Human Cartilage Aging and Osteoarthritis Shuanhu Zhou Ph

Human Cartilage Aging and Osteoarthritis Shuanhu Zhou, Ph. D. Department of Orthopedic Surgery Brigham and Women’s Hospital Harvard Medical School JBC Synergy Meeting September 29, 2017

Aging and OA Roos, E. M. & Arden, N. K. (2015) Strategies for the prevention of knee osteoarthritis Nat. Rev. Rheumatol. doi: 10. 1038/nrrheum. 2015. 135

Differences in the physiologic environment, metabolic rate, and cellular make-up of bone and cartilage Published by AAAS D J Huey et al. Science 2012; 338: 917 -921

Total Joint Replacement for Osteoarthritic Hip and knee http: //www. stoneclinic. com/knee-replacement

Cartilage tissue engineering for degenerative joint disease Advanced Drug Delivery Reviews, 58(2): 300– 322, 2006

Cooperation between TGF-b and Wnt pathways during chondrocyte differentiation of human MSCs. Control Wnt Demineralized Bone Promotes Chondrocyte Differentiation of Human MSCs Cultured in Collagen Sponges TGF-b 1 + Wnt Zhou S, et al. J Bone Miner Res. 2004 Mar; 19(3): 463 -70. Zhou S, et al. Cell Tissue Bank. 2005; 6(1): 33 -44. Current project: The effects of age on chondrocytogenesis and cartilage tissue engineering with human MSCs. Laney Burke/Shuanhu Zhou

Various clinical strategies regenerate cartilage using chondrocytes or MSCs: Autologous chondrocyte implantation (ACI), Microfracture involves subchondral bone penetration to release bone marrow that forms a stem cell–rich clot. D J Huey et al. Science 2012; 338: 917 -921

A randomized controlled trial comparing ACI and microfracture at 2 years follow-up reported that for both procedures, younger patients (18 -30 years of age) had better clinical outcomes than did older patients (31 -45 years, p=0. 007) Knutsen G, Engebretsen L, Ludvigsen TC, Drogset JO, Grontvedt T, Solheim E, et al. Autologous chondrocyte implantation compared with microfracture in the knee. J Bone Joint Surg. 2004; 86 A: 455– 64. )

Human Cartilage Aging and Osteoarthritis Images show young normal (age 40), old normal (age 76) and OA (age 88). Lotz M, Loeser RF. Effects of aging on articular cartilage homeostasis. Bone. 2012 Aug; 51(2): 241 -8.

Chondrocyte senescence  Changes in the total chondrocyte numbers of healthy and osteoarthritic articular cartilage with increasing age. K Bobacz et al. Ann Rheum Dis 2004; 63: 1618 -1622 Chris Bertucci/Shuanhu Zhou Martin JA, Brown TD, Heiner AD, Buckwalter JA. Chondrocyte senescence, joint loading and osteoarthritis. Clin Orthop Relat Res. 2004 Oct; (427 Suppl): S 96 -103.

Senescence-associated β-galactosidase staining in human hip articular cartilage AS, articular surface of the cartilage Chondrocytes close to the lesion site in osteoarthritic cartilage (Grade 2/3 ? ) Intact regions of osteoarthritic hip cartilage (Grade 0 ? ) Jo S. Price et al. Aging Cell (2002), 1, pp 57– 65 Normal (no disease) hip cartilage

Chondrocytes exhibit features of cellular senescence in the contexts of ageing and osteoarthritis Loeser, R. F. et. al. (2016) Ageing and the pathogenesis of osteoarthritis Nat. Rev. Rheumatol. doi: 10. 1038/nrrheum. 2016. 65

Local clearance of senescent cells attenuates the development of post-traumatic osteoarthritis and creates a pro-regenerative environment Male p 16 -3 MR mice undergoing ACLT were injected intra-articularly with vehicle (Veh) or senolytic molecule ganciclovir (GCV). p 16 -3 MR harbors a p 16 INK 4 a (Cdkn 2 a) promoter driving the expression of a fusion protein containing synthetic Renilla luciferase and monomeric red fluorescent protein domains, as well as a truncated form of herpes simplex virus 1 thymidine kinase (HSV-TK). Jeon OH, Kim C, Laberge RM, Demaria M, Rathod S, Vasserot AP, Chung JW, Kim DH, Poon Y, David N, Baker DJ, van Deursen JM, Campisi J, Elisseeff JH. Local clearance of senescent cells attenuates the development of post-traumatic osteoarthritis and creates a pro-regenerative environment. Nat Med. 2017 Jun; 23(6): 775 -781.

