Inhibition of sarco-endoplasmic reticulum calcium ATPases (serca) by polycyclic aromatic hydrocarbons: lack of evidence for direct effects on cloned rat enzymes

doi:10.1016/S0192-0561(96)00063-X

International Journal of Immunopharmacology

Volume 18, Issue 10, October 1996, Pages 589-598

International Journa...

https://doi.org/10.1016/S0192-0561(96)00063-X Get rights and content

Abstract

Previous studies have demonstrated that immunosuppressive polycyclic aromatic hydrocarbons (PAHs) disrupt Ca²⁺ homeostasis leading to inhibition of the Ca²⁺-dependent pathways of T cell and B cell activation. The sustained Ca²⁺-elevation produced by immunosuppressive PAHs may result from the inhibition of Ca²⁺-ATPases in the endoplasmic reticulum (SERCA). The purpose of the present study was to determine whether PAHs directly inhibit cloned SERCA enzymes, and whether there is any selectivity for certain isoforms. PAHs were examined for their effects on purified cloned rat SERCA enzymes, including SERCA 1, SERCA2a and SERCA3, transiently expressed in human embryonic kidney (HEK) cells. Results showed that known SERCA inhibitors, thapsigargin (100 nM) and 2,5-di(t-butyl)-1,4-benzohydroquinone (10 μmol), completely inhibited all rat SERCA isoforms, whereas 7,12-dimethylbenz(a)anthracene, benzo(a)pyrene, benzo(e)pyrene, anthracene, 3-methyl-cholanthrene, 9,10-dimethylanthracene and benz(a)anthracene at concentrations as high as 10 μmol appeared to have little inhibitory effect on any of the SERCA. The results demonstrating that PAHs do not inhibit cloned SERCA enzymes suggest that metabolism may be required for PAH-induced inhibition, or that other cellular elements, not present in the HEK transfection model, may be required for activity.

