We have evaluated the putative analgesic and anti-inflammatory activities of hispolon to clarify the pain and inflammation relieving effects. Two different analgesic testing methods were employed with the objective of identifying possible peripheral and central effects of the test substances. The acetic-acid writhing test is normally used to study the peripheral analgesic effects of drugs. Although this test is non-specific (e.g. anticholinergic, antihistaminic and other agents also show activity in the test), it is widely used for analgesic screening [26
]. In our study, we found that hispolon (10 and 20
mg/kg) exhibited antinociceptive effect in acetic-acid-induced writhing response (). This effect may be due to inhibition of the synthesis of the arachidonic acid metabolites [27
model of pain, formalin-induced paw pain has been well established as a valid model for analgesic study. The formalin test produces a distinct biphasic response and different analgesics may act differently in the early and late phases of this test. Therefore, the test can be used to clarify the possible mechanism of an antinociceptive effect of a proposed analgesic [28
]. Centrally acting drugs such as opioids inhibit both phases equally [26
], but peripherally acting drugs such as aspirin, Indo and dexamethasone only inhibit the late phase. The inhibitory effect of hispolon on the nociceptive response in the late phase of the formalin test suggested that the anti-nociceptive effect of the hispolon could be due to its peripheral action ().
The Carr test is highly sensitive to non-steroidal anti-inflammatory drugs, and has long been accepted as a useful phlogistic tool for investigating new drug therapies [29
]. The degree of swelling of the Carr-injected paws was maximal 3
h after injection. Statistical analysis revealed that hispolon and Indo significantly inhibited the development of edema 4
h after treatment (P
.001) (). They both showed anti-inflammatory effects in Carr-induced mice edema paw. It is well known that the third phase of the edema-induced by Carr, in which the edema reaches its highest volume, is characterized by the presence of prostaglandins and other compounds of slow reaction [30
]. It was found that the injection of Carr into the rat paw induces the liberation of bradykinin, which later induces the biosynthesis of prostaglandin and other autacoids, which are responsible for the formation of the inflammatory exudates. In addition, the classification of antinociceptive drugs is usually based on their mechanism of action either on the central nervous system or on the peripheral nervous system [31
In the studies of mechanism on the inflammation, l
-arginine-NO pathway has been proposed to play an important role in the Carr-induced inflammatory response [32
]. Our present results also confirm that Carr-induced paw edema model results in the production of NO. The expression of the inducible isoform of NO synthase has been proposed as an important mediator of inflammation [33
]. In our study, the level of NO was decreased significantly by treatment with 10 and 20
mg/kg hispolon. We suggest the mechanism of anti-inflammatory of hispolon may be through the l
-arginine-NO pathway since hispolon significantly inhibit the NO production ().
is a major mediator in inflammatory responses, inducing innate immune responses by activating T cells and macrophages, and stimulating secretion of other inflammatory cytokines [34
]. Also, TNF-α
is a mediator of Carr-induced inflammatory incapacitation, and is able to induce the further release of kinins and leukotrienes, which is suggested to have an important role in the maintenance of long-lasting nociceptive response [35
]. In this study, we found hispolon decreased the TNF-α
level in serum after Carr injection ().
The Carr-induced inflammatory response has been linked to neutrophils infiltration and the production of neutrophils-derived free radicals, such as hydrogen peroxide, superoxide and hydroxyl radicals, as well as the release of other neutrophils-derived mediators [36
]. Some researches demonstrate that inflammatory effect induced by Carr is associated with free radical. Free radical, prostaglandin and NO will be released when administrating with Carr for 1–6
]. The edema effect was raised to the maximum at the third hour [13
]. Janero demonstrated that MDA production is due to free-radical attack on plasma membrane [37
]. Thus, inflammatory effect would result in the accumulation of MDA. Glutathione is a known oxyradical scavenger. Enhances the level of Glutathione conducive toward favor reduces MDA the production. Cuzzocrea [39
] suggested that endogenous glutathione plays an important role against Carr-induced local inflammation. In this study, significantly increase in SOD, GRx and GPx activities with hispolon treatment was found (). Furthermore, there were significantly decreases in MDA level with hispolon treatment (). We assume that the suppression of MDA production is probably due to the increase of SOD, GRx and GPx activities.
In conclusion, these results suggested that hispolon possessed analgesic and anti-inflammatory effects. The anti-inflammatory mechanism of hispolon may be related to iNOS and it is associated with the increase in the activities of antioxidant enzymes (SOD, GPx and GRx). Hispolon may be used as a pharmacological agent in the prevention or treatment of disease in which free radical formation is a pathogenic factor.