Effect Of Valsartan On Myocardial Ca²⁺ Regulatory Proteins In Junior Rat With Heart Failure

[Background] Heart failure (HF) is a complicated clinical syndrome that leads to a disturbance of the normal pumping of blood to the peripheral organs to meet the metabolic demands of the body. For half a century, experts have been studying much about HF on pathogenesis and medicines, but the pathogenesis of chronic HF has still not been completely understood and the morbility and mortality of HF still increased. Patients with severe congestive heart failure (CHF), cardiac functionâ…¢-â…£class of NYHA, two-year mortality reached 50%. In pediatrics, HF is one of most common serious emergency cases and often result in death.So it's necessary to further explore the pathogenesis and look for effective medicine in children with HF.The homeostasis of Calcium(Ca²⁺) ions in cardiac muscle cell is important to maitain cardiac function.In cardiac excitation-contraction(E-C) coupling, a small amount of Ca²⁺ first enters through the L-type Ca²⁺ channel(LTCC) during membrane depolarization. This Ca²⁺ influx triggers a large-scale Ca²⁺ release through the Ca²⁺ release channel of the sarcoplasmic reticulum(SR), referred to as the ryanodine receptor(RyR), when the level of Ca²⁺ in cardiac muscle cell reaches 10-5mol/l, it binds to the troponin C within the myofilaments, which induces activation of the myofilaments and consequent muscle contraction. Relaxation is initiated by dissociation of Ca²⁺ from tropnin C, followed by its reuptake into the SR through phospholamban-regulated(PLB-regulated) Ca²⁺-ATPase (SERCA2a) (70﹪) and subsequent trans-sarcolemmal Ca²⁺ removal through the Na+/ Ca²⁺ exchanger(NCX)(28﹪). When the content of Ca²⁺ decreases to 10-7mol/l, myosin and actin separates, which resulting in the relaxation of cardiac muscle cell at last.Several lines of evdience have accumulated demonstrating that changes in activiy and express profile of Ca²⁺ handling proteins,in particular the SERCA2a are thought to cause an overall reduction in the amount of SR- Ca²⁺ available for contraction. In addition, abnormal Ca²⁺-leakage through the RyR2, has been demonstrated as a possible trigger for the development of heart failure.In those studies of heart failure, Ca²⁺-leak was found to be induced by the dissociation of FK506 binding protein (FKBP12.6) from the RyR2 as a result of the hyperphorylation of the RyR2.Data from our laboratory and others suggested that ?-adrenergic receptor (?-AR) blockade can correct the defective interaction of FKBP12.6 and RyR2 which is triggered by the protein kinase A (PKA)-mediated hyperphosphorylation of RyR2. Indeed this correction leads to a prevention of Ca²⁺-leak from SR. Angiotensinâ…¡receptor blocker(ARB) not only prevents hypertrophy and/or interstitial fibrosis in patients with HF, but also attenuates the downregulation of SERCA2a and improves intracellular Ca²⁺-handling.However, it donesn't know whether it can increase the affinity between Ca²⁺ and SERCA2a and correct the defective interaction of FKBP12.6 and RyR2. There are less reports about change of Ca²⁺ homeostasis in pediatric HF and the effect of ARB on SERCA2a and RyR2 so far.[Objective] To investigate the changes of Ca²⁺ regulatory proteins of SR, known as SERCA2a,PLB,RyR2 in junior rat with HF and the protective effect of angiotensinâ…¡receptor blockade(Valsartan) on these proteins.[Methods] The animal model of congestive heart failure was established by fistulation of abdominal aorta and inferior vena cava. Five weeks old rats were randomly divided into 3 groups: (1) HF group without treatment (n=15); (2) HF group treated with Valsartan (n=20); (3) Sham-operated group (n=15). Valstan was administered through direct gastric gavage after 8 weeks of operation, The high frequency ultrasound was performed after 4 weeks of treatment ,then the rats were killed, the atria were removed, the right and left ventricles were dissected, separated, and weighed. Some part of tissues were used to assess Cardiac histological changes after hematoxylin and eosin(HE) staining;SR was fractionated with velocity centrifugation, Serine Phosphate 16-PLB (Ser-16-PLB) and PLB were detected by immunohistochemical technique;Enzyme activity of SERCA2a in SR was detected with ultraviolet spectrophotometer(UV). the time course of Ca²⁺ uptake and leak were determined by fluorescent spectrophotometer.[Results] Compared with the sham-opreated group, left ventricular internal dimension diastole (LVIDd),left ventricular internal dimension systole(LVIDs),left ventricular end-diastolic volume(LVEDV),left ventricular end-systolic volume(LVESV),left ventricular ejection fraction(LVEF),left ventricular fractional shortening(LVFS)were all significantly increased (P<0.01), LVEF,LVFS were decreased (P<0.01) in HF group.LVIDd,LVIDs,LVEDV,LVESVwere all prominently decresed (P<0.01) ,LVEF,LVFS were inreased(P<0.01) in group of HF treated with Valsartan;Compared with the sham-operated group, the body weight decresed(P<0.01),left ventricular relative weigh(LVRW) and right ventricular relative weigh (RVRW) were all significantly increased(P<0.01)in HF group without treatment.LVRWand RVRW were decreased (P<0.01)in the group of HF treated with Valsartan when compared with that of HF group.Mean protein level (detected by Western blotting) of Ser-16-PLB in HF group were significantly lower(P<0.01) than that of sham-operated group, and it increased almost to normal level in HF group after treated with Valsartan.Enzyme activity of SERCA2a in HF group without treatment were significantly lower (P<0.01) than that of other groups.After adding ATP to the buffer including SR of three groups ,Ca²⁺ uptake were significantly decreased(P<0.01) in HF group,compaired with that of sham-operated group and group treated with Valsartan;After adding thapsigargin to the buffer of three groups,there were fewer Ca²⁺ leak in sham-poerated group and group treated with Valsartan (P<0.01),while after adding FK506 and thapsigargin together to the buffer including SR of three groups,there were markedly Ca²⁺ leak in sham-operated group and HF group treated with Valsartan (p<0.01),but there was no additional increase in Ca²⁺ leak in HF group compared with the value when only thapsigargin was added(P>0.05).[Conclusion] Heart function and enzyme activity of SERCA2a decrease in junior rat with heart failure, there are less Ca²⁺ uptake and more Ca²⁺ leak in HF. Valsartan not only improved Ca²⁺ uptake by increasing ser-16-PLB level, but also prevented protein kinase A(PKA) from hyperphosphorylating, rendering the SR less susceptible to Ca²⁺ leak,contributing to the recovery of contractility and relaxation of the cadiomyocyte.