenance of static tension in early callus by promoting osteogenesis and fracture healing. ß

α1 adrenergic receptor agonist, phenylephrine, actively contracts early rat rib fracture callus ex vivo

“…Preconditioning was necessary as callus displays a degree of hysteresis. 1 Once a steady force had been reached, tension was reduced from the designated basal tension to 0 and force was recorded. Importantly, although reductions in tension to reach this point varied between callus samples, changes in tension between solutions were always equal.…”

“…Release of tension typically produces passive reverse stress relaxation. 1 In eight callus preparations, PE was added to KH solution to make a final concentration of 10 À6 M PE and after 30 min tension was reduced from basal tension to absolute 0 and recorded until a steady-state force was produced. This force was compared to the passive response preceding PE addition, with additional force following PE exposure deemed to be due to ''active'' contraction.…”

“…1 Furthermore, cells of this callus in particular osteoprogenitor cells of early fibrous callus show significant expression of the essential contractile protein, alpha smooth muscle actin (aSMA), 2,3 which has recently been suggested as a possible marker for osteoprogenitor cells. 4 These characteristics suggest that early callus osteoprogenitor cells display a smooth muscle-like phenotype, like myofibroblasts.…”

“…3,7 As early fibrous callus appears to consist of contractile smooth muscle-like osteoprogenitor cells, it is suggested that they too have a functional role for tissue contraction in the healing of bone fractures. 1 We have previously isolated and quantified the passive viscoelastic phenomena of stress relaxation, reverse stress relaxation, and hysteresis in early callus ex vivo. 1 With this passive component isolated, it may be possible to identify any ''active'' smooth muscle-like contraction (i.e., force produced by myofilament cross-bridge cycling) in callus.…”

“…1 We have previously isolated and quantified the passive viscoelastic phenomena of stress relaxation, reverse stress relaxation, and hysteresis in early callus ex vivo. 1 With this passive component isolated, it may be possible to identify any ''active'' smooth muscle-like contraction (i.e., force produced by myofilament cross-bridge cycling) in callus. 8 The contribution of active force development can be measured as force produced above preconditioned stable passive force.…”