ICEOS: Nov. 19 th – Nov. 20 th École Polytechnique of Montreal (Canada) 2 Sainte-Justine Research Hospital Center (Canada) 3 Faculty of Dentistry, University of Montreal (Canada) 4 École des Mines de Saint-Étienne (France) Presenting author 1,2 M ÉNARD A-L, 2 G RIMARD G, 1,2,4 M ASSOL E, 2 L ONDONO I, 2,3 M OLDOVAN F, 1,2 V ILLEMURE I Static versus Dynamic Compression Applied and Subsequently Removed on Growing Rat Tails: Effects on Intervertebral Discs

2 Intro M&M Results Discussion Disclosures Authors Disclosure Informationa.Consultant b.Stock/Shareholder Presenter: Anne-Laure MénardNo Relationships Co-Authors: Guy Grimard(a,b) Emovi, Inc Élise MassolNo Relationships Irène LondonoNo Relationships Florina MoldovanNo Relationships Isabelle VillemureNo Relationships Presenter: Anne-Laure MénardNo Relationships Co-Authors: Guy Grimard(a,b) Emovi, Inc Élise MassolNo Relationships Irène LondonoNo Relationships Florina MoldovanNo Relationships Isabelle VillemureNo Relationships

3 Clinical context: fusionless devices  Research evidence: compression on the disc Mechanical loading: Causing factor of disc degeneration (Aronsson et al., 2011) Disc degeneration:  disc height (Wuertz et al., 2009)  proteoglycan in the nucleus (Sivan et al., 2014)  Compression-based fusionless devices (Skaggs et al., 2013) Span the intervertebral disc Can possibly be removed following spinal correction (Hunt et al., 2010) Intervertebral disc integrity in young growing individuals after physiological compression removal ? Intro M&M Results Discussion

4 Clinical research question 4 Staples : compression Growth plate (GP) (*) Is the intervertebral disc still healthy ? RESEARCH QUESTION Intro M&M Results Discussion Intervertebral disc Fusionless approaches Growth Device removal (*)

5 Hypothesis & Objectives 5 OBJECTIVES To assess and compare the effects of subsequently applying and removing static/dynamic compression on the intervertebral disc: (1) Structure: disc height (2) Composition: nucleus proteoglycan content Intro M&M Results Discussion HYPOTHESIS Dynamic compression better preserves long- term disc integrity and functionality

In vivo protocol: rat tail Growth modulation Growth resumption? Rat age in Days Loading (15 days) No loading (10 days) Surgery 6 Cd7 Proximal Distal Cd7 Control (n = 6) Sham (n = 6) Static (n = 6) Dynamic (n = 6) Static Compression Dynamic Compression Intro M&M Results Discussion (inspired by Walsh et al., 2004 ; Cancel et al., 2009 ; Valteau et al., 2011) Micro-loading device Dissection of 4-week group Dissection of 2-week group

Dissection & measures 7  Intervertebral disc height Intro M&M Results Discussion Analyzed discs Cd4Cd5 Cd6 Cd7Cd8 D 45 D 78  Collecting tissus 3D reconstruction of ex-vivo samples using microCT images Disc height = mean value of six images cut through vertebra

Intervertebral disc measurements Safranine-O staining (2.5X) Intensity level (ImageJ software) 8  Proteoglycan content in the nucleus (Nb pixels) proteoglycan Proteoglycan = (Nb pixels) nucleus Intro M&M Results Discussion  Nucleus/Annulus proportion: stereological method (Griffiths, 1993) (Nb points) nucleus Proportion = (Nb points) annulus Toluidine blue staining (2.5X) Grid (Gimp) + cell counter (ImageJ)

Intervertebral disc height 9 Both static and dynamic compressions  disc height Summary Intro M&M Results Discussion Disc height D 78 normalized with D 45 [μm/μm] * ** 4-week group2-week group p = & p =  One-way ANOVA, Tukey post-hoc comparisons, *p < 0.05 and **p < 0.01 Control Sham Static Dynamic Cd4Cd5 Cd6 Cd7Cd8 D 78

Intervertebral disc composition 10 No difference observed for nucleus/annulus proportion After loading removal, proteoglycan  for static but remained  for dynamic Intro M&M Results Discussion Nucleus/Annulus proportion of D 78 [μm/μm]Proteoglycan content in the nucleus for D 78 2-week group4-week group One-way ANOVA, Tukey post-hoc comparisons, **p < 0.01 ** 2-week group4-week group   ** Summary Control Sham Static Dynamic

Discussion 11 Intro M&M Results Discussion  Limitations One range of loading (magnitude/frequency) Rat discs: conservative model with more notochordal cells, therefore better adaptation capabilities  Future work Inflammatory pathways within disc  Physiological loading range Magnitude below 1.0MPa & frequency above 0.01Hz and below 1.0Hz preserve disc integrity (Iatridis et al., 2006) Dynamic loading: preserve nucleus PG content even following compression removal  PG in the nucleus COMPRESSION 2-week group REMOVING COMPRESSION 4-week group STATIC DYNAMIC

Funding & References 12 Intro M&M Results Discussion  Hunt KJ, Braun JT, Christensen BA. The Effect of Two Clinically Relevant Fusionless Scoliosis Implant Strategies on the Health of the Intervertebral Disc. Spine 2010;35:  Aronsson DD, Stokes IAF. Nonfusion Treatment of Adolescent Idiopathic Scoliosis by Growth Modulation and Remodeling. J Pediatr Orthop 2011;31: S  Wuertz K, Godburn K, MacLean JJ, Barbir A, Stinnett Donnelly J, Roughley PJ, Alini, M, Iatridis JC. In Vivo Remodeling of Intervertebral Discs in Response to Short- and Long-Term Dynamic Compression. J Orthop Res 2009;27:  Sivan SS, Wachtel E, Roughley PJ. Structure, Function, Aging and Turnover of Aggrecan in the Intervertebral Disc. Biochimica et Biophysica Acta 2014;1840:  Walsh AJ, Lotz JC. Biological Response of the Intervertebral Disc to Dynamic Loading. J Biomech 2004;37:  Cancel M, Grimard G, Thuillard-Crisinel D, Moldovan F, Villemure I. Effects of In Vivo Static Compressive Loading on Aggrecan and Type II and X Collagens in the Rat Growth Plate Extracellular Matrix. Bone 2009;44:  Valteau B, Grimard G, Londono I, Moldovan F, Villemure I. In Vivo Dynamic Bone Growth Modulation is Less Detrimental but as Effective as Static Growth Modulation. Bone 2011;49:  Griffiths G. Quantitative Aspects of Immunocytochemistry. In Fine Structure Immunocytochemistry. 1993:  Iatridis JC, MacLean JJ, Roughley PJ, Alini M. Effects of Mechanical Loading on Intervertebral Disc Metabolism In Vivo. J Bone Joint Surg Am 2006;88:  Funding sources:  References: