Research On The Initial And Recurrent Situation Of Modelled Rats With Stress Cardinmyopathy

[Background]Stress cardiomyopathy (SCM) is an increasingly recognized acute cardiac affliction which is characterized by transient left ventricular dysfunction with regional left ventricular wall motion abnormalities and mimic acute myocardial infarction with Electrocardiogram ST-T change and mild elevated cardiac enzymes, in the absence of significant angiographic coronary stenoses. Elderly women are the main victims of SCM and an episode of emotional or physical stress is believed to act as a trigger in the development of this syndrome. It first described in Japan by Sato in 1990 was coined on the basis of similarities between left ventricular morphologic features observed on left ventriculography and the shape of a fishing pot takotsubo used in Japan to trap octopuses,and also named "Takotsubo cardiomyopathy",it is also typically called as " transient left ventricular apical ballooning", "broken heart syndrome" or "ampulla cardiomyopathy" and so on. SCM is closely similar to acute coronary syndrome, accounting for 2% of sufferers who are suspected of having acute coronary syndrome,and therefore differential diagnosis is crucially important in the site of emergency care. SCM patients have generally a good short-term prognosis, with a rapid improvement of left ventricular systolic function in a period of days to few weeks, and 96% almost completely recover from their symptoms, However, In the acute phase it is still a fatal disease, As has been reported by medical experts such as Sharkey, approximately 5% of patients with SCM experience cardiac arrest. Several complications have been reported during the acute phase of SCM, including malignant arrhythmias, cardiogenic shock and ventricular rupture, and has been shown to be associated with In-hospital mortality rates range from 0 to 8%(0%-8%) and long-term mortality (0%-17%) along with recurrence rates (0%-11.4%). the most frequent chief complaint was chest pain (30%) over 4 years after initial presentation. SCM is a relatively new syndrome,the number of reported cases is increasing rapidly, but people still lack sufficient understanding and evidence for its pathogenesis and standard treatment. Therefore, further research and understanding into the disease is of great significance for clinic as well as social value.SCM is usually represented by a balloon-like apical akinesia with compensatory hyperkinesis of the basal segments, this is the typical form, accounting for about 85%, Such as the abnormal wall motion is involving non-apical, but in other parts will be the atypical SCM, Most atypical SCM is represented basal akinesis with of with normal contractility of the apical segments (inverted TTC), small part of them involve midventricular segments or other segments. regional wall motion abnormalities of recurrent SCM is not necessarily consistent with the initial presentation. The broad clinical spectrum suggests heterogeneous and multifactorial pathophysiologic mechanisms are involved. Hypotheses to explain the transient and reversible left ventricular dysfunction have included thefollowing:(1) sympathetic hyperactivity and direct catecholamine-mediated myocardial stunning; (2) estrogen deficiency, deficiency in cardioprotective; (3) Myocardial fatty acid metabolism is abnormal, the beta oxidation of fatty acid is inhibited, so that the deficiency of myocardial energy supply can affect the contractile function of the myocardium; (4) coronary artery spasm; (5)coronary microvascular dysfunction; (6) acute left ventricular outflow tract obstruction. among these, catecholamine mediated neurogenic myocardial toxicity is considered to be the most likely mechanism. It is known to be triggered by physical or emotional stress, which elevates the endogenous level of catecholamines; Wittstein et al. found that the serum epinephrine and norepinephrine were 7-34 times greater than normal, up to 2-3 times higher compared to patients presenting with acute myocardial infarction and Killip class III. In addition, exogenous administration of catecholamines such as epinephrine injection and disease conditions such as pheochromocytoma induce similar wall motion abnormalities, the above evidences were in favor of an important role for catecholamine.The mammalian left ventricle contains apical-basal gradients of β-adrenergic receptors and sympathetic innervation, with the apex characterized by highest β2-AR and lowest sympathetic nerve density. Therefore, it is speculated that the apical part is more sensitive to catecholamine and easily affected, and high levels of circulating epinephrine is playing a key role. high