Identification of nucleolin as a binding protein for midkine (MK) and heparin-binding growth associated molecule (HB-GAM).

Midkine (MK) is a heparin-binding growth/differentiation factor with a molecular weight of 13 kDa, and is structurally unrelated to fibroblast growth factors (FGF). We studied MK-binding proteins in order to clarify the action mechanism of MK. A 100-kDa protein was identified in PYS-2, 3T3, and L cells as an MK-binding protein by a ligand blot experiment. This MK-binding protein was purified by affinity chromatography on an MK-agarose column followed by SDS polyacrylamide gel electrophoresis. Sequence determination of N-terminal 23 amino acid residues revealed that the MK-binding protein was nucleolin, a major nucleolar protein, which functions as a shuttle protein between the nucleus and cytoplasm and is located also on the cell surface. Heparin-binding growth associated molecule (HB-GAM), which has 50% sequence identity with MK, fused to maltose-binding protein also bound to nucleolin. On the other hand, basic FGF (bFGF) scarcely bound to nucleolin in the absence of heparin, while both MK and bFGF bound weakly to nucleolin in the presence of heparin. Nuclear localization of MK was shown in hemangioma cells by immunohistochemical staining. These findings supported the hypothesis that parts of the MK and HB-GAM are translocated to the nucleus after binding with nucleolin.

Mapping and characterization of a retinoic acid-responsive enhancer of midkine, a novel heparin-binding growth/differentiation factor with neurotrophic activity.

MK is a gene that is activated by retinoic acid in embryonal carcinoma (EC) cells and is expressed temporarily during the mid-gestation period of mouse embryogenesis. Midkine, the product of the gene is a novel heparin-binding growth/differentiation factor with neurite outgrowth and neurotrophic activities. The regulatory DNA element in the retinoic acid-induced expression of the MK gene has been investigated. The 1.9 kb 5'-flanking region of the MK gene can mediate retinoic acid-responsive gene expression in F9 and HM-1 EC cells. Analysis of this region by deletion mutagenesis in F9 EC cells shows that there is a retinoic acid-responsive enhancer (designated as REM1) around 900 bp upstream from the transcription start site. This enhancer is composed of two sequence elements, which are located between -1006 and -895 and between -901 and -794. The core element of the upstream region (-971 to -955), whose deletion abolished the retinoic acid responsiveness, contained a sequence highly homologous to a binding site for retinoic acid receptors. Binding of a retinoic acid receptor heterodimer to this core element was verified by gel shift assay. Thus, retinoic acid and the receptor complex can directly induce the expression of a growth/differentiation factor gene.

Clinicopathological analysis of a patient with hypereosinophilic syndrome who demonstrated disseminated eosinophilic infiltration in various organs at autopsy.

A case (41 year old man) of idiopathic hypereosinophilic syndrome is presented. Typical eosinophilic endocarditis with restrictive cardiac failure and ventricular thrombosis was demonstrated by various clinical investigations, including cardiac pool scan and biopsy. The patient died after 22 months, and an autopsy revealed disseminated eosinophilic infiltration in the pancardium of the whole heart, aorta, main pulmonary artery, liver, and spleen.

High incidence of significant pathology in endomyocardial biopsy and familial occurrence in cases with arrhythmia and/or conduction disturbance.

In our series of studies of cardiomyopathy with endomyocardial biopsy in 1500 cases, we have noticed that there were a certain number of patients with arrhythmias and/or conduction disturbance who showed no evidence of hypertrophic or dilated form of caridiomyopathy but showed significant pathology in the biopsy. Significant pathology was determined when the biopsy findings from the right atrium and the right ventricle showed apparent pathology such as interstitial fibrosis, myocyte degeneration, and fragmentation of muscle bundles. Among 226 biopsied patients in whom arrhythmia and/or conduction disturbance were the main clinical feature, 85 patients (38%) demonstrated significant pathology in their biopsy. Familial occurrence was note-worthy among these patients. We propose that these cases should be grouped as nonhypertrophic, nondilated, and nonrestrictive cardiomyopathy, and suggest the term electric disturbance type of cardiomyopathy (ECM).

Effects of 4-week treatment with gastric carcinogens and enhancing agents on proliferation of gastric mucosa cells in rats.

Catechol and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) are gastric carcinogens in rats. Catechol, sodium chloride and bile salts have enhancing effects on gastric carcinogenesis induced by MNNG in rats. The effects of these compounds on proliferation of pyloric mucosa cells in male F344 rats were examined immunohistochemically using bromodeoxyuridine (BrdU) and anti-BrdU monoclonal antibody. Rats were given MNNG (83 micrograms/ml in their drinking water), catechol (0.8% in their diet), sodium taurocholate (0.3% in their diet), sodium taurodeoxycholate (0.3% in their diet), or sodium chloride (10% in their diet or by intragastric administration of 1 ml of saturated solution once a week) for 4 weeks. All these treatments markedly enhanced cell proliferation of the pyloric epithelium, suggesting the importance of enhanced cell proliferation in the development of gastric cancer.

