טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentStuemler Tanya
SubjectThe Effect of Heparanase Overexpression on Iron Metabolism
DepartmentDepartment of Biotechnology and Food Engineering
Supervisor Dr. Esther Meyron Holtz


Abstract

Inappropriately high or low levels of iron can have detrimental effects on the organism. Therefore, several regulators can sense iron and tightly regulate iron homeostasis accordingly. The systemic iron regulator is the peptide hormone hepcidin, which is highly expressed under high iron conditions. Hepcidin transcription is regulated by iron and inflammation. There are two distinct regulatory cascades sensing iron, one is mediated by transferrin, the molecule, which delivers iron to the cells, and the other by Bone morphogenetic protein 6 (BMP6), which reacts to high iron levels.BMP6 binds to the BMP receptor and signals through the phosphorylation of the Sons of Mothers Against Decapentaplegic (SMAD) proteins the last in the cascade then translocates to the nucleus and induces hepcidin transcription.

Recently heparin was identified to interfere with the iron sensing mechanism of hepcidin, mediating a non-iron related mis-regulation of iron homeostasis.  This brought us to hypothesize that heparanase (Hpa); an enzyme, which releases heparin sulfates (HS, endogenous heparin like molecules),may alter iron homeostasis.

Heparan sulfate of proteoglycans (HSPGs), the substrate of Hpa, are molecules, which are present on the extracellular matrix (ECM) and basement membranes (BM), where they facilitate many biological functions and mediate different cell-cell interactions.

Heparin (and its analogues) were found to interfere with BMP signalling through the HSPG/HS pathway. We therefore propose that manipulating Hpa may affect BMP6 binding to its receptor and thus the iron regulating BMP-signalling and iron homeostasis by reducing hepcidin levels.

To elucidate a possible connection between Hpa and iron homeostasis we analysed proteins involved in the BMP6-hepcidin pathway in Hpa overexpressing mice by qPCR and Western blot and compared the iron status of these mice to their normal controls.

Our results indicated that hepcidin levels were indeed reduced in the Hpa overexpressing mice, which may explain the iron overload we observed in the livers of these mice.

Hepcidin levels remained low despite of high BMP6, which is elevated due to the high liver iron. Elevated BMP6 should have induced Hepcidin expression, but did not, suggesting a reduced sensitivity to BMP6 of the BMP-pSMAD pathway induced hepcidin transcription. Over time BMP6 levels decreased despite the constantly rising iron levels, which may be a sign for hepatocyte-exhaustion in BMP6 production. Taken together, we propose a mechanism where BMP6 resistance initially is corrected by vast BMP6 overexpression, but eventually hepatocytes' capability of BMP6 production is diminished and the misregulation of iron homeostasis is not overcome.

Hpa and iron both being key players in inflammation and cancer onset and aggravation suggests that this likely interplay between the two warrants deeper understanding and investigation.