| Nitric oxide (NO) plays an important role in the regulation of cardiovascular function. The half-life of NO is very short, limiting its biological effects. In addition to the classic NO activation of the cGMP-dependent pathway, NO can also regulate cell function through protein S-nitrosylation, a redox dependent, thiol-based, reversible posttranslational protein modification that involves attachment of an NO moiety to a nucleophilic protein sulfhydryl group. Stamler and colleagues identified thiols as the storage substance for NO in 1992. Gabor proved that RSNO help to store NO from his study of the formation and degradation rate of RSNO under neutral pH.Protein S-nitrosylation not only leads to changes in protein structure and function, and prevent further irreversible thiol oxidation/nitration modification. Once protein is S-nitrosylated, the activity of protein is enhanced or inhibited, which is an important post-translational modification, mediating a wide range of functions by NO. RSNO serves as second messenger and effector as well, and plays a central role of NO. The s-nitrosylation mechanism help explain the wide range of NO effects in the cardiovascular system. The signal transduction proteins including ion channel proteins, G protein receptor proteins, and nuclear regulatory proteins contain sulfhydryl groups, transfering of thiols between proteins play a major role in regulating cell signaling involved in NO activity on the cardiovascular system.ROS (reactive oxygen species, ROS) and reactive nitrogen clusters (reactive nitrogen species, RNS) disequilibrium is closely related cardiovascular disease. ROS and RNS have chemical interactions or synergistic effects on the target protein of the signal pathway. Free radicals attack the polyunsaturated fatty acids (PUFA) in the tissue, inducing lipid peroxidation, and formation of malondialdehyde (MDA) etc. A variety of antioxidant mechanisms, such as superoxide dismutase (SOD) can prevent this type of injury. NO/redox disequilibrium reflected as the increased oxidation activity in the heart (xanthine oxidase), vascular (NADPH oxidase), red blood cells (hemoglobin oxidase), inhibit endogenous NO-based signaling pathways and promote cardiovascular failure. Remodeling of dilated cardiomyopathy can be reversed by targeted therapy of xanthine oxidase in rats. Therefore, the resumption of NO/redox balance offers a potentially effective method of maintaining cardiovascular function.The infants and young children with congenital heart disease (CHD) undergoing open-heart surgery, including ventricular septal defect without pulmonary hypertension (VSD group), ventricular septal defect with pulmonary hypertension (VSD+PH group) and Tetralogy of Fallot (TOF group) were included in our study. We evaluated the activity of perioperative plasma NO/cGMP pathway and RSNO and oxidative stress levels, as well as the perioperative hemodynamics. This study is to identify the changes and relationship between perioperative plasma RSNO levels and hemodynamics in infants and young children with CHD undergoing open-heart syrgery. A better understanding of the mechanism regulating protein S-nitrosylation and its role in cardioprotection will provide us new therapeutic opportunities and targets for interventions in cardiovascular diseases. Part I Perioperative hemodynamic changes in infants and young children undergoing open-heart surgeryAim:1. The clinical application of PiCCO in evaluating the hemodynamic changes in infants and young children after open-heart surgery.2. To analyze the respective hemodynamic characteristics in different types of congenital heart disease after open-heart surgery.Methods:1. Design and patientsThis is a prospective study. Consecutive 30 cases of 0-3 years old pediatric patients with congenital heart disease undergoing open heart surgery from August 2009 to June 2010 were included in this study. There were 10 cases with Tetralogy of Fallot,10 cases with VSD and pulmonary hypertension, and 10 cases with VSD without pulmonary hypertension. Exclusion criteria include severe coagulopathy, contraindications for femoral artery thermodilution catheterization, postoperative residual shunt and severe cardiac valve insufficiency, which may have impact on the results of this study.2. PiCCO determination of hemodynamicsPiCCO analyzer continuously measured hemodynamic data on 0h,4h,8h,12h,24h, 48h,72h after operation, including mean arterial pressure (MAP), heart rate (HR), cardiac index (CI), systemic vascular resistance index (SVRI), pulmonary vascular resistance index (PVRI), dedicated end-diastolic volume index (GEDVI), intrathoracic blood volume index (ITBVI), extravascular lung water index (EVLWI), the permeability of pulmonary vascular permeability index (PVPI).3. pHi measurement methods:tonometric catheter was connected to determine PgCO2 pHi=7.40-log PgCO2/PaCO24. LAP, CVP, gastric pHi, PAP, PVRI were measured and recorded on 0h,4h,8h, 12h,24h,48h,72h after surgery; inotropic score was calculated and recorded on 0h,4h, 8h,12h,24h,48h,72h after surgery:inotropic score=1x Dobutamine+1x Dopamine+ 10x Milrinone+100x epinephrine.5. Plasma CKMB activity, CKMB mass, troponin were tested before surgery, and on 4h,24h,48h,72h after syrgery.6. Postoperative course and clinical outcome(1) Plasma lactate levels and PaO2/FiO2 were record on 0h,4h,8h,12h,24h,48h, 72h after surgery.