Alexandra Alberta dos Santos

Rat Adipose Tissue-Derived Stem Cells Transplantation Attenuates Cardiac Dysfunction Post Infarction and Biopolymers Enhance Cell Retention

Background Cardiac cell transplantation is compromised by low cell retention and poor graft viability. Here, the effects of co-injecting adipose tissue-derived stem cells (ASCs) with biopolymers on cell cardiac retention, ventricular morphometry and performance were evaluated in a rat model of myocardial infarction (MI). Methodology/Principal Findings 99mTc-labeled ASCs (1×106 cells) isolated from isogenic Lewis rats were injected 24 hours post-MI using fibrin a, collagen (ASC/C), or culture medium (ASC/M) as vehicle, and cell body distribution was assessed 24 hours later by γ-emission counting of harvested organs. ASC/F and ASC/C groups retained significantly more cells in the myocardium than ASC/M (13.8±2.0 and 26.8±2.4% vs. 4.8±0.7%, respectively). Then, morphometric and direct cardiac functional parameters were evaluated 4 weeks post-MI cell injection. Left ventricle (LV) perimeter and percentage of interstitial collagen in the spare myocardium were significantly attenuated in all ASC-treated groups compared to the non-treated (NT) and control groups (culture medium, fibrin, or collagen alone). Direct hemodynamic assessment under pharmacological stress showed that stroke volume (SV) and left ventricle end-diastolic pressure were preserved in ASC-treated groups regardless of the vehicle used to deliver ASCs. Stroke work (SW), a global index of cardiac function, improved in ASC/M while it normalized when biopolymers were co-injected with ASCs. A positive correlation was observed between cardiac ASCs retention and preservation of SV and improvement in SW post-MI under hemodynamic stress. Conclusions We provided direct evidence that intramyocardial injection of ASCs mitigates the negative cardiac remodeling and preserves ventricular function post-MI in rats and these beneficial effects can be further enhanced by administrating co-injection of ASCs with biopolymers.

Exercise training inhibits inflammatory cytokines and more than prevents myocardial dysfunction in rats with sustained β‐adrenergic hyperactivity

Myocardial hypertrophy and dysfunction occur in response to excessive catecholaminergic drive. Adverse cardiac remodelling is associated with activation of proinflammatory cytokines in the myocardium. To test the hypothesis that exercise training can prevent myocardial dysfunction and production of proinflammatory cytokines induced by β‐adrenergic hyperactivity, male Wistar rats were assigned to one of the following four groups: sedentary non‐treated (Con); sedentary isoprenaline treated (Iso); exercised non‐treated (Ex); and exercised plus isoprenaline (Iso+Ex). Echocardiography, haemodynamic measurements and isolated papillary muscle were used for functional evaluations. Real‐time RT‐PCR and Western blot were used to quantify tumour necrosis factor α, interleukin‐6, interleukin‐10 and transforming growth factor β1 (TGF‐β1) in the tissue. NF‐κB expression in the nucleus was evaluated by immunohistochemical staining. The Iso rats showed a concentric hypertrophy of the left ventricle (LV). These animals exhibited marked increases in LV end‐diastolic pressure and impaired myocardial performance in vitro, with a reduction in the developed tension and maximal rate of tension increase and decrease, as well as worsened recruitment of the Frank–Starling mechanism. Both gene and protein levels of tumour necrosis factor α and interleukin‐6, as well as TGF‐β1 mRNA, were increased. In addition, the NF‐κB expression in the Iso group was significantly raised. In the Iso+Ex group, the exercise training had the following effects: (1) it prevented LV hypertrophy; (ii) it improved myocardial contractility; (3) it avoided the increase of proinflammatory cytokines and improved interleukin‐10 levels; and (4) it attenuated the increase of TGF‐β1 mRNA. Thus, exercise training in a model of β‐adrenergic hyperactivity can avoid the adverse remodelling of the LV and inhibit inflammatory cytokines. Moreover, the cardioprotection is related to beneficial effects on myocardial performance.

