Study Of Flurbiprofen Axetil Injection On Cardiac Sodium Channnel Under Normal And Ischemia-Reperfusion Conditions

BackgroudNon-steroidal anti-inflammatory drugs (NSAIDs) are widely used in anti-inflammation and analgesia; previous studies suggest that its effect mainly depends on its inhibition on cyclooxygenase (COX). However, some studies found that in addition to inhibiting COX, NSAIDs can play a variety of therapeutic effects and side effects, but the potential mechanisms are still unclear. Some NSAIDs have been shown to regulate the relevant ion channels, and these effects on ion channels appear to explain the clinical effects of relevant drugs, and such effects have no direct relationships with COX inhibition.Voltage-gated sodium channels play an important role in the initial stage of the depolarization during action potential (AP) in excitable cells, and thus regulating the sodium channels may become the important basis of the NSAIDs’treatment on pain and other excitatory state (such as epilepsy or cardiac arrhythmia).Flurbiprofen axetil (FA), the prodrug of flurbiprofen (non-acetylsalicylic acid salts), is lipophilic, and its injection type is widely used in clinical practice. FA injection is the composition of FA and lipid microspheres which can wrap FA. Lipid microspheres as a novel drug carrier system, they can make the FA injection has the characteristics of targeting (aggregated in the sites of inflammation), sustained efficacy and rapid onset of the drug. Although studies show that flurbiprofen may mediate its protective effects in the brain focal ischemia by inhibiting acid-sensitive ion channels, its effects on ion channels, particularly on the cardiac sodium channels remain unclear yet.ObjectivesThe present study was designed to investigate the features of the cardiac sodium channel (SCN5A) expressed in the human embryonic kidney (HEK)293cells, and to test its response to flurbiprofen axetil (FA). MethodsWild-type SCN5A gene and wild-type SCN1B gene were heterologously co-expressed in HEK293cells by transient co-transfection. Macroscopic voltage-gated INa was measured in the absence or presence of flurbiprofen axetil (0.15μM,0.5ml) after transfection with the standard whole-cell patch clamp technique in HEK293cells.ResultsThe transfection efficiency of sodium channel was about70%. Compared with no medication of FA, the sodium currents decreased at all the test potentials after using FA; at the test potential of-40mV, the peak sodium current decreased from-159.74±63.80(pA/pF) to-94.48±62.63(pA/pF)(n=7, p=0.012); the half-maximal voltage (V1/2) of the steady-state activation were-51.66±7.69mV and-58.19±2.45mV (n=7, p=0.048); V1/2of the steady-state inactivation were-83.55±3.21mV and85.25±4.78mV (n=7, p=0.041); channel recovery time constants ((?)) were33.86±23.18ms and67.71±58.58ms (n=7, p=0.048).ConclusionsFA decreased the peak sodium current and up-shifted the voltage-current curve; quickened the voltage-dependent activation of the sodium channel and promoted inactivation of voltage-dependent of the sodium channel; recovered slower after using FA. BackgroudNSAIDs are the most widely used drugs in clinical application, with anti-inflammatory, antipyretic, analgesic characteristics, and other effects. Although NSAIDs have a wide range of diverse structures, all have one thing in common:by inhibiting the COX activity, thereby preventing arachidonic acid ultimately from generating prostacyclin (PGI1), prostaglandin (PGE1, PGE2) and thromboxane A2(TXA2). In addition to inhibition of COX, NSAIDs can play a variety of therapeutic effects and side effects, but the potential mechanisms are still unclear. Some NSAIDs has been shown to regulate the relevant ion channels, and these effects on ion channels appear to explain the clinical effects of relevant drugs, and such effects have no direct relationships with COX inhibition.As the aforementioned studies show that FA decreased the peak sodium current and up-shifted the voltage-current curve; quickened the voltage-dependent activation of the sodium channel and promoted inactivation of voltage-dependent of the sodium channel; recovered slower after using FA. This kind of effect is similar to local anesthetics. Some studies suggest that FA can inhibit the inflammatory response during cardiopulmonary bypass, and thus reduce myocardial ischemia/reperfusion (I/R) injury. In addition to inhibiting the inflammatory response, whether there exists electrophysiological mechanisms of FA’s protection during myocardial I/R is not yet clear.ObjectivesThe study was designed to study the effects of flurbiprofen axetil (FA) management on the sodium current of the cardiomyocytes from the ventricles of the new-born rats at different stages of ischemia/reperfusion (I/R). MethodsThe cells were classified into four groups:group Ⅰ (the control group):the normal cardiomyocytes from new-born rats; group Ⅱ (the I/R group):the same cardiomyocytes that were treated under the condition of ischemia for three hours and reperfusion for one hour; group Ⅲ:the same cardiomyocytes that were treated with FA (0.15μM,0.5ml) during ischemia stage of I/R; group Ⅳ:the same cardiomyocytes that were treated with FA (0.15μM,0.5ml) during reperfusion stage of I/R. The sodium currents from the above four groups were recorded and compared by the whole-cell patch clamp technique.ResultsCompared with the control group, the sodium currents of all the cardiomyocytes from the I/R group at all the test potentials increased; at the test potential of-20mV, the peak sodium current increased from-375.47±70.31(pA/pF) to-557.11±127.86(n=5, p<0.05); the half-maximal voltage (V1/2) of the steady-state activation were-42.81±1.21mV and-31.49±2.40mV (n=5, p<0.05);1/2of the steady-state inactivation were-74.24±5.89mV and-68.70±6.26mV (n=5, p<0.05); channel recovery time constant were25.69±19.47ms and7.53±3.24ms (n=5, p<0.05); as for group Ⅲ and Ⅳ, the peak sodium current decreased from-375.47±70.31(pA/pF) to-208.80±121.44and-278.91±188.26(pA/pF)(n=5, p<0.05); V1/2of of the steady-state activation were-74.45±5.02mV and-75.49±3.32mV (n=5, p<0.05); V1/2of the steady-state inactivation were-58.77±3.83mV and-51.51±5.34mV (n=5, p<0.05); channel recovery time constants (τ) were824.98±16.41ms and26.30±11.82ms (n=5, p>0.05). The differences between the parameters from group Ⅲ and Ⅳ had no statistical significance.ConclusionsI/R treatment could increased the peak sodium current and down-shifted the voltage-current curve; slowed the time-dependent activation of the sodium channel and promoted inactivation of voltage-dependent of the sodium channel; recovered faster after the I/R management. FA management at both ischemia and reperfusion stage of I/R might inhibit changes of sodium current caused by I/R treatment.