Serological and genetic characterization of newly isolated Peaton virus in Japan. Brief report.

The viruses were isolated from the blood of sentinel cattle and Culicoides biting midges in the Kyushu district, southwestern Japan, in 1999 and identified by neutralization tests as Peaton (PEA) viruses. Before this study, PEA virus had been isolated in Australia only. The nucleotide identity of the nucleocapsid (N) protein encoded by the S segment ranged from 91.1 to 91.6% between the Australian and Japanese strains. A phylogenetic analysis of the N protein sequence revealed that the PEA virus strains are closely related to Aino (AIN) virus and suggested reassortment events for PEA and AIN viruses.

Activation of matrix metalloproteinase-2 is correlated with invasiveness in thymic epithelial tumors.

UNLABELLED: BACKGROUND AND OBJECTS: Matrix degradation, which is a critical event in the process of tumor invasion and metastasis, is considered to be caused by the action of proteolytic enzymes. METHODS: We examined the gelatinolytic activity of matrix metalloproteinase (MMP)-9, and the activity of active and inactive forms of MMP-2 in five thymi, five noninvasive thymomas, eight invasive thymomas, and five thymic carcinomas by quantitative gelatinolytic zymography. RESULTS: The gelatinolytic activity of active MMP-2 in five thymi was zero. The mean gelatinolytic activity of active MMP-2 was 0.020 +/- 0.015 in noninvasive thymoma, 0.084 +/- 0.098 in invasive thymoma and 0.246 +/- 0.194 in thymic carcinoma. The gelatinolytic activity of active MMP-2 correlated with the invasiveness of thymic epithelial tumors (Spearman rank correlation: r-value = 0.532). The gelatinolytic activity of active MMP-2 in three thymoma cases with microscopic capsular invasion was the same as that of noninvasive thymoma. When thymoma cases showing microscopic capsular invasion were classified into the "macroscopically noninvasive thymoma" group, the gelatinolytic activity of active MMP-2 correlated with the invasiveness of thymic epithelial tumors (Spearman rank correlation: r-value = 0.621). CONCLUSIONS: The gelatinolytic activity of active MMP-2 significantly correlated with the invasiveness in thymic epithelial tumors. J. Surg. Oncol. 2001;76:169-175.

Pocket oxygen tension in smokers and non-smokers with periodontal disease.

BACKGROUND: Smoking is associated with destructive periodontal disease. Pocket oxygen tension (pO2) is likely to be a major environmental determinant of the subgingival microflora, which is a primary etiological factor of the disease. This study aimed to compare the pocket pO2 in smokers and non-smokers with periodontal disease. METHODS: Pocket oxygen tension was compared in 27 smokers and 34 non-smokers by considering 2 confounding factors, probing depth and oxygen saturation of hemoglobin (S(O2)), in the gingiva. The pO2 was determined using oxygen microelectrode by polarographic method with an electronic compensation circuit for subgingival temperature. Gingival S(O2) was determined using tissue reflectance spectrophotometry. RESULTS: No significant difference was found in the modified gingival index and the plaque index between smokers and non-smokers. The pO2 was significantly lower in smokers (21.9+/-9.6 mmHg) than in non-smokers (33.4+/-8.4 mmHg). The difference was highly significant (P <0.0001) and was consistent when the confounding factors were considered. Correlation between the PO2 and probing depth approached statistical significance in smokers (r = -0.36, P = 0.0674) and significance in non-smokers (r = -0.41, P = 0.0174). Correlation of the PO2 to the gingival S(O2) was highly significant in non-smokers (r = 0.57, P = 0.0005), but no association was found in smokers (r = -0.08, P= 0.6975). CONCLUSIONS: These findings indicate that pO2 is lower in smokers than in non-smokers, and that the pO2 in smokers is not influenced by gingival oxygen sufficiency. The present study may provide the basis of understanding environmental factors possibly associated with microbial flora in the pockets of smokers.

