Chrysin Protects Secondary Myocardial Injury In Rats Subjected To Mechanical Trauma

Objective:Moderate to severe mechanical trauma causes cardiomyocyte apoptosis which contributes to posttraumatic cardiac injury.Previously study demonstrated that MT can cause body exhibit strongly oxidative reactive response and the overproduction of inflammatory cellular factors.Among the inflammatory cellular factors,TNF-? plays a causative role in cardiomyocyte apoptosis.Chrysin is a natural flavonoid contained in propolis,blue passion flower,and fruits.Several studies reported that chrysin has beneficial effects including anti-oxidant and anti-inflammatory activities.The present study was to elucidate whether chrysin alleviate MT-induced cardiac secondary injury and to study its possible mechanisms of action in vitro and in vivo models.Method:Dose dependence of chrysin-evoked responses on normal cardiomyocytes and the protective responses on TNF-?-treated cardiomyocytes were all detected by MTT,then the most appropriate chrysin concentration was determined.To identify the cardiac function,anesthetized rats were placed in supine position,the right common carotid artery intubation was adopted to record the Left Ventricular End Diastolic Pressure?LVEDP?,the Left Ventricular Developed Pressure?LVEDP?,the Mean Arterial Pressure?MAP?,left ventricular pressure rise and fall at the maximum rate of changes(±d P/d T_max).Myocardial tissues were made into frozen sections and stained with TUNEL,then DAPI/TUNEL fluorescent images were photoed and the apoptotic index was calculated.TNF-? assay kit was used to identify the TNF-? level both in vivo and vitro.DCFH-DA was used to estimate the intracellular generation of ROS.The dye is a non-polar compound that readily penetrates cells.Intracellular peroxides oxidize DCFH-DA to a highly fluorescent compound?DCF?.To observe the Ca²⁺ alteration,confocal laser scanning microscope photoed fluorescent images.Main results:1.Mechanical trauma model with 200 revolutions lead to myocardial injury LVDP ? MAP ? +d P/d T_max ?-d P/d T_max of rats in trauma group?200r?were all significantly decreased as compared to the sham group.2.In vitro,the most appropriate concentration of chrysin acted on cardiomyocytes was determinedCompared with the control group,incubation of normal cardiomyocytes with chrysin at 20 u M,40 u M or 80 u M respectively,chrysin dose dependently reduced the cell viability,where as,incubation with chrysin at 10 u M,the cell viability was not a significant change.The treatment of cardiomyocytes with TNF-? to mimic inflammatory conditions induced significantly decrease in the cell viability.Pretreatment with chrysin significantly increased the cell viability,compared with that in TNF-? treated group.3 Chrysin alleviated myocardial injury induced by mechanical trauma LVDP?MAP?+d P/d T_max?-d P/d T_max of rats in trauma group were all significantly decreased as compared to the sham group.LVDP?MAP?+d P/d T_max?-d P/d T_max of rats in trauma with chrysin-treated group were significantly increased,compared with that in the trauma group.4.Chrysin reduced myocardial apoptosis induced by mechanical trauma Myocardial apoptosis index of rats in trauma group was significantly increased as compared to the sham group.Pretreatment with 30mg/kg chrysin significantly attenuated the myocardial apoptosis index of rats as compared to the trauma group.5 Chrysin attenuated TNF-? levels in trauma plasma and also reduced TNF-? levels in LPS-stimulated monocytes TNF-? levels in trauma plasma was significantly higher as compared to sham group.Pretreatment with chrysin caused a significant decrease in TNF-? expression as compared to trauma group.Resting monocytes exhibited low levels of TNF-?,the treatment of monocytes with LPS to mimic inflammatory conditions induced an significantly increase in TNF-? expression,compared with that in control group.Pretreatment with chrysin significantly suppressed the LPS-stimulated expression of TNF-?,compared with that in LPS-treated group.6.Chrysin attenuated intracellular ROS generation in LPS-induced monocytes and also reduced intracellular ROS generation in TNF-?-induced myocardial cells The stimulation of monocytes with LPS to mimic inflammatory conditions induced significantly increase in intracellular ROS production,compared with that in control group.Treatment of chrysin prior to incubation with LPS significantly inhibited intracellular ROS generation,compared with that in LPS-treated group.There was an approximately increase in intracellular ROS production upon stimulation of the cardiomyocytes with TNF-?,compared with that in control group.Treatment of chrysin prior to incubation with TNF-? significantly inhibited intracellular ROS generation,compared with that in TNF-?-treated group.7.Chrysin attenuated intracellular Ca²⁺ overload induced by trauma plasma Stimulation with trauma plasma led to a significant increase of fluorescence intensity in myocardial cells,compared with that in control group.Pretreatment with chrysin prior to incubation with trauma plasma significantly attenuated the fluorescence intensity in myocardial cells,compared with that in trauma plasma group.Conclusion:These results indicate that chrysin alleviates cardiac function by inhibiting TNF-?-induced apoptosis via attenuates the intracellular ROS generation and Ca²⁺overload.We propose chrysin as a new therapeutic strategy for treating MT-induced cardiac secondary injury.