Skip to main content
SpringerLink
Log in

Glycogen synthase kinase 3 alpha/beta deletion induces precocious growth plate remodeling in mice

Journal of Molecular Medicine volume 99, pages 831–844 (2021)Cite this article

Abstract

Glycogen synthase kinase (GSK) 3 acts to negatively regulate multiple signaling pathways, including canonical Wnt signaling. The two mammalian GSK3 proteins (alpha and beta) are at least partially redundant. While Gsk3a KO mice are viable and display a metabolic phenotype, abnormal neuronal development, and accelerated aging, Gsk3b KO animals die late in embryogenesis or at birth. Selective Gsk3b KO in bone delays development of some bones, whereas cartilage-specific Gsk3b KO mice are normal except for elevated levels of GSK3A protein. However, the collective role of these two GSK3 proteins in cartilage was not evaluated. To address this, we generated tamoxifen-inducible, cartilage-specific Gsk3a/Gsk3b KO (described as “cDKO”) in juvenile mice and investigated their skeletal phenotypes. We found that cartilage-specific Gsk3a/Gsk3b deletion in young, skeletally immature mice causes precocious growth plate (GP) remodeling, culminating in shorter long bones and hence, growth retardation. These mice exhibit inefficient breathing patterns at later stages and fail to survive. The disrupted GP in cDKO mice showed progressive loss of cellular and proteoglycan components, and immunostaining for SOX9, while BGLAP (osteocalcin) and COL2A1 increased. In addition, we observed increased osteoclast recruitment and cell apoptosis. Surprisingly, changes in articular cartilage of cDKO mice were mild compared with the GP, signifying differential regulation of articular cartilage vs GP tissues. Taken together, these findings emphasize a crucial role of two GSK3 proteins in skeletal development, in particular in the maintenance and function of GP.

Key Messages

• Both GSK3 genes, together, are crucial regulators of growth plate remodeling.

• Cartilage-specific deletion of both GSK3 genes causes skeletal growth retardation.

• Deletion of both GSK3 genes decreases Sox9 levels and promotes chondrocyte apoptosis.

• Cartilage-specific GSK3 deletion in juvenile mice culminates in premature lethality.

• GSK3 deletion exhibits mild effects on articular cartilage compared to growth plate.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

39,95 €

Price includes VAT (Finland)

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Data Availability

The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

References

  1. Beurel E, Grieco SF, Jope RS (2015) Glycogen synthase kinase-3 (GSK3): Regulation, actions, and diseases. Pharmacol Ther 148:114–131

    Article  CAS  Google Scholar 

  2. Lal H, Ahmad F, Woodgett J, Force T (2015) The GSK-3 family as therapeutic target for myocardial diseases. Circ Res 116:138–149

    Article  CAS  Google Scholar 

  3. Kaidanovich-Beilin O, Lipina T, Takao K, Van Eede M, Hattori S, Lalibert C, Khan M, Okamoto K, Chambers J, Fletcher P et al (2009) Abnormalities in brain structure and behavior in GSK-3alpha mutant mice. Mol Brain 2:35

    Article  Google Scholar 

  4. Gillespie JR, Bush JR, Bell GI, Aubrey LA, Dupuis H, Ferron M, Kream B, DiMattia G, Patel S, Woodgett JR, Karsenty G, Hess DA, Beier F (2013) GSK-3β function in bone regulates skeletal development, whole-body metabolism, and male life span. Endocrinology 154:3702–3718

    Article  CAS  Google Scholar 

  5. Gillespie JR, Ulici V, Dupuis H, Higgs A, Dimattia A, Patel S, Woodgett JR, Beier F (2011) Deletion of glycogen synthase kinase-3β in cartilage results in up-regulation of glycogen synthase kinase-3α protein expression. Endocrinology 152:1755–1756

