[Surgical removal of native aortic valve thrombosis associated with acute myocardial infarction and protein C deficiency; report of a case].

Native aortic valve thrombosis is an uncommon event. We describe the case of a 76-year-old man who suffered acute myocardial infarction associated with native aortic valve thrombosis diagnosed by cardiac catheterization. Since the thrombus was localized on the right coronary cusp and occluded right coronary artery, rescue revascularization was performed using perfusion catheter, which was continuously engaged to the right coronary artery. Operation was immediately performed under cardiopulmonary bypass. After incision of ascending aorta, thrombus was removed easily and aortic valve was preserved without degenerative change. Histological study showed a typical thrombus without any specific findings. He had a good clinical course and discharged 9 days after the operation. He had no history of heart valve disease, left heart catheterization or bacterial endocarditis. Since laboratory data showed 41% in protein C antigen and 32% of protein C activity, he was diagnosed of protein C deficiency. Our report emphasize that this thrombus formation may be caused by protein C deficiency.

Gene regulation of melatonin and dopamine receptors during eye development.

To gain insight into the role of melatonin and dopamine in retinal development, gene expression of two melatonin receptors, MT1 and MT2, as well as five dopamine receptors, D1, D2, D3, D4 and D5, in the rat eye was analyzed by reverse transcription-polymerase chain reaction across various developmental stages. MT1 transcript levels reached maximum levels at embryonic day (E) 16 and then decreased gradually until reaching adult levels by postnatal day (P) 14. MT2 transcript levels similarly peaked at E16, but then decreased dramatically until birth to its lowest levels, which were maintained throughout the postnatal period. Thus, gene expression of both the MT1 and MT2 receptors showed a striking inverse correlation with maturation of the eye. In contrast to melatonin receptors, gene expression of all dopamine receptor subtypes, except for D3, showed only an increase as development proceeds with highest levels in adulthood. The D3 message was not detected throughout the developmental period examined. Gene expression of D1-like receptors, D1 and D5, showed a substantial increase to adult levels during the fetal period at E16 and E20, respectively. Transcript levels of D2-like receptors, D2 and D4, on the other hand, were not detected before birth but increased significantly to adult levels by P7 and P14, respectively. The present findings suggest the presence of unique developmental mechanisms by which transcription of various G protein-coupled receptors are regulated in the eye.

Focal enhancement of expression of c-Met/hepatocyte growth factor receptor in the myocardium in human myocardial infarction.

To determine the distribution and expression level of hepatocyte growth factor (HGF) specific receptor, c-Met, in human myocardial infarction. Autopsies of 13 patients who died without heart diseases (control) and 13 patients with a history of myocardial infarction (2 h to 10 years before death). The harvested myocardial tissues were stained with hematoxylin-eosin (H&E) and immunohistochemically stained for c-Met expression by the avidin-biotin-horseradish peroxidase complex method using an antibody to c-Met. C-Met expression was only slightly increased in control subjects and in noninfarcted myocardium of the test group. In contrast, high expression was noted in the peripheral region of the myocardial infarction and in some hypertrophic myocardial cells. C-Met was not expressed in the infarcted myocardium, but overexpression was noted in the surrounding myocardial cells of blood vessels and in the subendocardium and subepicardium in a band-like pattern. The expression level of c-Met was most enhanced at the time of appearance of coagulative necrosis and least in the myocardium of subjects with old infarcts. Our results indicate that HGF preferentially reaches the ischemic regions of the myocardium and has local and direct effects on the myocardium in patients with myocardial infarction.

Cyclical regulation of GnRH gene expression in GT1-7 GnRH-secreting neurons by melatonin.

