Expression and analysis of a cytolethal distending toxin (cdt) gene operon in Campylobacter lari.

The present study examines the expression of cytolethal distending toxin (cdt) gene encoding a cytotoxin in Campylobacter lari (n=6 urease-negative [UN] C. lari; n=4 urease-positive thermophilic Campylobacter [UPTC]). When reverse transcription polymerase chain reaction (RT-PCR) was carried out with 10 C. lari isolates using a primer pair to amplify the cdtB gene transcript segment, an approximate 260 bp RT-PCR amplicon was generated with all the isolates. In addition, cdtA, cdtB and cdtC gene operon was identified to be polycistronicly transcribed in the C. lari cells. The cdtB gene translation in the C. lari cells was also confirmed by Western blot analysis. Thus, the cdt gene operon in C. lari organisms, including UN C. lari and UPTC, was expressed at the transcriptional and translational levels in the cells. The present results suggest that all three cdt genes may be functional in the cells.

Comparative analysis of Campylobacter lari cytolethal distending toxin (CDT) effect on HeLa cells.

We aimed to clarify if Campylobacter lari exerts a cytolethal distending toxin (CDT) effect on HeLa cells. Campylobacter cell lysates (CCLys) from C. jejuni 81-176 and urease-positive thermophilic Campylobacter (UPTC) CF89-12 and UPTC NCTC12893 isolates were shown to exert a CDT effect on HeLa cells with morphological changes examined by Giemsa staining and microscopy. However, Campylobacter lari JCM2530(T) isolate showed no effect. In addition, Campylobacter cell culture supernatant wash gave low or absent toxic effects with both C. jejuni and C. lari organisms. When western blot analysis was carried out to clarify if there was a CDTB effect in the CCLys and soluble fractions from Campylobacter isolates, which had a CDT effect on HeLa cells or did not have any effect, anti-recombinant CjCDTB antibodies identified an immunoreactively positive signal at around approximately 25 kDa on all the C. lari isolates examined, as well as the C. jejuni 81116 strain. Thus, all the Campylobacter isolates including those without any CDT effect were shown to express CDTB at the translational level.

Common occurrence of everolimus-associated aphthous stomatitis in Japanese heart transplant recipients.

Mammalian target of rapamycin (mTOR) inhibitors display antiproliferative effects with less nephrotoxicity than calcineurin inhibitors. However, clinical use of mTOR inhibitors can be associated with a series of adverse events. We experienced cases of aphthous stomatitis associated with everolimus (EVL) in four Japanese heart transplant recipients treated at the target trough EVL blood level after a switch from mycophenolate mofetil between April and December 2007. All four patients developed aphthous stomatitis; three required reduction of the exposure and one, EVL discontinuation due to stomatitis as well as other side effects. All patients recovered from stomatitis after reduction or withdrawal of EVL. Thus, we considered that EVL-related stomatitis might occur commonly among the Japanese population. The proper dosage, effects, and frequency of the side effects of mTOR inhibitors may vary by ethnic population.

Cholera toxin production by the El Tor variant of Vibrio cholerae O1 compared to prototype El Tor and classical biotypes.

Vibrio cholerae O1 El Tor variant strains produced much more cholera toxin than did prototype El Tor strains. The amount of cholera toxin produced by El Tor variant strains both in vitro and in vivo was more or less equivalent to that produced by classical strains.

A highly sensitive and specific multiplex PCR assay for simultaneous detection of Vibrio cholerae, Vibrio parahaemolyticus and Vibrio vulnificus.

