Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The . gov means it’s official. Federal government websites often end in . gov or . mil VSports app下载. Before sharing sensitive information, make sure you’re on a federal government site. .

Https

The site is secure. The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely. V体育官网.

. 2010 Nov 4;116(18):3635-44.
doi: 10.1182/blood-2010-03-274571. Epub 2010 Aug 2.

In anemia of multiple myeloma, hepcidin is induced by increased bone morphogenetic protein 2

Ken Maes  1 , Elizabeta Nemeth, G David Roodman, Alissa Huston, Flavia Esteve, Cesar Freytes, Natalie Callander, Eirini Katodritou, Lisa Tussing-Humphreys, Seth Rivera, Karin Vanderkerken, Alan Lichtenstein, Tomas Ganz
Affiliations

In anemia of multiple myeloma, hepcidin is induced by increased bone morphogenetic protein 2

Ken Maes et al. Blood. .

"V体育安卓版" Abstract

Hepcidin is the principal iron-regulatory hormone and a pathogenic factor in anemia of inflammation. Patients with multiple myeloma (MM) frequently present with anemia. We showed that MM patients had increased serum hepcidin, which inversely correlated with hemoglobin, suggesting that hepcidin contributes to MM-related anemia VSports手机版. Searching for hepcidin-inducing cytokines in MM, we quantified the stimulation of hepcidin promoter-luciferase activity in HuH7 cells by MM sera. MM sera activated the hepcidin promoter significantly more than did normal sera. We then examined the role of bone morphogenetic proteins (BMPs) and interleukin-6 (IL-6), the major transcriptional regulators of hepcidin. Mutations in both BMP-responsive elements abrogated the activation dramatically, while mutations in the IL-6-responsive signal transducer and activator of transcription 3-binding site (STAT3-BS) had only a minor effect. Cotreatment with anti-BMP-2/4 or noggin-Fc blocked the promoter induction with all MM sera, anti-IL-6 blocked it with a minority of sera, whereas anti-BMP-4, -6, or -9 antibodies had no effect. BMP-2-immunodepleted MM sera had decreased promoter stimulatory capacity, and BMP-2 concentrations in MM sera were significantly higher than in normal sera. Our results demonstrate that BMP-2 is a major mediator of the hepcidin stimulatory activity of MM sera. .