The pineal hormone melatonin plays an important role in the neuroendocrine control of reproductive physiology, but its effects on hypothalamic GnRH neurons are not yet known. We have found that GT1-7 GnRH-secreting neurons express membrane-bound G protein-coupled melatonin receptors, mt1 (Mel-1a) and MT2 (Mel-1b) as well as the orphan nuclear receptors ROR alpha and RZR beta. Melatonin (1 nM) significantly downregulates GnRH mRNA levels in a 24-h cyclical manner, an effect that is specifically inhibited by the melatonin receptor antagonist luzindole (10 microM). Repression of GnRH gene expression by melatonin appears to occur at the transcriptional level and can be mapped to the GnRH neuron-specific enhancer located within the 5' regulatory region of the GnRH gene. Using transient transfection of GT1-7 cells, downregulation of GnRH gene expression by melatonin was further localized to five specific regions within the GnRH enhancer including -1827/-1819, -1780/-1772, -1746/-1738, -1736/-1728, and -1697/-1689. Interestingly, the region located at -1736/-1728 includes sequences that correspond to two direct repeats of hexameric consensus binding sites for members of the ROR/RZR orphan nuclear receptor family. To begin to dissect the mechanisms involved in the 24-h cyclical regulation of GnRH transcription, we have found that melatonin (10 nM) induces rapid internalization of membrane-bound mt1 receptors through a beta-arrestin 1-mediated mechanism. These results provide the first evidence that melatonin may mediate its neuroendocrine control on reproductive physiology through direct actions on the GnRH neurons of the hypothalamus, both at the level of GnRH gene expression and through the regulation of G protein-coupled melatonin receptors.

Extensive myocardial stunning showing transient regression of prolonged T wave inversion and prolonged sympathetic denervation.

A 69-year-old woman was admitted to the hospital with palpitations. Although left ventriculography showed extensive akinesis except in the basal hyperkinetic segment, coronary angiography showed normal coronary arteries. 123I-metaiodobenzylguanidine (MIBG) accumulation was obviously reduced in the anteroseptal, apical and inferior areas. Inverted T waves developed on day 3 and disappeared on day 104 after transient regression. Echocardiography showed normal left ventricular motion two weeks later. Ergonovine provocation test showed no vasospasm and thallium-201 showed no perfusion defect on day 46. Electrocardiography and MIBG returned to normal on day 216. These findings suggest prolonged sympathetic nerve injury in extensive myocardial stunning.

Dopaminergic and GABAergic amacrine cells are direct targets of melatonin: immunocytochemical study of mt1 melatonin receptor in guinea pig retina.

Distribution of the mt1 melatonin receptor in the guinea pig retina was immunocytochemically investigated using peptide-specific anti-mt1 receptor antibody. Western blots of the guinea pig retina showed a single band at approximately 37 kilodalton (kD) immunoreactive to the anti-mt1 antibody. The most intense immunoreactivity for the mt1 receptor was detected in the cell bodies of ganglion cells. Their dendrites and axons were also immunolabeled. Subpopulations of amacrine cells, the inner plexiform layer, and the outer plexiform layer also exhibited moderate to weak immunolabeling. The mt1-positive amacrine cells were located either at the vitreal border of the inner nuclear layer or displaced in the ganglion cell layer. Double immunolabeling using antibodies to the mt1 receptor and tyrosine hydroxylase revealed that the majority of dopaminergic amacrine cells showed mt1 immunoreactivity. Almost all the ICA type dopaminergic cells were mt1 positive while the 2CA type cells less frequently exhibited mt1 immunoreaction. By double immunolabeling for the mt1 receptor and GABA, more than 50% of the mt1-immunoreactive amacrine cells were shown to be GABAergic neurons. Approximately one-third of the GABAergic amacrine cells were immunolabeled for the mt1 receptor. The present results demonstrate expression of the mt1 receptor in diverse neuronal cell types in the guinea pig retina and provide the first evidence for the direct effect of melatonin on dopaminergic and GABAergic amacrine cells via the mt1 receptor.

Sequential changes of hepatocyte growth factor in the serum and enhanced c-Met expression in the myocardium in acute myocardial infarction.