levels of epinephrine through P 2-AR coupled inhibitory G protein (inhibitory G protein, Gi) signaling pathway play a negative inotropic effect. the density β2 receptor in apical is higher than the basel segments, so predominance of apical depression. In animal studies, high doses of epinephrine (β1, β2 and a-AR agonists), and isoproterenol (β1,β2-AR agonists), can successfully induce typical SCM animal models. Supporting for there is a certain relationship between the high density distribution of β2-AR and the typical SCM. Studies have shown that β1 can only be coupled with stimulatory G protein (G protein stimulatory, Gs) to produce a positive inotropic effect, while β2 receptor can be coupled Gs and Gi, Therefore norepinephrine cannot activate the Gi pathway, resulting in negative inotropy. Paur et al. Successfully induced typical SCM by injecting with high doses of epinephrine in rats. But equivalent high-dose intravenous norepinephrine (β1、 a-AR agonists) did not succeed in reproducing the model. However, Redfors et al. showed that rats received norepinephrine intraperitoneally can induce atypical SCM via experimental study.At present, there were frequently using isoproterenol and epinephrine to make the SCM model, Because of isoproterenol behaves like epinephrine, study on isoproterenol model accounted for the majority, however isoproterenol is a synthetic catecholamine derivate and not an endogenous catecholamine. Norepinephrine and epinephrine are endogenous catecholamine,but there is some controversy on the model of norepinephrine which rarely used to make the model; furthermore at home and abroad, the research on the recurrence of SCM is still blank; it has no clear answer that the left ventricular regional segmental changes of SCM whether or not depends on the gradient distribution of adrenergic receptor. In view of this, In this study, the SCM model and the recurrent model of rats were prepared by norepinephrine, meanwhile to explore the establishment of recurrent model induced by isoproterenol, comparing the difference with norepinephrine-induced, And the relationship between the patterns of SCM and the distribution of al, β1 and β2 receptors was also discussed. for further experimental foundation of studying the pathogenesis, clearing SCM patterns, preventing and treating the disease.Part I Study on the Doze of Norepinephrine to Make the Model of Rat Stress Cardinmyopathy[Objective]To explore the optimal dose of NA (norepinephrine) to prepare the model of stress cardiomyopathy (SCM) in rats, in order to lay the foundation for further experimental study.[Methods]30 male SD rats (300-350 g) were randomly divided into five groups received different doze of NA (1,2,3,4,5, n=6 in each group) intraperitoneally to study the the optimal dose, echocardiography was used to study wall motion 90 min post administration, Criteria for successful of modeling WAS left ventricular dysfunction accompanied of akinesia involving apical ("typical") or non-apical ("atypical"). After 1 week, echocardiography was used to observe the recovery of ventricular wall motion.[Results](1) In rats, NA at doses> 2 mg/kg induced severe SCM-like regional akinesia that completely resolved within 1 week. and in the 4 mg/Kg group a success rate of 66.67%(4/6 rats)was the highest, A total of 4 rats were successful (1 typical SCM rats, 3 atypical SCM rats).(2) With the increase of NA doses,2 h mortality, rat anal temperature and heart rate increased, half of rats died within 2 h in 5 mg/Kg group.[Conclusions](1) Models of typical and atypical SCM can be successfully prepared by intraperitoneal injection of NA, and 4 mg/Kg was the optimal dose.(2) The establishing method is simple and reproducible, supporting the pathogenesis of catecholamine-mediated cardiotoxic.PartⅡ Preparation of Initial Model and Rrecurrent Model with Stress Cardiomyopathy and Preliminary Exploration of Pathogenesis.[Objective]To explore the possibility of preparation for recurrent SCM model using noradrenaline (NA) and isoproterenol (ISO), and compare the characteristics of the two models. to provide a good animal model for the study of SCM in the future; and the relationship between the patterns of SCM in NA rats and the anatomical distribution of α1, β1 and β2 receptors was also performed to explore the pathogenesis of the disease.[Methods]60 male SD rats (300-350 g) were divided into two protocols for the study.