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Kindlin-2 preserves integrity of the articular cartilage to protect against osteoarthritis

Nature Aging volume 2, pages 332–347 (2022)Cite this article

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Abstract

Osteoarthritis (OA) is an aging-related degenerative joint disease with a poorly defined mechanism. Here we report that kindlin-2 is highly expressed in articular chondrocytes and downregulated in the degenerated cartilage of aged mice and patients with OA. Kindlin-2 deletion in articular chondrocytes leads to spontaneous OA and exacerbates instability-induced OA lesions in adult mice. Kindlin-2 deficiency promotes mitochondrial oxidative stress and activates Stat3, leading to Runx2-mediated chondrocyte catabolism. Pharmacological inhibition of Stat3 activation or genetic ablation of Stat3 in chondrocytes reverses aberrant accumulation of Runx2 and extracellular-matrix-degrading enzymes and limits OA deteriorations caused by kindlin-2 deficiency. Deleting Runx2 in chondrocytes reverses structural changes and OA lesions caused by kindlin-2 deletion without downregulating p-Stat3. Intra-articular injection of AAV5–kindlin-2 decelerates progression of aging- and instability-induced knee joint OA in mice. Collectively, we identify a pathway consisting of kindlin-2, Stat3 and Runx2 in articular chondrocytes that is responsible for maintaining articular cartilage integrity and define a potential therapeutic target for OA.

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Fig. 1: Inducible deletion of kindlin-2 in chondrocytes causes striking spontaneous OA in adult mice.
Fig. 2: Kindlin-2 deficiency in chondrocytes accelerates instability-induced OA.
Fig. 3: Kindlin-2 loss induces chondrocyte hypertrophic differentiation and catabolism through Stat3-dependent upregulation of Runx2.
Fig. 4: Kindlin-2 deficiency promotes oxidative stress, Stat3 activation and nuclear translocation in chondrocytes.
Fig. 5: Kindlin-2 expression is drastically reduced in human OA articular cartilages.
Fig. 6: Genetic deletion of Stat3 in chondrocyte corrects Runx2 accumulation and attenuates OA lesions in cKO mice.
Fig. 7: Genetical deletion of Runx2 in chondrocytes palliates spontaneous and DMM-induced OA defects in cKO mice.
Fig. 8: Intra-articular injection of AAV5–kindlin-2 protects against development of aging- and DMM-induced OA in mice.

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Data availability

All data supporting the findings of this study are available within the article and its Extended Data files or from the corresponding authors upon reasonable request. Source data are provided with this paper. RNA-seq data have been deposited in Sequence Read Archive under accession no. PRJNA773746.

References

  1. Hunter, D. J. & Bierma-Zeinstra, S. Osteoarthritis. Lancet 393, 1745–1759 (2019).

    Article  CAS  PubMed  Google Scholar 

  2. He, Y. et al. Pathogenesis of osteoarthritis: risk factors, regulatory pathways in chondrocytes, and experimental models. Biology https://doi.org/10.3390/biology9080194 (2020).

  3. Sun, M. M. & Beier, F. Chondrocyte hypertrophy in skeletal development, growth, and disease. Birth Defects Res. C Embryo Today 102, 74–82 (2014).

    Article  CAS  PubMed  Google Scholar 

  4. Chen, D. et al. Osteoarthritis: toward a comprehensive understanding of pathological mechanism. Bone Res. 5, 16044 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Song, E. K. et al. ITGBL1 modulates integrin activity to promote cartilage formation and protect against arthritis. Sci. Transl. Med. https://doi.org/10.1126/scitranslmed.aam7486 (2018).

  6. Charlier, E. et al. Toward diagnostic relevance of the αVβ5, αVβ3, and αVβ6 integrins in OA: expression within human cartilage and spinal osteophytes. Bone Res. 8, 35 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Wang, Q. et al. Dysregulated integrin αVβ3 and CD47 signaling promotes joint inflammation, cartilage breakdown, and progression of osteoarthritis. JCI Insight 4, e128616 (2019).

    Article  PubMed Central  Google Scholar 

  8. Zheng, Q. et al. Type X collagen gene regulation by Runx2 contributes directly to its hypertrophic chondrocyte-specific expression in vivo. J. Cell Biol. 162, 833–842 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Li, F. et al. Runx2 contributes to murine Col10a1 gene regulation through direct interaction with its cis-enhancer. J. Bone Miner. Res. 26, 2899–2910 (2011).

