Insulin-like growth factor (IGF)-II- mediated fibrosis in pathogenic lung conditions

Autoři: Sara M. Garrett aff001;  Eileen Hsu aff002;  Justin M. Thomas aff003;  Joseph M. Pilewski aff004;  Carol Feghali-Bostwick aff001
Působiště autorů: Division of Rheumatology, Department of Medicine, Medical University of South Carolina (MUSC), Charleston, South Carolina, United States of America aff001;  Mid Atlantic Permanente Medical Group, Mclean, Virginia, United States of America aff002;  Eisenhower Medical Center, Rancho Mirage, California, United States of America aff003;  Division of Pulmonary, Allergy, and Critical Care Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America aff004
Vyšlo v časopise: PLoS ONE 14(11)
Kategorie: Research Article
doi: 10.1371/journal.pone.0225422


Type 2 insulin-like growth factor (IGF-II) levels are increased in fibrosing lung diseases such as idiopathic pulmonary fibrosis (IPF) and scleroderma/systemic sclerosis-associated pulmonary fibrosis (SSc). Our goal was to investigate the contribution of IGF receptors to IGF-II-mediated fibrosis in these diseases and identify other potential mechanisms key to the fibrotic process. Cognate receptor gene and protein expression were analyzed with qRT-PCR and immunoblot in primary fibroblasts derived from lung tissues of normal donors (NL) and patients with IPF or SSc. Compared to NL, steady-state receptor gene expression was decreased in SSc but not in IPF. IGF-II stimulation differentially decreased receptor mRNA and protein levels in NL, IPF, and SSc fibroblasts. Neutralizing antibody, siRNA, and receptor inhibition targeting endogenous IGF-II and its primary receptors, type 1 IGF receptor (IGF1R), IGF2R, and insulin receptor (IR) resulted in loss of the IGF-II response. IGF-II tipped the TIMP:MMP balance, promoting a fibrotic environment both intracellularly and extracellularly. Differentiation of fibroblasts into myofibroblasts by IGF-II was blocked with a TGFβ1 receptor inhibitor. IGF-II also increased TGFβ2 and TGFβ3 expression, with subsequent activation of canonical SMAD2/3 signaling. Therefore, IGF-II promoted fibrosis through IGF1R, IR, and IGF1R/IR, differentiated fibroblasts into myofibroblasts, decreased protease production and extracellular matrix degradation, and stimulated expression of two TGFβ isoforms, suggesting that IGF-II exerts pro-fibrotic effects via multiple mechanisms.

Klíčová slova:

Collagens – Fibroblasts – Fibrosis – Gene expression – Protein expression – Pulmonary fibrosis – Small interfering RNAs – TGF-beta signaling cascade


1. Raghu G, Chen SY, Hou Q, Yeh WS, Collard HR. Incidence and prevalence of idiopathic pulmonary fibrosis in US adults 18–64 years old. Eur Respir J. 2016;48(1):179–86. doi: 10.1183/13993003.01653-2015 27126689.

2. Herzog EL, Mathur A, Tager AM, Feghali-Bostwick C, Schneider F, Varga J. Review: interstitial lung disease associated with systemic sclerosis and idiopathic pulmonary fibrosis: how similar and distinct? Arthritis Rheumatol. 2014;66(8):1967–78. doi: 10.1002/art.38702 24838199.

3. Martyanov V, Kim GJ, Hayes W, Du S, Ganguly BJ, Sy O, et al. Novel lung imaging biomarkers and skin gene expression subsetting in dasatinib treatment of systemic sclerosis-associated interstitial lung disease. PLoS One. 2017;12(11):e0187580. doi: 10.1371/journal.pone.0187580 29121645.

4. Hsu E, Shi H, Jordan RM, Lyons-Weiler J, Pilewski JM, Feghali-Bostwick CA. Lung tissues in patients with systemic sclerosis have gene expression patterns unique to pulmonary fibrosis and pulmonary hypertension. Arthritis Rheum. 2011;63(3):783–94. doi: 10.1002/art.30159 21360508.

