Phospholipase A2-mediated superoxide production of murine peritoneal macrophages induced by chrysotile stimulation

Our plain-English summary of Phospholipase A2-mediated superoxide production of murine peritoneal macrophages induced by chrysotile stimulation, by T. Nakajima, M. Ito, U. Tchoua, et al (The International Journal of Biochemistry & Cell Biology, 2000).

The study

• Authors: T. Nakajima, M. Ito, U. Tchoua, H. Tojo, M. Hashimoto
• Journal: The International Journal of Biochemistry & Cell Biology
• Published: 2000;32(7):779-87
• Cited by: 2 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

In the present study, we investigated how chrysotile-stimulated macrophages generate superoxide using murine peritoneal macrophages, with special attention to the modulatory role of phospholipase A(2) (PLA(2)). We examined the effects of the following inhibitors and antagonists for signaling molecules on the superoxide anion (O(2)(-)) production of chrysotile-stimulated macrophages: p-bromophenacyl bromide (pBPB) and mepacrine for PLA(2); islet-activating protein (IAP) for G-protein; H-7 for protein kinase C (PKC); AA861 for 5-lipoxygenase (5-LO); indomethacin for cyclo-oxygenase (COX); ETYA for both 5-LO and COX; hexanolamine PAF for platelet-activating factor (PAF). The PLA(2) and PKC inhibitors effectively inhibited the chrysotile-induced superoxide anion production of macrophages, but not the G-protein inhibitor, the 5-LO and COX inhibitors, and the PAF antagonist. We also examined the effects of the PLA(2) inhibitors on macrophages stimulated by phorbol 12-myristate 13-acetate (PMA) which directly activates PKC. The two structurally different PLA(2) inhibitors showed differential effects on the PMA-induced superoxide generation: pBPB inhibited it but mepacrine did not. These results suggested that (1) PLA(2) and PKC modulate the chrysotile-induced O(2) production, and (2) two different kinds of PLA(2) work upstream and downstream of PKC, but (3) G-protein, 5-LO and COX metabolites, and PAF have no modulatory role in the reaction.

In plain English

This laboratory experiment used macrophages, immune cells washed from the abdominal (peritoneal) cavity of male ICR mice. The authors exposed the cells to milled chrysotile asbestos and measured superoxide, a reactive form of oxygen, by light from its reaction with an added chemical, lucigenin. Some cells were first treated with chemicals blocking particular signaling molecules. SOD, an enzyme that destroys superoxide, completely quenched the chrysotile-induced light, which peaked about 40 minutes after stimulation. Two blockers of phospholipase A2 (pBPB and mepacrine) reduced superoxide production by 91 to 104% after both chrysotile and alkali (sodium hydroxide) stimulation. A protein kinase C blocker (H-7) inhibited production with all stimulants. With chrysotile, a G-protein blocker had no inhibitory effect, and blockers of 5-lipoxygenase, cyclo-oxygenase and platelet-activating factor made no significant difference. With PMA, which directly activates protein kinase C, pBPB inhibited superoxide production but mepacrine did not. The authors concluded that phospholipase A2 and protein kinase C are key enzymes, possibly with two kinds of phospholipase A2 acting before and after protein kinase C. They called for more detailed studies to identify the enzymes.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) experiment on mouse macrophages exposed to chrysotile, with signaling-molecule inhibitors
• Who was studied: Peritoneal macrophages lavaged from 8 to 9 week-old male ICR mice from Japan SLC. Eight to 12 samples were prepared from a single mouse per measurement series. The total number of mice is not stated.
• Exposure: Chrysotile reagent (Ishizu Chem. Inc.), ball-milled and sieved. The sieved fraction was 34% by number fibrous particles of 30 to 70 units length (units garbled in the text) and 66% fibrous or granular particles under 10 units. 6.6 mg of chrysotile was used per stimulation. Sodium hydroxide (alkali) and PMA were used as comparison stimulants. Superoxide was measured through chemiluminescence from its reaction with lucigenin.
• Main results:
  • SOD completely quenched chrysotile-induced light emission, indicating it was entirely due to superoxide production by the macrophages. Chrysotile without macrophages did not appear to produce superoxide (data not shown).
  • In chrysotile-stimulated macrophages, the light signal peaked at about 40 minutes and returned to basal level over the following 2 or 3 hours.
  • Alkali (sodium hydroxide) stimulation also induced superoxide production, with a time pattern resembling chrysotile stimulation, especially at pH 8.93.
  • The G-protein inhibitor IAP had no inhibitory effect on chrysotile-stimulated superoxide production.
  • Pretreatment with the phospholipase A2 inhibitors pBPB and mepacrine decreased superoxide production by 91 to 104% with both chrysotile and sodium hydroxide stimulation.
  • With PMA stimulation, pBPB showed an inhibitory effect but mepacrine did not. H-7 (protein kinase C inhibitor) inhibited superoxide production with all stimulants.
  • Pretreatment with AA861, indomethacin, ETYA or hexanolamine PAF produced no significant difference in chrysotile-induced superoxide production compared with control.
• The authors' conclusion: The authors concluded that phospholipase A2 is a key enzyme regulating chrysotile-induced superoxide generation and that protein kinase C is another key enzyme. They concluded that a mepacrine-insensitive phospholipase A2 works downstream of protein kinase C and a mepacrine-sensitive one upstream. They suggested that alkalinity in chrysotile may be responsible, and that there may be a pathway independent of the phospholipase C pathway.
• Limitations the authors state:
  • The authors did not directly examine the involvement of phospholipase C; they assumed its role based on another group's findings.
  • More detailed functional and structural studies are needed to identify precisely which types of phospholipase A2 are involved.
  • Their results cannot exclude an alternative mechanism in which chrysotile particles interact with glycoproteins on the macrophage membrane.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Chrysotile.

Medical terms in this study, explained in our Lexicon: Superoxide, Macrophage, Chrysotile.

Related studies

• IgG specifically enhances chrysotile asbestos-stimulated superoxide anion production by the alveolar macrophage — R.K. Scheule, A. Holian (1989)
• Asbestos effects on superoxide production. An in vitro study of hamster alveolar macrophages — B.W. Case, M.P. Ip, et al. (1986)
• IgG and IgA enhance the chrysotile-induced production of reactive oxygen metabolites by human polymorphonuclear leucocytes — M. Klockars, M. Hedenborg, et al. (1989)
• Brief inhalation of asbestos compromises superoxide production in cells from bronchoalveolar lavage — J.M. Petruska, J. Marsh, et al. (1990)
• Asbestos Induces Oxidative Stress and Activation of Nrf2 Signaling in Murine Macrophages: Chemopreventive Role of the Synthetic Lignan Secoisolariciresinol Diglucoside (LGM2605) — R.A. Pietrofesa, A. Velalopoulou, et al. (2016)
• Involvement of protein kinase C, phospholipase C, and protein tyrosine kinase pathways in oxygen radical generation by asbestos-stimulated alveolar macrophage — Y. Lim, S.H. Kim, et al. (1997)

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