    Article  CAS  PubMed  Google Scholar 

  10. Liao, L. et al. Deletion of Runx2 in articular chondrocytes decelerates the progression of DMM-induced osteoarthritis in adult mice. Sci. Rep. 7, 2371 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  11. Huang, J. et al. The microRNAs miR-204 and miR-211 maintain joint homeostasis and protect against osteoarthritis progression. Nat. Commun. 10, 2876 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  12. Catheline, S. E. et al. Chondrocyte-specific RUNX2 overexpression accelerates post-traumatic osteoarthritis progression in adult mice. J. Bone Miner. Res. 34, 1676–1689 (2019).

    Article  CAS  PubMed  Google Scholar 

  13. Rice, S. J. et al. Identification of a novel, methylation-dependent, RUNX2 regulatory region associated with osteoarthritis risk. Hum. Mol. Genet. 27, 3464–3474 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Chen, D., Kim, D. J., Shen, J., Zou, Z. & O’Keefe, R. J. Runx2 plays a central role in osteoarthritis development. J. Orthop. Translat. 23, 132–139 (2020).

    Article  PubMed  Google Scholar 

  15. Latourte, A. et al. Systemic inhibition of IL-6/Stat3 signalling protects against experimental osteoarthritis. Ann. Rheum. Dis. 76, 748–755 (2017).

    Article  CAS  PubMed  Google Scholar 

  16. Wang, S. W. & Sun, Y. M. The IL-6/JAK/STAT3 pathway: potential therapeutic strategies in treating colorectal cancer (review). Int. J. Oncol. 44, 1032–1040 (2014).

    Article  CAS  PubMed  Google Scholar 

  17. Miscia, S. et al. Tumor necrosis factor α (TNF-α) activates Jak1/Stat3-Stat5B signaling through TNFR-1 in human B cells. Cell Growth Differ. 13, 13–18 (2002).

    CAS  PubMed  Google Scholar 

  18. Kadry, Y. A. & Calderwood, D. A. Chapter 22: structural and signaling functions of integrins. Biochim. Biophys. Acta Biomembr. 1862, 183206 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Sun, Z., Costell, M. & Fassler, R. Integrin activation by talin, kindlin and mechanical forces. Nat. Cell Biol. 21, 25–31 (2019).

    Article  CAS  PubMed  Google Scholar 

  20. Michael, M. & Parsons, M. New perspectives on integrin-dependent adhesions. Curr. Opin. Cell Biol. 63, 31–37 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Rognoni, E., Ruppert, R. & Fassler, R. The kindlin family: functions, signaling properties and implications for human disease. J. Cell Sci. 129, 17–27 (2016).

    Article  CAS  PubMed  Google Scholar 

  22. Plow, E. F. & Qin, J. The kindlin family of adapter proteins. Circ. Res. 124, 202–204 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Svensson, L. et al. Leukocyte adhesion deficiency-III is caused by mutations in KINDLIN3 affecting integrin activation. Nat. Med. 15, 306–312 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Moser, M., Nieswandt, B., Ussar, S., Pozgajova, M. & Fassler, R. Kindlin-3 is essential for integrin activation and platelet aggregation. Nat. Med. 14, 325–330 (2008).

    Article  CAS  PubMed  Google Scholar 

  25. Schmidt, S. et al. Kindlin-3-mediated signaling from multiple integrin classes is required for osteoclast-mediated bone resorption. J. Cell Biol. 192, 883–897 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Malinin, N. L. et al. A point mutation in KINDLIN3 ablates activation of three integrin subfamilies in humans. Nat. Med. 15, 313–318 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Montanez, E. et al. Kindlin-2 controls bidirectional signaling of integrins. Genes Dev. 22, 1325–1330 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Zhan, J. & Zhang, H. Kindlins: roles in development and cancer progression. Int. J. Biochem. Cell Biol. 98, 93–103 (2018).

    Article  CAS  PubMed  Google Scholar 

  29. Wei, X. et al. Kindlin-2 mediates activation of TGF-β/Smad signaling and renal fibrosis. J. Am. Soc. Nephrol. https://doi.org/10.1681/ASN.2012101041 (2013).