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Polycyclic aromatic hydrocarbons (PAHs) are embryo- and cardiotoxic to fish that might be associated with improper intracellular Ca²⁺ management. Since sarco(endo)plasmic reticulum Ca²⁺-ATPase (SERCA) is a major regulator of intracellular Ca²⁺, the SERCA activity and the contractile properties of rainbow trout (Oncorhynchus mykiss) ventricle were measured in the presence of 3- and 4-cyclic PAHs. In unfractionated ventricular homogenates, acute exposure of SERCA to 0.1–1.0 μM phenanthrene (Phe), retene (Ret), fluoranthene (Flu), or pyrene (Pyr) resulted in concentration-dependent increase in SERCA activity, except for the Flu exposure, with maximal effects of 49.7–83 % at 1 μM. However, PAH mixture did not affect the contractile parameters of trout ventricular strips. Similarly, all PAHs, except Ret, increased the myotomal SERCA activity, but with lower effect (27.8–40.8 % at 1 μM). To investigate the putative chronic effects of PAHs on SERCA, the atp2a2a gene encoding trout cardiac SERCA was expressed in human embryonic kidney (HEK) cells. Culture of HEK cells in the presence of 0.3–1.0 μM Phe, Ret, Flu, and Pyr for 4 days suppressed SERCA expression in a concentration-dependent manner, with maximal inhibition of 49 %, 65 %, 39 % (P < 0.05), and 18 % (P > 0.05), respectively at 1 μM. Current findings indicate divergent effects of submicromolar PAH concentrations on SERCA: stimulation of SERCA activity in acute exposure and inhibition of SERCA expression in chronic exposure. The depressed expression of SERCA is likely to contribute to the embryo- and cardiotoxicity of PAHs by depressing muscle function and altering gene expression.
Organic chemicals from diesel exhaust particles affects intracellular calcium, inflammation and β-adrenoceptors in endothelial cells
2019, Toxicology Letters
Citation Excerpt :
Interestingly, diesel exhaust exposure seem to impair calcium-dependent vasomotor function in healthy men (Barath et al., 2010; Lucking et al., 2011); an effect that may relate to disturbed endothelial [Ca2+]i. Inflammatory effects of organic chemicals known to be present on DEP, seem to depend on increased [Ca2+]i (Mayati et al., 2014; Monteiro et al., 2008; N'Diaye et al., 2006; Zhao et al., 1996). We have previously shown that βARs may be involved in [Ca2+]i increase and induction of the pro-inflammatory chemokine CXCL8 in human bronchial epithelial BEAS-2B cells exposed to 1-NP, one of the dominating nitro-PAHs in DEP (Mayati et al., 2014).
Exposure to diesel exhaust particles (DEP) may contribute to endothelial dysfunction and cardiovascular disease. DEP, extractable organic material from DEP (DEP-EOM) and certain PAHs seem to trigger [Ca²⁺]_i increase as well as inflammation via GPCRs like βARs and PAR-2. In the present study we explored the involvement of βARs and PAR-2 in effects of DEP-EOM on [Ca²⁺]_i and expression of inflammation-associated genes in the endothelial cell-line HMEC-1. We exposed the human microvascular endothelial cell line HMEC-1 to DEP-EOM fractionated by sequential extraction with solvents of increasing polarity: n-hexane (n-Hex-EOM), dichloromethane (DCM-EOM), methanol (Methanol-EOM) and water (Water-EOM). While Methanol-EOM and Water-EOM had no marked effects, n-Hex-EOM and DCM-EOM enhanced [Ca²⁺]_i (2–3 times baseline) and expression of inflammation-associated genes (IL-1α, IL-1β, COX-2 and CXCL8; 2–15 times baseline) in HMEC-1. The expression of βARs (60–80% of baseline) and βAR-inhibitor carazolol suppressed the increase in [Ca²⁺]_i induced by both n-Hex- and DCM-EOM. Carazolol as well as the Ca²⁺-channel inhibitor SKF-96365 reduced the DCM-EOM-induced pro-inflammatory gene-expression. Overexpression of βARs increased DCM-EOM-induced [Ca²⁺]_i responses in HEK293 cells, while βAR-overexpression suppressed [Ca²⁺]_i responses from n-Hex-EOM. Furthermore, the PAR-2-inhibitor ENMD-1068 attenuated [Ca²⁺]_i responses to DCM-EOM, but not n-Hex-EOM in HMEC-1.
The results suggest that βAR and PAR-2 are partially involved in effects of complex mixtures of chemicals extracted from DEP on calcium signalling and inflammation-associated genes in the HMEC-1 endothelial cell-line.
Breast meat quality of broiler chickens can be affected by managing the level of nitric oxide
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Citation Excerpt :
