Much recent research has demonstrated that honokiol, a phenolic compound originally isolated from Magnolia officinalis, has potent anticancer activities; however, the detailed molecular mechanism of its anti-inflammatory activity has not yet been fully addressed. In this study we demonstrated that honokiol inhibited lipopolysaccharide (LPS)-induced tumor necrosis factor-alpha secretion in macrophages, without affecting the activity of the tumor necrosis factor-alpha converting enzyme. At the same time, honokiol not only inhibited nitric oxide expression in LPS-stimulated murine macrophages but also inhibited the LPS-induced phosphorylation of ERK1/2, JNK1/2, and p38. By means of confocal microscope analysis we demonstrated that phosphorylation and membrane translocation of protein kinase C-alpha, as well as NF-kappaB activation, were inhibited by honokiol in LPS-stimulated macrophages. Furthermore, it was found that honokiol neither antagonizes the binding of LPS to cells nor alters the cell surface expression of toll-like receptor 4 and CD14. Our current results have exhaustively described the anti-inflammatory properties of honokiol, which could lead to the possibility of its future pharmaceutical application in the realm of immunomodulation.

The assembly of four pore-forming ␣-subunits into tetramers is a prerequisite for the formation of functional K ؉ channels. A short carboxyl assembly domain (CAD) in the distal end of the cytoplasmic carboxyl terminus has been implicated in the assembly of Eag ␣-subunits, a subfamily of the ether-à-go-go K ؉ channel family. The precise role of CAD in the formation of Eag tetrameric channels, however, remains unclear. Moreover, it has not been determined whether other protein regions also contribute to the assembly of Eag subunits. We addressed these questions by studying the biophysical properties of a series of different rat Eag1 (rEag1) truncation mutants. Two truncation mutants without CAD (K848X and W823X) yielded functional phenotypes similar to those for wild-type (WT) rEag1 channels. Furthermore, nonfunctional rEag1 truncation mutants lacking the distal region of the carboxyl terminus displayed substantial dominant-negative effects on the functional expression of WT as well as K848X and W823X channels. Our co-immunoprecipitation studies further revealed that truncation mutants containing no CAD indeed displayed significant association with rEag1-WT subunits. Finally, surface biotinylation and protein glycosylation analyses demonstrated that progressive truncations of the carboxyl terminus resulted in aggravating disruptions of membrane trafficking and glycosylation of rEag1 proteins. Overall, our data suggest that the distal carboxyl terminus, including CAD, is dispensable for the assembly of rEag1 K ؉ channels but may instead be essential for ensuring proper protein biosynthesis. We propose that the S6 segment and the proximal carboxyl terminus may constitute the principal subunit recognition site for the assembly of rEag1 channels.

The ability to maintain optimal intracellular pH (pH i )is an essential requirement for all cells. A wealth of biological processes have been found to be sensitive to changes in pH i . These include enzyme activity [1], control of the cell volume [2], the regulation of cellular growth and differentiation [3], and the kinetic properties of Na ϩ , K ϩ , and Ca 2ϩ channels [4]. The pH i regulatory mechanism is especially important for the heart, since changes in pH i dramatically affect contractility and rhythm [5][6][7]. Moreover, it has been demonstrated that during myocardial ischemia, the pH i is substantially lowered [8], but during postischemia reperfusion, a rapid recovery of the pH i is observed [9]. Therefore these pH i disturbances have been claimed to be responsible for the reversible contractile dysfunction and malignant ventricular arrhythmias seen in cardiac myocytes [9].In animal cardiac cells, remarkable progress has been made in understanding the pH i regulating system over the past decade. It is well known now that the pH i in animal cardiac cells is kept within a narrow range (7.0-7.2) by a combination of active sarcolemmal transporters, i.e., intracellular acid-extrusion and acidloading carriers, and the passive intracellular buffering capacity [10, 11]. Na ϩ -H ϩ exchange (NHE) is important for pH i control in the animal heart, being one

Abstract. Hydrogen peroxide (H 2 O 2 ) and its metabolites have been shown to exert complex effects on the cardiac muscle during cardiac ischemia / reperfusion. The aim of the present study, by perfusing H 2 O 2 or / and different scavengers of oxygen free radicals (OFRs) into the human atrium, is to characterize the electropharmacological effects of H 2 O 2 and explore its possible underlying mechanism. Atrial tissues obtained from the heart of 19 patients undergoing corrective cardiac surgery were used. Transmembrane action potentials were recorded using the conventional microelectrode technique, and contraction of atrial fibers was evaluated in normal [K] o (4 mM) in the absence and presence of tested agents. H 2 O 2 (30 m M-3mM) had a biphasic effect on the contractile force (an increase, followed by a decrease), reduced the 0-phase depolarizing slope (dV/ dt), and prolonged the action potential duration (APD) in a concentrationdependent manner. However, even at a concentration as high as 3 mM, H 2 O 2 did not influence diastolic membrane potential (DMP). Pretreatment with N-(mercaptopropionyl)-glycine (N-MPG), a specific scavenger of the · OH free radical, significantly blocked the 3 mM H 2 O 2 -induced electromechanical changes, while the pretreatment with L-methionine (L-M), a specific scavenger of HOCl free radical, did not. Our data suggests that the toxic effects of H 2 O 2 are caused mainly through the generation of · OH, which is attributed to the electropharmacological inhibitory effects seen in the human atrium.