Peripheral denervation participates in heterotopic ossification in a spinal cord injury model

Abstract

We previously reported the development of a new acquired neurogenic HO (NHO) mouse model, combining spinal cord transection (SCI) and chemical muscle injury. Pathological mechanisms responsible for ectopic osteogenesis after central neurological damage are still to be elucidated. In this study, we first hypothesized that peripheral nervous system (PNS) might convey pathological signals from injured spinal cord to muscles in NHO mouse model. Secondly, we sought to determine whether SCI could lead to intramuscular modifications of BMP2 signaling pathways. Twenty one C57Bl6 mice were included in this protocol. Bilateral cardiotoxin (CTX) injection in hamstring muscles was associated with a two-stage surgical procedure, combining thoracic SCI with unilateral peripheral denervation. Volumes of HO (Bone Volume, BV) were measured 28 days after surgery using micro-computed tomography imaging techniques and histological analyses were made to confirm intramuscular osteogenesis. Volume comparisons were conducted between right and left hind limb of each animal, using a Wilcoxon signed rank test. Quantitative polymerase chain reaction (qPCR) was performed to explore intra muscular expression of BMP2, Alk3 and Id1. Nineteen mice survive the complete SCI and peripheral denervation procedure. When CTX injections were done right after surgery (n = 7), bilateral HO were detected in all animals after 28 days. Micro-CT measurements showed significantly increased BV in denervated paws (1.47 mm3 +/- 0.5) compared to contralateral sides (0.56 mm3 +/-0.4), p = 0.03. When peripheral denervation and CTX injections were performed after sham SCI surgery (n = 6), bilateral HO were present in three mice at day 28. Quantitative PCR analyses showed no changes in intra muscular BMP2 expression after SCI as compared to control mice (shamSCI). Peripheral denervation can be reliably added to spinal cord transection in NHO mouse model. This new experimental design confirms that neuro inflammatory mechanisms induced by central or peripheral nervous system injury plays a key role in triggering ectopic osteogenesis.

Eukarion-134 Attenuates Endoplasmic Reticulum Stress-Induced Mitochondrial Dysfunction in Human Skeletal Muscle Cells

“…Three amplifications, consisted of an initial denaturation cycle at 95 °C for 2 min, followed by 40 cycles of 20 s at 95 °C (denaturation), 20 s at optimal annealing temperature, and 20 s at 72 °C (extension), were performed for each primer, followed by a melt curve analysis. Threshold cycle for each target gene of interest was normalised to the housekeeping gene 18S (annealing temperature, 55.7 °C), which has been extensively used in muscle [ 18 , 19 , 20 ], and analysed using the delta-delta (2 −ΔΔCt ) method [ 21 ]. The sequences and annealing temperature of all mitochondrial- and ER stress pathway-associated primers used are provided in Table 1 and Table 2 , respectively.…”

Simultaneous Regeneration of Bone and Nerves Through Materials and Architectural Design: Are We There Yet?

“…[356] Although the mechanism of heterotopic ossification is still unclear, some studies have identified a close connection between the nervous system and the abnormally strong ossification ability. [357][358][359][360] When stimulation from the nervous system is applied under suitable conditions, the mediating pathway from nerve to bone is opened. Solving this riddle may offer a solution to the problem of insufficient bone formation in the repair of large clinical wounds.…”

Mechanism of traumatic heterotopic ossification: In search of injury‐induced osteogenic factors

“…30 NHO animal studies have demonstrated that peripheral denervation increased HO volume on the original SCI NHO model and peripheral denervation alone without SCI could also induce HO in about 50% of mice with local muscular injury. 31 Risk factors for NHO include artificial ventilation, immobilization and muscular spasticity. 26 Specifically, artificial ventilation alters blood homeostasis of electrolytes and acid-base balance, both of which will affect osteogenesis.…”

“…32 This could possibly be explained by the fact that BTA injection equals to a denervation model, and peripheral denervation has been previously shown to enhance NHO. 31 The aetiology of NHO is incompletely understood. A general hypothesis for NHO pathogenesis involves the release of osteogenic humoral factors following the breakdown of blood-brain barrier.…”