Toshio Nishikawa

Nitric Oxide Contributes to the Progression of Myocardial Damage in Experimental Autoimmune Myocarditis in Rats

BACKGROUNDnExcess amounts of NO produced by an inducible NO synthase (iNOS) in response to cytokines may be cytotoxic and can be destructive to tissue. We investigated the role of NO in the development of myocardial damage and the effects of aminoguanidine (AG), an inhibitor of iNOS, on experimental autoimmune myocarditis in rats.nnnMETHODS AND RESULTSnAutoimmune myocarditis was induced in 20 Lewis rats by injection of porcine cardiac myosin. Ten of the 20 rats were administered AG. The severity of myocarditis was evaluated by measuring the size of myocarditic lesion and serum levels of CK-MB. Serum NO levels were determined using the Cd/Cu method. Tissue specimens were immunohistochemically examined for iNOS and nitrotyrosine. Histopathological study revealed extensive myocardial destruction and massive inflammatory cell infiltration in AG-untreated rats but only focal mononuclear cell infiltration in AG-treated rats. The mean percent areas of inflammatory lesions in the untreated and treated rats were 56 +/- 13% and 3 +/- 2%, respectively (P < .001). NO levels were 102 +/- 23 and 25 +/- 9 IU/L, respectively (P < .01). CK-MB levels were 68 +/- 13 and 16 +/- 13 nmol/L, respectively (P < .01). Superoxide production as measured with an ex vivo monitoring system was also significantly decreased in the treated rats. Nitrotyrosine relating to the generation of peroxynitrite was detected through immunostaining in the inflammatory lesions of untreated rats but not in those of treated rats.nnnCONCLUSIONSnExcess amounts of NO produced by iNOS appear to contribute to the progression of myocardial damage in myocarditis. AG may prove to be useful in the treatment of myocarditis.

Tenascin-C Modulates Adhesion of Cardiomyocytes to Extracellular Matrix during Tissue Remodeling after Myocardial Infarction

Tenascin-C (TNC), an extracellular matrix glycoprotein, plays important roles in tissue remodeling. TNC is not normally expressed in adults but reappears under pathologic conditions. The present study was designed to clarify the contribution of TNC to ventricular remodeling after myocardial infarction. We examined the expression of TNC after experimental myocardial infarction in the rat by immunohistochemistry and in situ hybridization. Within 24 hours of permanent coronary ligation, interstitial fibroblasts in the border zone started to express TNC mRNA. The expression of TNC was down-regulated on Day 7 and was no longer apparent by Day 14 after infarction. During the healing process, TNC protein and TNC-producing cells were found at the edges of the residual myocardium. Some of the TNC-producing cells were immunoreactive for α-smooth muscle actin. In culture, TNC increased the number of cardiomyocytes attached to laminin but inhibited the formation of focal contacts at costameres. The results indicate that during the acute phase after myocardial infarction, interstitial cells in the border zone synthesize TNC, which may loosen the strong adhesion of surviving cardiomyocytes to connective tissue and thereby facilitate tissue reorganization.

Tenascin-C is a useful marker for disease activity in myocarditis

Tenascin‐C (TNC) is an extracellular matrix protein which appears at active sites of tissue remodelling during embryogenesis or cancer invasion. In normal heart, TNC is only present during the early stages of development but reappears in pathological states. This study examined the diagnostic value of TNC for assessing disease activity of myocarditis. Expression of TNC was examined in myosin‐induced autoimmune myocarditis mouse models. Sequential changes in amount, localization and the producing cells were analysed by reverse transcriptase–polymerase chain reaction, western blotting, immunohistochemistry and in situ hybridization and compared with the histological picture. The expression of TNC was upregulated at a very early stage of myocarditis. Immunostaining was detectable before cell infiltration and myocytolysis became histologically apparent, remained during the active stage while cell infiltration and necrosis continued, and disappeared in scar tissue with healing. TNC immunostaining was always observed at the periphery of necrotic or degenerating cardiomyocytes in foci of inflammation, the expression level correlating with histological evidence of inflammatory activity. Interstitial fibroblasts were the major source of TNC, expressing the large isoform containing alternative splicing sites. These data demonstrate that TNC is a useful marker for evaluation of disease activity in myocarditis. Copyright