A 68-year-old male with acute myocardial infarction (AMI) was admitted to the hospital with chest pain that had started 1 day earlier. The serum levels (ng/ml) of hepatocyte growth factor (HGF) were 1.06, 1.22, 1.05, 0.72 and 0.64 on days 2, 3, 4, 5 and 6 postinfarction, respectively. He died suddenly due to cardiopulmonary arrest on day 6. At autopsy, approximately 400 ml of bloody pericardial fluid, caused by rupture of the left ventricle, was detected and the c-Met expression in the myocardium was immunohistochemically found to be most intense in the border zone of the infarcted and non-infarcted region. Although there was no c-Met expression in the infarcted myocardium, it was increased in the myocardial cells surrounding the blood vessels. This is the first report to show sequential changes of HGF in the serum, as well as c-Met expression in the myocardium, in a patient with AMI.

Prognostic value of serum hepatocyte growth factor in patients with acute coronary syndromes.

The present study examined whether or not hepatocyte growth factor (HGF), an endothelium-specific growth factor that stimulates regeneration of the endothelium, is increased or has a prognostic significance in patients with acute coronary syndromes. HGF was measured in 106 patients with coronary artery disease (20 stable effort angina, 12 unstable angina without adverse events, 24 unstable angina with adverse events and 50 acute myocardial infarction) on admission and 21 normal volunteers. The measurements in all patients were recorded before administration of heparin, and in acute myocardial infarction patients they were recorded from days 2 to 6 after heparin discontinuation on day 1. HGF levels (ng/ml) were 0.30+/-0.06 for the controls, 0.31+/-0.08 for stable effort angina patients, 0.31+/-0.08 for unstable angina patients without adverse events, 0.40+/-0.20 for unstable angina patients with adverse events and in acute myocardial infarction patients they were 0.45+/-0.18 on day 0, 0.57+/-0.45 on day 2, 0.50+/-0.35 on day 3, 0.48+/-0.32 on day 4, 0.44+/-0.20 on day 5, and 0.38+/-0.14 on day 6. HGF plays a crucial role in the restoration of injured endothelial cells and is a predictor of adverse events in patients with acute coronary syndromes.

Expression of mt1 melatonin receptor in rat retina: evidence for multiple cell targets for melatonin.

Melatonin is synthesized in the retina at night and acts as a local modulator within this tissue by mediating the effects of darkness. We investigated the expression and localization of the mt1 (Mel1a) melatonin receptor in rat retina in order to disclose the cellular and molecular bases of melatonin's action in the mammalian retina. Western blotting of the mt1 receptor in rat retina exhibited a single immunoreactive band of approximately 37,000 mol. wt, which corresponds to the predicted molecular size of the receptor. The mt1 receptor was immunocytochemically localized to both the inner and outer plexiform layers. During postnatal development, retina from two-week-old rats showed the highest mt1 immunoreactivity; the outer plexiform layer and horizontal cell bodies were strongly immunolabeled, with weaker labeling in the inner plexiform layer. Expression of mt1 receptor messenger RNA in the rat retina was demonstrated by reverse transcription-polymerase chain reaction and in situ hybridization. mt1 receptor transcripts were localized to ganglion cells, amacrine cells and horizontal cells. These results suggest that melatonin influences retinal physiology by acting on multiple retinal cell types, including ganglion, amacrine and horizontal cells, via the mt1 receptor expressed in their processes.

Regional left ventricular dysfunction in a patient with severe prolonged anemia.

A 47-year-old woman with severe prolonged anemia developed heart failure. After treatment of the heart failure and anemia, she showed regional dysfunction of the left ventricular wall and myocardial fatty acid metabolism was disturbed in these sites. Coronary arteriography showed normal images. It took about 4 months to recover both left ventricular wall motion and fatty acid metabolism. Prolonged decrease of oxygen supply to the myocardium, which is caused by severe prolonged anemia, seemed to affect the myocardial function in this case, which could be another model of anemia-related myocardial dysfunction.

[Significance of hepatocyte growth factor measurement in patients with acute myocardial infarction].