(1) Research on preparation of the initial and recurrent model of SCM using NA. According to the results of the Part Ⅰ, the optimal dose of NA 4mg/KG was selected.45 rats were received intraperitoneal injection of 4mg/KG to make the initial model and 1 week after the recurrence modeling. After NA administration 90 min, the motion amplitude of each segment was compared among the apical segment, the midventricular segments and the basal segment of the left ventricular by M-mode Echocardiography, and measure fractional shortening (FS%), to judge model results. In the Initial of the successful model rats,5 typical rats and 5 atypical rats were sacrificed with myocardial tissue collected for pathological examination (assigned as initial model group). the remaining rats were fed normally, and reviewed ultrasound on the seventh day. The full recovery rats were taked out the total number of 2/3 rats by patterns and on the eighth day were used to make recurrent model. the successful recurrent rats were sacrificed with myocardial tissue collected for pathological examination, assigned as the recurrent model group. another 1/3 rats were used as a recovery group, and were injected with normal saline to be a recurrence control. andthen were sacrificed with myocardial tissue collected for pathological examination. The pathological changes and the expression of the receptor were observe by HE staining and immunohistochemical staining of adrenergic receptor al, β1 and β2 in the basal and apical segments collected from the left ventricle, and evaluate the animal model, comparison of pathological changes in acute phase and recovery phase, and the degree of myocardial injury between hyperkinesis segment and akinesia segment.A semi quantitative method was used to compare the expression ratio of al, β1 and β 2 receptor in cardiac basal segment and apical segment among the experimental groups by Image-ProPlus image software.5 rats were assigned as the normal control group to receive intraperitoneal injection of normal saline,and were sacrificed with myocardial tissue collected for pathological examination after completion of ultrasonic testing.(2) Study on the preparation of SCM model by using ISO.10 rats were received intraperitoneal injection with 50 mg/KG of ISO, Normal feeding after successful modeling, and the restored normal rats were used to make recurrent model on the eighth day.[Results](1) In NA initial model rats,26 rats were modelled successfully, the total success rate was 57.8%, atypical SCM 21 rats (20 inverted SCM rats; 1 midventricular SCM rats), accounting for 81%,5 typical SCM rats, accounting for 19%. all successfully model rats completely resolved within 1 week (FS% of each segment and the normal group compared with P> 0.05), the success rate of recurrent SCM in 10 atypical SCM rats was 30%(3/10), and they all transformed into typical SCM, transformation rate was 100%(3/3).(2) HE staining showed that there were different degrees of cardiac muscle fiber degeneration, edema, contraction band necrosis, and no obvious apoptosis and necrosis of myocardial cells, In initial modeling group, contraction band necrosis in the akinesia segments (typical SCM apical+atypical SCM basal) were more serious (Z= -2.145, P= 0.043< 0.05) than hyperkinesis segments (atypical SCM apical+ typical SCM basal); In the recovery group, there was a basic recovery of myocardial fiber degeneration and edema, with a small amount of inflammatory cell infiltration and a small part of the contraction band necrosis.(3) The expression ratio of al,β1 and β2 receptors with basal segment to apical segment in normal control group was 0.87,1.12,0.85 (median), and there was no significant difference in the ratio of other groups (P> 0.05).(4) 10 rats were received 50 mg/kg ISO with 6 rats successfully modelled, the success rate was 60%, all are typical of SCM, after 8 d,5 rats of them see successfully recurred, recurrence rate was 83%(5/6), The pattern were the same with initial model for typical SCM and pattern transformation rate was 0%.[Conclusions](1) both NA and ISO can be used to prepare initial or recurrent model of SCM.(2) In initial model, NA-model based on atypical SCM, ISO-model based on typical SCM, regional wall motion abnormalities of recurrent SCM is not necessarily consistent with the initial presentation. recurrence of NA-model can change the SCM patterns, but ISO-model unchanged.(3) Hypokinetic segment myocardial injury was more serious, But after 1 week, the ventricular wall motion function could be restored to normal, and the myocardial tissue was still visible sporadic contraction band necrosis.(4) Left ventricular adrenaline receptors exist gradient distribution of anatomy in rats, the advantage region of β2, al receptor was in the apical (higher density), the advantage region of β1 receptor was in the basal (higher density), the different gradient distribution is not directly impact on SCM patterns, specific mechanisms still need to elucidate.