  30. Sun, Y. et al. Kindlin-2 association with Rho GDP-dissociation inhibitor α suppresses Rac1 activation and podocyte injury. J. Am. Soc. Nephrol. 28, 3545–3562 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Zhu, K. et al. Kindlin-2 modulates MafA and β-catenin expression to regulate β-cell function and mass in mice. Nat. Commun. 11, 484 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Xue, X., Xue, S., Wan, W., Li, J. & Shi, H. HIF-1α interacts with kindlin-2 and influences breast cancer elasticity: a study based on shear wave elastography imaging. Cancer Med. 9, 4971–4979 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Guo, B., Gao, J., Zhan, J. & Zhang, H. Kindlin-2 interacts with and stabilizes EGFR and is required for EGF-induced breast cancer cell migration. Cancer Lett. 361, 271–281 (2015).

    Article  CAS  PubMed  Google Scholar 

  34. Guo, L. et al. Kindlin-2 links mechano-environment to proline synthesis and tumor growth. Nat. Commun. 10, 845 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  35. Guo, L. et al. PINCH-1 regulates mitochondrial dynamics to promote proline synthesis and tumor growth. Nat. Commun. 11, 4913 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Qin, L., Liu, W., Cao, H. & Xiao, G. Molecular mechanosensors in osteocytes. Bone Res. 8, 23 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. He, X. et al. Kindlin-2 deficiency induces fatal intestinal obstruction in mice. Theranostics 10, 6182–6200 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Qi, L. et al. Kindlin-2 suppresses transcription factor GATA4 through interaction with SUV39H1 to attenuate hypertrophy. Cell Death Dis. 10, 890 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  39. Gao, H. et al. Lipoatrophy and metabolic disturbance in mice with adipose-specific deletion of kindlin-2. JCI Insight 4, e128405 (2019).

    Article  PubMed Central  Google Scholar 

  40. Zhang, Z. et al. Kindlin-2 is essential for preserving integrity of the developing heart and preventing ventricular rupture. Circulation 139, 1554–1556 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  41. Wu, C. et al. Kindlin-2 controls TGF-β signalling and Sox9 expression to regulate chondrogenesis. Nat. Commun. 6, 7531 (2015).

    Article  PubMed  Google Scholar 

  42. Cao, H. et al. Focal adhesion protein kindlin-2 regulates bone homeostasis in mice. Bone Res. 8, 2 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Qin, L. et al. Kindlin-2 mediates mechanotransduction in bone by regulating expression of sclerostin in osteocytes. Commun. Biol. 4, 402 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Fu, X. et al. Kindlin-2 regulates skeletal homeostasis by modulating PTH1R in mice. Signal Transduct. Target Ther. 5, 297 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Dalagiorgou, G. et al. Mechanosensor polycystin-1 potentiates differentiation of human osteoblastic cells by upregulating Runx2 expression via induction of JAK2/STAT3 signaling axis. Cell. Mol. Life Sci. 74, 921–936 (2017).

    Article  CAS  PubMed  Google Scholar 

  46. Murphy, M. P. How mitochondria produce reactive oxygen species. Biochem. J. 417, 1–13 (2009).

    Article  CAS  PubMed  Google Scholar 

  47. Perez-Garcia, S. et al. Wnt and RUNX2 mediate cartilage breakdown by osteoarthritis synovial fibroblast-derived ADAMTS-7 and -12. J. Cell. Mol. Med. 23, 3974–3983 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Orfanidou, T., Iliopoulos, D., Malizos, K. N. & Tsezou, A. Involvement of SOX-9 and FGF-23 in RUNX-2 regulation in osteoarthritic chondrocytes. J. Cell. Mol. Med. 13, 3186–3194 (2009).

    Article  PubMed  Google Scholar 

  49. Li, D., Xie, G. & Wang, W. Reactive oxygen species: the 2-edged sword of osteoarthritis. Am. J. Med. Sci. 344, 486–490 (2012).

    Article  PubMed  Google Scholar 

  50. Henry, S. P. et al. Generation of aggrecan-CreERT2 knockin mice for inducible Cre activity in adult cartilage. Genesis 47, 805–814 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Aszodi, A., Hunziker, E. B., Brakebusch, C. & Fassler, R. β1 integrins regulate chondrocyte rotation, G1 progression, and cytokinesis. Genes Dev. 17, 2465–2479 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Raducanu, A., Hunziker, E. B., Drosse, I. & Aszodi, A. β1 integrin deficiency results in multiple abnormalities of the knee joint. J. Biol. Chem. 284, 23780–23792 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Franceschi, R. T., Romano, P. R. & Park, K. Y. Regulation of type I collagen synthesis by 1,25-dihydroxyvitamin D3 in human osteosarcoma cells. J. Biol. Chem. 263, 18938–18945 (1988).