A comparative ultrastructural study on the nature of interstitial cell constituents in idiopathic myocarditis and myocardial infarction.

In order to observe the characteristics of interstitial cellular proliferation in cases with myocarditis of possible viral origin, a systematic ultrastructural analysis of the cell components in the interstitium was made employing endomyocardial biopsy at the acute (0-10 days after onset), subacute (11-25 days), and convalescent stages (26-167 days) of the disease in ten cases. For comparison, myocardial specimens from cases with myocardial infarction were taken from early autopsy cases or from tissue obtained during surgery. The dominant interstitial cells observed in the acute stage of myocarditis were fibroblasts (38.3% +/- 17.5%, mean +/- SD), macrophages (23.3% +/- 12.1%), and lymphocytes (18.0% +/- 18.1%); at the convalescent stage, fibrocytes (44.6% +/- 20.2%) and fibroblasts (22.5% +/- 8.0%) were commonly seen. In myocardial infarction, the dominant cells were macrophages (35.0% +/- 16.0%) and neutrophils (26.0% +/- 13.9%) at the acute stage, and lymphocytes (30.0% +/- 17.8%), plasma cells (27.1% +/- 20.0%), and macrophages (26.4% +/- 11.4%) at the late stage. Thus, some differences between the two diseases could be recognized.

A clinicopathologic study on a cause of idiopathic cardiomyopathy and arrhythmia and conduction disturbance employing endomyocardial biopsy.

A comparative study of right ventricular histopathologic findings and clinical profiles in 174 cases with hypertrophic cardiomyopathy (HCM), 145 cases with dilated cardiomyopathy (DCM), and 241 cases with idiopathic arrhythmia and/or conduction disturbance with significant myocardial pathology [Electric disturbance type of cardiomyopathy (ECM)], totaling 560 cases, was made in order to clarify the role of viral myocarditis in these conditions. Postmyocarditis change (PMC) was defined by assessing serial biopsy findings in nine cases with acute myocarditis. The PMC was observed in three cases with HCM (1.7%), 22 with DCM (15%), and 21 with ECM (8.7%). The incidence of PMC was significantly higher in DCM and ECM than in HCM (P less than 0.001). The typical pattern of onset of acute myocarditis, i.e., high fever, upper respiratory infection, and gastrointestinal symptoms preceding cardiac symptoms within 10 days, was recognized in one case with HCM, eight with DCM, and 6 of 123 cases with ECM. The incidence was higher in DCM and ECM than in HCM (P less than 0.05). Of 46 cases with PMC, 12 (26%) showed this pattern, on the other hand only 3 of 377 cases (0.8%) without PMC did so (P less than 0.0001). Familial occurrence was not found in any of the 46 cases with PMC but was found in 38 of the 377 cases without PMC (P less than 0.01). This study indicates that viral myocarditis may often be the causative agent of idiopathic cardiomyopathy, especially DCM and ECM. It is also noticed that familial occurrence of the cases with PMC was never seen.

Long follow-up study in patients with prior myocarditis by radionuclide methods.

Ten patients with previous myocarditis were evaluated to determine cardiac conditions by T1-201 myocardial perfusion imaging and stress radionuclide ventriculography during the follow-up of 18-102 (average 56) months; the results were compared with those from ten sex- and age-matched controls. Exercise capacity by supine bicycle ergometer was reduced in patients with myocarditis. Their resting left ventricular ejection fraction (LVEF) was 57.5% +/- 3.9%, similar to that of controls. LVEF response to stress in myocarditis was abnormal with an increment of end-systolic volume, while in the controls LVEF increased significantly during stress. Seven of the eight patients with an abnormal ejection fraction response had constant T1-201 perfusion defects. This study indicates that latent left ventricular dysfunction is present in patients with prior myocarditis and that nuclear study is useful for long-term follow-up.

Clinical spectrum and endomyocardial biopsy findings in eosinophilic heart disease.

Fourteen cases of heart disease with hypereosinophilia were analyzed employing conventional cardiologic methods, including echocardiography, cardiac catheterization, and endomyocardial biopsy. The cases were divided into four types: Acute carditic (endocarditis, myocarditis, pericarditis; five cases); ventricular dilation (three cases); restrictive (three cases); electric disturbance (three cases). Biopsy revealed significant changes in all cases. In one case of the ventricular dilation type, endomyocardial fibrosis with myocardial degeneration was seen, and in another case mural thrombus formation was shown to be present. In three cases of the restrictive type, endomyocardial fibrosis (EMF) was observed. In two cases of the electric disturbance type, minor right ventricular myocardial degeneration was observed. In two of the three cases of the carditic type and in three of eight cases in other categories, postmyocarditic changes were observed. The course of the disease compared with the type of disorder revealed a short course in the carditic type and a longer course, ranging from 2 to 24 years, with one exception, in the other types. It is also confirmed that the various histopathologic changes can be related to particular clinical presentations. We have shown that the basic changes in eosinophilic heart disease are not restricted to the endomyocardium and that they occur in various parts of the heart causing more widespread manifestations. The more comprehensive term "eosinophilic heart disease" is a preferable description.

