Antonio Carlos Campos de Carvalho

Transendocardial, Autologous Bone Marrow Cell Transplantation for Severe, Chronic Ischemic Heart Failure

Background—This study evaluated the hypothesis that transendocardial injections of autologous mononuclear bone marrow cells in patients with end-stage ischemic heart disease could safely promote neovascularization and improve perfusion and myocardial contractility. Methods and Results—Twenty-one patients were enrolled in this prospective, nonrandomized, open-label study (first 14 patients, treatment; last 7 patients, control). Baseline evaluations included complete clinical and laboratory evaluations, exercise stress (ramp treadmill), 2D Doppler echocardiogram, single-photon emission computed tomography perfusion scan, and 24-hour Holter monitoring. Bone marrow mononuclear cells were harvested, isolated, washed, and resuspended in saline for injection by NOGA catheter (15 injections of 0.2 cc). Electromechanical mapping was used to identify viable myocardium (unipolar voltage ≥6.9 mV) for treatment. Treated and control patients underwent 2-month noninvasive follow-up, and treated patients alone underwent a 4-month invasive follow-up according to standard protocols and with the same procedures used as at baseline. Patient population demographics and exercise test variables did not differ significantly between the treatment and control groups; only serum creatinine and brain natriuretic peptide levels varied in laboratory evaluations at follow-up, being relatively higher in control patients. At 2 months, there was a significant reduction in total reversible defect and improvement in global left ventricular function within the treatment group and between the treatment and control groups (P =0.02) on quantitative single-photon emission computed tomography analysis. At 4 months, there was improvement in ejection fraction from a baseline of 20% to 29% (P =0.003) and a reduction in end-systolic volume (P =0.03) in the treated patients. Electromechanical mapping revealed significant mechanical improvement of the injected segments (P <0.0005) at 4 months after treatment. Conclusions—Thus, the present study demonstrates the relative safety of intramyocardial injections of bone marrow–derived stem cells in humans with severe heart failure and the potential for improving myocardial blood flow with associated enhancement of regional and global left ventricular function.

Improved Exercise Capacity and Ischemia 6 and 12 Months After Transendocardial Injection of Autologous Bone Marrow Mononuclear Cells for Ischemic Cardiomyopathy

Background—We recently reported the safety and feasibility of autologous bone marrow mononuclear cell (ABMMNC) injection into areas of ischemic myocardium in patients with end-stage ischemic cardiomyopathy. The present study evaluated the safety and efficacy of this therapy at 6- and 12-month follow-up. Methods and Results—Twenty patients with 6- and 12-month follow-up (11 treated subjects; 9 controls) were enrolled in this prospective, nonrandomized, open-label study. Complete clinical and laboratory evaluations as well as exercise stress (ramp treadmill), 2-dimensional Doppler echocardiography, single-photon emission computed tomography (SPECT) perfusion scanning, and 24-hour Holter monitoring were performed at baseline and follow-up. Transendocardial delivery of ABMMNCs was performed with the aid of electromechanical mapping to identify viable myocardium. Each patient received 15 ABMMNC injections of 0.2 mL each. At 6 and 12 months, total reversible defect, as measured by SPECT perfusion scanning, was significantly reduced in the treatment group as compared with the control group. At 12 months, exercise capacity was significantly improved in the treatment group. This improvement correlated well with monocyte, B-cell, hematopoietic progenitor cell, and early hemapoietic progenitor cell phenotypes. Conclusions—The 6- and 12-month follow-up data in this study suggest that transendocardial injection of ABMMNCs in patients with end-stage ischemic heart disease may produce a durable therapeutic effect and improve myocardial perfusion and exercise capacity.