    Article  CAS  PubMed  Google Scholar 

  54. Liao, L. et al. Runx2 is required for postnatal intervertebral disc tissue growth and development. J. Cell. Physiol. 234, 6679–6687 (2019).

    Article  CAS  PubMed  Google Scholar 

  55. Moh, A. et al. Role of STAT3 in liver regeneration: survival, DNA synthesis, inflammatory reaction and liver mass recovery. Lab. Invest. 87, 1018–1028 (2007).

    Article  CAS  PubMed  Google Scholar 

  56. Li, J. et al. Metformin limits osteoarthritis development and progression through activation of AMPK signalling. Ann. Rheum. Dis. 79, 635–645 (2020).

    Article  CAS  PubMed  Google Scholar 

  57. Chaplan, S. R., Bach, F. W., Pogrel, J. W., Chung, J. M. & Yaksh, T. L. Quantitative assessment of tactile allodynia in the rat paw. J. Neurosci. Methods 53, 55–63 (1994).

    Article  CAS  PubMed  Google Scholar 

  58. Gosset, M., Berenbaum, F., Thirion, S. & Jacques, C. Primary culture and phenotyping of murine chondrocytes. Nat. Protoc. 3, 1253–1260 (2008).

    Article  CAS  PubMed  Google Scholar 

  59. Valls-Lacalle, L. et al. Succinate dehydrogenase inhibition with malonate during reperfusion reduces infarct size by preventing mitochondrial permeability transition. Cardiovasc. Res. 109, 374–384 (2016).

    Article  CAS  PubMed  Google Scholar 

  60. Vaamonde-Garcia, C. et al. The mitochondrial inhibitor oligomycin induces an inflammatory response in the rat knee joint. BMC Musculoskelet. Disord. 18, 254 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  61. Xiao, G. et al. cooperative interactions between activating transcription Factor 4 and Runx2/Cbfa1 stimulate osteoblast-specific osteocalcin gene expression. J. Biol. Chem. 280, 30689–30696 (2005).

    Article  CAS  PubMed  Google Scholar 

  62. Lei, Y. et al. LIM domain proteins Pinch1/2 regulate chondrogenesis and bone mass in mice. Bone Res. 8, 37 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors acknowledge the assistance of Core Research Facilities of Southern University of Science and Technology. This work was supported, in part, by the National Key Research and Development Program of China grant (2019YFA0906004), the National Natural Science Foundation of China grants (81991513, 82022047, 81630066, 81870532 and 82972100), the Guangdong Provincial Science and Technology Innovation Council grant (2017B030301018) and the Science and Technology Innovation Commission of Shenzhen Municipal Government grants (JCYJ20180302174246105, JCYJ20180302174117738 and KQJSCX20180319114434843). The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript.

Author information

Author notes

  1. These authors contributed equally: Xiaohao Wu, Yumei Lai, Sheng Chen.

Authors and Affiliations

  1. Department of Biochemistry, School of Medicine, Southern University of Science and Technology, Guangdong Provincial Key Laboratory of Cell Microenvironment and Disease Research Shenzhen Key Laboratory of Cell Microenvironment, Shenzhen, China

    Xiaohao Wu, Chu Tao, Xuekun Fu, Huiling Cao & Guozhi Xiao

  2. Department of Orthopedic Surgery, Rush University Medical Center, Chicago, IL, USA

    Yumei Lai & Jun Li

  3. Department of Orthopedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China

    Sheng Chen, Wei Tong, Hongtao Tian & Zengwu Shao

  4. Department of Medical Laboratory, Tianjin First Center Hospital, Tianjin Medical University, Tianjin, China

    Chunlei Zhou

  5. Department of Orthopedic Surgery, New York University School of Medicine, New York, NY, USA

    Chuanju Liu

  6. Department of Cell Biology, New York University School of Medicine, New York, NY, USA

    Chuanju Liu

  7. Research Center for Human Tissues and Organs Degeneration, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China

    Di Chen

  8. Guangdong Provincial Key Laboratory of Bone and Joint Degeneration Diseases, Department of Cell Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China

    Xiaochun Bai

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Contributions

G.X., X.W., Y.L. and H.C. were responsible for study design. X.W., Y.L., S.C., C.Z., X.F., C.T., J.L., W.T., H.T., X.B. and G.X. were responsible for study conduct and data collection. X.W., Y.L. and G.X. analyzed data. G.X., X.W., C.L., Z.S., X.B. and D.C. interpreted data. G.X. and X.W. drafted the manuscript. X.W., H.C. and G.X. are responsible for the integrity of data analysis.