5. Garrett SM, Baker Frost D, Feghali-Bostwick C. The mighty fibroblast and its utility in scleroderma research. J Scleroderma Relat Disord. 2017;2(2):69–134. doi: 10.5301/jsrd.5000240 29270465.

6. O’Dwyer DN, Ashley SL, Moore BB. Influences of innate immunity, autophagy, and fibroblast activation in the pathogenesis of lung fibrosis. Am J Physiol Lung Cell Mol Physiol. 2016;311(3):L590–601. doi: 10.1152/ajplung.00221.2016 27474089.

7. Harrison NK, Cambrey AD, Myers AR, Southcott AM, Black CM, du Bois RM, et al. Insulin-like growth factor-I is partially responsible for fibroblast proliferation induced by bronchoalveolar lavage fluid from patients with systemic sclerosis. Clin Sci (Lond). 1994;86(2):141–8. doi: 10.1042/cs0860141 8143424.

8. Pilewski JM, Liu L, Henry AC, Knauer AV, Feghali-Bostwick CA. Insulin-like growth factor binding proteins 3 and 5 are overexpressed in idiopathic pulmonary fibrosis and contribute to extracellular matrix deposition. Am J Pathol. 2005;166(2):399–407. doi: 10.1016/S0002-9440(10)62263-8 15681824.

9. Yasuoka H, Zhou Z, Pilewski JM, Oury TD, Choi AM, Feghali-Bostwick CA. Insulin-like growth factor-binding protein-5 induces pulmonary fibrosis and triggers mononuclear cellular infiltration. Am J Pathol. 2006;169(5):1633–42. doi: 10.2353/ajpath.2006.060501 17071587.

10. Yasuoka H, Jukic DM, Zhou Z, Choi AM, Feghali-Bostwick CA. Insulin-like growth factor binding protein 5 induces skin fibrosis: A novel murine model for dermal fibrosis. Arthritis Rheum. 2006;54(9):3001–10. doi: 10.1002/art.22084 16947625.

11. Yasuoka H, Garrett SM, Nguyen XX, Artlett CM, Feghali-Bostwick CA. NADPH oxidase-mediated induction of reactive oxygen species and extracellular matrix deposition by insulin-like growth factor binding protein-5. Am J Physiol Lung Cell Mol Physiol. 2019;316(4):L644–L55. doi: 10.1152/ajplung.00106.2018 30810066.

12. Hsu E, Feghali-Bostwick CA. Insulin-like growth factor-II is increased in systemic sclerosis-associated pulmonary fibrosis and contributes to the fibrotic process via Jun N-terminal kinase- and phosphatidylinositol-3 kinase-dependent pathways. Am J Pathol. 2008;172(6):1580–90. doi: 10.2353/ajpath.2008.071021 18467708.

13. Nielsen FC. The molecular and cellular biology of insulin-like growth factor II. Prog Growth Factor Res. 1992;4(3):257–90. doi: 10.1016/0955-2235(92)90023-b 1307492.

14. Gallagher EJ, LeRoith D. Minireview: IGF, Insulin, and Cancer. Endocrinology. 2011;152(7):2546–51. doi: 10.1210/en.2011-0231 21540285.

15. Halje M, Nordin M, Bergman D, Engstrom W. Review: The effect of insulin-like growth factor II in the regulation of tumour cell growth in vitro and tumourigenesis in vivo. In Vivo. 2012;26(4):519–26. 22773563.

16. Bergman D, Halje M, Nordin M, Engstrom W. Insulin-like growth factor 2 in development and disease: a mini-review. Gerontology. 2013;59(3):240–9. doi: 10.1159/000343995 23257688.

17. Alvino CL, Ong SC, McNeil KA, Delaine C, Booker GW, Wallace JC, et al. Understanding the mechanism of insulin and insulin-like growth factor (IGF) receptor activation by IGF-II. PLoS One. 2011;6(11):e27488. doi: 10.1371/journal.pone.0027488 22140443.