Relationship of the substance P to indicators of host response in human gingival crevicular fluid.

BACKGROUND: The substance P (SP) level in human gingival crevicular fluid (GCF) was studied in relation to clinical periodontal variables and to various indicators of host response in the GCF. METHODS: GCF was collected from periodontal sites with gingival inflammation and shallow or moderately deep pocket in 48 subjects. The total amount of SP and the substances based on host response factors in a 30-s sample were determined by ELISA and enzymatic methods. RESULTS: Significant correlation was found between SP and probing depth (r= 0.637, p<0.001), while correlation was weak between SP and either gingival (r= 0.177, p=0.23) or plaque index (r=0.008, p=0.96). SP also showed significant correlation with the indicators of host response: prostaglandin E2, aspartate aminotransferase, alkaline phosphatase, myeloperoxidase, interleukin-1beta, tumor necrosis factor-alpha, interleukin-8 and monocyte chemoattractant protein-1 (r=0.434-0.867, p<0.01-0.001). CONCLUSION: These results indicate that neuropeptide SP in GCF may have a potential as an indicator of periodontal inflammation and the host response.

SCID mouse lymphogenous metastatic model of human lung cancer constructed using orthotopic inoculation of cancer cells.

An animal model of metastasis with features similar to those of lung cancer metastasis in humans is required for an understanding of the cellular and molecular mechanisms of human lung cancer metastasis. Ma-44 cell lines derived from human squamous cell lung cancer were percutaneously injected (2-3 x 10(4)) into the left lung of SCID mice. After orthotopic implantation, Ma-44 cell lines formed tumors in the left lung at a high rate (17/25, 68%), and many of those metastasized to mediastinal lymph nodes (13/17, 76%) by the 14th day, but not to other organs. After the ectopic implantation, the Ma-44 cell line inoculated subcutaneously (2-3 x 10(5)) formed a tumor at the inoculation site by the 28th day (all mice), but did not metastasize to any organs. The Ma-44 cell line inoculated intravenously (2-3 x 10(5)) formed metastases in the lungs (37/50, 74%), and these pulmonary metastases metastasized to the mediastinal lymph nodes at low rate (3/37, 8%) by the 14th day. The orthotopic sites of implantation are critical for the metastatic ability of transplanted tumors in SCID mice. Since non-small cell lung cancer (NSCLC) grows at the primary site in humans, lymphogenous metastasis occurs frequently, and blood-born metastasis occurs at moderate rate, our orthotopic SCID mice model was similar to the metastatic behavior of NSCLC in humans. Thus, this model may be useful for elucidating the mechanism of lymphogenous metastasis in human lung cancer, and testing the anti-metastatic efficacy of therapeutic agents in vivo.

Oxygen sufficiency in the gingiva of smokers and non-smokers with periodontal disease.

BACKGROUND: Epidemiological studies have demonstrated deteriorating effects of smoking on periodontal tissue. The aims of this study were to compare oxygen saturation of hemoglobin in the gingiva (GSo2) of smokers and non-smokers and to evaluate the chronic effect of smoking on gingival oxygen sufficiency. METHODS: GSo2 was determined using tissue reflectance spectrophotometry in 110 papillary gingival sites of 62 smokers and 100 sites of 60 non-smokers. RESULTS: No significant difference was found in GSo2 between smokers and non-smokers. In the model of ANOVA with covariates, age (P= 0.0048) and probing depth (P= 0.0012) had significant effects on GSo2. No significant effect was found in either smoking status (P= 0.3557) or the modified gingival index (MGI) (P= 0.3824). The interaction effect between smoking status and the MGI was highly significant (P = 0.0003) indicating that the effect of smoking status on the GSo2 should be compared at each level of the MGI score. GSo2 in healthy gingiva was significantly lower in smokers than non-smokers (P = 0.0014), while smokers showed higher GSo2 than non-smokers in moderately inflamed gingiva (P = 0.0356). The GSo2 in inflamed gingiva was significantly decreased compared with healthy gingiva in non-smokers (P = 0.0044), while smokers showed no significant difference between healthy and inflamed gingiva (P= 0.2772 to 0.8665). GSo2 in smokers was consistently and significantly lower than that of healthy gingiva of non-smokers (P = 0.0391 to 0.0004). CONCLUSIONS: Smokers exhibit possibly lower function of oxygen sufficiency in healthy gingiva and reduced ability to adapt the function in inflamed gingiva than non-smokers. This suggests that smokers have functional impairments in the gingival microcirculation.

