The biphasic effects of cyclopentenone prostaglandins, prostaglandin J2 and 15-deoxy-Δ12,14-prostaglandin J2 on proliferation and apoptosis in rat basophilic leukemia (RBL-2H3) cells
Introduction
Mast cells are regarded as critical effector cells in the inflammatory reaction that underlies immediate hypersensitivity-mediated clinical conditions such as bronchial asthma, pollinosis and atopic dermatitis. After activation by cross-linking of FcεRI IgE receptors, mast cells initiate the production and secretion of chemical mediators, such as histamine, lipid-derived PGs and leukotrienes, as well as several types of cytokines. The soluble mediators and cytokines recruit cells to the site of inflammation, resulting in an inflammatory response [1].
PGD2 is a major cyclooxygenase product in various kinds of cells, including mast cells [2], [3]. PGD2 has potent pharmacological actions such as bronchoconstriction, peripheral vasodilation, inhibition of platelet aggregation and neuromodulation, so on [4], [5]. Two different types of PGD2 synthesizing enzymes, PGD synthetase (PGDS) have been characterized, purified and cloned by Urade et al. [6]. One type of PGDS is a glutathione requiring enzyme, hematopoietic PGDS that is present in mast cells. PGD2 is readily dehydrated to the cyclopentenone prostaglandins of the J series (PGJ2 series), such as PGJ2 and 15d-PGJ2, via an albumin-independent mechanism [7], [8]. The PGJ2 series display several unique characteristics associated with cell proliferation and apoptosis. Specifically, a cytotoxic effect of Δ12-PGJ2 is readily induced after translocation, accumulation in nuclei, and binding to nuclear proteins [9]. In addition the electrophilic carbons in cyclopentenone rings of the 15d-PGJ2 can directly conjugate with a thiol residue and to modify the function of proteins [10]. Furthermore, 15d-PGJ2 is a natural ligand of the nuclear receptor, PPARγ. The mechanisms involved in PPARγ-dependent [11], [12] and PPARγ-independent cytotoxic effects [13] are unknown.
Because so little is known about the biological significance of the PGJ2 series in mast cells [14], [15], [16], we have studied the cytotoxic effects of the PGJ2 and 15d-PGJ2 in mast cells.
Apoptosis is the physiological process of cell death that occurs in all multicellular organisms. In mast cells, apoptosis is also a key process in the regulation of inflammation and proliferation. It is also responsible for maintaining a constant cell number in tissues under normal conditions. Mast cell hyperplasia occurs in several clinical conditions; for example, host responses to parasites and the chronic inflammatory conditions that are present during either Crohn’s disease or rheumatoid arthritis [17]. In contrast, mast cells also express Fas antigen and undergo apoptosis by direct activation of Fas antigen [18]. In many cases, apoptotic pathways are mediated via the p53 tumor suppressor protein [19]. The involvement of p53 in initiating apoptosis results in the up-regulation of proapoptotic members of the bcl-family, such as bax, in mitochondria [20]. The role of survival/apoptosis-regulated genes in mast cells activation is now being investigated [17].
In this study, the exogenous PGJ2 and 15d-PGJ2 both induced bifunctional effects on RBL-2H3 cells. Specifically, proliferation occurred at the lower concentrations and apoptosis was observed at the higher concentrations. These concentration-dependent bifunctional effects were independent of PPARγ and DP-receptor. And the apoptotic effects were mainly mediated by mitochondrial pathways, not involved in p53 accumulation.