The use of hepatocyte growth factor for acute myocardial infarction was investigated. Several other biochemical markers are already used for noninvasive detection of acute myocardial infarction. Hepatocyte growth factor, creatine phosphokinase (CK) and CK isozyme (CK-MB) levels were measured in 10 patients with stable effort angina after diagnostic catheterization, and in 21 patients with acute myocardial infarction twice a day for the first 3 days and once a day for the next 4 days. The time to reach the maximum level and the time to decline to less than half of the maximum level for each patient was also measured. Hepatocyte growth factor levels (ng/ml) were 0.3 +/- 0.1 for patients with angina pectoris, and 10.4 +/- 8.8 within 6 hours and 6.7 +/- 4.5 within 12 hours after the onset for patients with acute myocardial infarction. The time to reach the maximum (hours) and the time to decline to less than half of the maximum level (days) were 9.2 +/- 5.2 and 1.1 +/- 0.3 for hepatocyte growth factor, 19.5 +/- 7.2 and 2.3 +/- 1.1 for CK and 16.3 +/- 7.2 and 1.5 +/- 0.4 for CK-MB, respectively. The correlation coefficients between the maximum level of hepatocyte growth factor and the maximum levels of CK and CK-MB were 0.64 and 0.70, respectively. Hepatocyte growth factor is useful as a prognostic indicator and reflects the clinical course in patients with acute myocardial infarction.

Remodeling of pineal epithelium in the fetal rat as delineated by immunohistochemistry of laminin and cadherin.

Epithelial remodeling in the rat pineal during fetal development was immunohistochemically analyzed by using antibodies for laminin and cadherin as molecular markers of basal lamina and intercellular junctions, respectively. The proliferation and differentiation of pinealocytes were also investigated in relation to the advance of epithelial remodeling. The pineal anlage of embryonic day 16 is completely covered by basal lamina immunolabeled for laminin. After embryonic day 17, local dissolution of the basal lamina occurs on the epithelial folds, which develop predominantly in the rostral pineal wall. Some pineal cells migrate through these interruptions and form cellular aggregations outside the basal lamina. Cadherin immunostaining reveals focal dissolution of intercellular junctions in epithelial regions protruding into the pineal lumen. Dissolution of the basal lamina and intercellular junctions accompanied by cellular migration into the stromal tissue or into the pineal lumen continues until birth. The distribution of mitotic cells immunolabeled for BrdU is homogeneous throughout the organ during the fetal period, whereas that of differentiating pinealocytes immunoreactive for synaptophysin shows striking regional heterogeneity in close correlation with the remodeling of the pineal epithelium. The migrating cell populations located either outside the basal lamina or inside the pineal lumen are more liable to become synaptophysin-positive than the rest of the epithelium. These results suggest that epithelial remodeling in the fetal pineal is induced, at least in part, by epithelial infolding and that this remodeling promotes the differentiation of pinealocytes.

Hepatocyte growth factor(HGF): a new biochemical marker for acute myocardial infarction.

The purpose of this study was to investigate the use of hepatocyte growth factor as a biochemical marker for acute myocardial infarction. Several biochemical markers are used for noninvasive detection of acute myocardial infarction. However, hepatocyte growth factor has not been used previously for this purpose. We measured hepatocyte growth factor, creatine phosphokinase, and MB isoenzyme (CK-MB) levels in 6 patients with stable effort angina after diagnostic catheterization (controls) and in 12 patients with acute myocardial infarction (AMI). The measurements in the AMI patients were recorded twice a day for the first 3 days after onset of chest pain and once a day for the next 4 days. Furthermore, in each patient we evaluated the time to reach the maximum level and the time for the level to decline to less than half the maximum. Hepatocyte growth factor levels (ng/ml) were 0.3+/-0.1 for angina pectoris patients, and 15.7+/-9.1 within 6h and 12.5+/-4.6 within 12h after the onset for AMI patients, respectively. The correlation coefficients between hepatocyte growth factor and creatine phosphokinase and between hepatocyte growth factor and CK-MB were 0.68 and 0.74, respectively. The time to reach the maximum (h) and the time to decline to less than half of the maximum level (days) were 6.6+/-2.6 and 1.2 +/-0.2 for hepatocyte growth factor, 19.4+/-8.7 and 2.5+/-1.4 for creatine phosphokinase, and 16.6+/-7.7 and 1.5+/-0.4 for CK-MB, respectively. Hepatocyte growth factor is useful as a prognostic indicator and reflects the clinical course in patients with acute myocardial infarction.

