Human retinal pigment epithelium secretes a phospholipase A2 and contains two novel intracellular phospholipases A2

Abstract

The sensitivity of different phospholipase A2 (PLA2)-active fractions eluted from cation-exchange chromatography to para-bromophenacylbromide (pBPB), Ca2+-EGTA, DTT, heat, and H2SO4 indicates that human cultured retinal pigment epithelial (hRPE) cells probably contain two different intracellular PLA2 enzymes. Control experiments using "back-and-forth" thin-layer chromatography confirmed that, in our assay conditions, the generation of free fatty acids originated solely from PLA2 activity. Together with immunoblot experiments where no cross-reactivity was observed between the hRPE cytosolic PLA2 enzymes and several antisera directed against secretory PLA2s (sPLA2s) and cytosolic PLA2 (cPLA2), these findings suggest that intracellular hRPE PLA2s are different from well-known sPLA2s, cPLA2, and Ca2+-independent PLA2s. We also report an additional hRPE-PLA2 enzyme that is secreted and that exhibits sensitivity to pBPB, Ca2+-EGTA, DTT, heat, and H2SO4, which is characteristic of sPLA2 enzymes. This approximately 22-kDa PLA2 cross-reacted weakly with an antiserum directed against porcine pancreatic group I sPLA2 but strongly with an antiserum directed against N-terminal residues 1-14 of human synovial group II sPLA2, suggesting that this extracellular enzyme is a member of the sPLA2 class of enzymes. We thus conclude that there are three distinct PLA2 enzymes in cultured hRPE cells, including two novel intracellular PLA2s and a 22-kDa secreted sPLA2 enzyme.

Gabapentin Attenuates Ocular Inflammation: In vitro and In vivo Studies

“…There is only a little knowledge about PLA 2 expression in other parts of the eye, in addition to the retina (Castagnet and Giusto, 1993; Van Themsche et al, 2001; Kolko et al, 2007). cPLA has been identified in the human cornea (Landreville et al, 2004), while, in the conjunctiva, cPLA 2 is mainly localized in the surface of the epithelium, probably participating in the protection against risks caused by mechanical wear and tear stress (Helin et al, 2008).…”

Expression and location of mRNAs encoding multiple forms of secretory phospholipase A2 in the rat retina

“…sPLA 2 activity has been described for the retina (Castagnet and Giusto, 1993;Hanasaki and Arita, 1999), and there seems to be enhanced sPLA 2 activity in retinal ischemia and light damage (Jung and Reme, 1994;Rego et al, 1996). Furthermore, different PLA 2 s are present in the retinal pigment epithelium (Jacob et al, 1997;Van Themsche et al, 2001). To date, no PLA 2 -specific receptors have been cloned or characterized in the retina.…”

Human secretory phospholipase A2, group IB in normal eyes and in eye diseases

“…PLA 2 activity has been shown to be present in the RPE (Van Themsche et al. 2001) and our previous study revealed immunohistochemical expression of rGIB in rat RPE (Kolko et al.…”

“…PLA 2 have only been investigated in the human cornea and no knowledge of PLA 2 expression in other parts of the human eye exists. However, indirect evidence points to PLA 2 activity in the outer layers of the retina (Castagnet & Giusto 1993; Van Themsche et al. 2001).…”