Intracellular phospholipase A₂ expression and location in human macrophages: Influence of synthetic material surface chemistry

Abstract

Phospholipase A(2) (PLA(2)) enzymes participate in a potent inflammatory pathway through the liberation of arachidonic acid upon hydrolysis of membrane glycerophospholipids. The presence of implanted polycarbonate-urethane (PCNU) materials, used in several medical applications, has the ability to influence inflammatory responses of human macrophages that are recruited to a tissue-material interface; however, the specific inflammatory pathways that are activated upon macrophage attachment to PCNU are largely unknown. Previous studies suggested the participation of PLA(2) pathways in material degradation with the use of chemical inhibitors, such as aristolochic acid (ARIST), however not accurately defining the specific PLA(2) enzymes involved. The current study aimed to establish specific groups of PLA(2) involved in the macrophage foreign body response to PCNU. ARIST was assessed for specific effects on secretory PLA(2) (sPLA(2)) protein expression and non-specific effects on key proteins, beta-actin and monocyte-specific esterase, implicated in the macrophage attack on PCNU materials. Macrophage attachment to PCNU materials induced increased intracellular expression of cytosolic PLA(2) (cPLA(2)), but not sPLA(2), relative to tissue culture polystyrene (TCPS) as detected by immunoblot analysis, demonstrating an early and delayed stimulation during the time course of increased cPLA(2) protein expression. Laser scanning confocal microscopy images indicated a change in location of cPLA(2) in macrophages adherent to PCNU surfaces compared to TCPS. This study has illustrated changes in macrophage cPLA(2) expression in response to cell-attachment to PCNU surfaces, demonstrating that the macrophage foreign body response to biomaterials induces a potent inflammatory pathway, which may lead to tissue damage near the site of material implantation.

Amyloid-β peptide induces temporal membrane biphasic changes in astrocytes through cytosolic phospholipase A2

“…5B, right lower ) observed in Aβ 42 -induced cPLA 2 activation are similar to those reported GP distribution in macrophage membranes at 37°C [34]. Since macrophages are relatively active immune cells in which cPLA 2 can be frequently activated [38], we speculate that the GP distributions in macrophage membranes reported by Gaus et al, [34] might be affected in part by cPLA 2 activation.…”

Probing the Local Polarity of Alkylammonium Formate Ionic Liquids and Their Mixtures with Water by Using a Carbonyl Carotenoid

“…These results indicate that although the sample preparation included a 24 h serum‐free culture period, a significant amount of FBS protein had adsorbed to each surface (with greater amounts onto HDI431); during the serum‐free period and sample collection, these proteins were desorbed from the surface and into the CM. This apparent increase of FBS protein in the HDI431 CM sample would be expected since HDI431 is a much more hydrophobic surface, as indicated by water contact angle analysis,18, 31 thus promoting protein adsorption via hydrophobic interactions. Interference of serum proteins in proteomic analysis has certainly been noted as a problem that is difficult to overcome 32.…”

“…These differences between TCPS and PCNU biomaterials were reflected in varied MDM phenotypes, and although there is a considerable understanding of MDM behaviour in typical in vitro cell culture systems, the MDM–PCNU interface will undoubtedly produce an altered CM microenvironment. Our knowledge of signalling pathways involved and the cytokine milieu is becoming more detailed, with studies illustrating roles for esterase enzymes, as outlined above, phospholipase A 2 ,17, 18 protein kinase C,19, 30 as well as cytoskeletal proteins21, 28 which modulate how MDM spread on biomaterial surfaces and form focal contacts between the cell and the surface. Studies of the CM profiles at cell–biomaterial interfaces will enable us to further understand the foreign body response.…”

“…In the current study, TCPS has been used as a ‘control’ surface in terms of its lack of ability to be degraded by MDM and also as a reference material to compare MDM behaviour on model PCNU materials. Although a recent study demonstrated that TCPS and polydimethylsiloxane materials appear to be ‘stressful’ to cells in culture,44 studies have also shown that PCNU materials have the capacity to induce a greater inflammatory response compared to that of TCPS, as assessed through activation of the phospholipase A 2 pathway17 and phospholipase A 2 enzyme expression 18. Despite this difference, there was still a basal level of phospholipase A 2 protein expression and activity in TCPS‐adherent macrophages; however, it is not known whether this basal level would be sufficient to cause a damaging inflammatory response.…”

“…The destructive capacity of inflammatory MDM toward model polyurethanes has been demonstrated through illustrating biodegradation visually, using scanning electron microscopy, and quantitatively by measuring products liberated from radiolabelled PCNUs 12–14. Numerous signalling and enzymatic pathways have been assessed for their involvement in these degradative processes,15 demonstrating specific roles of serine proteases/esterases,16 phospholipase A 2 17, 18 and protein kinase C 19. Furthermore, cytoskeletal remodelling has been demonstrated, by our group7, 20 and others,21 as a central regulator of MDM and FBGC responses to polyurethane biomaterials.…”