(2) Complications were recorded, such as low cardiac output syndrome, arrhythmia, pulmonary hypertension and crisis, capillary leak syndrome, atelectasis, pleural effusion, bleeding, infection, pneumothorax, non-invasive CPAP, re-intubation, etc.(3) The duration of mechanical ventilation, ICU stay, hospital stay were recorded and compared between groups.Results:1. The patients'demographics(1) There were 10 patients in VSD group with an age of 15.6±3.2 months and a weight of 9.9±1kg; There were 10 patients in VSD+PH group with an age of 11.4±3.2 months and a weight of 7.1±0.8kg; There were 10 patients in TOF group with an age of 13.6±2.8 months and a weight of 8.7±0.9kg.(2) There was no significant difference of age among the three groups (P>0.05); the body weight of patients in VSD group and the TOF group was significantly higher than that of VSD+PH group (P<0.05); the cardiopulmonary bypass time, aortic clamping time was significantly shorter in VSD group than in VSD+PH and TOF group (P<0.05); the duration of mechanical ventilation, ICU stay, hospital stay was significantly shorter in VSD group than in VSD+PH and TOF group (P<0.05).2. The CKMB activity, CKMB mass, troponin levels were significantly higher in TOF and VSD+PH group than that of VSD group. Spearman correlation analysis showed CKMB activity, CKMB mass, troponin was positively correlated with SVRI, PVRI, inotropic score, lactate and negatively correlated with CI, ITBVI, GEDVI, PaO2/FiO2.3. The cardiac index in TOF patients decreased significantly to the lowest nadir on 8h-12h after surgery, and was significantly improved on 24h after surgery. The CI of TOF group was significantly lower than the VSD group accopanied with significantly higher SVRI and inotropic score as well as higher lactate levels.4. VSD+PH group of patients have significantly higher mPAP and PVRI than that of VSD and TOF group, while the mPAP and PVRI of TOF group showed no significant difference with the VSD group.5. The GEDVI and ITBVI was significantly lower in TOF and VSD+PH group than VSD group during the early postoperative period.6. The EVLWI was significantly higher in VSD+PH group than TOF group at 48h after surgery, whereas the EVLWI was significantly higher in TOF group than VSD group.Conclusions:1. The patients in TOF and the VSD+PH group showed more severe myocardial damage than in VSD group patients, with decreased CI, increased SVRI, poor tissue perfusion, higher inotropic score.2. The GEDVI and ITBVI was significantly lower in TOF and VSD+PH group than in VSD group in the early period after surgery.3. VSD+PH group of patients have significantly higher mPAP and PVRI than that of VSD and TOF group, while the mPAP and PVRI of TOF group showed no significant difference with the VSD group.4. The EVLWI was significantly higher in VSD+PH group than TOF group at 48h after surgery, whereas the EVLWI was significantly higher in TOF group than VSD group.5. Spearman rank correlation test also showed a positive correlation between EVLWI and PVPI. There was a positive correlation between EVLWI and ITBVI and GEDVI, and a negative correlation with PaO2/FiO2. The PVPI was negatively correlated with CI, ITBVI and GEDVI decreased with the increased pulmonary vascular permeability, and showed a negative effect on PaO2/FiO2.Part?The perioperative activity of plasma NO/cGMP pathway and nitrosothiols and oxidative stress with hemodynamics in infants and young children undergoing open-heart surgeryAim:1. To identify the activity of perioperative NO/cGMP pathway and plasma nitrosothiols in infants and young children undergoing open-heart surgery.2. To identify the activity of perioperative oxidative stress levels in infants and young children undergoing open-heart surgery.3. To evaluate the relationship between plasma nitrosothiols and NO/cGMP pathway and redox activity in infants and young children undergoing open-heart surgery.4. Statistical analysis of perioperative plasma RSNO levels and hemodynamic data in infants and young children with congenital heart disease undergoing open-heart surgery, to underly the molecular mechanisms of protein nitrosylation/oxidation balance in regulating cardiovascular function in infants and young children undergoing open-heart surgery.Methods:1. Design and patients Same as part?.2. The collection of blood samplesBlood samples were collected on:1) before surgery; 2) 4h post-op; 3) 24h post-op; 4) 48h post-op; 5) 72h post-op; 3ml blood samples (per lml of blood containing heparin 10u) were collected, and instant centrifugated at 4?,3000 r/min for 10 minutes. Plasma was separated and stored in liquid nitrogen and light protected. The blood was drawn before diet to avoid its impact on nitrite and nitrate.3. Plasma RSNO, cGMP, NO2-/NO3-, MDA, SOD levels were measured.4. The mathmatic correction of the experimental data Corrected value= measured value x preoperative HCT/sample HCT5. Statistical analysisThe corrected experimental data were expressed as mean±standard error. A paired sign test was used to compare between groups. Spearman rank correlation test was used to evaluate the correlations between plasma RSNO, cGMP, NO2-/NO3-, MDA, SOD levels. P<0.05 was of significant difference. These statistical analyses were completed by the 15.0 statistical software SPSS for windows.Results:1. The 4h post-op plasma RSNO level were significantly higher compared with preoperative RSNO level in VSD group (190±21 pA VS 129±16 pA, P<0.05). There were no significant differences of the plasma RSNO levels between other groups as well as all other time points (P>0.05).2. Four hours after operation, the cGMP levels were significantly lower in VSD+ PH and TOF group than in VSD group(p<0.05). 