AIMS: To develop an effective multiplex PCR for simultaneous and rapid detection of Vibrio cholerae, Vibrio vulnificus and Vibrio parahaemolyticus, the three most important Vibrio species that can cause devastating health hazards among human. METHODS AND RESULTS: Species-specific PCR primers were designed based on toxR gene for V. cholerae and V. parahaemolyticus, and vvhA gene for V. vulnificus. The multiplex PCR was validated with 488 Vibrio strains including 322 V. cholerae, 12 V. vulnificus, and 82 V. parahaemolyticus, 20 other Vibrio species and 17 other bacterial species associated with human diseases. It could detect the three target bacteria without any ambiguity even among closely related species. It showed good efficiency in detection of co-existing target species in the same sample. The detection limit of all the target species was ten cells per PCR tube. CONCLUSIONS: Specificity and sensitivity of the multiplex PCR is 100% each and sufficient for simultaneous detection of these potentially pathogenic Vibrio species in clinical and environmental samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This simple, rapid and cost-effective method can be applicable in a prediction system to prevent disease outbreak by these Vibrio species and can be considered as an effective tool for both epidemiologist and ecologist.

Identification of Vibrio harveyi as a causative bacterium for a tail rot disease of sea bream Sparus aurata from research hatchery in Malta.

A bacterial disease was reported from gilthead sea bream (Sparus aurata) within a hatchery environment in Malta. Symptoms included complete erosion of tail, infection in the eye, mucous secretion and frequent mortality. A total of 540 strains were initially isolated in marine agar from different infected body parts and culture water sources. Subsequently 100 isolates were randomly selected, identified biochemically and all were found to be Vibrio harveyi-related organisms; finally from 100 isolates a total of 13 numbers were randomly selected and accurately identified as V. harveyi by 16S rRNA gene sequencing and species-specific PCR. Ribotyping of these strains with HindIII revealed total of six clusters. In vivo challenge study with representative isolates from each cluster proved two clusters each were highly pathogenic, moderately pathogenic and non-pathogenic. All 13 isolates were positive for hemolysin gene, a potential virulence factor. Further analysis revealed probably a single copy of this gene was encoded in all isolates, although not in the same locus in the genome. Although V. harveyi was reported to be an important pathogen for many aquatic organisms, to our knowledge this might be the first report of disease caused by V. harveyi and their systematic study in the sea bream hatchery from Malta.

Development of simple and rapid PCR-fingerprinting methods for Vibrio cholerae on the basis of genetic diversity of the superintegron.

AIMS: To develop simple and rapid PCR-fingerprinting methods for Vibrio cholerae O1 (El Tor and classical biotypes) and O139 serogroup strains which cause major cholera epidemics, on the basis of the diversity of superintegron (SI) carried by these strains. METHODS AND RESULTS: PCR-restriction fragment length polymorphism (PCR-RFLP) assay was developed targeting region between integrase gene in the SI and its nearby ORF, followed by BglI digestion. Besides, a V. cholerae repeat-amplified fragment length polymorphism (VCR-AFLP) assay was also developed. In the PCR-RFLP, 94 El Tor, 29 classical and 54 O139 strains produced nine, three and six different DNA fingerprints, respectively. On the other hand, VCR-AFLP distinguished these El Tor, classical and O139 strains into five, nine and two DNA fingerprints, respectively. Combining both assays the El Tor, classical and O139 strains could be differentiated into 11, 10 and seven different types, respectively. In a comparative study, pulsed-field gel electrophoresis (PFGE) showed similar differentiation for El Tor (11 types), but lower discrimination for O139 (two types) and classical strains (five types). CONCLUSIONS: The PCR assays based on SI diversity can be used as a useful typing tool for epidemiological studies of V. cholerae. SIGNIFICANCE AND IMPACT OF STUDY: This newly developed method is more discriminatory, simple, rapid and cost-effective in comparison with PFGE, and thus can be widely applicable.

Development of a haemolysin gene-based multiplex PCR for simultaneous detection of Vibrio campbellii, Vibrio harveyi and Vibrio parahaemolyticus.

