The Influence Of Volume Overload After Artificial Shunt Of Left To Right On Rat Right Ventricle Remodeling

Objective Remodeling concerns the two components of the cardiovascular system. The structure of both the myocardium and the vessels, including the coronary vessels, is indeed able to change under the influence of external factors, such as ischemia and mechanical overload. Cardiac remodeling is triggered by mechanical stretch; nevertheless, there are also several different factors including ischemia, hormones, and vasoactive peptides, which can modify the effects of the mechanical factor. The most common clinical situation during which remodeling is known to occur is a rather complex mixture of ischemia, stretch due to pressure overload, stress due a myocardial scar, and increased plasma levels of hormones or vasoactive peptides.From a biological point of view, CR is determined by 1 ) the general process of adaptation which allows both the myocyte and the collagen network to adapt to new working conditions; 2) ventricular fibrosis, i.e., increased collagen concentration, which is multifactorial and caused by senescence, ischemia, various hormones, and/or inflammatory processes; 3) cell death, a parameter linked to fibrosis, which is usually due to necrosis and apoptosis and occurs in nearly all models of CR. The process of adaptation is associated with various changes in genetic expression, including a general activation that causes hypertrophy, isogenic shifts which result in the appearance of a slow isomyosin, and a new Na⁺-K⁺-ATPase with a low affinity for sodium, reactivation of genes encoding for atrial natriuretic fator and the renin-angiotensin system, and a diminished concentration of sarcoplasmic reticulum Ca²⁺-ATPase, (?)-adrenergic receptors, and the potassium channel responsible for transient outward current.From a clinical point of view, fibrosis is for the moment a major marker for cardiac failure and a crucial determinant of myocardial heterogeneity, increasing diastolic stiffness, and the propensity for reentry arrhythmias. In addition, systolic dysfunction is facilitated by slowing of the calcium transient and the downregulation of the entire adrenergic system. Modifications of intraceliular calcium movements are the main determinants of the triggered activity and automaticity that cause arrhythmias and alterations in relaxation.Pathologic remodeling is a maladaptive process that, left unimpeded, progresses to left ventricular failure. Remodeling is most often the consequence of hemodynamic overload and the resulting abnormal wall stresses. At the molecular and cellular levels, remodeling involves myocyte hypertrophy, changes in the quantity and quality of the interstitial matrix, alterations in gene expression with regression to fetal phenotypes, and the death of myocytes by apoptosis or necrosis.However, most of the results of the above were mainly concerned of the left ventricle, and as the consequence of myocardial infarction(MI).But in the area of pediatrics,the congenital heart diseases(CHD) were very common and the function of the ritht ventricle was essential for the therapy and prognosis. So the goal of this study tries to simplifya complex problem and to identify pathways for future research.To evaluate the the influence of volume and pressure overload on rat right ventricle remodeling, we developed a special model which connect the arteria carotis communnis and venae jugularis externa by using a special technique, making a artificial shunt of left to right, which result in volume and pressure overload. By using various methods, from different point of view, we can study the remodeling in the right ventricle of volume and pressure overload rat.Methods We develeped four experimental groups and four controlled groups, each has 10 weigh/sex matched male Wistar rats, weight 150～200g. The experimental groups undergone a special surgery which made a artificial shunt of left to right by connect the arteria carotis communnisand venae jugularis externa of the right side, result in volume overload.After 1, 2, 4, 8weeks later, The animals were anesthetized with pentobarbital (60 mg/Kg, ip), placed over a heating pad, and tracheostomized formechanical