OSTEOARTHRITIS Goldring M, Marcu K. Arthr Res Therapy. 11: 224, 2009

The two sides of the chondrocyte

Osteoarthritis (OA) Osteoarthritis (human knee) OA grade of cartilage Effects of OA grade on cartilage. (A) Regions having sufficient homogeneity were partitioned and graded clinically by the surgeons according to the Outerbridge classification: Grade 0, normal appearance; Grade 1, with softening and swelling; Grade 2, minor fragmentation and fissuring involving an area of less than half an inch; Grade 3, fragmentation and fissuring in an area greater than half an inch; Grade 4, exposed subchondral bone. (B) Histological staining photomicrographs show partitioned grades of cartilage discarded from knee replacement for OA of a 57 -yearold male subject (Toluidine blue-O staining, scale bars: 100 mm). Zhou S, Thornhill TS, Meng F, Xie L, Wright J, Glowacki J. Influence of osteoarthritis grade on molecular signature of human cartilage. J Orthop Res. 2016 Mar; 34(3): 454 -62.

Effects of OA grade on molecular signatures in chondrocytes. (a, b) Inflammatory mediators and receptors, (c) Proteolytic enzymes. Zhou S, Thornhill TS, Meng F, Xie L, Wright J, Glowacki J. Influence of osteoarthritis grade on molecular signature of human cartilage. J Orthop Res. 2016 Mar; 34(3): 454 -62.

Osteoclast-like phenotype in chondrocytes of human articular cartilage Zhou S, Thornhill TS, Meng F, Xie L, Wright J, Glowacki J. Influence of osteoarthritis grade on molecular signature of human cartilage. J Orthop Res. 2016 Mar; 34(3): 454 -62.

Inhibition of osteoclasts (by a bisphonate , Zoledronate ) prevents cartilage loss and pain in a rat model of degenerative joint disease (monosodium iodoacetate (MIA) model) Sham/Control C. Early sub-chronic zoledronate treatment when initiated 2 weeks after MIA treated joints Pre-emptive chronic treatment with zoledronate at the same day 1 of MIA. Strassle BW, et al. , Osteoarthritis Cartilage. 2010; 18(10): 1319 -28 Zoledronate (disodium tetrahydrate): subcutaneous (s. c. ) administration.

The role of proinflammatory cytokines in the pathophysiology of OA Bisphonates OPG ? Kapoor, M. et al. (2010) Role of proinflammatory cytokines in the pathophysiology of osteoarthritis Nat. Rev. Rheumatol. doi: 10. 1038/nrrheum. 2010. 196

JBC Pilot & Feasibility Microgrant: We plan to study the influence of OA disease severity on gene expression profiles of human cartilage by RNA-Seq analysis. Total RNA will be isolated from discarded human cartilages that will be partitioned and stratified by OA disease severity from Outerbridge Grade 0 to Grade 3. We shall quantitatively assess the influence of OA grade on gene expression profiles in human cartilage from subjects with osteoarthritis. In order to reduce the variability by age, cartilage will be studied from eligible male and female, age range 55 -65 years, subjects undergoing TKA. . Expected Results: The outcomes of these RNA-seq analyses of gene expression profiling in human cartilage may reveal novel mechanisms/pathways or therapeutic targets for osteoarthritis.

In Vitro Effect of IL-1 b (24 hr) on Gene Expression in Normal Human Chondrocytes

The effect of OA grade on Wnt-related genes in human articular cartilage The effects of Wnt/b-catenin activator SB 216763 on gene expression in human chondrocytes Wnts are senescence-associated secretory phenotype (SASP) factors in OA?

Intra-articular treatment with the Wnt inhibitor sclerostin sustains the FCSC pool and regenerates cartilage in a Temporomandibular joint (TMJ) injury model. fibrocartilage stem cells (FCSCs) Sclerostin (SOST) Embree MC, Chen M, Pylawka S, Kong D, Iwaoka GM, Kalajzic I, Yao H, Shi C, Sun D, Sheu TJ, Koslovsky DA, Koch A, Mao JJ. Exploiting endogenous fibrocartilage stem cells to regenerate cartilage and repair joint injury. Nat Commun. 2016 Oct 10; 7: 13073.

Cartilage injury causes activation of WNT signalling through downregulation of antagonists such as FRZB and upregulation of several agonists such as WNT 16 and WNT 8. WNT 16 buffers the canonical WNT activation to homeostatic levels through its capacity to directly support a weak activation and preventing excessive activation induced by other ligands. Giovanna Nalesso et al. Ann Rheum Dis doi: 10. 1136/annrheumdis-2015 -208577 © 2016 by BMJ Publishing Group Ltd and European League Against Rheumatism

Tissue-autonomous mechanisms of OA progression Cartilage Chondrocyte MMPs Cat. K ADAMTS-5 TNF-a, ILs, IFN-g, M-CSF Wnts ? ? Senescent Chondrocyte RANKL Apoptosis Chondroclast? Or Osteoclast-like BAX/Bcl-2 Bone

Acknowledgments Julie Glowacki, Ph. D Li Xie, Ph. D Xiaonan Liang, MD Jeff Lange, MD Thomas Thornhill, MD John Wright, MD Anna Gorska, MS Chris Bertucci, Ph. D Laney Burke, MS These studies were supported by grants from NIH, JBC Pilot, BWH BRI and Department of Orthopedic Surgery. No conflicts-of-interest on this subject.