Autocrine loop through cholecystokinin-B/gastrin receptors involved in growth of human leukemia cells.

The cholecystokinin (CCK)-B/gastrin receptor binds two brain-gut hormones, CCK and gastrin, with high affinities. These peptides have a trophic effect on gastrointestinal cells expressing the receptor in vivo as well as in vitro. Recently, this receptor mRNA was reported to be expressed in immunocytes localized in the lamina propria of normal rat stomach mucosa. Here, we studied the receptor expression in human hematopoietic cells in order to determine whether they play a role in cell growth. The CCK-B/gastrin receptor mRNA was detectable in the polymorphonuclear (PMN) cells but not in the mononuclear cells of normal peripheral white blood cells by reverse transcription-polymerase chain reaction. The receptor transcript was, however, expressed in human leukemia cell lines (14 of 18 cell lines tested) derived from not only myeloid, but also T- and B- lymphoid lineages. The CCK-B/gastrin receptors on several leukemia cell lines were shown to be biologically active by demonstrating ligand-dependent cell proliferation in serum-deprived medium. Interestingly, a human CCK-B/gastrin receptor specific antagonist, YM022, but not its stereotype isoform, selectively inhibited the DNA synthesis of THP-1, MOLT-16, MOLT-14, and CCRF-CEM in the absence of exogenous peptide ligands. Further investigation revealed that these leukemia cell lines and normal PMN cells also expressed gastrin mRNA. These results suggest that growth of human leukemia cells is promoted by an autocrine mechanism through the CCK-B/gastrin receptors.

Antiproliferative effect of a novel cholecystokinin-B/gastrin receptor antagonist, YM022.

Cholecystokinin (CCK)-B and gastrin receptors are expressed on a variety of human tumor cells. Recently, we have demonstrated that the human brain CCK-B receptors are identical to the gastrin receptors derived from the stomach mucosa, and that the brain-gut peptides, CCK-8 and gastrin I are mitogenic for mouse NIH 3T3 fibroblasts expressing human CCK-B/gastrin receptors (N-hCCKBR). In this report, we evaluated the antiproliferative potency of CCK-B/gastrin receptor antagonists by using N-hCCKBR cells. Among several antagonists, a benzodiazepine derivative, YM022 had the most potent activities in competing with [125I]CCK-8 or [125I]gastrin I binding, inhibition of CCK-8- or gastrin I-induced phosphoinositide hydrolysis and increasing cytoplasmic free calcium. Interestingly, a potent antagonist for rat CCK-B/gastrin receptors did not have such activities in N-hCCKBR cells. YM022 inhibited the CCK-8- or gastrin I-induced [methyl-3H]thymidine incorporation of N-hCCKBR cells in a dose-dependent manner. In the absence of exogenous peptide ligands, YM022 also inhibited the proliferation of several human cancer cell lines expressing the genes for both gastrin and its receptor. These results suggest that YM022 could intervene in the autocrine stimulation of human tumor cell lines through CCK-B/gastrin receptors. N-hCCKBR cells are an excellent tool to screen for novel human CCK-B/gastrin receptor antagonists possessing antiproliferative activity for human cancer cells.

[Cholecystokinin-B/gastrin receptor is a novel molecular probe for human small cell lung cancer].