AIM: To develop a haemolysin (hly) gene-based species-specific multiplex PCR for simple and rapid detection of Vibrio campbellii, V. harveyi and V. parahaemolyticus. METHODS AND RESULTS: The complete hly genes of three V. campbellii strains isolated from diseased shrimps were sequenced and species-specific PCR primers were designed based on these sequences and the registered hly gene sequences of Vibrio harveyi and Vibrio parahaemolyticus. Specificity and sensitivity of the multiplex PCR was validated with 27 V. campbellii, 16 V. harveyi, and 69 V. parahaemolyticus, 18 other Vibrio species, one Photobacterium damselae and nine other bacterial species. The detection limits of all the three target species were in between 10 and 100 cells per PCR tube. CONCLUSIONS: Specificity and sensitivity of the multiplex PCR is 100% each and sufficient to be considered as an effective tool in a prediction system to prevent potential disease outbreak by these Vibrio species. SIGNIFICANCE AND IMPACT OF THE STUDY: Because there is lack of simple, rapid and cost-effective method to differentiate these closely related V. campbellii, V. harveyi and V. parahaemolyticus species, the multiplex PCR developed in this study will be very effective in epidemiological, ecological and economical points of view.

Genotoxicity studies on licorice flavonoid oil (LFO).

Licorice flavonoid oil (LFO) is a new functional food ingredient. In this study, the genotoxicity of LFO was investigated using a test battery of three different methods. In a reverse mutation assay using four Salmonella typhimurium strains and Escherichia coli, LFO did not increase the number of revertant colonies in any tester strain with or without metabolic activation by rat liver S9 mix. In a chromosomal aberration test using Chinese hamster lung (CHL/IU) cells, LFO did not induce any chromosomal aberrations either in the short period test without rat liver S9 mix or in the continuous treatment (24 h or 48 h) test. However, in the short-period test with rat liver S9 mix, LFO induced structural chromosomal aberrations at concentrations higher than 0.6 mg/mL. A bone marrow micronucleus test using male F344 rats was initially conducted. The animals were dosed by oral gavage at doses up to 5000 mg/kg/day. No significant or dose-dependent increases in the frequency of micronucleated polychromatic erythrocytes (MNPCE) were observed and the high dose suppressed the ratio of polychromatic erythrocytes (PCE) to total erythrocytes. Subsequently, a liver and peripheral blood micronucleus test using male F344 rats was conducted. No micronuclei induction either in hepatocytes or PCE was observed even at the highest dose of 5000 mg/kg/day. From the findings obtained from the genotoxicity assays performed in this study and the published pharmacokinetic studies of LFO, it appears unlikely that dietary consumption of LFO will present any genotoxic hazard to humans.

Serial changes in plasma brain natriuretic peptide concentration at the infarct and non-infarct sites in patients with left ventricular remodelling after myocardial infarction.

OBJECTIVES: To clarify the role of infarct and non-infarct sites on left ventricular (LV) remodelling after myocardial infarction by measuring brain natriuretic peptide (BNP) from each site. METHODS AND RESULTS: BNP from the aorta and the anterior interventricular vein (AIV) was measured in 45 patients with first anterior myocardial infarction at one, six, and 18 months. The LV was significantly dilated (> 10 ml/m(2) of end diastolic volume from one to 18 months) in 20 patients (remodelling (R) group) but not in 25 others (non-remodelling (NR) group). Patient characteristics and LV functions did not differ significantly at one month but plasma BNP concentration was higher in group R than in group NR (336 (288) v 116 (106) pg/ml, p < 0.01), predicting the degree of LV dilatation. The difference in BNP concentration between the aortic root and AIV (DeltaBNP), reflecting BNP secreted from the infarct site, did not differ at one month. In both groups BNP and DeltaBNP significantly decreased from one to six months (p < 0.05) and decreased from six months to 18 months, but the change was not significant. BNP and DeltaBNP were significantly higher in group R than in group NR after six months, when LV dilatation was not evident in both groups. CONCLUSION: Enhanced BNP secretion at one month in the non-infarct and infarct ventricular sites predicts subsequent LV dilatation (that is, remodelling). The slower process of LV remodelling decreased BNP secretion at both sites. Thus, BNP concentration should be useful for monitoring ventricular remodelling after infarction.

