Combinatrial treatment of anti-High Mobility Group Box-1 monoclonal antibody and epothilone B improves functional recovery after spinal cord contusion injury
Introduction
Spinal cord injury (SCI) disrupts neural networks and causes motor and sensory functional loss, bringing physical and mental suffering on patients. Following the direct damage to the spine which may results from earthquake, traffic accident, fall or violence, local immune response-induced secondary damage triggers a sequential process including inflammation, necrosis and apoptosis, leading to further damage to the spinal cord (Aidemise, 2011). For this reason, reducing secondary damage is considered as a promising therapeutic approach for SCI.
High Mobility Group Box-1 (HMGB1) is one of the damage-associated molecular patterns and has been reported to play an important role in the secondary injury following central nervous system (CNS) injuries: once released from damaged cells, it can bind to cell surface receptors, such as the receptor for advanced glycation end products, toll-like receptor (TLR) 2 and TLR4, which induce blood brain barrier (BBB) disruption and inflammatory response (Hayakawa et al., 2010; Lotze and Tracey, 2005; Zhang et al., 2011). We have previously shown that administration of anti-HMGB1 monoclonal antibody (mAb) after SCI in the early acute phase attenuate secondary damage and promoted motor functional recovery in mouse model (Nakajo et al., 2019; Uezono et al., 2018). Compared with untreated mice, those received anti-HMGB1 mAb showed an increase in neurite sprouting from spared axons, reduced-spinal cord swelling and -neuronal apoptosis, indicating decreased blood-spinal cord barrier (BSCB) disruption.
Since mice treated with anti-HMGB1 mAb showed only partial recovery of motor function after SCI, aiming for a further improvement, we previously transplanted human induced pluripotent stem cells-derived neural stem cells (hiPSC-NSCs) into mouse spinal cord after anti-HMGB1 mAb administration (Uezono et al., 2018). As a result, neurons differentiated from hiPSC-NSCs integrated into host neural circuit and further improved the anti-HMGB1 mAb-derived functional recovery after SCI. However, the use of iPSC-derived cells still retains concerns such as teratoma formation, cell preparation and financial burden, which makes it unsuitable for current clinical application (Gutierrez-Aranda et al., 2010; Huang et al., 2019; Seki and Fukuda, 2015).
In right of these facts, we attempted to find a simple, economical and clinically applicable method that can enhance the therapeutic effect of anti-HMGB1 mAb for SCI. We thus focused, in this study, on epothilone B (Epo B), which has been known as an anticancer drug approved by U.S. Food and Drug Administration and has been shown to be effective in rat model of SCI (Forli, 2014; Ruschel et al., 2015). Epo B binds to β-tubulin subunits in microtubules and increase microtubule stability, affecting the microtubule-based functions such as cell division, cell migration and neurite outgrowth (Pagano et al., 2012; Ruschel et al., 2015). Previous study has reported that administration of low doses of Epo B promoted axon elongation and improved motor function in a rat model of spinal cord dorsal hemisection (Ruschel et al., 2015).
In the present study, we examined the therapeutic effect of a combination of anti-HMGB1 mAb and Epo B treatments in contusion SCI mouse model. In contrast to hemisection SCI model (Pagano et al., 2012; Ruschel et al., 2015), we found no effect of Epo B treatment alone on sprouting of remaining axons and locomotion recovery after contusion SCI. However, the combination therapy of anti-HMGB1 mAb and Epo B significantly improved the axonal outgrowth and motor functional recovery compared with either treatment alone, suggesting that anti-HMGB1 mAb-mediated suppression of contusion injury-induced secondary damage is prerequisite for Epo B to reveal the therapeutic effect. Therefore, it is conceivable that both anti-HMGB1 mAb-mediated preservation of lesion area and Epo B-induced axonal sprouting are necessary for efficient functional recovery after contusion SCI.
Section snippets
Animals
This study was exploratory and all mouse experiments were conducted in accordance with guidelines of the Kyushu University Center for Animal Resources and Development. A total of 55 female C57BL/6J mice (aged 8–10 weeks, weight = 18 g–22 g, Japan SLC, Research Resource Identifiers (RRID): IMSR_JAX:000664) were used in this study. All mice were arbitrarily assigned to experimental groups and no randomization was performed. For dose determination analysis, 9 mice were randomly divided into three
Combinationatrial treatment of anti-HMGB1 mAb and Epo B improve locomotion recovery after SCI
Before we investigate the effect of the combinatorial administration of anti-HMGB1 mAb and Epo B, we first examined whether intraperitoneal (i.p.) injection of Epo B has any adverse effects. In a previous study using rat SCI model, Ruschel et al. adopted i.p. injection with 0.75 mg/kg BW of Epo B at day 1 and 15 after SCI (Ruschel et al., 2015). Since the concentration of Epo B in rat spinal cord is 40 ng/g 1 week after i.p injected at 0.75 mg/kg BW (Ruschel et al., 2015), while that in mouse
Discussion
In the present study, we have found that combination of Epo B and anti-HMGB1 mAb treatment improved the intraspinal neuron survival, increased axons outgrowth, and improved locomotion recovery after SCI. Administration of Epo B enhanced axon elongation only when it is combined with anti-HMGB1 mAb, which may be due to the decreased inflammatory response by anti-HMGB1 mAb. We also performed ablation experiment and have found that survived intraspinal neurons are critical for functional recovery.