Hemodynamic studies and response to a combined therapy of nitroglycerin ointment and dopamine in patients with acute myocarditis.

Left ventricular performance was assessed in ten patients with acute myocarditis during the acute stage (within the first 3 days after the onset of cardiac symptoms) and during the convalescent phase (3-5 weeks later). In nine, the diagnosis was confirmed by endomyocardial biopsy during the acute stage, when left ventricular function was markedly reduced, i.e., cardiac index 1.53 +/- 0.33 l/min/m2 (mean +/- SD), left ventricular stroke work index 14.1 +/- 3.9 g/beat/m2, pulmonary capillary pressure 25.9 +/- 4.3 mm Hg, and systemic vascular resistance 27.9 +/- 9.4 R units. During convalescence, seven patients improved clinically with normal cardiac function and in three it remained impaired. Cardiovascular response to nitroglycerin ointment in three patients and a combination of dopamine and nitroglycerin ointment in five were compared. Nitroglycerin ointment reduced pulmonary capillary pressure by an average of 28.5% (P less than 0.05) and systemic vascular resistance by 26.8%, while the cardiac index increased by 34.1% (P less than 0.05) and left ventricular stroke work index by 30.3% (P less than 0.01). The combined therapy was more effective, with a reduction of pulmonary capillary pressure by 30.4% (P less than 0.05) and systemic vascular resistance by 27.5% (P less than 0.05), accompanying an increase in cardiac index by 53.5% (P less than 0.005) and left ventricular stroke work index by 63.5% (P less than 0.01), with little change in the double products. This study suggests that serial hemodynamic monitoring accurately evaluates cardiac function, selection of appropriate therapy, and the hemodynamic effects of treatment in patients with acute myocarditis.(ABSTRACT TRUNCATED AT 250 WORDS)

Eosinophilic disorders affecting the myocardium and endocardium: a review.

A wide range of disorders give rise to eosinophil counts greater than 1.5 X 10(9)/l (hypereosinophilia) and cardiac injury. The best known of these is eosinophilic endomyocardial disease (Löffler's endomyocardial fibrosis), which occurs as a major complication of the idiopathic hypereosinophilic syndrome. Here the heart damage appears to be a direct result of tissue injury produced by toxic eosinophil granule proteins within the heart. However, it is not known what causes the eosinophilia in these patients, why the eosinophils degranulate, or why the endocardium is especially susceptible to this type of injury. A number of parasitic infections may give rise to eosinophilic myocarditis. This is usually the result of the presence of the parasites within the myocardium where they die within inflammatory lesions, which may be extensive. Occasionally, drug reactions and rejection of a transplanted heart may produce eosinophilic myocarditis. Allergic granulomatosis and vasculitis (the Churg-Strauss syndrome), which gives rise to granulomas involving the myocardium, and eosinophilic (hypersensitivity) myocarditis usually respond rapidly to treatment with steroids. However, diffuse myocardial involvement may lead to heart failure, and some of these patients may later develop dilated cardiomyopathy. It is concluded that the heart may be affected by a variety of diseases in which eosinophils are a prominent component in the inflammatory cell infiltrates. Eosinophils themselves may contribute to some of the myocardial cell injury which occurs in these diseases, and attempts to limit this with steroids may be worthwhile in some patients.

Histopathological findings of acute and convalescent myocarditis obtained by serial endomyocardial biopsy.

Serial endomyocardial biopsy findings in 20 cases with acute myocarditis of possible viral origin were analyzed. The histopathological findings were analyzed during the acute (0-10 days after the onset: 8.3 +/- 1.9 days; 6 cases), the subacute (11-21 days: 18.2 +/- 2.2 days, 6 cases), and the convalescent stages (22-167 days: 54.5 +/- 45.4 days; 8 cases). The incidence and severity of various changes of the cardiac myocytes and interstitial changes were analyzed and compared at each stage of the disease. In the acute stage, interstitial cell infiltrations were composed of fibroblasts, macrophages and lymphocytes, taken out in descending order. In the convalescent stage, interstitial cell infiltration showed a marked increase and was replaced by fibrocytes. In the subacute stage, transitional changes between the acute and convalescent stages were observable. A controlled myocardium in 21 cases with myocardial infarction which were compared at various stages revealed that in the acute stage, neutrophils were most prominent, and in the subacute and convalescent stages, macrophages were most prominent and plasma cells were most often observed. The time course changes of the histopathological findings in acute myocarditis were as follows: In the acute stage, Interstitial cell infiltration which is composed of fibroblasts, macrophages and lymphocytes, fragmentation of the muscle bundles, myocytolytic changes, swelling and scarcity of the cytoplasm and swelling of nuclei, variation in size of the myocytes, disarrangement of the muscle bundles, interstitial edema, increased glycogen deposition in the myocytes, abnormal branching of the myocytes, and interstitial fibrosis were observable. In the convalescent stage, most of the above described findings were still observable except for the myocytolytic change, swelling of myocytes and interstitial edema.(ABSTRACT TRUNCATED AT 250 WORDS)