The healing process of infarct-related plaques. Insights from 18 months of serial angioscopic follow-up.

OBJECTIVES: To clarify the healing process of disrupted culprit plaques of acute myocardial infarction (MI), we serially observed the culprit plaques for 18 months after the onset of acute MI by angioscopy. BACKGROUND: Although it has been reported that disruption of the yellow plaque and subsequent thrombosis cause acute MI and that the thrombogenicity of the plaque lasts for a month, the healing process of the plaque after disruption has not been clarified. METHODS: Eighty-five patients with acute MI were prospectively and consecutively enrolled. Angioscopic studies were performed immediately and at 1, 6 and 18 months after successful reperfusion. The prevalence of yellow plaques and thrombus was examined. The color grade of the plaque was determined as 0 (white), 1 (light yellow), 2 (yellow) or 3 (bright yellow). RESULTS: Although yellow plaque was present at the culprit lesion in most patients throughout follow-up, its color grade was reduced from one to six months (1.9 +/- 0.6 vs. 1.1 +/- 0.7, p = 0.0003) after reperfusion, especially in the patients without hyperlipidemia (HL). The incidence of thrombus was 92.5% immediately after reperfusion, which was reduced significantly to 63.8%, 4.8% and 11.8% at 1, 6 and 18 months, respectively. The incidence of thrombus (77.8% vs. 45.0%, p = 0.03) at one month was higher in the patients with diabetes mellitus (DM). CONCLUSIONS: The healing process of yellow plaques at the culprit lesions of MI was detected by angioscopy as reductions of color grade and thrombogenicity at six months and partially at one month after the onset of acute MI. This healing process appears to deteriorate by complicating cases of DM or HL.

Benidipine, a long-acting Ca channel blocker, limits infarct size via bradykinin- and NO-dependent mechanisms in canine hearts.

Amlodipine increases NO levels in coronary vessels and aorta via bradykinin-dependent mechanisms in vitro. We have previously reported that a long-acting Ca channel blocker, benidipine, increases cardiac NO levels in ischemic canine hearts, suggesting that benidipine may also protect against ischemia and reperfusion injury via bradykinin- and NO-dependent mechanisms. We examined this possibility. In open chest dogs, the left anterior descending coronary artery was perfused with blood through a bypass tube and was occluded for 90 min followed by 6 hours of reperfusion. Infarct size was assessed by TTC staining at 6 hours of reperfusion. When benidipine doses of 50, 100, and 200 ng/kg/min were infused via the bypass tube between 10 min prior to the onset of ischemia and after 60 min of reperfusion, systemic blood pressure did not change significantly. Infarct size decreased with the administration of benidipine (50, 100, and 200 ng/kg/min) when compared to the untreated condition (24.8+/-2.5, 17.3+/-3.1, and 16.5+/-2.0 vs. 43.4+/-5.6%, respectively) associated with the increased release of NO and bradykinin in the coronary venous blood upon reperfusion. Myeloperoxidase activity of the myocardium increased after 6 hours of reperfusion, which was attenuated by benidipine. The limitation of infarct size and the increase in myeloperoxidase activity were completely blunted by either L-NAME or HOE140. There were no significant differences in collateral blood flow assessed by the microsphere method after 45 min of ischemia for any of the groups. Thus, we conclude that the Ca channel blocker, benidipine, limits infarct size via bradykinin- and NO-dependent mechanisms.

cDNA array hybridization reveals cardiac gene expression in acute ischemic murine hearts.