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Correspondence to Xiaochun Bai, Huiling Cao or Guozhi Xiao.

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Extended data

Extended Data Fig. 1 K2 is highly expressed in mouse articular cartilage.

(a-b) Safranin O & Fast Green (SO&FG) and immunofluorescent (IF) staining of serial sections of mouse knee joint cartilage (a). Green double-headed arrow indicates hyaline cartilage; Red double-headed arrow indicates the calcified cartilage. White dashed lines indicate the cartilage surfaces. Red dashed line indicates the tide mark. Scale bar: 50 μm. Quantification of Kindlin-1-, 2- and -3-positive cells in articular cartilage (b). n = 4 biologically independent animals per group and results from one representative replicate are shown in a. Quantitative data are shown as mean ± standard deviation in b. The exact p values are shown. One-way ANOVA with Tukey’s multiple comparisons was used for statistical analysis.

Source data

Extended Data Fig. 2 Generation and characterizations of K2 cKO mice.

(a) Breeding strategy. (b) PCR genotyping using tail DNA. K2flox KO, ~300bp; K2flox WT, ~200bp; AggrecanCreERT2, ~650bp. (c) Representative images of SO&FG-stained and fluorescent images showing the tdTomato expression in knee joint sections from AggrecanCreERT2; Ai9 mice at 4 weeks after tamoxifen injections. S: synovium; C: cartilage. White dashed lines indicate the cartilage surfaces. Scale bar: 50 μm. (d, e) IF staining for expression of K2 in control and cKO synovium (d). Scale bar: 50 μm. Percentage of K2-positive cells in control and cKO synovium (e). n = 8 mice per group. (f) μCT scans of knee joints. Scale bar, 1.0 mm. n = 6 mice per group. (g) Representative images of Alcian blue/hematoxylin and eosin-stained knee sections. Higher magnification images of the boxed areas (right panels) are shown. Scale bar: 400 μm. n = 6 mice per group. (h-i) OARSI score. n = 6 mice per group. (j, k) Immunohistochemical (IHC) staining. Scale bar: 40 μm. n = 9 mice per group. (l-o) IF staining. Scale bar: 40 μm. n = 10 mice per group. Experiments in b and c were repeated at least three times independently with similar results. Quantitative data are shown as mean ± standard deviation in e, h, i, k, m and o. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Extended Data Fig. 3 Effects of K2 deletion in mouse primary articular chondrocytes.

(a) Isolation of primary articular chondrocytes from adult K2fl/fl; AggrecanCreERT2 male mice. (b-e) Western blotting analyses of expression of K2, p-Stat3/t-Stat3 and Runx2 in primary articular chondrocytes. Primary articular chondrocytes were isolated from 2-month-old K2fl/fl; AggrecanCreERT2 male mice and cultured in 6-well plates. The cells were then treated with 4-hydroxytamoxifen (cKO) and vehicle (control) for 48h. n = 3 biologically independent experiments for each group. (f) IF staining. Scale bar: 40 μm. (g) Representative images of primary articular chondrocytes attachment and spreading on type II collagen-coated surfaces at 6 and 24 h after seeding. Higher magnification images are shown in red dashed boxes. Scale bar: 50 μm. Primary articular chondrocytes were isolated from control and K2 cKO mice at 8 weeks after tamoxifen injections. Experiments in a, b, f and g were repeated at least three times independently with similar results. Quantitative data are shown as mean ± standard deviation in c, d and e. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Extended Data Fig. 4 siRNA knockdown / overexpression of Stat3 and Runx2 in chondrocytes.

(a) Quantitative data for Figure 3e. (b, c) siRNA knockdown of Runx2 and Stat3 in ATDC5 cells. siRNA #1380 (for Runx2) and #1962 (for Stat3) were used for subsequent experiments. (d) Quantitative data for Figure 3r. (e) Quantitative data for Figure 3s. (f) ATDC5 cells were transfected with si-NC or si-Stat3, followed by western blotting for expression of Stat3 and Runx2. (g) ATDC5 cells were transfected with empty vector (EV) and Stat3 expression vector (Stat3), followed by western blotting for expression of t-Stat3, p-Stat3 and Runx2. All experiments were repeated at least three times independently with similar results. Quantitative data are shown as mean ± standard deviation. The exact p values are shown in figures. In a, two-sided unpaired t-test was used for statistical analysis. In d and e, one-way ANOVA with Tukey’s multiple comparisons was used for statistical analysis.