Author contributions
Y.Z. and K.N.: conception/design, manuscript writing; Y.Z.: performed the experiments, data analysis; Y.Z. and T.N.: behavior analysis; D.W. and M. Nishibori: contributed to the purification of anti-HMGB1 mAb; N.U., T.Y. and M. Nakajo: experiment support; Y.Z., N.U., T.N., and K.N.: results discussion. K.N.: financial support, project supervising, and final approval of manuscript.
Disclosure
The authors declare no conflict of interest.
Acknowledgements
We appreciate all members in K. Nakashima lab for experimental help and valuable discussion. We would like to thank M. Amago at the Research Support Center, Research Center for Human Disease Modeling, Kyushu University Graduate School of Medical Sciences for technical assistance. This work was supported by the Suzuken Memorial Foundation (to KN), a Grant-in-Aid for Scientific Research on Innovative Areas JP16H06527 (to KN), JP16K21734 (to KN), a Grant-in-Aid for Challenging Research
References (48)
- et al.
Gait analysis of adult paraplegic rats after spinal cord repair
Exp. Neurol.
(1997) - et al.
Conditional genetic deletion of PTEN after a spinal cord injury enhances regenerative growth of CST axons and motor function recovery in mice
Exp. Neurol.
(2015) - et al.
Reorganization of intact descending motor circuits to replace lost connections after injury
Neurotherapeutics
(2016) - et al.
Methods to assess the development and recovery of locomotor function after spinal cord injury in rats
Exp. Neurol.
(1993) - et al.
Behavioral and histological outcomes following graded spinal cord contusion injury in the C57Bl/6 mouse
Exp. Neurol.
(2001) - et al.
Therapeutic time window of anti-high mobility group box-1 antibody administration in mouse model of spinal cord injury
Neurosci. Res.
(2019) - et al.
Differences in cytokine gene expression profile between acute and secondary injury in adult rat spinal cord
Exp. Neurol.
(2003) - et al.
Pretreatment with a γ-secretase inhibitor prevents tumor-like overgrowth in human iPSC-derived transplants for spinal cord injury
Stem Cell Reports
(2016) - et al.
Epothilone B inhibits migration of glioblastoma cells by inducing microtubule catastrophes and affecting EB1 accumulation at microtubule plus ends
Biochem. Pharmacol.
(2012) - et al.
Systemic epothilone D improves hindlimb function after spinal cord contusion injury in rats
Exp. Neurol.
(2018)
Induction of pluripotent stem cells from adult human fibroblasts by defined factors
cell
Fatty acid synthesis is indispensable for survival of human pluripotent stem cells
iScience
Epothilone B speeds corneal nerve regrowth and functional recovery through microtubule stabilization and increased nerve beading
Sci. Rep.
Engrafted neural stem/progenitor cells promote functional recovery through synapse reorganization with spared host neurons after spinal cord injury
Stem Cell Reports
Neurons derived from transplanted neural stem cells restore disrupted neuronal circuitry in a mouse model of spinal cord injury
J. Clin. Invest.
Role of NMDA and non-NMDA ionotropic glutamate receptors in traumatic spinal cord axonal injury
J. Neurosci.
Secondary injury mechanisms in traumatic spinal cord injury: a nugget of this multiply cascade
Acta Neurobiol. Exp. (Wars)
The injured spinal cord spontaneously forms a new intraspinal circuit in adult rats
Nat. Neurosci.
Basso Mouse Scale for locomotion detects differences in recovery after spinal cord injury in five common mouse strains
J. Neurotrauma
Cell Death, Repair, and Recovery of Function After Spinal Cord Contusion Injuries in Rats, Neurobiology of Spinal Cord Injury
Spinal cord repair: advances in biology and technology
Nat. Med.
Reducing pericyte-derived scarring promotes recovery after spinal cord injury
Cell
Exercise training after spinal cord injury selectively alters synaptic properties in neurons in adult mouse spinal cord
J. Neurotrauma
Epothilones: from discovery to clinical trials
Curr. Top. Med. Chem.
Cited by (5)
High-Mobility Group Box 1 in Spinal Cord Injury and Its Potential Role in Brain Functional Remodeling After Spinal Cord Injury
2023, Cellular and Molecular NeurobiologyReview: The role of HMGB1 in spinal cord injury
2023, Frontiers in ImmunologyPromising Advances in Pharmacotherapy for Patients with Spinal Cord Injury—A Review of Studies Performed In Vivo with Modern Drugs
2022, Journal of Clinical Medicine