組織間隙纖維化為慢性腎衰竭常見的特徵，其成因為過度活化的肌成纖維細胞造成細胞外基質堆積及交互連結於組織間隙中。組織纖維化是一種膠原蛋白失去動態平衡的疾病，其中包含了膠原蛋白束的排列、膠原蛋白的製造及膠原蛋白的降解。Discoidin 結構域接受器1型 (DDR1)是酪胺酸激酶的膠原蛋白接受器，其表現量會在由單側輸尿管結紮所誘導的腎臟纖維化中高度表達。起初，我們發現TGF-β1會增加DDR1在NRK-49F細胞的表現量，為了探討DDR1是否或如何參與在膠原蛋白的動態平衡中，我們建立了DDR1基因減弱在NRK-49F的穩定細胞株。我們發現DDR1基因減弱並不會影響TGF-β1所誘導的α-平滑肌肌動蛋白表現量，其為肌成纖維細胞活化的指標蛋白，另一方面，我們將NRK-49F細胞種在膠原蛋白膠裡面去觀察細胞收縮力，也沒有得到有結論性的結果，然而有趣的是，我們觀察到DDR1基因減弱會促使強烈的膠原蛋白降解，除此之外，DDR1基因減弱會促進足體的形成，而足體是一種聚集的F-肌動蛋白且有強力降解細胞外基質的結構，免疫螢光染色的結果也顯示了MMP2 有位在DDR1基因減弱所誘導的足體中，再者，我們發現了隨著NRK-49F細胞培養的時間越久，應力纖維的形成會增加而足體的形成會減少，暗示著應力纖維與足體存在著互相排斥的現象，而DDR1基因減弱會降低點狀黏著點的長度及大小，似乎與增加足體的形成有關係。總體來說，我們發現DDR1抑制了足體的形成和膠原蛋白的降解，然而其中的機制仍需要進一步去研究。

The common feature of chronic kidney disease (CKD) is interstitial fibrosis, as reflected by aberrant extracellular matrix (ECM) deposition and crosslinking in the interstitium due to the myofibroblast activation. Fibrosis is the imbalance of collagen homeostasis, including the collagen fiber organization, collagen production and collagen degradation. Discoidin domain receptor 1 (DDR1), one of the two collagen receptors with tyrosine kinase, was upregulated in unilateral ureteral obstruction-induced renal fibrosis. In in vitro study, we found that TGF-β1 induced upregulation of DDR1 in NRK49F cells. In order to investigate whether and how DDR1 involves in the collagen homeostasis, we established stable clones of NRK49F cells harboring short hairpin DNAi of DDR1. We found that knockdown of DDR1 did not affect TGF-β1-induced α-smooth muscle actin expression, a typical marker of myofibroblast. On the other hand, experiments by culturing NRK49F cells in collagen gel could not provide conclusive results in terms of cell contractility. Interestingly, we found that knockdown of DDR1 markedly enhanced collagen degradation. In addition, knockdown of DDR1 promoted the formation of podosome, an F-actin rich structure to actively digest extracellular matrix. Immunofluorescence studies further confirmed that MMP-2 was localized in podosome in DDR1 knockdown clone. We found the podosome structure was gradually lost with time by plating cells on culture dish from 4 to 24 hours, meanwhile cells start to spread out and developed stress fibers, which supported mutual exclusive effect of stress fiber and podosome formation. Knockdown of DDR1 reduced the size and length of focal adhesion, which may be associated with the increasing of podosome formation. In summary, we demonstrated that DDR1 inhibits the podosome formation and collagen digestion. However, the mechanism whereby DDR1 inhibits podosome formation remains to be further investigated.

1. Chen, S.J., et al., Stimulation of type I collagen transcription in human skin fibroblasts by TGF-beta: involvement of Smad 3. J Invest Dermatol, 1999. 112(1): p. 49-57.
2. Georges, P.C., et al., Increased stiffness of the rat liver precedes matrix deposition: implications for fibrosis. Am J Physiol Gastrointest Liver Physiol, 2007. 293(6): p. G1147-54.
3. Yuan, Q., R.J. Tan, and Y. Liu, Myofibroblast in kidney fibrosis: origin, activation, and regulation. Adv Exp Med Biol, 2019. 1165: p. 253-283.
4. Kuppe, C., et al., Decoding myofibroblast origins in human kidney fibrosis. Nature, 2021. 589(7841): p. 281-286.
5. Hynes, R.O., Integrins: bidirectional, allosteric signaling machines. Cell, 2002. 110(6): p. 673-87.
6. Hamidi, H., M. Pietilä, and J. Ivaska, The complexity of integrins in cancer and new scopes for therapeutic targeting. Br J Cancer, 2016. 115(9): p. 1017-1023.