PURPOSE: Although cDNA array technique has recently become available in the cardiovascular field, it has not yet been established what kind of genes in the myocardium are expressed by acute ischemia. Since many substances contribute to the pathophysiology of acute ischemic hearts, we investigated transcription responses of murine hearts to ischemia using cDNA array representing 18,376 genes. METHODS AND RESULTS: In 29 male mice, we ligated the proximal site of the left coronary artery for 60 min. In 14 mice, we performed the sham operation without the ligation of the left coronary artery. After 60 min, the hearts were excised to obtain mRNA, and we performed cDNA array analysis. In 18,376 cDNA, 2 known genes were upregulated over 10-fold, 11 known genes were upregulated 5.0- to 9.9-fold, and 32 unknown genes were upregulated over 5.0-fold compared to sham-operated controls. In contrast, 11 known genes and 7 unknown genes were downregulated to levels below 0.2-fold. For 9 of the 13 known genes of which expression was increased as analyzed by cDNA array, subsequent Northern blot analysis also revealed an increase in expression. CONCLUSION: Using cDNA array analysis we found that cardiac expression of 24 known and 39 unknown genes was modulated by acute ischemic stress, and appeared to be related to the pathophysiology of ischemic hearts. These results show that cDNA array analysis may provide a new molecular insight to the pathophysiology of acute ischemic hearts.

Nifedipine limits infarct size via NO-dependent mechanisms in dogs.

OBJECTIVES: Amlodipine increases NO levels in coronary vessels and aorta via bradykinin-dependent mechanisms in vitro. We have previously reported that nifedipine increases cardiac NO levels in the ischemic canine hearts, suggesting that nifedipine may also have protective effects against ischemia and reperfusion injury, because the enhancement of NO production limits infarct size. We tested whether nifedipine limits infarct size via NO-dependent mechanisms. METHODS: In open chest dogs, the left anterior descending coronary artery was perfused with blood through a bypass tube and occluded for 90 min followed by 6 hours of reperfusion. Infarct size was assessed at 6 hours of reperfusion. Nifedipine of 3 or 6 microg/kg/min was infused into the bypass tube between 10 min prior to the onset of ischemia and 60 min of reperfusion. RESULTS: Neither systemic blood pressure nor heart rate changed during infusion of nifedipine. Infarct size was reduced by the administration of nifedipine (3 or 6 microg/kg/min) compared with the untreated condition (25.6+/-2.6 and 19.1+/-3.5 vs. 43.4+/-5.6%, respectively), which was completely blunted by L-NAME (45.0+/-3.6 and 45.4+/-4.2 vs. 47.9+/-3.9% in the nifedipine (3 or 6 microg/kg/min) with L-NAME groups vs. the L-NAME group). Myeloperoxidase activity of the myocardium increased after 6 hours of reperfusion, which was attenuated by nifedipine. The limitation of infarct size and the attenuation in myeloperoxidase actiivity were completely blunted by L-NAME. There were no significant differences in collateral blood flow at 45 min of ischemia between each group. CONCLUSIONS: We conclude that the Ca channel blocker, nifedipine, limits infarct size via NO-dependent mechanisms.

Differential subcellular actions of ACE inhibitors and AT(1) receptor antagonists on cardiac remodeling induced by chronic inhibition of NO synthesis in rats.