Apoptosis of neutrophils and their elimination by Kupffer cells in rat liver.

The fate of neutrophils in the peripheral circulation is poorly understood. In this study, the role of Kupffer cells in eliminating aged and apoptotic neutrophils was investigated. Liver, spleen, lung, and blood samples from Wistar rats were examined by light and electron microscopy, the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) method, and immunohistochemistry after the intravenous injection of OK-432, a streptococcal preparation. Neutrophils were trapped predominantly in the periportal and midzonal regions of hepatic lobules and were in contact with endothelial cells and Kupffer cells, or were surrounded by Kupffer cells. The trapping of neutrophils peaked after 6 hours. Apoptotic neutrophils, with or without buds, were found in the lumen of hepatic sinusoids as early as 6 hours, reached maximal levels after 12 hours, and represented greater than 60% of the total number of neutrophils in the liver. The presence of apoptotic neutrophils was correlated with the degree of neutrophil phagocytosis. Double-staining showed that TUNEL-positive neutrophils were phagocytosed or encircled by ED1- or ED2-positive Kupffer cells. In contrast, apoptosis and phagocytosis of neutrophils were rare in the spleen, lung, and peripheral blood. These results suggested that the appearance of apoptotic neutrophils in the hepatic sinusoids and their rapid clearance by Kupffer cells occurs after the invasion of bacteria (i.e., bacteremia or bacteriotoxemia) or the release of inflammatory mediators into the blood stream. These findings have important implications for the regulation of neutrophil homeostasis, the limitation of inflammation and tissue injury, and provide insight into the physiological removal of circulating, senescent neutrophils.

Polymorphic ventricular tachycardia induced by intracoronary injection of ioxaglate in a patient with borderline QT prolongation.

A 68-year-old female patient with borderline QT prolongation developed polymorphic ventricular tachycardia soon after intracoronary injection of ioxaglate. There was no coronary stenosis or vasospasm. To investigate abnormality of the ventricular repolarization, we recorded the monophasic action potential (MAP) during coronary angiography. Ioxaglate prolonged the MAP duration regionally in the perfused area and produced temporal dispersion of ventricular repolarization.

Expression of class II MHC molecules in the rat pineal gland during development and effects of treatment with carbon tetrachloride.

Cells expressing major histocompatibility complex (MHC) class II (Ia) antigen have been examined during the development of rat pineals and in the pineal gland of adult rats treated with carbon tetrachloride. Cells positive for MHC class II are first detected in the pineal gland of the 7-day-old rat. These positive cells increase in number gradually during development, MHC class II immunoreactivity reaching adult levels at 4 weeks after birth. The MHC class II antigen is intensely labeled on the cell surface, and labeled cells are distributed throughout the organ, several positive cells being gathered into groups. The positive cells are small (7-12 microm in diameter), irregular in shape, and frequently exhibit one or more processes. At the electron-microscopic level, the cytoplasm of positive cells contains few organelles, variously sized empty vacuoles, and a few electron-dense lysosome-like structures. Pinealocytes with synaptic ribbons have been found adjacent to immunoreactive cells. Double-immunoperoxidase staining for MRC OX6, MRC OX42, and ED1 results in OX6(-)/ED1(+)/OX42(+), OX6(-)/ED1(-)/OX42(+), and OX6(+)/ED1(-)/OX42(- )cells. These findings suggest that OX6-positive cells in the pineal can be considered as peripheral dendritic cells. The number of cells expressing MHC class II (Ia) antigen significantly increases in the pineal gland of rats after treatment with carbon tetrachloride (P<0.005). Our results indicate that at least some of the OX6-positive cells migrate into the gland from the circulation under these conditions.

