Anti-Inflammatory Activity of Oleanolic Acid by Inhibition of Secretory Phospholipase A₂

Abstract

Oleanolic acid, a triterpenoid known for its anti-inflammatory properties, is commonly present in several medicinal plants. The present study evaluated the effect of oleanolic acid on sPLA₂, a key enzyme in inflammatory reactions. Oleanolic acid inhibited sPLA₂ activities of human synovial fluid (HSF), human pleural fluid (HPF) and Vipera russelli (VRV-PL-V) and Naja naja (NN-PL-I) snake venoms in a concentration-dependent manner. The IC₅₀ values of sPLA₂ from these sources ranged from 3.08 to 7.78 μM. Increasing calcium (Ca²⁺) concentrations from 2.5 to 15 mM and substrate concentration up to 180 nM did not affect the level of inhibition. Oleanolic acid enhanced the relative intrinsic fluorescence intensity of sPLA₂ (VRV-PL-V). In the presence of oleanolic acid, an apparent shift in the far UV-CD spectrum of sPLA₂ was observed. These studies indicate direct interaction with the enzyme and formation of an sPLA₂-oleanolic acid complex. The complex formed resulted in irreversible inhibition of sPLA₂. Oleanolic acid inhibited indirect hemolytic activity and mouse paw edema induced by sPLA₂. Inhibition of in vitro and in vivo sPLA₂ activity by oleanolic acid explains the observed anti-inflammatory properties of several oleanolic acid-containing medicinal plants.

Abbreviations

COX.1/2:cyclooxygenase-1/2

HPF:human pleural fluid

HSF:human synovial fluid

LOX:lipoxygenase

NN-PL-I:Naja naja phospholipase-I

PAF:platelet activation factor

p-BPB:para-bromophenacyl bromide

sPLA₂:secretory phospholipase A₂

UV-CD:ultraviolet circular dichroism

VRV-PL-V:Vipera russelli venom phospholipase-V

Key words

oleanolic acid - human synovial fluid - human pleural fluid - anti-inflammatory - sPLA₂ - snake venom PLA₂