3. Plasma NO2-/NO3- levels were significantly decreased after operation in 3 groups.4. Plasma MDA levels were significantly increased in VSD group at 4h after operation; preoperatively, the plasma MDA levels were significantly higher in VSD+ PH group than in TOF group (P<0.05).5. Plasma SOD levels were significantlydecreased in TOF group at 4h after operation, and were significantly lower than VSD+PH group.6. The relationship between perioperative plasma nitrosothiols and NO/cGMP pathway activity and redox levels:(1) Spearman rank correlation test of plasma RSNO levels of all patients at all time point with NO/cGMP pathway activity (cGMP, NO2-/NO3-) and redox molecular level (MDA, SOD):1) There was a positive correlation between plasma RSNO and cGMP levels, the correlation coefficient r=0.22, P=0.0078.2) Plasma RSNO levels positively correlated with plasma MDA level, the correlation coefficient r=0.35, P=0.0001.3) cGMP levels in plasma were positively correlated with MDA levels, the correlation coefficient r=0.22, P=0.0059.(2) Spearman rank correlation test of plasma RSNO levels of all time point with NO/cGMP pathway activity and redox molecular level (MDA, SOD) in each group:1) VSD Group:there is a positive correlation between plasma RSNO and cGMP levels (correlation coefficient=0.45, P=0.0015); RSNO levels were positively correlated with MDA level (correlation coefficient=0.42, P=0.0026); cGMP levels were positively correlated with MDA level (correlation coefficient-0.31,P=0.03).2) VSD+PH group:plasma RSNO level and MDA level positively correlated (correlation coefficient= 0.49, P= 0.0004), plasma SOD levels were positively correlated with cGMP levels (correlation coefficient=0.37, P=0.0091). 3) There was no correlations in TOF group (P>0.05).7. The relationship between perioperative plasma nitrosothiols and hemodynamics:(1) Spearman rank correlation test showed that among 3 groups of congenital heart disease undergoing open-heart surgery for all time points, the plasma RSNO levels showed no significant correlation with all the hemodynamic data.(2) Regardless of time point, we compared the relationship between plasma RSNO levels and hemodynamic data in VSD, VSD+PH, TOF group respectively, the results showed:1) VSD group:plasma RSNO levels were negatively correlated with PVRI (correlation coefficient r=-0.66, P=0.0052).2) VSD+PH group:plasma RSNO levels were negatively correlated with CI (correlation coefficient r=-0.37, P=0.0230).3) TOF group:there was no significant correlation between plasma RSNO levels and hemodynamic data.(3) We compared the relationship between plasma RSNO levels and hemodynamic data in VSD, VSD+PH, TOF group of patients in different time points, the results showed:1) VSD group:?0h post-op:plasma RSNO levels positively correlated with pHi (correlation coefficient r=0.52, P=0.027);?4h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?24h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?48h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?72h post-op:plasma RSNO levels positively correlated with PVPI (correlation coefficient r=0.58, P=0.022).2) VSD+PH group:?0h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?4h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?24h post-op:plasma RSNO levels negatively correlated with mPAP (correlation coefficient r=-0.91, P=0.011);?48h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?72h post-op:plasma RSNO levels showed no correlation with the hemodynamics.3) TOF group:?0h post-op:plasma RSNO levels positively correlated with pHi (correlation coefficient r=0.94, P=0.005);?4h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?24h post-op:plasma RSNO levels negatively correlated with CKMB activity (correlation coefficient r=-0.67, P=0.05);?48h post-op:plasma RSNO levels showed no correlation with the hemodynamics;?72h post-op:plasma RSNO levels positively correlated with PVPI (correlation coefficient r =0.76, P=0.031).Conclusions:1. There was transient NO/cGMP pathway inhibition in VSD, VSD+PH, TOF group of patients early after open heart surgery.2. This study shows that plasma RSNO levels has a positive linear relationship with NO/cGMP pathway and oxide stress.3. Plasma RSNO plays an important role in regulating perioperative heart function and pulmonary vascular resistance in infants and young children with congenital heart disease undergoing open-heart surgery. |
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