Left Ventricle Radio-frequency Ablation in the Rat: A New Model of Heart Failure due to Myocardial Infarction Homogeneous in Size and Low in Mortality

BACKGROUND The purpose of the current study was to create a model of myocardial infarction (MI) that is homogeneous in size with a low immediate (24 hours) mortality. METHODS AND RESULTS Male and female rats (n = 256) underwent left ventricle (LV) ablation (Ab) by a radiofrequency current (1000 kHz; 12 watts for 12 seconds) to promote a MI. A transmural MI occurred in all rats. Post-Ab complex arrhythmias were frequent (atrioventricular block, ventricular tachycardia, and fibrillation), which rapidly and spontaneously reverted to sinus rhythm. Among 66 male rats, immediate mortality occurred in 7.5%. Small MI size dispersion was characterized by smaller variability following Ab (x +/- SD: 45 +/- 8%) when compared with coronary occlusion (Oc; 40 +/- 19%). The histopathologic evaluations identified lesions similar to those which occurred following Oc, with scarring complete at 4 weeks. The hemodynamic and Doppler echocardiograms showed comparable increases in LV dimension, end-diastolic pressure, and pulmonary water content 1 and 4 weeks post-MI. Papillary muscle mechanics 6 weeks post-MI had matched inotropic and lusitropic dysfunction. CONCLUSIONS LV Ab gave rise to a MI within a narrow size limit and with a low immediate mortality. LV Ab resulted in histopathologic evolution, ventricular dilation, and dysfunction, impairment in myocardial mechanics, and congestive outcome that reproduced a MI from Oc.

Bone marrow cell therapy prevents infarct expansion and improves border zone remodeling after coronary occlusion in rats

BACKGROUND Since the cell therapy benefits for myocardial infarction are mainly related to infarct reduction by regenerating lost myocardium or increasing survival of tissues at risk, we evaluated the effects of bone marrow-derived mononuclear cells (MNC), implanted after the completion of necrosis, on infarct progression and cardiac remodeling. METHODS After 48 h of induction of myocardial infarction (MI), Lewis-inbred rats were injected with 6 × 10(6) cells (MI+MNC) or saline (MI). After six weeks, scar dimension, ventricular morphology and function were analyzed by echocardiography followed by histomorphology of the infarcted and border zones. RESULTS After therapy, the relative size of the infarct was smaller in MI+MNC (37 ± 1% of the left ventricle) than in MI (43 ± 1%). While the MI group exhibited parallel elongation of the infarcted (31.6 ± 3.8% increase) and reminiscent ventricular portions (33.5 ± 3.7%), MNC therapy preserved the initial infarct length. Infarcted walls were thicker (979 ± 31 mm) in the MNC group than in the untreated group (709 ± 41 mm), also demonstrating an absence of infarct expansion. In the border zones, MNC led to increased capillary densities and capillary/myocyte ratios. The cardiac systolic function remained depressed in MI, but improved by 19 ± 5% in MI+MNC which reduced the incidence of pulmonary arterial hypertension (37.5% in MI and 6.25% in MI+MNC). CONCLUSION MNC therapy prevented the infarct expansion and thinning related to cardiac remodeling and was associated with an improvement of border zone microcirculation: as a result, MNC therapy reduced typical MI dysfunctional repercussions.

Swimming training attenuates remodeling, contractile dysfunction and congestive heart failure in rats with moderate and large myocardial infarctions.

1 The aim of the present study was to evaluate the effect of swimming on myocardial remodelling after myocardial infarction (MI) in female rats induced by coronary occlusion, which was not performed in sham rats. 2 Rats were divided in six groups, three sedentary (sham (SSh; n = 14), moderate infarct (SMI; n = 8) and large infarct (SLI; n = 10)) and three trained (sham (TSh; n = 16), moderate infarct (TMI; n = 9) and large infarct (TLI; n = 8)) groups. Training (8 weeks, 60 min/day, 5 days/week) was initiated 4 weeks after MI or sham operation. Training did not affect mortality rate, but attenuated the increases in atrial/bodyweight (SSh: 0.07 ± 0.02; TSh: 0.07 ± 0.02; SMI: 0.11 ± 0.03; TMI: 0.09 ± 0.03; SLI: 0.17 ± 0.09; TLI: 0.10 ± 0.05 mg/g) and right ventricular/bodyweight (SSh: 0.15 ± 0.02; TSh: 0.17 ± 0.02; SMI: 0.17 ± 0.07; TMI: 0.20 ± 0.03; SLI: 0.29 ± 0.13; TLI: 0.22 ± 0.08 mg/g) ratios. Myocardial infarction increased pulmonary and myocardial water content in infarcted sedentary animals, whereas no changes were observed in trained infarcted rats. Sedentary infarcted rats showed inotropic and lusitropic depression proportional to the size of the infarct (SSh > SMI > SLI), whereas no differences were noted in trained rats (TLI = TMI = TSh). Indeed, in sedentary rats there was depression of +dT/dt (SSh: 68 ± 25; TSh: 72 ± 21; SMI: 53 ± 20; TMI: 77 ± 30; SLI: 33 ± 15; TLI: 57 ± 22 g/mm2 per s) and –dT/dt (SSh: 33 ± 13; TSh: 36 ± 11; SMI: 24 ± 5; TMI: 35 ± 11; SLI: 15 ± 4; TLI: 32 ± 11 g/mm2 per s) compared with trained rats. 3 In conclusion, swimming clearly favoured post‐MI cardiac remodelling, attenuated myocardial hypertrophy, contractile and relaxation dysfunction and prevented pulmonary congestion.