Source data

Extended Data Fig. 5 Immunofluorescent analysis of articular chondrocytes in K2 cKO mice.

(a) IF staining for expression of K2, 9EG7, p-p38, p-Erk and p-Jak2 in articular chondrocytes from control and cKO mice at 6 months after tamoxifen injections. White dashed lines indicate the cartilage areas. Scale bar: 50 μm. (b-d) Percentage of positive-stained cells in control and cKO articular chondrocytes. For all experiments, n = 6 biologically independent animals per group and results from one representative replicate are shown in a. Quantitative data are shown as mean ± standard deviation in b-d. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Extended Data Fig. 6 Systemic inhibition of Stat3 activation limits OA lesions caused by K2 loss.

(a) A schematic diagram illustrating the experimental design. (b) Stattic improves restriction of movement of knee joint caused by K2 loss in DMM mice. (c) SO&FG (left panels), IF (middle panels) and H&E (right panels) staining of knee joint sections from cKO mice treated with PBS or Stattic for 2 months after DMM surgery. Scale bar, 50 μm. (d-g) OARSI score (d), cartilage area (e), osteophyte score (f) and synovitis score (g) were analyzed using histological sections. For all experiments, n = 8 biologically independent animals per group and results from one representative replicate are shown in b, c. Quantitative data are shown as mean ± standard deviation in d-g. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Extended Data Fig. 7 Breeding strategy and PCR genotyping.

(a) Breeding strategy for generating the K2fl/fl; Stat3fl/fl; AggrecanCreERT2 (KSdKO) mice. (b) PCR genotyping using tail DNA. K2flox KO, ~300bp; K2flox WT, ~200bp; Stat3flox KO, 187bp; Stat3flox WT, 146bp; AggrecanCreERT2, ~650bp. Primer sets are listed in Supplementary Table 4. Experiments in b were repeated at least three times independently with similar results.

Extended Data Fig. 8 Effect of Stat3 haploinsufficiency in chondrocytes on cartilage loss caused by K2 loss.

(a) A schematic diagram illustrating the experimental design. (b) SO&FG-stained sections knee joints of K2fl/fl; AggrecanCreERT2 mice and K2fl/fl; Stat3fl/+; AggrecanCreERT2 mice subjected to sham or DMM surgeries and tamoxifen injections as described in (a). Scale bar: 50 μm. (c) Quantification of cartilage loss. n = 8 biologically independent animals per group and results from one representative replicate are shown in b. Quantitative data are shown as mean ± standard deviation in c. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Extended Data Fig. 9 Effect of Runx2 haploinsufficiency in chondrocytes on cartilage loss caused by K2 loss.

(a) A schematic diagram illustrating the experimental design. (b) Alcian blue & orange G-stained sections of knee joints of K2fl/fl; AggrecanCreERT2 mice and K2fl/fl; Runx2fl/+; AggrecanCreERT2 mice subjected to sham or DMM surgeries and tamoxifen injections as described in (a). Scale bar: 50 μm. (c) Quantification of cartilage loss. n = 6 biologically independent animals per group and results from one representative replicate are shown in b. Quantitative data are shown as mean ± standard deviation in c. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Extended Data Fig. 10 Effect of Stat3/Runx2 double haploinsufficiency in chondrocytes on DMM-induced OA lesions.

(a) A schematic diagram illustrating the experimental design. (b) SO&FG-stained sections of the knee joints of Stat3fl/+; AggrecanCreERT2, Runx2fl/+; AggrecanCreERT2 and Stat3fl/+; Runx2fl/+; AggrecanCreERT2 mice subjected to DMM/sham surgery and tamoxifen (TAM)/corn oil injections as described in (a). Scale bar: 50 μm. (c) OARSI score. (d) Loss of cartilage. For all experiments, n = 6 biologically independent animals per group and results from one representative replicate are shown in b. Quantitative data are shown as mean ± standard deviation in c, d. The exact p values are shown in figures. Two-sided unpaired t-test was used for statistical analysis.

Source data

Supplementary information

Source data

Source Data Fig. 1

Statistical source data for Fig. 1.

Source Data Fig. 2

Statistical source data for Fig. 2.

Source Data Fig. 3

Statistical source data for Fig. 3.

Source Data Fig. 3

Unprocessed western blots for Fig. 3.

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Wu, X., Lai, Y., Chen, S. et al. Kindlin-2 preserves integrity of the articular cartilage to protect against osteoarthritis. Nat Aging 2, 332–347 (2022). https://doi.org/10.1038/s43587-021-00165-w

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