No significant differences in pH values of chicken breast meat were found after incubating with 3 different solutions for 24 h (P > 0.05, Figure 1). The SERCA and RyR are known to be susceptible to NO-induced nitrosylation and oxidation due to their high amounts of cysteine (Zhao et al., 1996; Stamler et al., 2008). Chronic low-frequency stimulation in fast-twitch muscles of rabbit inactivated SERCA due to protein oxidation and peroxynitrite-mediated tyrosine nitration (Klebl et al., 1998).
The objective of this study was to investigate the effects of nitric oxide (NO) on the quality of broiler chicken meat during postmortem storage. Eighteen broiler chickens were slaughtered and breast meat was randomly assigned to 1 of 3 treatments including the control group, nitric oxide synthase (NOS) inhibitor group, and NO enhancer group. The breast samples were incubated with water, NO enhancer, and NO inhibitor for 1 d and then stored at 4°C under atmospheric conditions for 4 and 7 d. Left side of breast meat was used to determine protein oxidation, lipid oxidation, water-holding capacity, and pH, whereas the right side was used for the measurement of color and drip loss. Breast meat from NO enhancer group showed the lowest water-holding capacity during refrigerated storage, whereas drip losses were not significantly (P > 0.05) different among 3 treatments. Lipid oxidation showed a significant difference (P < 0.05) only at 7 d in which NOS inhibitor group showed the highest, whereas the NO enhancer group showed the lowest levels of lipid oxidation. Carbonyl content in NO enhancer group was significantly higher than the control and NOS inhibitor treatment at 1 and 4 d of refrigerated storage. Lightness of breast meat in the NOS inhibitor group was greater, whereas redness was lower than the control and NO enhancer group at 4 and 7 d of refrigerated storage. The ultimate pH of 3 treatments was not significantly (P > 0.05) different from one another. These data indicate that NO could play a significant role in modulating the quality of fresh broiler breast meat during refrigerated storage.
Induction of intracellular calcium concentration by environmental benzo(a)pyrene involves a β2-adrenergic receptor/adenylyl cyclase/Epac-1/inositol 1,4,5-trisphosphate pathway in endothelial cells
2012, Journal of Biological Chemistry
Citation Excerpt :
Moreover, knockdown AhR expression failed to counteract [Ca2+]i signal triggered by B(a)P (7). Mechanisms that contribute to [Ca2+]i modulation by PAHs have been linked to inhibition of sarcoendoplasmic reticulum calcium ATPase (9) or to activation of protein-tyrosine kinases (10), ryanodine receptor (11), store-operated calcium channel, or inositol 1,4,5-trisphosphate (IP3) receptor (5, 12). However, the initial events that trigger calcium signaling in response to PAH exposure still remain unknown.
Polycyclic aromatic hydrocarbons (PAHs) such as benzo(a)pyrene (B(a)P) are widely distributed environmental contaminants, known as potent ligands of the aryl hydrocarbon receptor (AhR). These chemicals trigger an early and transient increase of intracellular calcium concentration ([Ca²⁺]_i), required for AhR-related effects of PAHs. The mechanisms involved in this calcium mobilization were investigated in the present study. We demonstrated that B(a)P-mediated [Ca²⁺]_i induction was prevented in endothelial HMEC-1 cells by counteracting β2-adrenoreceptor (β2ADR) activity using pharmacological antagonists, anti-β2ADR antibodies, or siRNA-mediated knockdown of β2ADR expression; by contrast, it was strongly potentiated by β2ADR overexpression in human kidney HEK293 cells. B(a)P was shown, moreover, to directly bind to β2ADR, as assessed by in vitro binding assays and molecular modeling. Pharmacological inhibition and/or siRNA-mediated silencing of various signaling actors acting downstream of β2ADR in a sequential manner, such as G protein, adenylyl cyclase, Epac-1 protein, and inositol 1,4,5-trisphosphate (IP₃)/IP₃ receptor, were next demonstrated to prevent B(a)P-induced calcium signal. Inhibition or knockdown of these signaling elements, as well as the use of chemical β-blockers, were finally shown to counteract B(a)P-mediated induction of cytochrome P-450 1B1, a prototypical AhR target gene. Taken together, our results show that B(a)P binds directly to β2ADR and consequently utilizes β2ADR machinery to mobilize [Ca²⁺]_i, through activation of a G protein/adenylyl cyclase/cAMP/Epac-1/IP₃ pathway. This β2ADR-dependent signaling pathway activated by PAHs may likely be crucial for PAH-mediated up-regulation of AhR target genes, thus suggesting a contribution of β2ADR to the health-threatening effects of these environmental pollutants.
