Identification of myocilin as a blood plasma protein and analysis of its role in leukocyte adhesion to endothelial cell monolayers

PLoS One. 2018 Dec 17;13(12):e0209364. doi: 10.1371/journal.pone.0209364. eCollection 2018.

Abstract

Myocilin is an extracellular glycoprotein with a poorly understood biological function and typically known because of its association with glaucoma. In this study, we analyzed the expression and biological activity of human myocilin in some non-ocular tissues. Western immunoblot showed the presence of myocilin in blood plasma as well as in liver and lymphoid tissues (thymus and lymph node). Quantitative PCR confirmed the expression of MYOC in these lymphoid organs and revealed that its mRNA is also present in T-lymphocytes and leukocytes. In addition, detection of 30 kDa C-terminal myocilin fragments in thymus and liver suggested that myocilin undergoes an in vivo proteolytic processing that might regulate its biological activity. The presence of myocilin in blood was further corroborated by peptide mass fingerprinting of the HPLC-isolated protein, and gross estimation of its concentration by Western immunoblot indicated that it is a medium-abundance serum protein with an approximate concentration of 0.85 mg/ml (15.5 μM). Finally, in vitro analyses indicated that myocilin acts as an anti-adhesive protein for human circulating leukocytes incubated with endothelial cell monolayers. Altogether, these data provide insightful information on new biological properties of myocilin and suggest its putative role as a blood matricellular protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Aged, 80 and over
  • Blood Proteins / analysis
  • Blood Proteins / physiology*
  • Blotting, Western
  • Cell Adhesion*
  • Cytoskeletal Proteins / blood
  • Cytoskeletal Proteins / physiology*
  • Eye Proteins / blood
  • Eye Proteins / physiology*
  • Female
  • Glycoproteins / blood
  • Glycoproteins / physiology*
  • HEK293 Cells
  • Healthy Volunteers
  • Human Umbilical Vein Endothelial Cells / physiology*
  • Humans
  • Leukocytes / physiology*
  • Liver / metabolism
  • Male
  • Middle Aged
  • Proteolysis
  • RNA, Messenger / analysis
  • Real-Time Polymerase Chain Reaction
  • Thymus Gland / metabolism

Substances

  • Blood Proteins
  • Cytoskeletal Proteins
  • Eye Proteins
  • Glycoproteins
  • RNA, Messenger
  • trabecular meshwork-induced glucocorticoid response protein

Grants and funding

This study has been supported by research grants (to JE) from the Regional Ministry of Science and Technology of the Board of the Communities of “Castilla-La Mancha” (http://www.educa.jccm.es/es, PEII-2014-002-P to J.E.) and the “Instituto de Salud Carlos III/FEDER” (http://www.isciii.es/, RD12/0034/0003, RD16/0008/0019, PI11/00662 and PI15/01193, all to J.E.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.