Ultrastructural features of the endomyocardium in patients with eosinophilic heart disease. An endomyocardial biopsy study.

In a series of studies with endomyocardial biopsy, 7 adult cases with cardiac disease and eosinophilia were studied clinically and electro-microscopy. Degranulation of the eosinophils in the peripheral blood was observed ultrastructurally in 3 of the 4 cases studied. The clinical expression of the 7 patients were, restrictive cardiomyopathy in 2, dilated cardiomyopathy in 2 and sick sinus syndrome in 1 and others in 2. Endocardial thickening was observed in 5 cases, one of whom showed a marked cellular infiltration with macrophages, plasma cells, lymphocytes and mast cells. One other case showed cell debris of degranulated eosinophils. Degeneration of the myocytes was manifested by an increase in Z-bands of the myofibrils as well as streaming (1 case), disarrangement of the myofibrils, and mitochondrial change (1 case) which was characterized by giant mitochondria (1 case) as well as by a numerical increase in mitochondria. Increase of atrial granules and mitochondria in a right atrial biopsy of a case with sick sinus syndrome was noteworthy. Basal lamina layering of the capillaries of the myocardium and pyknosis of an endothelial cell cytoplasm were also noted in each case. These observations may reveal that various disease processes are taking place in the endocardium and adjacent myocardium.

Ventriculographic findings in the convalescent stage in eleven cases with acute myocarditis.

In order to clarify the sequelae of 11 patients who suffered from acute myocarditis of viral origin and were diagnosed by endomyocardial biopsy in the early and the convalescent stage of the disease, left ventriculograms were obtained from 17 to 167 (46.5 +/- 41.2, mean +/- SD) days after the onset of the disease were analyzed. The data were compared with the biopsy findings in the convalescent stage. The mean left ventricular ejection fraction (LVEF) was 57.5 +/- 10.8% and in 3 cases the LVEF was less than 50%. In 2 cases, apparent asyneresis was observed. In all cases coronary arteriographic findings did not show significant stenosis. It is concluded that after viral myocarditis, increase in left ventricular end-diastolic volume, depressed ejection fraction and asyneresis occur in some cases and it can be said that a development from myocarditis to dilated cardiomyopathy occurs.

Long-term follow-up of electrocardiographic findings in patients with acute myocarditis proven by endomyocardial biopsy.

Sixteen cases of acute myocarditis, proven by endomyocardial biopsy, of possible viral origin which were described in a previous paper have been followed up for up to 5 years. The cases were divided into 3 groups according to the electrocardiographic (ECG) patterns: with conduction disturbance at the early stage (Group I, 9 cases), without conduction disturbance (Group II, 4 cases); early death cases in which the follow-up could not be carried out (Group III, 3 cases). In Group I, 2 cases revealed persistent complete A-V block, 4 showed incomplete recovery of right bundle branch and/or fascicular blocks, and 3 showed complete recovery of the ECG abnormalities. In Group II, all 4 cases showed a pseudoinfarction pattern which consisted of abnormal Q waves, poor R wave progression and ST elevation. These findings disappeared during the follow-up period. All patients of Groups I and II are still alive after a follow-up period of up to 5 years.

Early clinical profiles of cases with histopathologically proven acute idiopathic myocarditis and a proposal for diagnostic criteria.

The early clinical features, primary care, treatment and short-term prognosis in 15 cases of acute myocarditis where diagnostic confirmation was made by endomyocardial biopsy or autopsy were analyzed. Characteristically, idiopathic myocarditis of possible viral etiology revealed preceding symptoms which consisted of flu-like symptoms, i.e., fever, upper respiratory infection (sore throat, cough), myalgia or arthralgia, general malaise, and gastrointestinal disorders (vomiting, anorexia, nausea, abdominal pain and soft stool). A severe cardiac or generalized disease condition may follow. Depending upon the progress of intensive medical and cardiac care, the patients' prognosis is not always poor. Diagnostic criteria based upon our own experience have been constructed as a proposal.