    Article  CAS  Google Scholar 

  6. Cuzzocrea S, Mazzon E, Di Paola R, Muià C, Crisafulli C, Dugo L, Collin M, Britti D, Caputi AP, Thiemermann C (2006) Glycogen synthase kinase-3β inhibition attenuates the degree of arthritis caused by type II collagen in the mouse. Clin Immunol 120:57–67

    Article  CAS  Google Scholar 

  7. Yost C, Torres M, Miller JR, Huang E, Kimelman D, Moon RT (1996) The axis-inducing activity, stability, and subcellular distribution of β-catenin is regulated in Xenopus embryos by glycogen synthase kinase 3. Genes Dev 10:1443–1454

    Article  CAS  Google Scholar 

  8. Kaushal JB, Sankhwar P, Kumari S, Popli P, Shukla V, Hussain MK, Hajela K, Dwivedi A (2017) The regulation of Hh/Gli1 signaling cascade involves Gsk3β- mediated mechanism in estrogen-derived endometrial hyperplasia. Sci Rep 7. https://doi.org/10.1038/s41598-017-06370-1

  9. Nelson ER, Levi B, Sorkin M, James AW, Liu KJ, Quarto N, Longaker MT (2011) Role of GSK-3β in the osteogenic differentiation of palatal mesenchyme. PLoS One 6:10

    PubMed Central  Google Scholar 

  10. Sawakami K, Robling AG, Ai M, Pitner ND, Liu D, Warden SJ, Li J, Maye P, Rowe DW, Duncan RL, Warman ML, Turner CH (2006) The Wnt co-receptor LRP5 is essential for skeletal mechanotransduction but not for the anabolic bone response to parathyroid hormone treatment. J Biol Chem 281:23698–23711

    Article  CAS  Google Scholar 

  11. Yuasa T, Kondo N, Yasuhara R, Shimono K, Mackem S, Pacifici M, Iwamoto M, Enomoto-Iwamoto M (2009) Transient activation of Wnt/β-catenin signaling induces abnormal growth plate closure and articular cartilage thickening in postnatal mice. Am J Pathol 175:1993–2003

    Article  CAS  Google Scholar 

  12. Maeda Y, Nakamura E, Nguyen MT, Suva LJ, Swain FL, Razzaque MS, Mackem S, Lanske B (2007) Indian Hedgehog produced by postnatal chondrocytes is essential for maintaining a growth plate and trabecular bone. Proc Natl Acad Sci U S A 104:6382–6387

    Article  CAS  Google Scholar 

  13. Hirai T, Chagin AS, Kobayashi T, Mackem S, Kronenberg HM (2011) Parathyroid hormone/parathyroid hormone related protein receptor signaling is required for maintenance of the growth plate in postnatal life. Proc Natl Acad Sci U S A 108:191–196

    Article  CAS  Google Scholar 

  14. Maeda Y, Schipani E, Densmore MJ, Lanske B (2010) Partial rescue of postnatal growth plate abnormalities in Ihh mutants by expression of a constitutively active PTH/PTHrP receptor. Bone 46:472–478

    Article  CAS  Google Scholar 

  15. Hoeflich KP, Luo J, Rubie EA, Tsao MS, Jin O, Woodgett JR (2000) Requirement for glycogen synthase kinase-3β in cell survival and NF-κB activation. Nature 406:86–90

    Article  CAS  Google Scholar 

  16. MacAulay K, Doble BW, Patel S, Hansotia T, Sinclair EM, Drucker DJ, Nagy A, Woodgett JR (2007) Glycogen Synthase Kinase 3α- Specific Regulation of Murine Hepatic Glycogen Metabolism. Cell Metab 6:329–337

    Article  CAS  Google Scholar 

  17. Patel S, Doble BW, MacAulay K, Sinclair EM, Drucker DJ, Woodgett JR (2008) Tissue-Specific Role of Glycogen Synthase Kinase 3 in Glucose Homeostasis and Insulin Action. Mol Cell Biol 28:6314–6328