18. Back K, Brannmark C, Stralfors P, Arnqvist HJ. Differential effects of IGF-I, IGF-II and insulin in human preadipocytes and adipocytes—role of insulin and IGF-I receptors. Mol Cell Endocrinol. 2011;339(1–2):130–5. doi: 10.1016/j.mce.2011.04.005 21524684.

19. Krizkova K, Chrudinova M, Povalova A, Selicharova I, Collinsova M, Vanek V, et al. Insulin-Insulin-like Growth Factors Hybrids as Molecular Probes of Hormone:Receptor Binding Specificity. Biochemistry. 2016;55(21):2903–13. doi: 10.1021/acs.biochem.6b00140 27171135.

20. Pandini G, Frasca F, Mineo R, Sciacca L, Vigneri R, Belfiore A. Insulin/insulin-like growth factor I hybrid receptors have different biological characteristics depending on the insulin receptor isoform involved. J Biol Chem. 2002;277(42):39684–95. doi: 10.1074/jbc.M202766200 12138094.

21. Scott CD, Kiess W. Soluble M6P/IGFIIR in the circulation. Best Pract Res Clin Endocrinol Metab. 2015;29(5):723–33. doi: 10.1016/j.beem.2015.08.001 26522457.

22. Garrett SM, Zhao Q, Feghali-Bostwick C. Induction of a Th17 Phenotype in Human Skin-A Mimic of Dermal Inflammatory Diseases. Methods Protoc. 2019;2(2). doi: 10.3390/mps2020045 31164624.

23. Hetzel M, Bachem M, Anders D, Trischler G, Faehling M. Different effects of growth factors on proliferation and matrix production of normal and fibrotic human lung fibroblasts. Lung. 2005;183(4):225–37. doi: 10.1007/s00408-004-2534-z 16211459.

24. Pardo A, Selman M. Matrix metalloproteases in aberrant fibrotic tissue remodeling. Proc Am Thorac Soc. 2006;3(4):383–8. doi: 10.1513/pats.200601-012TK 16738205.

25. Menou A, Duitman J, Crestani B. The impaired proteases and anti-proteases balance in Idiopathic Pulmonary Fibrosis. Matrix Biol. 2018;68–69:382–403. doi: 10.1016/j.matbio.2018.03.001 29518524.

26. Murphy G. Tissue inhibitors of metalloproteinases. Genome Biol. 2011;12(11):233. doi: 10.1186/gb-2011-12-11-233 22078297.

27. Khalil N, O’Connor R, Unruh H, Warren P, Kemp A, Greenberg A. Enhanced expression and immunohistochemical distribution of transforming growth factor-beta in idiopathic pulmonary fibrosis. Chest. 1991;99(3 Suppl):65S–6S. doi: 10.1378/chest.99.3_supplement.65s-a 1997280.

28. Broekelmann TJ, Limper AH, Colby TV, McDonald JA. Transforming growth factor beta 1 is present at sites of extracellular matrix gene expression in human pulmonary fibrosis. Proc Natl Acad Sci U S A. 1991;88(15):6642–6. doi: 10.1073/pnas.88.15.6642 1862087.

29. Dantas AT, Goncalves SM, de Almeida AR, Goncalves RS, Sampaio MC, Vilar KM, et al. Reassessing the Role of the Active TGF-beta1 as a Biomarker in Systemic Sclerosis: Association of Serum Levels with Clinical Manifestations. Dis Markers. 2016;2016:6064830. doi: 10.1155/2016/6064830 27965520.

30. Jelaska A, Korn JH. Role of apoptosis and transforming growth factor beta1 in fibroblast selection and activation in systemic sclerosis. Arthritis Rheum. 2000;43(10):2230–9. doi: 10.1002/1529-0131(200010)43:10<2230::AID-ANR10>3.0.CO;2-8 11037882.

31. Louvi A, Accili D, Efstratiadis A. Growth-promoting interaction of IGF-II with the insulin receptor during mouse embryonic development. Dev Biol. 1997;189(1):33–48. doi: 10.1006/dbio.1997.8666 9281335.

