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115 Plods and lox participate in vascular smooth muscle cell calcification
  1. Eva Jover1,
  2. Ana Silvente2,
  3. Francisco Marín2,
  4. Carmen María Puche2,
  5. Mariano Valdés2,
  6. Diana Hernández-Romero2,
  7. José Martínez-González3,
  8. Mar Orriols4,
  9. Carlos Manuel Martinez4,
  10. Cristina Rodriguez5
  1. 1University of Bristol, Translational Health Sciences, Research Floor Level 7, BRI, Upper Maudlin Street, Bristol, UK
  2. 2Hospital Clínico Universitario Virgen de la Arrixaca, Universidad de Murcia, IMIB-Arrixaca, Spain
  3. 3Instituto de Investigaciones Biomédicas de Barcelona (IIBB-CSIC), IIB-Sant Pau, Barcelona, Spain
  4. 4CIBER de Enfermedades Cardiovasculares (CIBERCV), Spain
  5. 5Institut de Recerca del Hospital de la Santa Creu i Sant Pau-ICCC, IIB-Sant Pau. Barcelona, Spain


Background and aims Vascular smooth muscle cells (VSMC) transdifferentiate into osteoblast-like cells during vascular calcification inducing an active remodelling and calcification of the extracellular matrix (ECM). Intracellular and extracellular enzymes, such as lysyl hydroxylase 1 (PLOD1) or lysyl oxidase (LOX), contribute to ECM maturation and stabilisation towards collagen cross-linking. We aimed to assess the contribution of these enzymes to hyperphosphatemia (HPM)-induced calcification.

Methods and results Human and murine VSMC (hVSMC and mVSMC) were differentiated into functional osteoblast-like cells by HPM conditioning. ECM mineralization was demonstrated by von Kossa and calcium colorimetry or Alizarin Red staining, indicating the presence of calcium phosphates. Osteoblast markers were up-regulated at the transcript and protein levels and that was associated with the early induction of collagen-related genes such as LOX, PLOD1 and collagen type I. In addition, deposition of ECM insoluble collagen was enhanced by HPM. HPM-induced calcification and osteoblast transdifferentiation in mVSMC was limited by chemical inhibition of PLOD (2,2’-dipyridil). Moreover, mVSMC harvested from transgenic mice over-expressing LOX (TgLOX) exhibited an increase in HPM-dependent calcification and osteoblast commitment compared with wild-type cells. HPM-induced calcification was additionally studied ex vivo by using aortic rings harvested from wild-type or TgLOX mice, thus confirming our in vitro results. Therefore, we explored if regulation of PLOD1 and LOX could inhibit hVSMC calcification. Conversely, both β-aminopropionitrile (BAPN; LOX inhibitor), and LOX knockdown abrogated hVSMC calcification and transdifferentiation. Interestingly, deposition of mature collagen was significantly diminished by BAPN as shown by confocal imaging and in agreement with a consistent reduction of cross-linked collagen. The clinical impact of these results was supported by the positive association between the levels of both LOX expression and vascular calcification in human atherosclerotic lesions. Likewise, 2,2’-dipyridil (PLOD inhibitor) and PLOD1 knockdown impaired HPM-induced ECM mineralization and osteoblast commitment.

Conclusions Our findings identify LOX and PLOD as critical players in vascular calcification and highlight the importance of ECM remodelling in this process.

  • vascular calcification
  • extracellular matrix
  • PLOD1
  • LOX
  • BAPN
  • dipyridyl

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