Nonmuscle and Smooth Muscle Myosin Heavy Chain Expression in Rejected Cardiac Allografts A Study in Rat and Monkey Models

BACKGROUNDnDiagnosis of acute rejection and graft arteriosclerosis (chronic rejection) is critical to the success of cardiac transplantation, but accurate diagnosis is often difficult. We have reported that there are three types of vascular myosin heavy chain (MHC) isoforms: SM1, SM2, and SMemb. SM2 is specifically expressed in differentiated smooth muscle cells (SMCs). SMemb is a nonmuscle-type MHC abundantly expressed in SMCs of fetal aorta.nnnMETHODS AND RESULTSnTo evaluate the usefulness of MHC expression for diagnosis and analysis of acute and chronic rejection, heterotopic cardiac transplantation was performed in rats and monkeys. Immunohistochemistry, electron microscopy, and Northern blot assay were performed to evaluate MHC expression. SMemb was expressed in spindle-shaped cells located in acutely rejected myocardium in the rats and monkeys. These cells were also observed in areas lacking cellular infiltration. These SMemb-positive cells were activated fibroblasts or myofibroblasts. SMemb mRNA was enhanced parallel to the progression of acute rejection. In the coronary arteries of chronically rejected allografts, enhanced SMemb and reduced SM2 expression was observed in both thickened intima and media. The reduced medial SM2 expression was observed before the intimal thickening occurred. These cells were phenotypically modulated SMCs.nnnCONCLUSIONSnAltered expression of MHC isoforms is a sensitive indicator in the diagnosis of acute and chronic cardiac rejection. The pathophysiology of this alteration in MHC isoform expression should be studied further to elucidate the pathogenesis of cardiac rejection.

Diagnostic utility of tenascin‐C for evaluation of the activity of human acute myocarditis

Tenascin‐C (TN‐C) is an extracellular matrix protein that is expressed transiently in close association with tissue remodelling in various body sites. In the heart, TN‐C is only present during early stages of development, is not expressed in the normal adult, but reappears in pathological states. The purpose of this study was to analyse the expression of TN‐C in myocardial tissue from myocarditis patients, and to evaluate the diagnostic value of immunostaining for TN‐C in the assessment of inflammatory activity in biopsy specimens. A total of 113 biopsy specimens obtained from 32 patients with a clinical diagnosis of acute myocarditis were examined by immunohistochemistry and in situ hybridization for TN‐C. The immunostaining was semi‐quantified and compared with histological diagnosis according to the Dallas criteria. Furthermore, serial biopsies from 22 patients were taken during convalescence, and sequential changes in TN‐C levels were analysed. Expression of TN‐C was specifically detected in endomyocardial biopsy specimens from patients with active‐stage inflammation, and disappeared in healed stages. The degree of expression of TN‐C correlated with the severity of histological lesions. These data suggest that TN‐C reflects disease activity in cases of human myocarditis. Immunostaining for TN‐C could enhance the sensitivity and accuracy of diagnosis using biopsy specimens. Copyright

Hypertrophic cardiomyopathy in Noonan syndrome

Noonan syndrome, a well‐known multiple congenital anomalies syndrome, is frequently accompanied by cardiovascular diseases including hypertrophic cardiomyopathy (HCM). The incidence of HCM in Noonan syndrome is approximately 20–30% and one‐third of cases reveal ventricular outflow obstruction. HCM in Noonan syndrome is occasionally associated with a congenital heart defect, whereas classic HCM seldom accompanies cardiac malformations. Asymmetric septal hypertrophy and symmetric septal hypertrophy (concentric hypertrophy) can be observed both in HCM with Noonan syndrome and in classic HCM. but apical hypertrophy has not been reported in Noonan syndrome yet, although it appears in classic HCM. Congestive heart failure is the major cause of death in patients with HCM in Noonan syndrome, but cases of sudden death have also been reported. The histopathologic findings of ventricular myocardial tissue in HCM with Noonan syndrome are similar to those in classic HCM.