Cloning of CCK-B/gastrin receptor cDNAs showed that they possess the feature of serpentine G protein coupled receptors. In this study, the receptor mRNA was detected selectively in all SCLCs (10 cases) with a RT-PCR assay. By contrast, it was detectable in only one of thirteen squamous cell carcinomas or twenty-one adenocarcinomas of lung. Thus, CCK-B/gastrin receptor has been demonstrated to provide a novel molecular marker for the diagnosis of small cell lung cancer (SCLC) by using biopsy specimens.

Cholecystokinin-B/gastrin receptor: a novel molecular probe for human small cell lung cancer.

The brain-gut hormones, gastrin and cholecystokinin, have a trophic effect on the gastrointestinal mucosa in vivo and promote the growth of several neoplastic cell lines. In this study, cholecystokinin-B/gastrin receptor has been demonstrated to provide a novel molecular marker for the diagnosis of small cell lung cancer by using biopsy specimens. Physiological expression of the receptor mRNA is detectable in particular areas of the human brain, stomach, and pancreas but not in the lung. The receptor mRNA was detected selectively in all small cell lung cancer (10 cases) with a RT-PCR assay. By contrast, it was detectable in only 1 of 13 squamous cell carcinomas or 21 adenocarcinomas of the lung. Thus, the cholecystokinin-B/gastrin receptor could be an attractive therapeutic target for small cell lung cancer.

A new type of staphylococcal exfoliative toxin from a Staphylococcus aureus strain isolated from a horse with phlegmon.

A new type of staphylococcal exfoliative toxin (sET) was isolated from the culture filtrate of a Staphylococcus aureus strain isolated from a horse with skin infection including phlegmon. The new sET was purified by precipitation with 80% saturated ammonium sulfate, column chromatography on DEAE-cellulofine A-500, gel filtration on a Sephadex G-75 column, and polyacrylamide gel electrophoresis (7.5% polyacrylamide). The new sET elicited general exfoliation of the epidermis with the so-called Nikolsky sign when inoculated into both 3-day-old mice and 1-day-old chicks, whereas sETA and sETB from human strains of S. aureus caused exfoliation in a 3-day-old mouse alone and shET from a porcine strain of Staphylococcus hyicus caused exfoliation in 1-day-old chicks alone. Intraepidermal splitting was observed at the granular layer of the epidermis of mice inoculated with the new sET as well as those inoculated with sETA. Exfoliation at the germinative layer of the epidermis was also observed in the chicks inoculated with the new sET as well as those inoculated with shET. The new sET was serologically different from sETA, sETB, and shET and showed the same molecular weight on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was thermolabile and lost its toxicity after being heated at 60 degrees C for 15 min. We propose that the new sET be designated as sETC.

Intracellular distribution of cardiac beta-adrenoceptors in SHR and WKY.

To compare the intracellular distribution of beta-adrenoceptors in isolated myocytes of SHR with that of age-matched WKY and to examine changes in the distribution with aging, we measured the number of cell surface and total beta-adrenoceptors in 6-(young groups) and 37-week old (old groups) SHR and WKY. The number of surface beta-adrenoceptors was significantly lower in 6-week old SHR compared with that in age-matched WKY. But there was no difference in the number of total beta-adrenoceptors between the two groups. The number of surface beta-adrenoceptors in old groups was significantly reduced compared with that in the young groups, in both SHR and WKY. However, the number of total beta-adrenoceptors in the old groups did not show any difference from that in the young groups. Isoproterenol-stimulated c-AMP formation in 6-week old SHR was significantly lower than that of WKY of the same age. Isoproterenol-stimulated c-AMP formation in the old groups was significantly reduced than that of the young groups in both SHR and WKY. These results suggested that (1) there may be a difference in the intracellular distribution of cardiac beta-adrenoceptors between 6-week old SHR and WKY, (2) the intracellular distribution of cardiac beta-adrenoceptors may be changed with aging, and (3) intracellular distribution might be related to the difference in c-AMP formation to beta-agonist.

