| Objectives:To investigate the application of sequential therapy of Recombinant human brain natriuretic peptide(rhBNP)and furoxime in HFREF rats by determining the expression of aquaporin-2(AQP2)and the concentration of AQP2 in the urine of reduced ejection fraction(HFREF)rats.To observe the effect of rhBNP on diuretic effect and its formation mechanism,so as to provide a theoretical basis for clinical application of rhBNP to improve HFREF fluid retention and the best effect of diuretic resistance.Methods:We established the model of CHF after acute myocardial infarction by ligating the left anterior descending of male Wistar rats.Four weeks after the operation,the survival rats were detected by ultrasound(Ejection Fraction,EF<55%),suggesting that the model was successfully established.The successfully established model rats were randomly divided into the model group,the rhBNP group,the furosemide group,and the rhBNP+furosemide group.Another sham-operation group was set up.The blood and urine samples were collected at 4 weeks after intervention,and the 24-hour liquid intake and output of the rats were also recorded at 2 and 4 weeks after intervention.Plasma detection:Angiotensin II(AngII),Aldosterone(ALD),Arginine Vasopressin(AVP),and N-terminal pro brain natriuretic peptide(NT-proBNP).Urine detection: concentration of AQP2 and creatinine.The heart function of rats was evaluated by ultrasonic examination at 4 weeks after intervention.Meanwhile,the expression of AQP2 in rats’ kidney was detected by immunohistochemistry(ICH),western blot(WB)and Real-time quantitative Polymerase Chain Reaction(RT-PCR).Results:1.Concentration of AngII,ALD,AVP and NT-proBNP in rats’ plasma: 4weeks after the intervention,the 2-week rhBNP+ 2-week furosemide group and the rhBNP+ furosemide group were lower than the model group and the 2-week furosemide +2-week rhBNP group(P<0.05),but were higher than that of the sham-operation group(P <0.05).For the first three indicators : the 2-week furosemide + 2-week rhBNP group was higher than that of the model group(P <0.05);no statistical difference was found between the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group(P > 0.05).For the last indicator : no statistical differences were found between the following comparisons: the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group,the 2-week furosemide + 2-week rhBNP group and the model group(P>0.05).2.Excretion rate of AQP2(urine AQP2 concentration/creatinine)in the urine of rats: 4 weeks after intervention,the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group were lower than that of the model group and the 2-week furosemide + 2-week rhBNP group(P<0.05),but were higher than the sham-operation group(P < 0.05);the 2-week furosemide + 2-week rhBNP group was higher than the model group(P < 0.05);no significant difference was found between the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group(P > 0.05).3.Difference of the 24-hour intake and output in rats: 2 weeks after intervention,the 2-week furosemide + 2-week rhBNP group and the rhBNP+furosemide group were higher than that of the 2-week rhBNP+ 2-week furosemide group(P < 0.05).No significant difference was found between the 2-week furosemide + 2-week rhBNP group and the rhBNP+furosemide group(P > 0.05).The first three groups were higher than the sham-operation group and the model group(P < 0.05),the sham-operation group was higher than the model group,and the difference was statistically significant(P < 0.05).4weeks after intervention,the rhBNP+furosemide group was higher than that of the 2-week rhBNP+ 2-week furosemide group and the 2-week furosemide + 2-week rhBNP group(P <0.05);the 2-week furosemide + 2-week rhBNP group was higher than the 2-week rhBNP+2-week furosemide group(P < 0.05).Meanwhile,the first three groups were higher than the sham-operation group and the model group(P < 0.05).At the same time,the sham-operation group was higher than the model group,and the difference was statistically significant(P < 0.05).4.Ultrasound results of rats at 4 weeks after intervention: Comparison of left ventricular end diastolic diameter(LVEDD)and left ventricular end systolic diameter(LVESD): the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group were lower than that of the model group and the 2-week furosemide + 2-week rhBNP group(P < 0.05),but were higher than the sham-operation group(P<0.05).However,no statistical differences were found between the following comparisons: the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group,the 2-week furosemide + 2-week rhBNP group and the model group(P>0.05).Comparison of ejection fraction(EF)and shortening fraction(FS): the 2-week rhBNP+2-week furosemide group and the rhBNP+furosemide group were higher than that of themodel group and the 2-week furosemide + 2-week rhBNP group(P<0.05),but were lower than that of the sham-operation group(P<0.05).However,no statistical significant differences were found between the following comparisons: the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group,the 2-week furosemide + 2-week rhBNP group and the model group(P > 0.05).5.The expression of AQP2 protein or mRNA levels in rats’ kidney at 4weeks after intervention: the 2-week rhBNP+ 2-week furosemide group and the rhBNP+furosemide group were lower than that of the model group and the 2-week furosemide + 2-week rhBNP group(P < 0.05),but were higher than the sham-operation group(P < 0.05).The 2-week furosemide + 2-week rhBNP group was higher than the model group(P < 0.05);no significant difference was found between the 2-week rhBNP+2-week furosemide group and the rhBNP+furosemide group(P > 0.05).Conclusions:The combination of rhBNP and furosemide was used in rats with heart failure with reduced ejection fraction.rhBNP could antagonize the over activation of the neuroendocrine system and reduce the excretion rate of urine AQP2 as well as the expression of protein and mRNA of AQP2,decrease the renal tubular reabsorption of sodium and water,enhance the diuretic effect of furosemide,and improve the retention of water and sodium and heart function.The earlier use it,the greater benefit obtain. |
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