References

• 1 Uhl W, Buchler M, Nevalainen T J, Deller A, Beger H G. Serum phospholipase A₂ in patients with multiple injuries.  J Trauma. 1990;  30 1285-90
• 2 Nevalainen T J, Gronroos J M, Kortesuo P T. Pancreatic and synovial type phospholipases A₂ in serum samples from patients with severe acute pancreatitis.  Gout. 1993;  34 1133-6
• 3 Haapamaki M M, Gronroos J M, Nurmi H, Alanen K, Kallajoki M, Nevalainen T J. Gene expression of group II phospholipase A₂ in intestine in ulcerative colitis.  Gout. 1997;  40 95-100
• 4 Anderson S, Sjursen W, Laegreid A, Volden G, Johansen B. Elevated expression of human non-pancreatic phospholipase A₂ in psoriatic tissue.  Inflammation. 1994;  18 1-12
• 5 Nanda B L, Nataraju A, Rajesh R, Rangappa K S, Shekar M A, Vishwanath B S. PLA₂ mediated arachidonate free radicals: PLA₂ inhibition and neutralization of free radicals by anti-oxidants: a new role as anti-inflammatory molecule.  Curr Top Med Chem. 2007;  7 765-77
• 6 Krug H F, Mattern D, Bidault J, Ninio E. Effects of organometals on cellular signaling II Inhibition of reincorporation of free arachidonic acid and influence on paf-acether synthesis by triethyllead.  Environ Health Perspect. 1994;  102 331-4
• 7 Vane J R, Botting R M. Anti-inflammatory drugs and their mechanism of action.  Inflamm Res. 1998;  47 78-87
• 8 Sadashiva M P, Nataraju A, Mallesha H, Rajesh R, Vishwanath B S, Rangappa K S. Synthesis and evaluation of trimethoxyphenyl isoxazolidines as inhibitors of secretory phospholipase A₂ with anti-inflammatory activity.  Int J Mol Med. 2005;  16 895-904
• 9 Monti M C, Casapullo A, Riccio R, Paloma L G. Further insights on the structural aspects of PLA₂ inhibition by hydroxybutenolide-containing natural products: a comparative study on petrosaspongiolides M–R.  Bioorg Med Chem. 2003;  12 1467-74
• 10 Stefanski E, Pruzanski W, Sternby B, Vadas P. Purification of a soluble phopsholipase A2 from synovial fluid in rheumatoid arthritis.  J Biochem. 1986;  100 1297-303
• 11 Kasturi S, Gowda T V. Purification and characterization of a major phopsholipase A₂ from Russell’s viper (Vipera russellii) venom.  Toxicon. 1989;  27 229-37
• 12 Rudrammaji L M, Gowda T V. Purification and characterisation of three acidic, cytotoxic phospholipase A₂ from Indian cobra (Naja naja ) venom.  Toxicon. 1988;  36 921-32
• 13 Lowry O H, Rosebrough N J, Farr A L, Randall R J. Protein measurement with the Folin phenol reagent.  J Biol Chem. 1951;  193 265-75
• 14 Patriarca P, Beckerdite S, Elsbach P. Phospholipases and phospholipid turnover in Escherichia coli spheroplasts.  Biochim Biophys Acta. 1972;  260 593-600
• 15 Vishwanath B S, Frey F J, Bradbury M J, Dallman M F, Frey B M. Glucocorticoid deficiency increases phospholipse A₂ activity in rats.  J Clin Invest. 1993;  92 1974-80
• 16 Boman H G, Kaletta U. Chromatography of rattlesnake venom: a separation of three phosphodiesterases.  Biochim Biophys Acta. 1957;  24 619-31
• 17 Yamakawa M M, Hokama N Z. Fractionation of Sakishima habu (T. elegans) venom and lethal hemorrhagic and edema forming activity of the fractions. In: Ohsaka A, Hayashi K, Sawai Y, editors Animal, plant and microbial toxins. Vol.1 New York; Plenum Press 1976: 97
• 18 Nataraju A, Raghavendra Gowda C D, Rajesh R, Vishwanath B S. Group IIA secretory PLA₂ inhibition by ursolic acid: a potent anti-inflammatory molecule.  Curr Top Med Chem. 2007;  7 801-9
• 19 Calixto J B, Campos M M, Otuki M F, Santos A R. Anti-inflammatory compounds of plant origin. Part II. Modulation of pro-inflammatory cytokines, chemokines and adhesion molecules.  Planta Med. 2004;  70 93-103
• 20 Calixto J B, Otuki M F, Santos A R. Anti-inflammatory compounds of plant origin. Part I. Action on arachidonic acid pathway, nitric oxide and nuclear factor kappa B (NF-kappaB).  Planta Med. 2003;  69 973-83
• 21 Kim H P, Son K H, Chang H W, Kang S S. Anti-inflammatory plant flavonoids and cellular action mechanism.  J Pharmacol Sci. 2004;  96 229-45
• 22 Davidson F F, Dennis E A, Powell M, Glenney J R. Inhibition of phospholipase A₂ by ”lipocortins” and calpactins. An effect of binding to substrate phospholipids.  J Biol Chem. 1987;  262 1698-705
• 23 Fawzy A A, Vishwanath B S, Franson R C. Inhibition of human non-pancreatic phospholipases A₂ by retinoids and flavonoids: Mechanism of action.  Agents Actions. 1988;  25 394-400
• 24 Jameel N M, Shekar M A, Vishwanath B S. Alpha-lipoic acid an inhibitor of secretory phospholipase A₂ with anti-inflammatory activity.  Life Sci. 2006;  80 46-53
• 25 Vishwanath B S, Kini R M, Gowda T V. Characterization of three edema inducing phospholipase A₂ enzymes from habu (Trimeresurus flavovoridis) venom and their interaction with the alkaloid aristolochic acid.  Toxicon. 1987;  25 501-15
• 26 Lin Y H, Huang W N, Lee S C, Wu W G. Heparin reduces the alpha-helical content of cobra basic phospholipase A(2) and promotes its complex formation.  Int J Biol Macromol. 2000;  27 71-6
• 27 Marshall L A, Blazek E, Chang J. Characterization of an in vivo model for evaluation of phospholipase A₂ inhibitors.  Fed Proc. 1987;  46 854-62