Section snippets
Assay of cell proliferation and viability
The rat mast cell line RBL-2H3 was kindly supplied from Dr. Michael A. Beaven (National Institutes of Health). The incorporation of 5-bromo-2′-deoxyuridine (BrdU) in place of thymidine is carried out to monitor cell proliferation. Briefly, RBL-2H3 cells (2×104 cells/200 μL/well) were cultured with BrdU in 96-well culture plate for 24 hr at 37°, and BrdU assay was carried out according to manufacture’s protocol. The optical density was then measured on a microplate reader with a wavelength of 450
Biphasic effects of PGJ2 and 15d-PGJ2 on proliferation of RBL-2H3 cells
The incorporation of BrdU in place of thymidine is measured to monitor cell proliferation. Cell proliferation was enhanced by PGJ2 at 1 μM (1.23-fold compared with the control) and by 15d-PGJ2 at 3 μM (1.27-fold), showing that the potency of cell proliferative effect induced by 15d-PGJ2 was slightly less than that of PGJ2 (Fig. 1). Incubation of RBL-2H3 cells with high concentrations of either PGJ2 or 15d-PGJ2 caused significant cytotoxic effects. Cell proliferation was suppressed by 48.4±9.6%,
Discussion
Mast cells play key roles in allergy and inflammatory responses via synthesis and release of inflammatory mediators. Some of the mediators are preformed and stored in secretory granules. Others such as the PGs, which are arachidonic acid metabolites, are generated de novo. PGJ2 and 15d-PGJ2, dehydration of PGD2, are known as cytotoxic inducer [7]. They also have been identified as ligands for PPARγ and regulate adipocyte differentiation and inflammatory response in macrophages [12].
In this
Acknowledgements
We are grateful to Dr. Takashi Inui (Tsu City College) for helpful suggestions and Dr. Yo Sato (RIKEN Harima Institute) for assistance with the TUNEL assay, as well as critical suggestions. We also thank Dr. Yasushi Fujitani (Takeda Chemical Industry, Ltd.) for critical suggestions, and Prof. Junya Tanaka (Ehime University, Dept. Physiology) for assistance with the fluorescence staining assay.
References (45)
- et al.
Mast cells contain spleen-type prostaglandin D synthetase
J. Biol. Chem.
(1990) - et al.
15-Deoxy-Δ12,14-prostaglandin J2. A prostaglandin D2 metabolite generated during inflammatory processes
J. Biol. Chem.
(2002) - et al.
15-Deoxy-Δ12,14-prostaglandin J2 induces apoptosis of human hepatic myofibroblasts. A pathway involving oxidative stress independently of peroxisome-proliferator-activated receptors
J. Biol. Chem.
(2001) - et al.
Peroxisome proliferator-activated receptors are expressed in mouse bone marrow-derived mast cells
FEBS Lett.
(2000) - et al.
High-performance liquid chromatographic determination of plasma and brain histamine without previous purification of biological samples: cation-exchange chromatography coupled with post-column derivatization fluorometry
J. Chromatogr.
(1985) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assay
J. Immunol. Methods
(1983)- et al.
Single-step method of RNA isolation by acid guanidinium thiocyanate–phenol–chloroform extraction
Anal. Biochem.
(1987) - et al.
Human colorectal cancer cells efficiently conjugate the cyclopentenone prostaglandin, prostaglandin J2, to glutathione
Biochim. Biophys. Acta
(2002) - et al.
A cyclopentenone prostaglandin activates mesangial MAP kinase independently of PPARγ
Biochem. Biophys. Res. Commun.
(2001) - et al.
p53 is not required for regulation of apoptosis or radioprotection by interleukin-3
Blood
(1997)
Apoptosis in cancer therapy: crossing the threshold
Cell
c-kit ligand mediates increased expression of cytosolic phospholipase A2, prostaglandin D2 synthase and increased IgE-dependent prostaglandin D2 generation in immature mouse mast cells
J. Biol. Chem.
Cyclopentenone prostaglandins as potential inducers of intracellular oxidative stress
J. Biol. Chem.
Regulation of mast cell survival by IgE
Immunity
Mast cells
Physiol. Rev.
Prostaglandin D2 generation after activation of rat and human mast cells with anti-IgE
J. Immunol.
Anaphylactic- and calcium-dependent generation of prostaglandin D2 (PGD2), thromboxane B2, and other cyclooxygenase products of arachidonic acid by dispersed human lung cells and relationship to histamine release
J. Immunol.
Prostaglandin D2 in sleep–wake regulation: recent progress and perspectives
Neuroscientist
Prostaglandin D2 as a mediator of allergic asthma
Science
Prostaglandin J2—anti-tumour and anti-viral activities and the mechanisms involved
Eicosanoids
Site and mechanism of growth inhibition by prostaglandins. II. Temperature-dependent transfer of a cyclopentenone prostaglandin to nuclei
J. Pharmacol. Exp. Ther.
The cyclopentenone 15-deoxy-Δ12,14-prostaglandin J2 binds to and activates H-Ras
Proc. Natl. Acad. Sci. U.S.A.