Chronic inhibition of NO synthesis induces cardiac hypertrophy independent of systemic blood pressure (SBP) by increasing protein synthesis in vivo. We examined whether ACE inhibitors (ACEIs) enalapril and temocapril and angiotensin II type-I receptor antagonists (angiotensin receptor blockers [ARBs]) losartan and CS-866 can block cardiac hypertrophy and whether changes in activation of 70-kDa S6 kinase (p70S6K) or extracellular signal-regulated protein kinase (ERK) are involved. The following 13 groups were studied: untreated Wistar-Kyoto rats and rats treated with NO synthase inhibitor Nomega-nitro-L-arginine methyl ester (L-NAME), D-NAME (the inactive isomer of L-NAME), L-NAME plus hydralazine, L-NAME plus enalapril (3 mg. kg(-1). d(-1)) or temocapril (1 or 10 mg. kg(-1). d(-1)), L-NAME plus losartan (10 mg. kg(-1). d(-1)) or CS-866 (1 or 10 mg. kg(-1). d(-1)), L-NAME plus temocapril-CS866 in combination (1 or 10 mg. kg(-1). d(-1)), and L-NAME plus rapamycin (0.5 mg. kg(-1). d(-1)). After 8 weeks of each experiment, ratios of coronary wall to lumen (wall/lumen) and left ventricular weight to body weight (LVW/BW) were quantified. L-NAME increased SBP, wall/lumen, and LVW/BW compared with that of control. ACEIs, ARBs, and hydralazine equally canceled the increase in SBP induced by L-NAME. However, ACEIs and ARBs equally (but not hydralazine) attenuated increase in wall/lumen and LVW/BW induced by L-NAME. The L-NAME group showed both p70S6K and ERK activation in myocardium (2.2-fold and 1.8-fold versus control, respectively). ACEIs inactivated p70S6K and ARBs inactivated ERK in myocardium, but hydralazine did not change activation of either kinase. Thus, ACEIs and ARBs modulate different intracellular signaling pathways, inhibiting p70S6K or ERK, respectively, to elicit equal reduction of cardiac hypertrophy induced by chronic inhibition of NO synthesis in vivo.

Role of cellular acidosis in production of nitric oxide in canine ischemic myocardium.

We tested the hypothesis that cellular acidosis modulates the production of nitric oxide (NO) in ischemic hearts. In canine hearts, we decreased coronary blood flow (CBF) to one third of the control by reduction of coronary perfusion pressure (105+/-3 to 41+/-5 mmHg), and thereafter we maintained CBF constant (89.8+/-1.6 to 30.0+/-0.5 ml/100 g/min) with an intracoronary administration of either saline, atropine, rauwolscine, HOE140, 8-sulfophenyltheophylline (8SPT), NaHCO3, or HOE642 (the inhibitor of Na+/H+ exchange). The cardiac NO levels defined as the differences of the nitrate and nitrite levels between coronary venous and arterial blood increased in the saline administration (2.9+/-0.2 to 12.7+/-1.7 micromol/l), and the extents of increases were identical in the condition of either saline, atropine, rauwolscine, HOE140 or 8SPT administration. In the condition with either NaHCO3 or HOE642, the increases in the cardiac NO levels were blunted (4.5+/-0.7 and 4.8+/-0.4 micromol/l, respectively). Cyclic GMP content of epicardial coronary artery in the ischemic area increased, which was also attenuated by either NaHCO3 or HOE642. We confirmed the acidosis-induced NO production in a more severe ischemic myocardium, and also showed that cellular acidosis produced by infusion of HCl increased NO production in non-ischemic myocardium. We conclude that cellular acidosis and subsequent activation of Na+/H+ exchanges modulate production of endogenous NO in canine ischemic myocardium.

Cardioprotective effect afforded by transient exposure to phosphodiesterase III inhibitors: the role of protein kinase A and p38 mitogen-activated protein kinase.