Beta-adrenergic receptors in ischemic and nonischemic canine myocardium: relation to ventricular fibrillation and effects of pretreatment with propranolol and hexamethonium.

To explore possible roles of the sympathetic nervous system (especially the beta-adrenergic receptor and cAMP system) in the pathogenesis of ventricular fibrillation (VF), the authors examined changes in the number of myocardial beta-adrenergic receptors and the cAMP level in animals showing VF after experimental induction of myocardial ischemia. In animals that developed VF approximately 15 min after coronary ligation, the ischemic myocardium had a significantly larger number of beta-adrenergic receptors (90 +/- 8 fmol/mg protein) and a significantly higher level of cAMP (1.4 +/- 0.17 nmol/g weight) compared with the nonischemic area where the number of beta-receptors was 68 +/- 7 fmol/mg protein and the cAMP level was 0.80 +/- 0.20 nmol/g weight. In animals in which VF was electrically induced 20 min after coronary ligation, no elevation was recorded in the ischemic area in terms of the number of beta-adrenergic receptors (49 +/- 6 fmol/mg protein) and the level of cAMP (0.79 +/- 0.06 nmol/g weight). When animals were pretreated with a beta-blocker or a ganglion blocker, coronary ligation did not result in VF. In the ischemic area of these animals, no significant change was noted in the number of beta-adrenergic receptors (54 +/- 7 fmol/mg protein for animals pretreated with a beta-blocker, 56 +/- 3 fmol/mg protein for animals pretreated with a ganglion blocker). These results suggest that there is a strong correlation between the pathogenesis of VF (observed about 15 min after coronary ligation) and the increase of the number of beta-adrenergic receptors and cAMP levels.

[Blood flow patterns in the left ventricle in patients with myocardial infarction and ventricular aneurysm: evaluation using real-time two-dimensional Doppler echocardiography].

To evaluate how the intraventricular blood flow is affected by the size of a left ventricular aneurysm and ventricular dysfunction, systolic left ventricular blood flow patterns were evaluated using two-dimensional Doppler flow images (real-time 2-D Doppler echo). The subjects consisted of 10 normal controls, 35 patients with anteroseptal infarction, two patients with inferior infarction and five patients with anteroseptal-inferior infarctions. The systolic period was divided into three subsets; early, mid- and end-systole. Forty-two patients with myocardial infarction were classified into three groups according to the left ventricular inflow patterns on real-time 2-D Doppler echo using the apical left ventricular long-axis approach; i.e., inflow signals confined to early systole (Group I), visualized up to mid-systole (Group II) and end-systole (Group III). Left ventricular end-diastolic dimension (LVDd), left ventricular end-systolic dimension (LVDs), and % non-contractile circumference (delta L) were calculated by the same echocardiographic approach. Ejection fraction (EF) was calculated by left ventricular cineangiography using the Simpson's method. The left ventricular inflow Doppler signals in the normal controls and Group I turned in the apex and then directed toward the left ventricular outflow tract during late diastole and early systole. Significant differences in EF were observed among the three groups. EF in Group I, II and III was 53 +/- 9%, 41 +/- 8% and 29 +/- 7%, respectively. However, LVDd, LVDs and delta L had the largest values in Group III and the smallest values in Group I. LVDd, LVDs and delta L were smallest in Group I and largest in Group III. In the normal controls, the left ventricular inflow signals proceeded to the apex and directed toward the left ventricular outflow tract in the early systolic period. Various changes in the inflow pattern were observed in patients with myocardial infarction and severe wall motion abnormalities, including delayed timing in proceeding from the apex to the left ventricular outflow tract, stagnant blood at the apex and further inflow of blood toward the apex even during end-systole. The patients with sustained inflow during late systole had hypofunction of the left ventricle as demonstrated by smaller EF and larger LVDd, LVDs, and delta L. In conclusion, the observation of intracardiac blood flows by real-time 2-D Doppler echo is of help in evaluating the severity of myocardial infarction.