Bone Marrow Multipotent Mesenchymal Stromal Cells Do Not Reduce Fibrosis or Improve Function in a Rat Model of Severe Chronic Liver Injury

The objective of our study was to evaluate the therapeutic potential of bone marrow mesenchymal stromal cells (MSC) in a rat model of severe chronic liver injury. Fourteen female Wistar rats were fed exclusively an alcoholic liquid diet and received intraperitoneal injections of carbon tetrachloride every other day during 15 weeks. After this period, eight animals (MSC group) had 1 × 107 cells injected into the portal vein while six animals (placebo group) received vehicle. Blood analysis was performed to evaluate alanine aminotransferase (ALT), aspartate aminotransferase (AST), and albumin before cell therapy and 1 and 2 months after cell or placebo infusion. Fibrosis was evaluated before and 1 month after cell or placebo injection by liver biopsies. Two months after cell delivery, animals were sacrificed and histological analysis of the livers was performed. Fibrosis was quantified by histomorphometry. Biopsies obtained before cell infusion showed intense collagen deposition and septa interconnecting regenerative nodules. One month after cell injection, this result was unaltered and differences in fibrosis quantification were not found between MSC and placebo groups. ALT and AST returned to normal values 2 weeks after cell or placebo infusion, without significant differences between experimental groups. Two months after cell or placebo injection, albumin had also returned to normal values and histological results were maintained, again without differences between MSC and placebo groups. Therefore, under our experimental conditions, MSC were unable to reduce fibrosis or improve liver function in a rat model of severe chronic liver injury.

Perspectives on Trypanosoma cruzi–Induced Heart Disease (Chagas Disease)

Chagas disease is caused by the parasite Trypanosoma cruzi. It is a common cause of heart disease in endemic areas of Latin America. The year 2009 marks the 100th anniversary of the discovery of T cruzi infection and Chagas disease by the Brazilian physician Carlos Chagas. Chagasic cardiomyopathy develops in from 10% to 30% of persons who are chronically infected with this parasite. Echocardiography and magnetic resonance imaging (MRI) are important modalities in the evaluation and prognostication of individuals with chagasic heart disease. The etiology of chagasic heart disease likely is multifactorial. Parasite persistence, autoimmunity, and microvascular abnormalities have been studied extensively as possible pathogenic mechanisms. Experimental studies suggest that alterations in cardiac gap junctions may be etiologic in the pathogenesis of conduction abnormalities. The diagnosis of chronic Chagas disease is made by serology. The treatment of this infection has shortcomings that need to be addressed. Cardiac transplantation and bone marrow stem cell therapy for persons with Chagas disease have received increasing research attention in recent years.

Safety of Dobutamine-Atropine Stress Echocardiography: A Prospective Experience of 4033 Consecutive Studies ☆ ☆☆

Dobutamine-atropine stress echocardiography (DASE) is an established method and has been shown to be accurate for the detection of coronary artery disease. Still, there are few large clinical studies that analyze the safety of DASE in general or the safety of performing it on an ambulatory basis. Most studies use a target heart rate as the primary end point regardless of whether asymptomatic ischemia occurs. Such studies have shown a serious cardiac event rate of approximately 0.3%. We prospectively studied 4,033 consecutive patients on an ambulatory basis and in the hospital with the use of DASE from July 1991 to December 1998. All tests were performed by an experienced physician, and all clinical and DASE data were stored in a large database organized at the beginning of the study. Dobutamine was infused in scalar doses of 5, 10, 20, 30, and 40 microg/kg per minute in 3-minute stages. Development of a new wall motion abnormality, achievement of 85% of target heart, and end of the DASE infusion protocol were used as an end point. If 85% of the target heart rate was not achieved, atropine was infused up to 1 mg in the absence of myocardial ischemia, which was used in 1,280 studies. There were 3,645 diagnostic tests, and 388 (10%) were found to be nondiagnostic. This result was due to poor image quality in 115 (3%), end of protocol in negative-submaximal examinations in 124 (3%), and limiting side effects in 149 (4%). Thirty-seven percent of the tests showed positive results for myocardial ischemia. Major test-related cardiac complications occurred in 10 (0.25%) patients and included 1 ventricular fibrillation, 1 case of myocardial infarction, and 8 cases of sustained ventricular tachycardia. Atropine poisoning was observed in 5 (0.12%) patients. No deaths occurred as a direct or indirect consequence of DASE. We conclude that dobutamine-atropine stress echocardiography is a reasonably safe method for detection of coronary artery disease in the hospital or in an ambulatory basis. The use of new wall motion abnormality as 1 of the end points may prevent further ischemia-related complications.

Transplanted bone marrow cells repair heart tissue and reduce myocarditis in chronic chagasic mice.