ventilation with oxygen-enriched air at 60 cpm, with a tidal volume of 1 ml/100 g. Respiratory rate and tidal volume were adjusted to keep arterial blood gases and pH within physiological limits. The heart was exposed through a median sternotomy, and the pericardium was widely opened. RV pressures were measured with a 2-Fr high-fidelity micromanometer inserted through the RV free-wall into the RV cavity and through pulmonary artery valves into the pulmonary trunk. After complete instrumentation, the animal preparation was allowed to stabilize for 15 min before the beginning of the experimental protocols. Hemodynamic recordings were made with respiration suspended at end-expiration. RV pressure were measured at end-diastole and peak systole. Then took out the heart and measure the weight, then separated the tissue of right ventricle and studied by using various methods. (1)Anatomic level: Observe the ratios of right ventricle/(left ventricle + septum) [RV/ (LV+S) ],right ventricle/body weight (RV/BW) , which reflecting the changes of right ventricle. (2)Cellular level: Observe the myocardium by the meaning of histotomy and histomorphology. Analysis the ratio of myocyte length to myocyte width and myocyte cross-sectional area by using a special computer software. (3)We performed RT-PCR to examine the changes while the right ventricle remodeling. The total mRNA were extracted by the methods described before, and the different primers were designed as Entrez PubMed, according to the issures before.1) The changes of mRNA expression ofβ-myosin heavy chain, which refleting the protein production of the myocyte.2) The changes of mRNA expression of rennin-angiotension II system(RAS), andβ-adrenergic receptors, which reflecting the changes of hormone system.3) The changes of mRNA expression of type I collagen, type III collagen, fibronectin I , which attribute to the fibrosis of the right ventricle.4) The changes of mRNA expression of MMPs-9 and TIMPs-1, which present the balance of matrix.5) The changes of mRNA expression of IGF-1 .which present the hyperplasia of the myocytes.6) The changes of mRNA expression of BNP, which present the sress of the ventricle wall.7) The changes of mRNA expression of bcl-2, which present the apoptosis of myocyte. The controlled groups undergone the same operation except connecting the arteria carotis communnis and venae jugularis externa of the right side. And were fed under the conditions.Result The right ventricle pressure was elevated and maintained relatively stable at the 1^st , 2^nd , 4^th and 8^th week after surgery. There was no statistical significance among the experimental groups. The obstruction of the arteria carotis communnis to venae jugularis externa occurred in two animals at 4^th week and 8^th week each, they were excluded. No animals occurred heart failure. (1)There's increment of the ratio of right ventricle weight to the body weight (RV mg/BW g) after 4 weeks, but until 8^th week it has statistical significance(0. 64±0.05 vs 0.43±0.03, P<0. 01) ; At the same time, the ratios of right ventricle/(left ventricle + septum) [RV/( LV+S ) ] increased to (0. 36 ± 0. 04 vs 0. 21 ± 0. 02 , P<0. 05) ; (2)No heart failure occurred in all groups. At 4^th week, the ratio of myocyte length to myocyte width and myocyte cross-sectional area changed obviously. The ratio of myocyte length to myocyte width decreased from 7.1 ± 0.3: 1 vs 6.5± 0.3: l(P<0.05), meanwhile the myocyte cross-sectional area increases from (250.4 ± 17.7) μm² to (328.9 ± 12.5) μm²(P<0. 05). At the 8^th week, the changes still existed, ratio of myocyte length to myocyte width (6.4 ± 0.4) vs (7.2 ± 0.3), P<0.05, meanwhile the myocyte cross-sectional area (324.8±8.9) μm² vs (250.4 ± 22.1) μm², P<0.05. But there's no statistical significance between 4^th and 8^th week. (3) The mRNA expression of β myosin heavy chain increased remarkably at 1^st week after surgery(1. 37 ± 0.21 vs 0.77 ± 0.15, p<0. 001, average of densitometry). At the 2^nd week, the changes still existed, but there were no difference from the l^st week (1.39 ± 0.22 vs 0.79 ± 0.17, p<0.01, average of densitometry) .At the 4^th and 8^th week, the gene expression of β myosin heavy chain decreased to normal level (0. 