EVI1 expression associated with a 3q26 anomaly in a leukemia cell line derived from the blast crisis of chronic myeloid leukemia.

Two leukemia cell lines, TS9;22 and YS9;22, were established from different individuals with Philadelphia chromosome (Ph)-positive chronic myeloid leukemia in blast crisis. The reverse transcript-polymerase chain reaction (RT-PCR) technique revealed that both cell lines expressed GATA-1, GATA-2, and the stem cell leukemia (SCL) gene, consistent with a megakaryocyte lineage. Chromosome analysis revealed that TS9;22 cells show the Ph translocation without abnormality of chromosome 3. In contrast, YS9;22 cells show the Ph translocation and dic(3)(q26;p12). Northern analysis revealed that YS9;22 cells express the EVI1 (ecotropic virus integration-1) gene, possibly because of the chromosomal translocation in the 3q26 region; TS9;22 cells do not express EVI1. However, no rearrangements were detected over 600 kb upstream or over 900 kb downstream of EVI1 in the YS9;22 cell line, suggesting a different mechanism of EVI1 activation from that in leukemia cells with either a t(3;3)(q21;q26) or inv(3)(q21q26). These results indicate that EVI1 expression in YS9;22 cells is linked to the 3q26 abnormality and that EVI1 activation plays an oncogenic role in the blastic transformation of chronic myeloid leukemia.

Expression of neuron-specific enolase in the developing rat retina as revealed by immunocytochemistry.

Expression of neuron-specific enolase (NSE) in retinal neurons was immunocytochemically investigated during the development of the rat retina. At embryonic day 14 (E14), the first immunoreaction of NSE was identified in the pigment epithelium. NSE-positive ganglion cells occurred at the inner surface of the retina by E15. Horizontal cells and photoreceptor cells became stainable for NSE in the outer portion of the neuroblastic layer as early as E17. At E20, when the majority of ganglion cells were intensely positive for NSE, immunoreactive amacrine cells first appeared at the outer surface of the developing inner plexiform layer. It was not until postnatal day 7 (P7) that NSE-positive bipolar cells occurred in the middle of the inner nuclear layer. At this stage, most of the photoreceptor cells located in the outer nuclear layer were immunolabeled, whereas the ectopic photoreceptor cells in the inner nuclear layer were devoid of immunoreaction. Most identifiable retinal neurons became strongly immunostained for NSE by P14. Our results indicate that the NSE expression of retinal neurons occurs just after their migration to the final location and prior to establishing the synaptic structures. In this paper, the characteristic sequence in which different types of retinal neurons exhibit NSE immunoreaction is discussed in the light of certain autoradiographic data on the sequence of retinal cell genesis.

Analysis of the heterogeneity within bovine pineal gland by immunohistochemistry and in situ hybridization.

In the present study, we demonstrate a cortical and medullary arrangement of parenchymal cells in the bovine pineal gland by using antibodies for neuron-specific enolase, synaptophysin, and hydroxyindole O-methyltransferase (HIOMT) as markers of pinealocytes, and glial fibrillary acidic protein (GFAP) as a marker of interstitial (glial) cells. Furthermore, by means of probes specific for HIOMT mRNA, we have examined possible differences in melatonin synthesis between the cortex and the medulla. Immunoreactive pinealocytes for each antigen investigated are more densely distributed in the cortex than in the medulla. In the cortex, GFAP-positive interstitial cells have large intenselystained somata endowed with several long, thin cytoplasmic processes, whereas in the medulla, they display smaller, less intensely labeled perikarya from which numerous fine short processes emerge. Golgi staining has confirmed these morphological differences between the interstitial cells in the cortex and those in the medulla. An analysis using confocal laser microscopy together with in situ hybridization for HIOMT mRNA has shown that the expression of mRNA transcripts in the cortex is more intense than that in the medulla. The expression of the HIOMT gene in a cluster of cells in the medial habenular nucleus is lower than that in pinealocytes of the pineal organ proper.