Myocardial remodeling after large infarcts in rat converts post rest-potentiation in force decay

Miocárdio Remodelado después de Grandes Infartos en Ratas Convierte Potenciación Post-pausa en Disminucion de la Fuerza Danilo Sales Bocalini1, Leonardo dos-Santos4, Ednei Luiz Antonio1, Alexandra Alberta dos Santos1, Ana Paula Davel3, Luciana Venturini Rossoni2, Dalton Valentim Vassallo5, Paulo José Ferreira Tucci1 Departamento de Medicina Divisão de Cardiologia Universidade Federal de São Paulo1; Departamento de Fisiologia e Biofísica Instituto de Ciências Biomédicas Universidade de São Paulo2, São Paulo, SP; Departamento de Biologia Estrutural e Funcional Instituto de Biologia Universidade Estadual de Campinas3, Campinas, SP; Departamento de Ciências Fisiológicas Universidade Federal do Espírito Santo4; Departamento de Ciências Morfofuncionais Escola Superior de Ciências da Santa Casa de Misericórdia5, Vitória, ES, BrasilBACKGROUND Post-rest contraction (PRC) of cardiac muscle provides indirect information about the intracellular calcium handling. OBJECTIVE Our aim was to study the behavior of PRC, and its underlying mechanisms, in rats with myocardial infarction. METHODS Six weeks after coronary occlusion, the contractility of papillary muscles (PM) obtained from sham-operated (C, n=17), moderate infarcted (MMI, n=10) and large infarcted (LMI, n=14) rats was evaluated, following rest intervals of 10 to 60 seconds before and after incubation with lithium chloride (Li(+)) substituting sodium chloride or ryanodine (Ry). Protein expression of SR Ca(2+)-ATPase (SERCA2), Na(+)/Ca(2+) exchanger (NCX), phospholamban (PLB) and phospho-Ser(16)-PLB were analyzed by Western blotting. RESULTS MMI exhibited reduced PRC potentiation when compared to C. Opposing the normal potentiation for C, post-rest decays of force were observed in LMI muscles. In addition, Ry blocked PRC decay or potentiation observed in LMI and C; Li(+) inhibited NCX and converted PRC decay to potentiation in LMI. Although MMI and LMI presented decreased SERCA2 (72±7% and 47±9% of Control, respectively) and phospho-Ser(16)-PLB (75±5% and 46±11%, respectively) protein expression, overexpression of NCX (175±20%) was only observed in LMI muscles. CONCLUSION Our results showed, for the first time ever, that myocardial remodeling after MI in rats may change the regular potentiation to post-rest decay by affecting myocyte Ca(2+) handling proteins.FUNDAMENTO: La Contraccion pos pausa (CPP) del musculo cardiaco provee informaciones indirectas sobre la manejo del calcio intracelular. OBJETIVO: Nuestro objetivo fue estudiar el comportamiento de la CPP y sus mecanismos subyacentes en Ratas con infarto de miocardio. METODOS: Seis semanas despues de la oclusion coronaria, la contractilidad de los Musculos Papilares (MP) obtenidos a partir de Ratas sometidos a falsa cirurgia (C, n = 17), con infarto moderado (MMI, n = 10) y gran infarto (LMI, n = 14), fue evaluada despues de pausas de estimulos de 10 a 60 segundos antes y despues de la incubacion con cloruro de litio (Li+) en substitucion del cloruro de sodio o rianodina (Ry). La expresion proteica de SR Ca(2+)-ATPasa (SERCA2), intercambiador Na+/Ca2+ (NCX), fosfolamban (PLB) y fosfo-Ser (16)-PLB fue analizada por Western blotting. RESULTADOS: Los Ratas MMI presentaron potenciacion de CPP reducida en comparacion a los Ratas C. En oposicion a la potenciacion normal para Ratas C, fueron observadas decaimientos de fuerza post-reposo en los musculos de Ratas LMI. Ademas de eso, la Ry bloqueo la decaimiento o potenciacion de PRC observada en Ratas LMI y C; el Li+ inhibio el NCX y convirtio la decaimiento en potenciacion de CPP en Ratas LMI. Aunque los Ratas MMI y LMI hayan presentado disminucion en el SERCA2 (72 ± 7% y 47 ± 9% de Ratas control, respectivamente) y expresion proteica de fosfo-Ser16-PLB (75 ± 5% y 46 ± 11%, respectivamente), la superexpresion del NCX (175 ± 20%) solo fue observada en los musculos de Ratas LMI. CONCLUSION: Nuestros resultados mostraron, por primera vez, que el remodelado miocardico post-IAM en Ratas puede cambiar la potenciacion regular para decaimiento post-reposo, afectando las proteinas de manejo del Ca(2+) en miocitos.