BACKGROUND: Phosphodiesterase III inhibitors (PDEIII-Is) improve the hemodynamic status of heart failure via inotropic/vasodilatory effects attributable to the increase in intracellular cAMP level. Direct cardioprotection by PDEIII-Is and its underlying mechanisms, however, have not been identified. We tested the infarct size-limiting effect of PDEIII-Is and the roles of cAMP, protein kinase (PK) A, PKC, and mitogen-activated protein kinase (MAPK) families in open-chest dogs. Methods and Results-- Milrinone, olprinone (PDEIII-Is), or dibutyryl-cAMP (db-cAMP) was injected intravenously 30 minutes before 90-minute ischemia, followed by 6 hours of reperfusion. Olprinone was also examined with an intracoronary cotreatment with a PKA inhibitor (H89), a PKC inhibitor (GF109203X), an extracellular signal-regulated kinase kinase (MEK) inhibitor (PD98059), or a p38 MAPK inhibitor (SB203580) throughout the preischemic period. Either PDEIII-Is or db-cAMP caused substantial hemodynamic changes, which returned to control levels in 30 minutes. Collateral flow and percent risk area were identical for all groups. Both PDEIII-Is and db-cAMP increased myocardial p38 MAPK activity during the preischemic period, which was blocked by H89, but not by GF109203X. Both PDEIII-Is and db-cAMP reduced infarct size (19.1+/-4.1%, 17.5+/-3.3%, and 20.3+/-4.8%, respectively, versus 36.1+/-6.2% control, P<0.05 each). Furthermore, the effect of olprinone was blunted by either H89 (35.5+/-6.4%) or SB203580 (32.6+/-5.9%), but not by GF109203X or PD98059. H89, GF109203X, PD98059, or SB203580 alone did not influence infarct size. CONCLUSIONS: Pretreatment with PDEIII-Is has cardioprotective effects via cAMP-, PKA-, and p38 MAPK-dependent but PKC-independent mechanisms in canine hearts.

Extensive development of vulnerable plaques as a pan-coronary process in patients with myocardial infarction: an angioscopic study.

OBJECTIVES: To test our hypothesis that the development of vulnerable plaques is not limited to the culprit lesions, but is a pan-coronary process, we directly observed all three major coronary arteries by angioscopy and evaluated the prevalence of yellow plaques in patients with myocardial infarction (MI). BACKGROUND: Although pathologic studies have suggested that the disruption of atheromatous plaque plays a major role in the development of acute MI, the prevalence of yellow plaques in the whole coronary arteries of patients with MI has not been clarified. METHODS: Thirty-two patients undergoing follow-up catheterization one month after the onset of MI were prospectively and consecutively enrolled in this study. The prevalence of yellow plaques and thrombus in the major coronary arteries was successfully evaluated in 20 patients (58 coronary arteries, 21 culprit lesions) by coronary angioscopy. The diameter stenosis (DS) of the culprit lesions and the maximal diameter stenosis (maxDS) of nonculprit segments were angiographically measured for each coronary artery. RESULTS: The DS of the culprit lesions and maxDS were 27 +/- 17% and 19 +/- 13%, respectively. Yellow plaques and thrombus were detected in 19 (90%) and 17 (81%) of 21 culprit lesions, respectively. Yellow plaques were equally prevalent in the infarct-related and non-infarct-related coronary arteries (3.7 +/- 1.6 vs. 3.4 +/- 1.8 plaques/artery). However, thrombus was only detected in the nonculprit segments of one (2%) coronary artery. CONCLUSIONS: In patients with MI, all three major coronary arteries are widely diseased and have multiple yellow though nondisrupted plaques. Acute MI may represent the pan-coronary process of vulnerable plaque development.

[Improvement of method to estimate guidance by pharmacists and trial to obtain standard pharmaceutical management and guidance services program].

The level of understanding of taking medicine was examined at the start and the end of medicine-taking guidance, based on the estimation by pharmacists and inpatients. Six items for the estimation, such as "the way of taking medicines" were evaluated using a five-grade system. The significant improvement was observed in all items, suggesting that inpatients' understanding is improved by the guidance. A markedly positive correlation was found between the estimation by inpatients and that by pharmacists (p < 0.001). This confirms the appropriateness of estimation by pharmacists. Inpatients whose estimation was significantly different from that by pharmacists (difference of two grades or more) were examined to identify clear and potential problems with taking medicine. The problems were classified into such seven categories as "types of medicine". For each item with differences in estimation (two grades or more), problems were biased in some specific categories. On the basis of such a bias, a flow chart was prepared to clarify problems. In addition, a standard pharmaceutical management and guidance services program was drafted, in which measures in observation, treatment and education against problems by pharmacists were described according to the frequency of occurrence. Information about the program available on the Internet enables its use by other hospitals.