79 + 0. 10 vs 0. 76±0. 08, p>0. 05; 0.84 ± 0.16 vs 0.78±0. 14, p>0. 05. separately , average of densitometry) (4) The changes of mRNA expression of rennin-angiotension II system(RAS) were increased obviously at the 1^st week(0. 51±0.18 vs 0. 35 ± 0. 09, p<0. 05, average of densitometry). There was statistically different between the 1^st and 2^nd week(0.77 ± 0. 16 vs 0.51 ± 0.18, p<0. 05, average of densitometry). At the 4^th and 8^th week, there's no differences compared to before. (5) The changes of mRNA expression of β-adrenergic receptors were decreased at the 1^st week(0.47±0. 13 vs 0.80 ± 0. 12, p<0. 05, average of densitometry). At the 2^nd, 4^th and 8^th week, the changes still existed, but there were no difference among them and all of them had no difference from the 1^st week. (6) IGF-1 mRNA expression raised from the 1^st week, but until the 2^nd week, the difference had the statistical significance(0. 24 ± 0. 06)vs (0. 14± 0. 03), p<0. 01, average of densitometry; At the 4^th week, the result was still higher than the 2^nd week (0.35 ± 0.07) vs (0.24 ± 0.06), p<0.05, average of densitometry; At the 8^th week, returned to the level simmiliar to the l^st week (0.15 ± 0.05) vs (0.15 ± 0.04), p>0.05, average of densitometry.(7) BNP mRNA expression raised from the 1^st week (0.71 ±0.17) vs (0.55 ± 0.27), p<0. 01, average of densitometry; And still to be higher at the 2^nd week (0.82 ± 0.15) vs (0.71 ± 0.17), p<0. 05, average of densitometry; At the 4^th and 8^th week, it returned to the level simmiliar to the 1^st week. (8) The changes of mRNA expression of type I collage, type III collagen were statistically different between the experimental groups and controlled groups at 2^nd week (type I collagen: 0. 93±0. 18 vs 0. 79 ± 0. 07, p<0. 05; type III collagen: 0. 43 ± 0. 07 vs 0. 36± 0. 07, p<0.05. average of densitometry). There was no difference between the experimental groups and controlled groups at the 1^st , 4^th and 8^th week. And the expression of fibronectin I mRNA was increased at the 1^st week, preceded the accumulation of fibrosis (fibronectin I : 0. 26 ± 0. 06 vs 0. 20 ± 0. 05, p<0. 05, average of densitometry). At the 2^nd week, it returned to controlled group level. (9) The mRNA expression of MMPs-9 raised at the 2^nd week, (0.54± 0.19) vs (0.27 ± 0.10), p<0. 01, average of densitometry. At the 4^th week,(0.61±0.20) vs (0.31±0.12), p<0. 01, average of densitometry. At the 8^th week, (0. 53 ± 0. 20) vs (0. 33 ± 0. 16), p<0.05, average of densitometry; At the same time, TIMPs-1 had simmiliar changes, 2^nd week[ (0.44±0.21) vs (0. 20±0. 09), p<0. 01, average of densitometry], 4^th week[ (0. 44±0. 14) vs (0.27±0.11), p<0. 01, average of densitometry], 8^th week[ (0.47±0.21) vs (0.25±0. 16), p<0.05, average of densitometry]. (10)bcl-2 elevated at the l^st and 2^nd week(the 1^st week 0.28±0.07 vs 0.16±0.03, p<0. 01, the 2^nd week 0. 27±0. 07 vs 0. 15±0. 04, p<0. 01, average of densitometry). No changed occurred in the later 4 weeks. There' s no difference between the 4^th and 8^th week.Conclusion (1)Volume and pressure overload can cause myocardial remodeling in the rat right ventricle, include the myocyte and the interstitial connective tissue. (2) Some genes, such asβmyosin heavy chain, rennin-angiotension II system(RAS),β-adrenergic receptors, IGF-1, et al, would changed very soon according to the changes of environment, Such adaptation would do benefit for the heart function. (3)The interstitial connective tissue also changed in the earlier stage, include the type I collagen,type III collagen and fibronectin I . The reason would be not only the increase of gene expression, but also the decrease of degeneration of the fibnogen. (4) The gene expression of bcl-2 elevated at the 1^st week, indicate the increment of apoptosis involved in the myocardial remodeling. (5)The balance of MMPs and TIMPs remains static in our experiment, and no heart failure occurred, this would be one reason. (6)So we can see at defferent time-point, the phase of right ventricle myocardial remodeling was defferent. Therefore, we could take defferent method to interrupt the progression of myocardial remodeling, these sings would be benefit to the myocardial remodeling therapy.