Cell therapy prevents structural, functional and molecular remodeling of remote non-infarcted myocardium

BACKGROUND/OBJECTIVES Therapy using bone marrow (BM) cells has been tested experimentally and clinically due to the potential ability to restore cardiac function by regenerating lost myocytes or increasing the survival of tissues at risk after myocardial infarction (MI). In this study we aimed to evaluate whether BM-derived mononuclear cell (MNC) implantation can positively influence the post-MI structural remodeling, contractility and Ca(2+)-handling proteins of the remote non-infarcted tissue in rats. METHODS AND RESULTS After 48 h of MI induction, saline or BM-MNC were injected. Six weeks later, MI scars were slightly smaller and thicker, and cardiac dilatation was just partially prevented by cell therapy. However, the cardiac performance under hemodynamic stress was totally preserved in the BM-MNC treated group if compared to the untreated group, associated with normal contractility of remote myocardium as analyzed in vitro. The impaired post-rest potentiation of contractile force, associated with decreased protein expression of the sarcoplasmic reticulum Ca(2+)-ATPase and phosphorylated-phospholamban and overexpression of Na(+)/Ca(2+) exchanger, were prevented by BM-MNC, indicating preservation of the Ca(2+) handling. Finally, pathological changes on remodeled remote tissue such as myocyte hypertrophy, interstitial fibrosis and capillary rarefaction were also mitigated by cell therapy. CONCLUSIONS BM-MNC therapy was able to prevent cardiac structural and molecular remodeling after MI, avoiding pathological changes on Ca(2+)-handling proteins and preserving contractile behavior of the viable myocardium, which could be the major contributor to the improvements of global cardiac performance after cell transplantation despite that scar tissue still exists.

Cardiac implications after myocardial infarction in rats previously undergoing physical exercise.

BACKGROUND Few studies have analyzed the cardiac effects of exercise prior to coronary occlusion. OBJECTIVE To evaluate the effects of myocardial infarction in rats undergoing physical exercise. METHODS Female rats underwent swimming exercise or were kept sedentary for eight weeks and were randomized to coronary occlusion or sham surgery, in one of the following four groups: Sedentary (S), exercise (E), Sedentary myocardial infarction (SMI) and Exercise myocardial infarction (EMI). After six weeks, their biometrics, Doppler echocardiography, hemodynamics and myocardial mechanics were analyzed. RESULTS No cardioprotection was observed in EMI animals and there was no difference in infarct size (%LV) between EMI (38.50 ± 4.60%) and SMI (36.58 ± 4.11%). Water content of the lung (%) of SMI (80 ± 0.59) and EMI (80 ± 0.57) was higher than that of S (78 ± 0.15) and E (78 ± 0.57) groups. Left ventricular systolic pressure (mmHg) (S: 130 ± 5, E: 118 ± 8; SMI: 91 ± 3; EMI: 98 ± 3) and the first positive time derivative (mmHg) positive pressure (S: 8216 ± 385; E: 8437 ± 572; SMI: 4674 ± 455; EMI: 5080 ± 412) of S and E were higher than those of SMI and EMI. The transverse fractional shortening (%) of SMI (27 ± 2) and EMI (25 ± 2) were similar and lower than that of E (65 ± 2) and S (69 ± 2). The E/A ratio was higher in SMI (5.14 ± 0.61) and EMI (4.73 ± 0.57) compared to S (2.96 ± 0.24) and E (2.83 ± 0 21). In studies of isolated papillary muscle, depression of the contractile capacity observed was similar to that of SMI and EMI, and there was no change in myocardial stiffness. CONCLUSION Previous training by swimming did not attenuate cardiac implications due to myocardial infarction.

