The three isoforms of hepcidin in human serum and their processing determined by liquid chromatography-tandem mass spectrometry (LC-tandem MS)

Abstract

Hepcidin, the iron regulatory hormone, has three isoforms; -20, -22 and -25. While hepcidin-25 has been studied extensively, the physiological significance of other isoforms remains poorly understood. Using a quantitative method based on liquid chromatography-tandem mass spectrometry (LC-tandem MS) developed by our group, we quantified hepcidin isoforms in human serum to elucidate their characteristics, and investigated the role of hepatocytes in isoform processing. Hepcidin isoforms in serum obtained from 40 healthy volunteers were quantified. Synthetic hepcidin peptides were added to healthy serum, and to HepG2 culture media, and hepcidin isoform concentrations determined. All three hepcidin isoforms were detected in human serum; however, hepcidin-25 concentrations were highest. The three hepcidin isoforms showed a strong positive correlation with each other and with serum ferritin. Additionally, while hepcidin-20 was strongly correlated with serum creatinine, the other isoforms were not. Hepcidin-20 and -25 levels were also increased in chronic kidney disease (CKD) serum. Hepcidin-22 rapidly degraded into hepcidin-20, whereas hepcidin-25 remained relatively stable. Finally, hepcidin-22 degradation into hepcidin-20 was accelerated in the presence of HepG2. This method has enabled us to reveal fundamental characteristics of the three hepcidin isoforms in serum and may be a powerful tool for quantifying hepcidin isoform expression and processing.

High pretransplant hepcidin levels are associated with poor overall survival and delayed platelet engraftment after allogeneic hematopoietic stem cell transplantation

“…Because pretransplant serum ferritin levels have been demonstrated to be an independent risk factor for the survival after allo‐HSCT , we compared the significance of pretransplant serum ferritin and hepcidin‐25 levels on the outcome of allo‐HSCT in our cohort. Consistent with previous reports , pretransplant serum hepcidin‐25 levels were positively correlated with levels of serum ferritin ( R = 0.57, P < 0.001, Fig. S1).…”

“…Among these factors, iron strongly induces hepcidin production. Since expression of both hepcidin and ferritin are upregulated by systemic iron overload , it follows that serum hepcidin‐25 levels are positively correlated with those of serum ferritin . Consistently, we observed a positive correlation between pretransplant serum levels of ferritin and hepcidin‐25 (Figure S1).…”

Balance of cardiac and systemic hepcidin and its role in heart physiology and pathology

“…It exists in three main isoforms: hepcidin-25, -22, and -20, of which hepcidin-25 is considered the main physiological actor in iron homeostasis. 5 It is produced by hepatocytes mainly in response to iron-mediated pathways. 6 Iron-mediated pathways induce signaling through bone morphogenetic protein 6 (BMP6), HFE protein, and transferrin receptor 2 (TFR2).…”

Iron Balance and the Role of Hepcidin in Chronic Kidney Disease

“…26 They are much more prominent in urine than in serum, and hepcidin-20 appears to increase out of proportion to total hepcidin in chronic kidney disease. 27,28 Neither the mechanisms by which the smaller hepcidin forms are generated nor their modulation by renal impairment are understood.…”