    Article  CAS  Google Scholar 

  18. Terpstra L, Prud'homme J, Arabian A, Takeda S, Karsenty G, Dedhar S, St-Arnaud R (2003) Reduced chondrocyte proliferation and chondrodysplasia in mice lacking the integrin-linked kinase in chondrocytes. J Cell Biol 162:139–148

    Article  CAS  Google Scholar 

  19. Wang G, Woods A, Agoston H, Ulici V, Glogauer M, Beier F (2007) Genetic ablation of Rac1 in cartilage results in chondrodysplasia. Dev Biol 306:612–623

    Article  CAS  Google Scholar 

  20. Suzuki D, Yamada A, Amano T, Yasuhara R, Kimura A, Sakahara M, Tsumaki N, Takeda S, Tamura M, Nakamura M, Wada N, Nohno T, Shiroishi T, Aiba A, Kamijo R (2009) Essential mesenchymal role of small GTPase Rac1 in interdigital programmed cell death during limb development. Dev Biol 335:396–406

    Article  CAS  Google Scholar 

  21. Fang H, Huang L, Welch I, Norley C, Holdsworth DW, Beier F, Cai D (2018) Early Changes of Articular Cartilage and Subchondral Bone in The DMM Mouse Model of Osteoarthritis. Sci Rep 8:2855

    Article  Google Scholar 

  22. Dupuis H, Pest MA, Hadzic E, Vo TX, Hardy DB, Beier F (2019) Exposure to the rxr agonist sr11237 in early life causes disturbed skeletal morphogenesis in a rat model. Int J Mol Sci 20. https://doi.org/10.3390/ijms20205198

  23. Pest MA, Russell BA, Zhang YW, Jeong JW, Beier F (2014) Disturbed cartilage and joint homeostasis resulting from a loss of mitogen-inducible gene 6 in a mouse model of joint dysfunction. Arthritis Rheumat 66:2816–2827

    Article  CAS  Google Scholar 

  24. James CG, Stanton LA, Agoston H, Ulici V, Underhill TM, Beier F (2010) Genome-wide analyses of gene expression during mouse endochondral ossification. PLoS One 5:e8693. https://doi.org/10.1371/journal.pone.0008693

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Akiyama H, Lyons JP, Mori-Akiyama Y, Yang X, Zhang R, Zhang Z, Deng JM, Taketo MM, Nakamura T, Behringer RR, McCrea P, de Crombrugghe B (2004) Interactions between Sox9 and beta-catenin control chondrocyte differentiation. Genes Dev 18:1072–1087

    Article  CAS  Google Scholar 

  26. Jing Y, Jing J, Wang K, Chan K, Harris SE, Hinton RJ, Feng JQ (2018) Vital roles of beta-catenin in trans-differentiation of chondrocytes to bone cells. Int J Biol Sci 14:1–9

    Article  CAS  Google Scholar 

  27. Houben A, Kostanova-Poliakova D, Weissenböck M, Graf J, Teufel S, von der Mark K, Hartmann C (2016) Beta-catenin activity in late hypertrophic chondrocytes locally orchestrates osteoblastogenesis and osteoclastogenesis. Dev (Cambridge) 143:3826–3838

    CAS  Google Scholar 

  28. Doble BW, Patel S, Wood GA, Kockeritz LK, Woodgett JR (2007) Functional Redundancy of GSK-3α and GSK-3β in Wnt/β-Catenin Signaling Shown by Using an Allelic Series of Embryonic Stem Cell Lines. Dev Cell 12:957–971

    Article  CAS  Google Scholar 

  29. Holzer T, Probst K, Etich J, Auler M, Georgieva VS, Bluhm B, Frie C, Heilig J, Niehoff A, Nüchel J (2019) Respiratory chain inactivation links cartilage mediated growth retardation to mitochondrial diseases. J Cell Biol 218:1853–1870