32. Zaina S, Newton RV, Paul MR, Graham CF. Local reduction of organ size in transgenic mice expressing a soluble insulin-like growth factor II/mannose-6-phosphate receptor. Endocrinology. 1998;139(9):3886–95. doi: 10.1210/endo.139.9.6200 9724044.

33. Brown J, Jones EY, Forbes BE. Interactions of IGF-II with the IGF2R/cation-independent mannose-6-phosphate receptor mechanism and biological outcomes. Vitam Horm. 2009;80:699–719. doi: 10.1016/S0083-6729(08)00625-0 19251056.

34. Nielsen FC, Ostergaard L, Nielsen J, Christiansen J. Growth-dependent translation of IGF-II mRNA by a rapamycin-sensitive pathway. Nature. 1995;377(6547):358–62. doi: 10.1038/377358a0 7566093.

35. Osipo C, Dorman S, Frankfater A. Loss of insulin-like growth factor II receptor expression promotes growth in cancer by increasing intracellular signaling from both IGF-I and insulin receptors. Exp Cell Res. 2001;264(2):388–96. doi: 10.1006/excr.2000.5121 11262195.

36. Wang ZQ, Fung MR, Barlow DP, Wagner EF. Regulation of embryonic growth and lysosomal targeting by the imprinted Igf2/Mpr gene. Nature. 1994;372(6505):464–7. doi: 10.1038/372464a0 7984240.

37. Lau MM, Stewart CE, Liu Z, Bhatt H, Rotwein P, Stewart CL. Loss of the imprinted IGF2/cation-independent mannose 6-phosphate receptor results in fetal overgrowth and perinatal lethality. Genes Dev. 1994;8(24):2953–63. doi: 10.1101/gad.8.24.2953 8001817.

38. Ludwig T, Eggenschwiler J, Fisher P, D’Ercole AJ, Davenport ML, Efstratiadis A. Mouse mutants lacking the type 2 IGF receptor (IGF2R) are rescued from perinatal lethality in Igf2 and Igf1r null backgrounds. Dev Biol. 1996;177(2):517–35. doi: 10.1006/dbio.1996.0182 8806828.

39. Li Y, Meng G, Huang L, Guo QN. Hypomethylation of the P3 promoter is associated with up-regulation of IGF2 expression in human osteosarcoma. Hum Pathol. 2009;40(10):1441–7. doi: 10.1016/j.humpath.2009.03.003 19427670.

40. Tarrago D, Aguilera I, Melero J, Wichmann I, Nunez-Roldan A, Sanchez B. Identification of cation-independent mannose 6-phosphate receptor/insulin-like growth factor type-2 receptor as a novel target of autoantibodies. Immunology. 1999;98(4):652–62. doi: 10.1046/j.1365-2567.1999.00889.x 10594701.

41. Slaaby R, Schaffer L, Lautrup-Larsen I, Andersen AS, Shaw AC, Mathiasen IS, et al. Hybrid receptors formed by insulin receptor (IR) and insulin-like growth factor I receptor (IGF-IR) have low insulin and high IGF-1 affinity irrespective of the IR splice variant. J Biol Chem. 2006;281(36):25869–74. doi: 10.1074/jbc.M605189200 16831875.

42. Siddle K, Urso B, Niesler CA, Cope DL, Molina L, Surinya KH, et al. Specificity in ligand binding and intracellular signalling by insulin and insulin-like growth factor receptors. Biochem Soc Trans. 2001;29(Pt 4):513–25. doi: 10.1042/bst0290513 11498020.

43. Keppler S, Weibetabach S, Langer C, Knop S, Pischimarov J, Kull M, et al. Rare SNPs in receptor tyrosine kinases are negative outcome predictors in multiple myeloma. Oncotarget. 2016;7(25):38762–74. doi: 10.18632/oncotarget.9607 27246973.

