Inhibition of secretory phospholipase A2 enzyme by bilirubin: A new role as endogenous anti-inflammatory molecule

Abstract

Bilirubin is a powerful antioxidant that suppresses the inflammatory process. However its interaction with proinflammatory PLA(2) enzyme is not known. Inhibition of several secretory phospholipase A(2) (sPLA(2)) enzyme activities by bilirubin was studied using (14)C-oleate labeled Escherichia coli as substrate. Bilirubin inhibits purified sPLA(2) enzyme from Vipera russellii and Naja naja venom and partially purified sPLA(2) enzymes from human ascitic fluid, pleural fluid and normal serum in a dose dependent manner. IC(50) values calculated for these enzymes ranges from 1.75 to 10.5 microM. Inflammatory human sPLA(2) enzymes are more sensitive to inhibition by bilirubin than snake venom sPLA(2)s. Inhibition of sPLA(2) activity by bilirubin is independent of calcium concentration. Increasing substrate concentration (upto 180 nmol) did not relieve the inhibition of sPLA(2) by bilirubin and it is irreversible. Bilirubin quenched the relative fluorescence intensity of sPLA(2) in a dose dependent manner in the same concentration range at which in vitro sPLA(2) inhibition was observed. In the presence of bilirubin, apparent shift in the far UV-CD spectra of sPLA(2) was observed, indicating a direct interaction with the enzyme. Inhibition of sPLA(2) induced mouse paw edema by bilirubin confirms its sPLA(2) inhibitory activity in vivo also. These findings indicate that inhibition of sPLA(2) by bilirubin is mediated by direct interaction with the enzyme and bilirubin may act as an endogenous regulator of sPLA(2) enzyme activity.

A biophysical approach to phospholipase A2 activity and inhibition by anti-inflammatory drugs

“…Each NSAID was studied individually regarding its ability to modify the enzyme's secondary structure. The binding of ligands can affect the structure of enzymes and it has been previously reported the ability of inhibitors to change PLA 2 conformation through the increase or decrease in the α-helical content [3,32,33]. It is visible that PLA 2 CD spectra after addition of the NSAIDs show minor differences comparing with that from the enzyme alone, indicating lack of major changes in the secondary structure (Fig.…”

α-lipoic acid: An inhibitor of secretory phospholipase A2 with anti-inflammatory activity

“…PLA 2 from these fluids were partially purified according to previously described methods (Franson et al, 1990;Jameel et al, 2005). Briefly, fluids were centrifuged at 1000 ×g for 15 min to sediment cells and debris.…”

“…Inhibition studies, in vitro interaction studies and edema activity were designed as described earlier (Jameel et al, 2005). For PLA 2 assay, the amount of protein was chosen such that 10-15% hydrolysis of substrate was obtained at 37°C for 60 min.…”

Increased Serum Phospholipase A2 Activity After Non-Heart-Beating Donor Liver Transplantation and Association With Ischemia-Reperfusion Injury

“…This suggests that the liver is either producing or regulating the synthesis of some natural inhibitors. Here bilirubin may play a role since it has been shown to be an endogenous regulator of sPLA 2 enzyme activity [28]. Other proteins that could inhibit sPLA 2 activity are members of the annexin family [29] or liver proteins of the type of uteroglobin/ Clara cell secretory protein or mammaglobin [30,31].…”