A progressive destruction of the myocardium occurs in approximately 30% of Trypanosoma cruzi-infected individuals, causing chronic chagasic cardiomyopathy, a disease so far without effective treatment. Syngeneic bone marrow cell transplantation has been shown to cause repair and improvement of heart function in a number of studies in patients and animal models of ischemic cardiopathy. The effects of bone marrow transplant in a mouse model of chronic chagasic cardiomyopathy, in the presence of the disease causal agent, ie, the T. cruzi, are described herein. Bone marrow cells injected intravenously into chronic chagasic mice migrated to the heart and caused a significant reduction in the inflammatory infiltrates and in the interstitial fibrosis characteristics of chronic chagasic cardiomyopathy. The beneficial effects were observed up to 6 months after bone marrow cell transplantation. A massive apoptosis of myocardial inflammatory cells was observed after the therapy with bone marrow cells. Transplanted bone marrow cells obtained from chagasic mice and from normal mice had similar effects in terms of mediating chagasic heart repair. These results show that bone marrow cell transplantation is effective for treatment of chronic chagasic myocarditis and indicate that autologous bone marrow transplant may be used as an efficient therapy for patients with chronic chagasic cardiomyopathy.

Physiologic multivalvular regurgitation during pregnancy: a longitudinal Doppler echocardiographic study☆

Valvular function, assessed by Doppler technique, has not been extensively investigated during normal pregnancy. To prospectively study this feature, 18 normal pregnant women were followed during their pregnancies and puerperium, with serial clinical and pulsed-continuous Doppler echocardiographic examinations. In four gestational periods and the puerperium, we analysed: (a) ventricular and atrial dimensions, as well as valve annular diameters; (b) prevalence and characteristics of trivial valvular regurgitations. During pregnancy, slight but significant increases of the four cardiac chamber dimensions and valve annular diameters were observed, except for the aortic ring. The prevalence of physiologic valvular regurgitation in early pregnancy (mitral, 0%; tricuspid, 38.9%; pulmonary, 22.2%; aortic, 0%), was similar to a control group of 18 healthy non-pregnant women. As pregnancy evolved, there was a progressive and significant increase of multivalvular regurgitation, maximal at full-term (mitral, 27.8%; tricuspid, 94.4%; pulmonary, 94.4%, P < 0.05 vs. early pregnancy). Aortic regurgitation was not detected in any stage of pregnancy. In the puerperium, mitral regurgitation resolved, but tricuspid and pulmonary regurgitation were still significantly prevalent (83.3% and 66.7%, respectively, P < 0.05 vs. early pregnancy). It is concluded that physiologic multivalvular regurgitation is frequent in pregnancy, mainly involving right-sided valves in late gestational periods, occasionally persisting in the early puerperium. Chamber enlargement, valve annular dilatation, and increased prevalence of trivial valve regurgitation are time-related events during normal pregnancy, resulting from a reversible cardiac remodeling process induced by physiologic volume overload. These aspects should be considered for a correct interpretation of Doppler echocardiographic findings in pregnant women with suspected heart disease.

Cardioprotective properties of humoral factors released from rat hearts subject to ischemic preconditioning.