44. Liu TC, Hsieh MJ, Liu MC, Chiang WL, Tsao TC, Yang SF. The Clinical Significance of the Insulin-Like Growth Factor-1 Receptor Polymorphism in Non-Small-Cell Lung Cancer with Epidermal Growth Factor Receptor Mutation. Int J Mol Sci. 2016;17(5). doi: 10.3390/ijms17050763 27213344.

45. Pineiro-Hermida S, Lopez IP, Alfaro-Arnedo E, Torrens R, Iniguez M, Alvarez-Erviti L, et al. IGF1R deficiency attenuates acute inflammatory response in a bleomycin-induced lung injury mouse model. Sci Rep. 2017;7(1):4290. doi: 10.1038/s41598-017-04561-4 28655914.

46. Choi JE, Lee SS, Sunde DA, Huizar I, Haugk KL, Thannickal VJ, et al. Insulin-like growth factor-I receptor blockade improves outcome in mouse model of lung injury. Am J Respir Crit Care Med. 2009;179(3):212–9. doi: 10.1164/rccm.200802-228OC 19011156.

47. Alberts B, Johnson A, Lewis J. Molecular Biology of the Cell: The Extracellular Matrix of Animals. 4th ed. New York: Garland Science; 2002.

48. Kim SY, Lee JH, Kim HJ, Park MK, Huh JW, Ro JY, et al. Mesenchymal stem cell-conditioned media recovers lung fibroblasts from cigarette smoke-induced damage. Am J Physiol Lung Cell Mol Physiol. 2012;302(9):L891–908. doi: 10.1152/ajplung.00288.2011 22307909.

49. Park AM, Hayakawa S, Honda E, Mine Y, Yoshida K, Munakata H. Conditioned media from lung cancer cell line A549 and PC9 inactivate pulmonary fibroblasts by regulating protein phosphorylation. Arch Biochem Biophys. 2012;518(2):133–41. doi: 10.1016/ 22209754.

50. Page-McCaw A, Ewald AJ, Werb Z. Matrix metalloproteinases and the regulation of tissue remodelling. Nat Rev Mol Cell Biol. 2007;8(3):221–33. doi: 10.1038/nrm2125 17318226.

51. Fowlkes JL, Thrailkill KM, Serra DM, Suzuki K, Nagase H. Matrix metalloproteinases as insulin-like growth factor binding protein-degrading proteinases. Prog Growth Factor Res. 1995;6(2–4):255–63. doi: 10.1016/0955-2235(95)00017-8 8817668.

52. Craig VJ, Zhang L, Hagood JS, Owen CA. Matrix metalloproteinases as therapeutic targets for idiopathic pulmonary fibrosis. Am J Respir Cell Mol Biol. 2015;53(5):585–600. doi: 10.1165/rcmb.2015-0020TR 26121236.

53. Yamashita CM, Radisky DC, Aschner Y, Downey GP. The importance of matrix metalloproteinase-3 in respiratory disorders. Expert Rev Respir Med. 2014;8(4):411–21. doi: 10.1586/17476348.2014.909288 24869454.

54. Bullard KM, Lund L, Mudgett JS, Mellin TN, Hunt TK, Murphy B, et al. Impaired wound contraction in stromelysin-1-deficient mice. Ann Surg. 1999;230(2):260–5. doi: 10.1097/00000658-199908000-00017 10450741.

55. Bullard KM, Mudgett J, Scheuenstuhl H, Hunt TK, Banda MJ. Stromelysin-1-deficient fibroblasts display impaired contraction in vitro. J Surg Res. 1999;84(1):31–4. doi: 10.1006/jsre.1999.5599 10334885.

56. Arpino V, Brock M, Gill SE. The role of TIMPs in regulation of extracellular matrix proteolysis. Matrix Biol. 2015;44–46:247–54. doi: 10.1016/j.matbio.2015.03.005 25805621.

57. Madtes DK, Elston AL, Kaback LA, Clark JG. Selective induction of tissue inhibitor of metalloproteinase-1 in bleomycin-induced pulmonary fibrosis. Am J Respir Cell Mol Biol. 2001;24(5):599–607. doi: 10.1165/ajrcmb.24.5.4192 11350830.