Antisense Bcl-x oligonucleotide induces apoptosis and prevents arterial neointimal formation in murine cardiac allografts

OBJECTIVEnCardiac allograft arteriosclerosis, which limits long-term survival of recipients, cannot be prevented by conservative therapies. The arteriopathy is characterized by diffuse intimal thickening comprised of proliferative smooth muscle cells (SMCs). Cell death is a prominent feature of atherosclerosis; Bcl-x is one of the anti-apoptotic mediators.nnnMETHODSnTo test the hypothesis that antisense bcl-x oligodeoxynucleotide (ODN) is effective in preventing intimal hyperplasia through enhancing apoptosis after cardiac transplantation, we performed single intraluminal delivery of antisense bcl-x ODN into murine cardiac allografts (n = 9). DBA/2 (H-2d) hearts were transplanted into B10.D2 (H-2d) mice. Sense bcl-x ODN (n = 8) and no treatment (n = 8) studies were also performed.nnnRESULTSnAllografts were harvested at 4 weeks after transplantation; all allografts kept beating throughout the period. Coronary intimal thickening had developed in nontreated and sense ODN transfected allografts at 4 weeks after transplantation with enhanced expression of Bcl-x and cell adhesion molecules, and suppressed apoptosis. However, antisense bcl-x ODN prevented neointimal formation through enhanced apoptosis.nnnCONCLUSIONnThese results indicate that apoptosis of vascular SMCs induced by Bcl-x is associated with initial hyperplasia after heart transplantation. Antisense bcl-x ODN inhibits SMC proliferation by inducing apoptosis in graft coronary arteries.

Intravascular stents for management of pulmonary artery and right ventricular outflow obstruction.

SummaryThis study was performed to determine the efficacy of balloon-expandable stents in the treatment of branch pulmonary artery-stenoses and conduit stenosis in children. A total of eight stainless steel stents were implanted in seven patients. Three patients had tetralogy of Fallot with pulmonary artery stenosis following total correction, one patient had conduit stenosis following correction of transposition of the great arteries, one patient had intra-cardiac conduit stenosis after septation for single left ventricle, and two patients had pulmonary artery stenosis after Fontan operation. Six stents were placed in the branch pulmonary arteries, one in the extracardiac conduit, and one in the intracardiac conduit. The mean age at implantation was 13 ± 3 years and the mean weight 37 ± 12kg. Follow-up time ranged from 0.3–2 years. The diameter of pulmonary arteries with stenoses increased from 5.6 ± 2.2 mm to 10.6 ± 1.8 mm (n = 7). The systolic pressure gradient decreased from 56 ± 26 mmHg to 22 ± 16 mmHg (n = 5). No embolization or thrombotic event has been noted. One stent placed in the intracardiac conduit was compressed and fractured. These data indicate that balloon-expandable stents are useful in the treatment of pulmonary artery branch stenoses and extracardiac conduit stenosis in children. The use of stents for intracardiac stenosis may result in stent fracture.

An autopsied case of Williams syndrome complicated by moyamoya disease

An 18 year old girl with typical clinical features of Williams syndrome suddenly died of intracerebral hemorrhage due to moyamoya disease. Autopsy revealed vascular abnormalities, such as supravalvular aortic stenosis (SAS) and an abnormal complicated cerebrovascular network in the cerebral arteries. The arterial wall of the SAS lesion consisted of thickened medial tissue showing elastic disorganization with prominence of the smooth muscle cells. The narrowed vessels of the circle of Willis showed intimal thickening with an extremely wavy internal elastic lamina and marked thinning of the media. To our knowledge, this is the first report of moyamoya disease associated with Williams syndrome.

In Situ Morphology of the Heart and Great Vessels in Fetal and Newborn Rats

ABSTRACT. Morphology of the cardiac chambers and great vessels of fetal and neonatal rats was studied using the whole body freezing technique and by sectioning through the short axis of the heart with a freezing microtome. Compared to the fetal heart, the neonatal heart showed rapid change 2 to 8 days after birth. The ventricular sinus septum was straight in the fetus and became concave to the left ventricle after birth. The right ventricular wall was as thick as the left in the fetus and became thinner rapidly after birth. At the same time, the right ventricular cavity dilated. The right and left pulmonary arteries and pulmonary veins were small in the fetus and enlarged soon after birth. At the same time, the foramen ovale was closed and the diameters of the inferior vena cava and descending aorta were diminished. One-half-mm thick sections were cut serially and then photographed. Ventricular volumes and masses were calculated from summation of the areas of each chambers. Left ventricular mass per body weight increased rapidly after birth, whereas right ventricular mass per body weight remained constant from 0 to 8 days after birth.