Myocardial protection can be achieved by transfer of coronary effluent from ischemically preconditioned to non-preconditioned hearts. This study was designed to test the hypothesis that preconditioned effluent from rat hearts purified by Sep-Pak C-18 cartridges could induce remote cardioprotection against ischemia/reperfusion (I/R) injury through the activation of protein kinase C signaling pathway. Buffer-perfused rat hearts were subject to 30 min ischemia and 60 min reperfusion. The myocardial I/R injury was assessed by postischemic contractile function recovery and infarct size. The protective effect of coronary effluent collected during ischemic preconditioning (IPC) was tested in non-preconditioned hearts in presence or absence of a PKC inhibitor, chelerythrine. Infarct size was 17 ± 2% in preconditioned versus 37 ± 1% in control hearts (P < 0.001). Hearts perfused with fresh preconditioned effluent had infarct sizes of 16 ± 3% versus 36 ± 1% in hearts treated with non-preconditioned effluent. The cardioprotective effect was lost when the effluent was left at room temperature during 24 h (infarct size, 40 ± 3%) or heated to 70°C (26 ± 4%, P < 0.05) or 100°C (39 ± 1%, P < 0.001). The lyophilized effluent was stable for 30 days, and its purification in a Sep-Pak C-18 column resulted in a hydrophobic fraction that reduced the infarct size to 17 ± 2% versus 38 ± 2% for the hydrophilic fraction. Chelerythrine (100 μM) inhibited the reduction of infarct size induced by IPC (35 ± 4%) or hydrophobic fraction (37 ± 3%). Recovery of the contractile function at reperfusion was higher in preconditioned group (74 ± 6% versus 17 ± 7% in control, P < 0.001) and hydrophobic fraction (66 ± 7% versus 8 ± 4% in hydrophilic fraction, P < 0.001). Similarly, chelerythrine was able to abrogate the contractile function recovery (12 ± 6%, P < 0.001 versus preconditioned group and 19 ± 7%, P < 0.001 versus hydrophobic fraction). In conclusion, the cardioprotective factors released in the coronary effluent by IPC are thermolabile hydrophobic substances with molecular weights higher than 3.5 kDa and acting through PKC activation.

Symptomatic cardiac involvement in juvenile rheumatoid arthritis

In a retrospective study of 172 patients with juvenile rheumatoid arthritis, symptomatic cardiac involvement occurred in 13 (7.6%) patients (11 systemic and 2 polyarticular). There was predominance of the male sex and in most patients the involvement occurred in the initial years of the disease. Pericarditis occurred in seven patients; perimyocarditis in four and myocarditis in two patients. In the follow-up, one of the patients with pericarditis died of an arrhythmia during pericardiocentesis for cardiac tamponade. Among the patients with myocarditis, three died of septicemia during active disease. One of these three patients had myocarditis associated with cardiac tamponade. Among the 172 patients with juvenile rheumatoid arthritis, five children died; four belonged to the symptomatic cardiac involvement group (P less than 0.001). Cardiac involvement, in particular myocarditis and cardiac tamponade, can be regarded as a factor of worse prognosis.

Optimized labeling of bone marrow mesenchymal cells with superparamagnetic iron oxide nanoparticles and in vivo visualization by magnetic resonance imaging

BackgroundStem cell therapy has emerged as a promising addition to traditional treatments for a number of diseases. However, harnessing the therapeutic potential of stem cells requires an understanding of their fate in vivo. Non-invasive cell tracking can provide knowledge about mechanisms responsible for functional improvement of host tissue. Superparamagnetic iron oxide nanoparticles (SPIONs) have been used to label and visualize various cell types with magnetic resonance imaging (MRI). In this study we performed experiments designed to investigate the biological properties, including proliferation, viability and differentiation capacity of mesenchymal cells (MSCs) labeled with clinically approved SPIONs.ResultsRat and mouse MSCs were isolated, cultured, and incubated with dextran-covered SPIONs (ferumoxide) alone or with poly-L-lysine (PLL) or protamine chlorhydrate for 4 or 24 hrs. Labeling efficiency was evaluated by dextran immunocytochemistry and MRI. Cell proliferation and viability were evaluated in vitro with Ki67 immunocytochemistry and live/dead assays. Ferumoxide-labeled MSCs could be induced to differentiate to adipocytes, osteocytes and chondrocytes. We analyzed ferumoxide retention in MSCs with or without mitomycin C pretreatment. Approximately 95% MSCs were labeled when incubated with ferumoxide for 4 or 24 hrs in the presence of PLL or protamine, whereas labeling of MSCs incubated with ferumoxide alone was poor. Proliferative capacity was maintained in MSCs incubated with ferumoxide and PLL for 4 hrs, however, after 24 hrs it was reduced. MSCs incubated with ferumoxide and protamine were efficiently visualized by MRI; they maintained proliferation and viability for up to 7 days and remained competent to differentiate. After 21 days MSCs pretreated with mitomycin C still showed a large number of ferumoxide-labeled cells.ConclusionsThe efficient and long lasting uptake and retention of SPIONs by MSCs using a protocol employing ferumoxide and protamine may be applicable to patients, since both ferumoxides and protamine are approved for human use.