58. Lee MH, Atkinson S, Murphy G. Identification of the extracellular matrix (ECM) binding motifs of tissue inhibitor of metalloproteinases (TIMP)-3 and effective transfer to TIMP-1. J Biol Chem. 2007;282(9):6887–98. doi: 10.1074/jbc.M610490200 17202148.

59. Brew K, Nagase H. The tissue inhibitors of metalloproteinases (TIMPs): an ancient family with structural and functional diversity. Biochim Biophys Acta. 2010;1803(1):55–71. doi: 10.1016/j.bbamcr.2010.01.003 20080133.

60. Zhang Q, Guo Y, Dong R, Dai R, Zhou M. Suppressor of cytokine signaling 1-modulated metalloproteinases and tissue inhibitor of metalloproteinase in pulmonary fibrosis. Mol Med Rep. 2015;12(3):3855–61. doi: 10.3892/mmr.2015.3810 25997387.

61. Melendez-Zajgla J, Del Pozo L, Ceballos G, Maldonado V. Tissue inhibitor of metalloproteinases-4. The road less traveled. Mol Cancer. 2008;7:85. doi: 10.1186/1476-4598-7-85 19025595.

62. Kubo H. Extracellular Vesicles in Lung Disease. Chest. 2018;153(1):210–6. doi: 10.1016/j.chest.2017.06.026 28684288.

63. Chung B, Hinek A, Keating S, Weksberg R, Shah V, Blaser S, et al. Overgrowth with increased proliferation of fibroblast and matrix metalloproteinase activity related to reduced TIMP1: a newly recognized syndrome? Am J Med Genet A. 2012;158A(10):2373–81. doi: 10.1002/ajmg.a.35570 22965799.

64. Rogers MA, Kalter V, Strowitzki M, Schneider M, Lichter P. IGF2 knockdown in two colorectal cancer cell lines decreases survival, adhesion and modulates survival-associated genes. Tumour Biol. 2016;37(9):12485–95. doi: 10.1007/s13277-016-5115-x 27337954.

65. Vogel C, Marcotte EM. Insights into the regulation of protein abundance from proteomic and transcriptomic analyses. Nat Rev Genet. 2012;13(4):227–32. doi: 10.1038/nrg3185 22411467.

66. Grotendorst GR, Rahmanie H, Duncan MR. Combinatorial signaling pathways determine fibroblast proliferation and myofibroblast differentiation. FASEB J. 2004;18(3):469–79. doi: 10.1096/fj.03-0699com 15003992.

67. Talior-Volodarsky I, Arora PD, Wang Y, Zeltz C, Connelly KA, Gullberg D, et al. Glycated Collagen Induces alpha11 Integrin Expression Through TGF-beta2 and Smad3. J Cell Physiol. 2015;230(2):327–36. doi: 10.1002/jcp.24708 24962729.

68. Shin JY, Beckett JD, Bagirzadeh R, Creamer TJ, Shah AA, McMahan Z, et al. Epigenetic activation and memory at a TGFB2 enhancer in systemic sclerosis. Sci Transl Med. 2019;11(497). doi: 10.1126/scitranslmed.aaw0790 31217334.

69. Bukowska J, Kopcewicz M, Kur-Piotrowska A, Szostek-Mioduchowska AZ, Walendzik K, Gawronska-Kozak B. Effect of TGFbeta1, TGFbeta3 and keratinocyte conditioned media on functional characteristics of dermal fibroblasts derived from reparative (Balb/c) and regenerative (Foxn1 deficient; nude) mouse models. Cell Tissue Res. 2018;374(1):149–63. doi: 10.1007/s00441-018-2836-8 29637306.

70. Susol E, Rands AL, Herrick A, McHugh N, Barrett JH, Ollier WE, et al. Association of markers for TGFbeta3, TGFbeta2 and TIMP1 with systemic sclerosis. Rheumatology (Oxford). 2000;39(12):1332–6. doi: 10.1093/rheumatology/39.12.1332 11136875.

Článek vyšel v časopise


2019 Číslo 11