Polyamines affect histamine synthesis during early stages of IL‐3‐induced bone marrow cell differentiation

Abstract

Mast cells synthesize and store histamine, a key immunomodulatory mediator. Polyamines are essential for every living cell. Previously, we detected an antagonistic relationship between the metabolisms of these amines in established mast cell and basophilic cell lines. Here, we used the IL-3-driven mouse bone marrow-derived mast cell (BMMC) culture system to further investigate this antagonism in a mast cell model of deeper physiological significance. Polyamines and histamine levels followed opposite profiles along the bone marrow cell cultures leading to BMMCs. alpha-Difluoromethylornithine (DFMO)-induced polyamine depletion resulted in an upregulation of histidine decarboxylase (HDC, the histamine-synthesizing enzyme) expression and activity, accompanied by increased histamine levels, specifically during early stages of these cell cultures, where an active histamine synthesis process occurs. In contrast, DFMO did not induce any effect in either HDC activity or histamine levels of differentiated BMMCs or C57.1 mast cells, that exhibit a nearly inactive histamine synthesis rate. Sequence-specific DNA methylation analysis revealed that the DFMO-induced HDC mRNA upregulation observed in early bone marrow cell cultures is not attributable to a demethylation of the gene promoter caused by the pharmacological polyamine depletion. Taken together, the results support an inverse relationship between histamine and polyamine metabolisms during the bone marrow cell cultures leading to BMMCs and, moreover, suggest that the regulation of the histamine synthesis occurring during the early stages of these cultures depends on the concentrations of polyamines.

Polyamines Are Present in Mast Cell Secretory Granules and Are Important for Granule Homeostasis

“…Polyamine depletion may be achieved by using DFMO, an irreversible inhibitor of ODC [38]. In a recent study we treated BMMCs with DFMO and obtained a large general reduction of both putrescine and spermidine [28], an effect in line with other works [39]. However, the possibility that polyamines are present in the mast cell secretory granules and the potential impact of polyamine presence on granule homeostasis has not been addressed previously.…”

“…The intracellular content of putrescine, histamine, serotonin, spermidine and spermine was simultaneously determined by fluorimetry after separation of their dansyl derivatives by reversed-phase HPLC as previously described [28]. Amines in isolated mast cell granules were assayed by this method as well.…”

“…Protein expression levels for mMCP-6, mMC-CPA, acetyl-histone H3, COX-IV and M6PRBP1 were assayed by Western blot as described previously [28]. Primary antibodies were used at a dilution of 1∶2000 (anti-mMCP-6 and anti-mMC-CPA), 1∶1000 (anti-COX-IV and anti-M6PRBP1) and 1∶5000 (anti-acetyl-histone H3), and secundary antibody (HRP-conjugated anti-rabbit IgG; GE Healthcare) at 1∶5000.…”

Deficiency of ovarian ornithine decarboxylase contributes to aging‐related egg aneuploidy in mice

“…To determine whether inhibition of oocyte ODC activity affects mouse oocyte in vitro maturation (IVM), we employed adifluoromethylornithine (DFMO), an enzyme-activated irreversible inhibitor for eukaryotic ODC (Metcalf et al, 1978;Poulin et al, 1992;Pegg, 2006). Immature mouse oocytes (aka 'GV oocytes' for the presence of intact germinal vesicle) from young CF1 mice (7-8 weeks of age) were cultured in medium (control) or medium containing 5 mM DFMO (Garcia-Faroldi et al, 2009). DFMO in oocyte IVM medium did not affect spindle assembly (not shown) or first polar body emission (776/ 945 or 82%, compared with 676/843 or 80.2% for control), consistent with our observation in frog oocytes (Zhou et al, 2009).…”

Vesicular Polyamine Transporter Mediates Vesicular Storage and Release of Polyamine from Mast Cells

“…Quantification of Polyamine and Histamine Release-Polyamines were quantified by dansylation and HPLC as reported previously with some modifications (4,38,39). Briefly, 400 l of supernatant was incubated with 150 nM 1,8-diaminooctane as an internal standard and 4.3% perchloric acid for 30 min on ice.…”