    Article  CAS  Google Scholar 

  30. Tamamura Y, Otani T, Kanatani N, Koyama E, Kitagaki J, Komori T, Yamada Y, Costantini F, Wakisaka S, Pacifici M, Iwamoto M, Enomoto-Iwamoto M (2005) Developmental regulation of Wnt/β-catenin signals is required for growth plate assembly, cartilage integrity, and endochondral ossification. J Biol Chem 280:19185–19195

    Article  CAS  Google Scholar 

  31. Guidotti S, Minguzzi M, Platano D, Santi S, Trisolino G, Filardo G, Mariani E, Borzì RM (2017) Glycogen Synthase Kinase-3β Inhibition Links Mitochondrial Dysfunction, Extracellular Matrix Remodelling and Terminal Differentiation in Chondrocytes. Sci Rep 7. https://doi.org/10.1038/s41598-017-12129-5

  32. Kobayashi T, Chung UI, Schipani E, Starbuck M, Karsenty G, Katagiri T, Goad DL, Lanske B, Kronenberg HM (2002) PTHrP and Indian hedgehog control differentiation of growth plate chondrocytes at multiple steps. Development 129:2977–2986

    Article  CAS  Google Scholar 

  33. Kobayashi T, Soegiarto DW, Yang Y, Lanske B, Schipani E, McMahon AP, Kronenberg HM (2005) Indian hedgehog stimulates periarticular chondrocyte differentiation to regulate growth plate length independently of PTHrP. J Clin Inv 115:1734–1742

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank the following funding sources for financial support. S.K.B. received a postdoctoral fellowship award from CONNECT! NSERC CREATE Program in Soft Connective Tissue Regeneration/Therapy. S.K.B. and C.P. were also supported by the Transdisciplinary Training Award from Collaborative Program in Musculoskeletal Health Research (CMHR) at Western’s Bone and Joint Institute. F.B holds the Canada Research Chair in Musculoskeletal Research and is the recipient of a Foundation Grant from the Canadian Institutes of Health Research (Grant #332438). We thank all members of our lab for valuable discussions and encouragement, in particular Julia Bowering for tissue sectioning services.

Code availability (software application or custom code)

Not applicable

Author information

Authors and Affiliations

  1. Department of Physiology and Pharmacology, Schulich School of Medicine & Dentistry, Western University, London, ON, N6A 5C1, Canada

    Supinder Kour Bali, Dawn Bryce, Carina Prein & Frank Beier

  2. Western Bone and Joint Institute, Western University, London, ON, N6A 5C1, Canada

    Supinder Kour Bali, Dawn Bryce, Carina Prein & Frank Beier

  3. Lunenfeld-Tanenbaum Research Institute, Sinai Health, Toronto, ON, M5G 1X5, Canada

    James R. Woodgett

Authors
  1. Supinder Kour Bali
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Dawn Bryce
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Carina Prein
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. James R. Woodgett
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Frank Beier
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Conceptualization and methodology, S.K.B. and F.B.; investigation, S.K.B., D.B., C.P.; analyses, S.K.B., C.P. and F.B.; manuscript preparation and review, S.K.B., D.B., J.R.W., and F.B. All authors approved submission of manuscript.

Corresponding author

Correspondence to Frank Beier.

Ethics declarations

Conflict of interest

The authors declare no conflict of interests related to this study. The authors have no financial relationship with the sponsor of this research, the Canadian Institutes of Health Research (with the exception of grant funding).

Ethics approval

All experimental mice were handled in accordance with the guidelines from the Canadian Council on Animal Care, and experiments were approved by the Animal Use Subcommittee at The University of Western Ontario.

Consent to participate

Not applicable

Consent for publication

Not applicable

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

ESM 1

(DOCX 17709 kb)

Rights and permissions

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bali, S.K., Bryce, D., Prein, C. et al. Glycogen synthase kinase 3 alpha/beta deletion induces precocious growth plate remodeling in mice. J Mol Med 99, 831–844 (2021). https://doi.org/10.1007/s00109-021-02049-3

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00109-021-02049-3

Keywords

Search

Navigation