Digitoxin prolongs survival of female rats with heart failure due to large myocardial infarction.

BACKGROUND We analyzed whether digitoxin affects the survival of rats with congestive heart failure. METHODS AND RESULTS The influence of digitoxin (0.1 mg.100 g.day, orally) on the survival of infarcted female rats (n=170) randomized as Control Infarcted (CI, n=85) or Digitoxin (D, n=85) was evaluated for 280 days. Mean survival was 235+/-7 days for CI and 255+/-5 days for D (log-rank test: P=.0602). Digitoxin did not affect survival in rats with congestive heart failure from myocardial infarction <40% of the left ventricle, but did prolong survival in rats with infarction >or=40%. The log-rank test defined higher mortality (P=.0161) in CI >40% (56%) than in D >40% (34%), with a hazard ratio of 2.03. Pulmonary water content and papillary muscle mechanics were analyzed in CI (n=7) and D (n=14) survivors. Significant differences were observed regarding pulmonary water content (CI: 82+/-0.3; D: 80+/-0.3%; P=.0014), developed tension (CI: 2.7+/-0.3; D: 3.8+/-0.3g/mm(2); P=.0286) and +dT/dt (CI: 24+/-3; D: 39+/-4 mg mm(2).s; P=.0109). CONCLUSION In conclusion, long-term digitoxin administration reduced cardiac impairment after myocardium infarction, attenuated myocardial dysfunction, reduced pulmonary congestion, and provided the first evidence regarding the efficiency of digitoxin in prolonging survival in experimental cardiac failure.

Are there gender differences in left ventricular remodeling after myocardial infarction in rats

Objective An unclear issue is whether gender may influence at cardiac remodeling after myocardial infarction (MI). We evaluated left ventricle remodeling in female and male rats post-MI. Methods Rats were submitted to anterior descending coronary occlusion. Echocardiographic evaluations were performed on the first and sixth week post-occlusion to determine myocardial infarction size and left ventricle systolic function (FAC, fractional area change). Pulsed Doppler was applied to analyze left ventricle diastolic function using the following parameters: E wave, A wave, E/A ratio. Two-way ANOVA was applied for comparisons, complemented by the Bonferroni test. A P≤=0.05 was considered significant. Results There were no significant differences between genders for morphometric parameters on first (MI [Female (FE): 44.0±5.0 vs. Male (MA): 42.0±3.0%]; diastolic [FE: 0.04±0.003 vs. MA: 0.037±0.005, mm/g] and systolic [FE: 0.03±0.0004 vs. MA: 0.028±0.005, mm/g] diameters of left ventricle) and sixth (MI [FE: 44.0±5.0 vs. MA: 42.0±3.0, %]; diastolic [FE: 0.043±0.01 vs. MA: 0.034±0.005, mm/g] and systolic [FE: 0.035±0.01 vs. MA: 0.027±0.005, mm/g] of LV) week. Similar findings were reported for left ventricle functional parameters on first (FAC [FE: 34.0±6.0 vs. MA: 32.0±4.0, %]; wave E [FE: 70.0±18.0 vs. MA: 73.0±14.0, cm/s]; wave A [FE: 20.0±12.0 vs. MA: 28.0±13.0, cm/s]; E/A [FE: 4.9±3.4 vs. MA: 3.3±1.8]) and sixth (FAC [FE: 29.0±7.0 vs. MA: 31.0±7.0, %]; wave E [FE: 85.0±18.0 vs. MA: 87.0±20.0, cm/s]; wave A [FE: 20.0±11.0 vs. MA: 28.0±17.0, cm/s]; E/A [FE: 6.2±4.0 vs. MA: 4.6±3.4]) week. Conclusion Gender does not influence left ventricle remodeling post-MI in rats.