[Comparison of two-dimensional echocardiography with 201T1-single photon emission computed tomography for evaluating the myocardial infarct zone].

The reliability of measuring the myocardial infarct zone by two-dimensional echocardiography (2 DE) was compared with that by regional myocardial blood flow as evaluated by single photon emission computed tomography (SPECT) in 47 patients with old myocardial infarction, with ventricular aneurysm (An group; n = 15), and without ventricular aneurysm (Non-An group; n = 32). Short-axis images of the left ventricle at the level of the mitral valve, the papillary muscles, and the apex were obtained both by 2DE and SPECT. The left ventricular wall was divided into 36 segments in 2DE and 40 segments in SPECT with reference points at the posterior end of the right side of the interventricular septum. The segments in which the radial shortening on 2DE and the 201Tl uptake index on SPECT were below one standard deviation from the means of 10 normal subjects were defined as those with abnormal wall motion and hypoperfused areas, respectively. The relationships between these findings were studied. The extent of apical movement was measured by left ventricular cineangiography in each case, and was compared between the An and Non-An groups. 1. Wall motion abnormalities on 2DE and hypoperfusion on SPECT showed a correspondence of 81% in the An group and 78% in the Non-An group at the level of the mitral valve, and 78% in the An group and 76% in the Non-An group at the papillary muscle level. However, a better correspondence was observed in the An group (84%) as compared to the Non-An group (64%) at the level of the apex. 2. Apical movement assessed by cineangiography showed more extensive changes in the Non-An group than in the An group (2.3 +/- 0.9 mm vs 4.1 +/- 1.7 mm in the RAO view, 2.4 +/- 1.9 mm vs 5.3 +/- 2.1 mm in the LAO view). From these observations, it was suspected that the cause of disparity between radial shortening and the 201Tl uptake index at the level of the apex is related to the cardiac movement of the apex toward the base during systole. 3. Since wall motion abnormalities demonstrated by radial shortening (2DE) and hypoperfusion indicated by the 201Tl uptake index (SPECT) generally corresponded well, 2DE was thought to be a useful method for evaluating myocardial infarct zone.

[Two-dimensional echocardiography in ventricular septal rupture after acute myocardial infarction].

We studied the echocardiographic findings of 11 patients with proven ventricular septal defect following acute myocardial infarction. There were seven men and four women whose ages ranged from 48 to 77 years, with an average of 66 years. Nine patients had acute anterior and two acute inferior myocardial infarctions. Two-dimensional echocardiography (2DE) was performed for eight patients and M-mode echocardiography for all 11 patients. In all eight patients with apical four-chamber view, in whom four had additional apical short-axis view, the septal defect was directly visualized, but it was not detected by M-mode echocardiography. The defect was visualized in the apical region of the septum in all eight patients by the apical four-chamber view. The anteroapical region of the septum was the site in three with anterior infarction and the inferoapical region in one with inferior infarction by the apical short-axis view. In five of the eight patients who underwent 2DE, surgical or autopsy confirmation of the defects was obtained, with a complete agreement with the echocardiographic findings. In two patients with echocardiographic findings of septal defects, the perforations were confirmed at surgery. Two cases with aneurysmal bulges of thin septum into the right ventricle had the thin necrotic muscle in the anteroapical regions. One patient with a cystic bulge into the septum showed an irregular tear in the inferoapical region of the septum at surgery. In eight patients, the left ventricular wall motion was assessed by 2DE. Six patients revealed hyperkinetic motion in the non-infarcted areas of the basal septum or posterior wall, and these cases had good prognosis. We concluded that 2DE is a sensitive, prompt and safe technique for diagnosing and observing the risk of complicating septal defects in acute myocardial infarction. In this respect, both the apical four-chamber and short-axis views should be utilized for the topographic diagnosis of the defect.