7. Yeh, Y.C., et al., Transforming growth factor-β1 induces Smad3-dependent β1 integrin gene expression in epithelial-to-mesenchymal transition during chronic tubulointerstitial fibrosis. Am J Pathol, 2010. 177(4): p. 1743-54.
8. Vogel, W., et al., The discoidin domain receptor tyrosine kinases are activated by collagen. Mol Cell, 1997. 1(1): p. 13-23.
9. Alves, F., et al., Distinct structural characteristics of discoidin I subfamily receptor tyrosine kinases and complementary expression in human cancer. Oncogene, 1995. 10(3): p. 609-18.
10. Faraci-Orf, E., C. McFadden, and W.F. Vogel, DDR1 signaling is essential to sustain Stat5 function during lactogenesis. J Cell Biochem, 2006. 97(1): p. 109-21.
11. Avivi-Green, C., M. Singal, and W.F. Vogel, Discoidin domain receptor 1-deficient mice are resistant to bleomycin-induced lung fibrosis. Am J Respir Crit Care Med, 2006. 174(4): p. 420-7.
12. Nokin, M.J., et al., Inhibition of DDR1 enhances in vivo chemosensitivity in KRAS-mutant lung adenocarcinoma. JCI Insight, 2020. 5(15).
13. Quan, J., et al., Identification of receptor tyrosine kinase, discoidin domain receptor 1 (DDR1), as a potential biomarker for serous ovarian cancer. Int J Mol Sci, 2011. 12(2): p. 971-82.
14. Guerrot, D., et al., Discoidin domain receptor 1 is a major mediator of inflammation and fibrosis in obstructive nephropathy. Am J Pathol, 2011. 179(1): p. 83-91.
15. Chiusa, M., et al., The extracellular matrix receptor Discoidin domain receptor 1 regulates collagen transcription by translocating to the nucleus. J Am Soc Nephrol, 2019. 30(9): p. 1605-1624.
16. Ge, C., et al., Discoidin receptor 2 controls bone formation and marrow adipogenesis. J Bone Miner Res, 2016. 31(12): p. 2193-2203.
17. Olaso, E., et al., Impaired dermal wound healing in discoidin domain receptor 2-deficient mice associated with defective extracellular matrix remodeling. Fibrogenesis Tissue Repair, 2011. 4(1): p. 5.
18. Ren, T., et al., Discoidin domain receptor 2 (DDR2) promotes breast cancer cell metastasis and the mechanism implicates epithelial-mesenchymal transition programme under hypoxia. J Pathol, 2014. 234(4): p. 526-37.
19. Li, X., et al., Deletion of discoidin domain receptor 2 attenuates renal interstitial fibrosis in a murine unilateral ureteral obstruction model. Ren Fail, 2019. 41(1): p. 481-488.
20. Tarone, G., et al., Rous sarcoma virus-transformed fibroblasts adhere primarily at discrete protrusions of the ventral membrane called podosomes. Exp Cell Res, 1985. 159(1): p. 141-57.
21. Kuo, S.L., et al., Biogenesis of podosome rosettes through fission. Sci Rep, 2018. 8(1): p. 524.
22. Cougoule, C., et al., Three-dimensional migration of macrophages requires Hck for podosome organization and extracellular matrix proteolysis. Blood, 2010. 115(7): p. 1444-52.
23. Rottiers, P., et al., TGFbeta-induced endothelial podosomes mediate basement membrane collagen degradation in arterial vessels. J Cell Sci, 2009. 122(Pt 23): p. 4311-8.
24. Zhang, Y., et al., Autophagy promotes osteoclast podosome disassembly and cell motility athrough the interaction of kindlin3 with LC3. Cell Signal, 2020. 67: p. 109505.
25. Hammoud, A.A., et al., DDR2 induces linear invadosome to promote angiogenesis in a fibrillar type I collagen context. bioRxiv, 2020: p. 2020.04.11.036764.
26. Bayer, S.V., et al., DDR2 controls breast tumor stiffness and metastasis by regulating integrin mediated mechanotransduction in CAFs. Elife, 2019. 8.
27. Staudinger, L.A., et al., Interactions between the discoidin domain receptor 1 and β1 integrin regulate attachment to collagen. Biol Open, 2013. 2(11): p. 1148-59.