PubMed Disclaimer

Figures

Figure 1
Figure 1
Correlation between serum hepcidin levels and hemoglobin concentrations. We used 25 MM patient serum samples for which hemoglobin content and serum hepcidin concentration were previously measured.,, (A) Serum hepcidin levels in MM patients (n = 25) were significantly higher than in healthy individuals (n = 15). Boxes represent median and 25-75 percentiles, and whiskers represent 10 and 90 percentiles. Circles are the outliers. Statistical significance was determined with the Mann-Whitney rank-sum test; *P < .01, compared with normal serum. (B) Patients with normal renal function (n = 16) were included to investigate the relationship between serum hepcidin levels and hemoglobin concentrations. Hemoglobin concentrations and serum hepcidin inversely correlate (R2 = 0.4277, P = .006, Pearson correlation).
Figure 2
Figure 2
Scheme of mutations introduced in HAMP promoter. Boxed areas are the putative transcription factor binding sites for BMP and IL-6 pathways. Mutated nucleotides are shown in bold capitals. Mutations in BRE1 and -2 were previously described in Island et al and the STAT3-BS mutation in Truksa et al.
Figure 3
Figure 3
Dose-dependent effect of recombinant human cytokines on HAMP promoter activity. HuH7 cells were cotransfected with WT-HAMP promoter-firefly luciferase construct and pTK-RL construct and treated with increasing doses of recombinant human cytokines. After treatment, cells were lysed and luciferase activity was measured. Results for individual cytokines (A-F) are expressed as fold induction over untreated control (firefly/renilla ratio of the experimental condition divided by the firefly/renilla ratio of untreated control). Dots and error bars represent mean ± SD of at least 3 independent experiments executed in duplicate. Statistical significance was determined with the Mann-Whitney rank-sum test, and *P < .05 compared with untreated control.
Figure 4
Figure 4
Effect of mutations in STAT3-BS or BREs on HAMP promoter response to cytokines. HuH7 cells were transfected with WT-HAMP, mSTAT, or mBRE1/2 promoter-firefly luciferase construct together with pTK-RL construct and treated with the indicated concentration of recombinant human cytokines. (A) IL-6 signaling was abrogated by the mutation in the STAT3-BS, but not by the mutation in both BREs. In contrast, BMP signaling was disrupted by the mutation in both BREs, but not by the STAT3-BS mutation. Bars represent mean ± SD of at least 3 independent experiments executed in duplicate. Statistical significance was determined with the Student t test or Mann-Whitney rank-sum test. *P < .05, compared with untreated control (optimem/WT-HAMP); †P < .05; and ††P < .001, compared with the WT-HAMP construct within the same cytokine group. (B) IL-6 and BMP-9 act synergistically on the HAMP promoter. (C) The mutations in BREs abrogated the synergy, while IL-6 induction was preserved. (D) The mutation of the STAT3-BS did not abrogate the synergy or the BMP-9 induction. Dots and error bars represent mean ± SD. Results are expressed as fold induction over untreated control.
Figure 5
Figure 5
Effect of MM patient sera on HAMP promoter activity. HuH7 cells were cotransfected with WT-HAMP, mSTAT, mBRE1/2, or mAll3 promoter-firefly luciferase construct, together with pTK-RL construct, and treated with 10% serum in Optimem. After 6 hours, cells were lysed and luciferase activity was measured. (A) Treatment with MM patient sera caused a significantly higher induction of WT-HAMP promoter activity, compared with sera from healthy individuals. Results are expressed as fold induction over untreated control. Statistical significance was determined with the Mann-Whitney rank-sum test. Boxes represent median and 25-75 percentiles, and whiskers represent 10 and 90 percentiles. Circles are outliers; *P < .05, compared with normal sera. (B) Mutation of BREs abrogated induction by MM sera dramatically, while the STAT3-BS mutation had a statistically less significant effect. Results are expressed as fold induction over untreated control (construct/optimem). Bars represent mean ± SD. Statistical significance was determined with 1-way ANOVA to compare the same sera for the different mutations. To compare MM patient sera with normal sera within the same construct, the Mann-Whitney rank-sum test was used; *P < .05, compared with untreated control (construct/optimem), and †P < .05, compared with WT-HAMP promoter activity for the same sera.
Figure 6
Figure 6
Effect of cytokine inhibitors on the induction of HAMP promoter activity by MM patient sera. HuH7 cells were cotransfected with WT-HAMP promoter-firefly luciferase construct and pTK-RL construct. Next, cells were treated with indicated doses of cytokines or 10% serum with or without cytokine inhibitors. After treatment, cells were lysed and luciferase activity was measured. (A) Specificity array of the cytokine inhibitors. All results are expressed as percentage promoter activity. Bars represent mean ± SD of at least 3 independent experiments executed in duplicate. Statistical significance was determined with the Student t test or Mann-Whitney rank-sum test; *P < .05, and †P < .001, compared with the cytokine-only group (cytokine/vehicle). (B) The effect of cytokine inhibitors on the induction of the WT-HAMP promoter by MM or healthy sera. Anti–BMP-2/4 and noggin-Fc significantly reversed the stimulatory effect of MM sera. Results are expressed as fold induction over untreated control (vehicle/optimem). Bars represent mean ± SD. Statistical significance was determined with 1-way ANOVA to compare the same sera for the different cytokine inhibitors. To compare MM patient sera with normal sera within the same group, the Mann-Whitney rank-sum test was used; *P < .05, compared with untreated control (vehicle/optimem), and †P < .05, compared with serum only group (vehicle/serum).
Figure 7
Figure 7
BMP-2 is present in MM sera. (A) MM sera (n = 3) were immunodepleted with anti–BMP-2/4 or isotype-control antibody. Depleted sera were used to treat HuH7 cells that had been transfected with WT-HAMP promoter-luciferase construct. After treatment, cells were lysed and luciferase activity was measured. Bars represent mean ± SD. Statistical significance was determined with the Student t test; *P < .001. (B) ELISA assay was used to measure BMP-2 values in normal (n = 9) and MM sera (n = 23). As expected, MM sera contained higher amounts of BMP-2, then normal sera. Bars represent mean ± SD. The Mann-Whitney rank-sum test was used to determine statistical significance, and *P < .001.
See this image and copyright information in PMC

Comment in

References (V体育2025版)

    1. Kyle RA, Rajkumar SV. Multiple myeloma. Blood. 2008;111(6):2962–2972. - PMC - PubMed
    1. Vanderkerken K, Asosingh K, Croucher P, Van Camp B. Multiple myeloma biology: lessons from the 5TMM models. Immunol Rev. 2003;194:196–206. - PubMed
    1. Kyle RA, Rajkumar SV. Multiple myeloma. N Engl J Med. 2004;351(18):1860–1873. - PubMed
    1. Kyle RA, Gertz MA, Witzig TE, et al. Review of 1027 patients with newly diagnosed multiple myeloma. Mayo Clin Proc. 2003;78(1):21–33. - PubMed
    1. Cucuianu A, Patiu M, Rusu A. Hepcidin and multiple myeloma related anemia. Med Hypotheses. 2006;66(2):352–354. - PubMed

Publication types

MeSH terms