Inhibition by polycyclic aromatic hydrocarbons of ATPase activities in Sebastiscus marmoratus larvae in Relationship with the development of early life stages
2011, Marine Environmental Research
Citation Excerpt :
The role of Ca2+-ATPase in osmoregulation during early fish life stages needs further investigation. Some studies have shown that PAHs modulate intracellular Ca2+ concentration by acting on the endoplasmic reticulum Ca2+-ATPases (Zhao et al., 1996; Reynaud et al., 2003). 3-methylcholanthrene (3-MC) acts on the endoplasmic reticulum to increase intracellular calcium levels in carp (Cyprinus carpio) phagocytes (Reynaud et al., 2001) and leucocytes (Reynaud et al., 2003), which suggests that 3-MC directly acts on Ca2+-ATPases (Reynaud et al., 2001).
Sebastiscus marmoratus larvae were exposed to waterborne polycyclic aromatic hydrocarbons (PAHs) containing 3–5 rings, benzo[a]pyrene (BaP), pyrene (Py) or phenanthrene (Phe), respectively at 0.01, 0.1 and 1 μg L⁻¹. Cumulative mortality, frequency of dorsal curvature and rate of pericardial and yolk sac edema in larvae treated for 8 days were significantly increased in a dose-dependent manner. All three PAHs resulted in reduction of the lower jaw, and the extent of reduction increased with increasing concentrations of the PAHs. Na⁺/K⁺-ATPase and Ca²⁺-ATPase activity in larvae exposed to the three PAHs were all significantly inhibited in a dose-dependent manner. Analysis using the Pearson correlation coefficient indicated a significant correlation between the rate of the dorsal curvature and edema and the inhibition of Na⁺/K⁺-ATPase and Ca²⁺-ATPase activity, suggesting that the developmental defects caused by PAHs were related to their inhibition of Na⁺/K⁺-ATPase and Ca²⁺-ATPase activity.
The involvement of secondary signaling molecules in cytochrome P-450 1A1-mediated inducible nitric oxide synthase expression in benzo(a)pyrene-treated rat polymorphonuclear leukocytes
2007, Life Sciences
Benzo(a)pyrene induces cytochrome P-450 1A1 (CYP1A1) expression in rat polymorphonuclear leukocytes (PMNs) that upregulates expression of inducible nitric oxide synthase (iNOS). In the present study, the involvement of secondary signaling molecules in CYP1A1-mediated augmentation of iNOS expression in benzo(a)pyrene-treated rat PMNs was investigated. PMNs were isolated from the peripheral blood of controls and benzo(a)pyrene-treated rats. The expression and/or activity of CYP1A1, iNOS, tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and intracellular calcium ([Ca²⁺]i) concentrations were measured in control and benzo(a)pyrene-treated rat PMNs with and without alpha-naphthoflavone, aminoguanidine, genistein, pyrrolidine dithiocarbamate (PDTC), felodipine, or SB202190 pre-treatment. A significant elevation in CYP1A1 and [Ca²⁺]i was observed in benzo(a)pyrene-treated rat PMNs, which was significantly restored by alpha-naphthoflavone or genistein. Neither PDTC, SB202190, nor aminoguanidine altered the benzo(a)pyrene-mediated increase in [Ca²⁺]i. Although felodipine reduced the benzo(a)pyrene-mediated increase in [Ca²⁺]i, no significant change was observed in CYP1A1 expression and activity. Benzo(a)pyrene-augmented iNOS expression and activity in PMNs were significantly reverse by felodipine, genistein, or PDTC. Benzo(a)pyrene also induced TNF-α and IL-1β production in PMNs, which was significantly reversed by genistein. The results demonstrated the involvement of [Ca²⁺]i, tyrosine kinase, inflammatory cytokines, and NF-κB in CYP1A1-mediated iNOS expression in benzo(a)pyrene-treated rat PMNs.

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Inhibition of sarco-endoplasmic reticulum calcium ATPases (serca) by polycyclic aromatic hydrocarbons: lack of evidence for direct effects on cloned rat enzymes

Abstract

Int. J. Immunopharmacol.

Toxicol. Appl. Pharmacol.

Fund. Appl. Toxicol.

Analyt. Biochem.

Toxicol. Appl. Pharmacol.

Toxicol. Appl. Pharmacol.

Cell

J. Biol. Chem.

Toxicol. Appl. Pharmacol.

Toxicol. Appl. Pharmacol.

J. Biol. Chem.

J. Biol. Chem.

FEBS Lett.

Toxicol. Appl. Pharmacol.