Cited by (27)
The chemistry, biology and pharmacology of the cyclopentenone prostaglandins
2020, Prostaglandins and Other Lipid MediatorsCitation Excerpt :Another example of a CyPG-induced mitochondrial apoptosis pathway has been reported. PGJ2 and 15d-PGJ2 induced apoptosis in RBL-2H3 cells at a concentrations of 3 and 10 μM in a PPAR-gamma independent manner [36]. Evidence was also found showing Inhibition of caspase activity.
15-Deoxy-Δ<sup>12,14</sup>-prostaglandin J<inf>2</inf> activates PI3K-Akt signaling in human breast cancer cells through covalent modification of the tumor suppressor PTEN at cysteine 136
2018, Cancer LettersCitation Excerpt :In addition, it has been reported that 15d-PGJ2 contributes to mouse skin tumor formation [66]. Although aforementioned events induced by 15d-PGJ2 vary depending on tissue and cell types as well as concentrations and duration of exposure to it, many studies have shown that this cyclopentenone PG promotes cell proliferation at relatively low concentrations while it exhibits cytotoxic effects at a higher concentration [61, 62, 67, 68]. Our previous study demonstrated that 15d-PGJ2 induces COX-2 expression via Akt signaling [8].
15-Deoxy-δ<sup>12,14</sup>-prostaglandin J<inf>2</inf> induces expression of 15-hydroxyprostaglandin dehydrogenase through Elk-1 activation in human breast cancer MDA-MB-231 cells
2014, Mutation Research - Fundamental and Molecular Mechanisms of MutagenesisCitation Excerpt :15d-PGJ2 has been known to have dual effects on cell survival, apoptosis and angiogenesis. This cyclopentenone prostaglandin induces apoptosis in rat basophilic leukemia (RBL-2H3) [47]. In addition, it attenuated the expression and activity of matrix metalloproteinase (MMP)-2, -7 and -9, and subsequently the invasiveness of pancreatic and colorectal cancer cells [20,21].
Role of mitochondria in programmed cell death mediated by arachidonic acid-derived eicosanoids
2013, MitochondrionCitation Excerpt :PGJ2 and 15d-PGJ2 induce caspase-dependent apoptosis via the inhibition of IκBα degradation in granulocytes (Ward et al., 2002). In contrast, in basophilic leukemia cells and myofibroblasts, PGJ2-induced apoptosis is primarily mediated by the activation of caspase-3 and -9 (Emi and Maeyama, 2004; Li et al., 2001). On the other hand, in neuroblastoma cells, 15d-PGJ2 induces the accumulation of p53 and the activation of the caspase cascade mediated by Fas and the Fas ligand (Kondo et al., 2002).
Identification, organ expression and ligand-dependent expression levels of peroxisome proliferator activated receptors in grass carp (Ctenopharyngodon idella)
2012, Comparative Biochemistry and Physiology - C Toxicology and PharmacologyCitation Excerpt :In addition, 1 mg kg− 1 15d-PGJ2 showed no effect on PPARα and PPARγ among the three tissues of grass carp. While this compound increased the transcriptional activity of murine chimera system (Forman et al., 1995; Kliewer et al., 1995) and rat basophilic leukemia cells PPARγ (Emi and Maeyama, 2004), its effect on fish PPARβ is still unknown. In primarily cultured zebrafish hepatocytes, 15d-PGJ2 (0.3 μM) could induce the PPARα and γ expressions (Ibabe et al., 2005).
The elmiric acids: Biologically active anandamide analogs
2008, NeuropharmacologyCitation Excerpt :This latter prostaglandin has been suggested to be an important promoter of the resolution phase of inflammation possibly through activation of PPAR-gamma, however, this last point is being actively debated (Nosjean and Boutin, 2002). Related to this debate are the reports that mediators other than PPAR-g have been implicated in some of the actions of 15-deoxy-delta-12, 14-PGJ2 (Emi and Maeyama, 2004; Trivedi et al., 2006). Thus, this putative mechanism for EMA action will no doubt undergo many modifications as new data become available.