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Sodium-glucose cotransporter 2 (SGLT2) is imporant in glucose reabsorption. SGLT2 inhibitors suppress renal glucose reabsorption, therefore reducing blood glucose levels in patients with type 2 diabetes. We and others have developed several SGLT2 inhibitors starting from phlorizin, a natural product. Using cryo-electron microscopy, we present the structures of human (h)SGLT2-MAP17 complexed with five natural or synthetic inhibitors. The four synthetic inhibitors (including canagliflozin) bind the transporter in the outward conformations, while phlorizin binds it in the inward conformation. The phlorizin-hSGLT2 interaction exhibits biphasic kinetics, suggesting that phlorizin alternately binds to the extracellular and intracellular sides. The Na + -bound outward-facing and unbound inward-open structures of hSGLT2-MAP17 suggest that the MAP17-associated bundle domain functions as a scaffold, with the hash domain rotating around the Na + -binding site. Thus, Na + binding stabilizes the outward-facing conformation, and its release promotes state transition to inward-open conformation, exhibiting a role of Na + in symport mechanism. These results provide structural evidence for the Na + -coupled alternating-access mechanism proposed for the transporter family., (© 2023. The Author(s).)

MND-2119 is a self-emulsifying formulation of highly purified eicosapentaenoic acid ethyl ester (EPA-E) designed to be administered once daily due to improved absorption compared with the nonself-emulsifying formulation. In these studies, MND-2119 was administered to healthy adult males in single or multiple doses. In the single administration study, MND-2119 (0.5-4 g) was administered under fed and fasted conditions to evaluate MND-2119 pharmacokinetics and safety under these conditions. This study showed that C max and AUC 0-72h of plasma EPA concentration after single administration were higher under fed conditions than under fasted conditions, for all doses. In the multiple administration study, subjects received either MND-2119 (0.5-4 g) immediately after breakfast or EPA-E (0.9 g) immediately after breakfast and dinner for 11 days to compare pharmacokinetics and safety of MND-2119 to EPA-E. In this study, the rate of rise in C min of the plasma EPA concentration with MND-2119 decreased from days 6 to 8 after administration and was thought to have reached a steady state on day 11. The mean C ss,max of MND-2119 administered as 1 g once daily, and the mean C ss,min and the mean AUC ss,0-24h of MND-2119 administered as 2 g once daily were higher than those of EPA-E administered as 0.9 g twice daily. No safety-related issues occurred in either study. These results suggest that MND-2119 administered once daily may achieve equivalent or higher plasma EPA concentrations compared to the nonself-emulsifying formulation administered twice daily., (© 2022, The American College of Clinical Pharmacology.)

Background: In Japan, eicosapentaenoic acid ethyl ester (EPA-E) is administered twice-daily or three-times-daily for dyslipidemia. We have developed MND-2119, a novel self-emulsifying formulation of highly purified EPA-E, which can be administered once-daily., Objective: The objective of this study was to assess non-inferiority in the efficacy of MND-2119 in patients with hypertriglyceridemia compared with highly purified EPA-E., Methods: In this multicenter, 12-week, double-blind study, patients with high triglyceride (TG levels between ≥ 150 and < 500 mg/dL) undergoing lifestyle modification were randomized to MND-2119 2 g/day (n=145), MND-2119 4 g/day (n=145), EPA-E 1.8 g/day (n=145) or EPA-E 2.7 g/day (n=145). The primary endpoint was percentage change in TG levels from baseline to end of treatment., Results: MND-2119 2, 4 g/day and EPA-E 1.8, 2.7 g/day reduced TG levels from baseline by -10.09%, -15.51%, -9.30%, and -8.80%, respectively. The TG reduction rate of MND-2119 2 g/day was non-inferior to that of EPA-E 1.8 g/day (LS mean difference: -0.42, 95%CI: -5.76 to 4.91). Moreover, the TG reduction rate of MND-2119 4 g/day was superior to that of MND-2119 2 g/day (LS mean difference: -5.74, 95%CI: -10.59 to -0.89). There were no remarkable safety differences between MND-2119 2 g/day and EPA-E 1.8 g/day and between MND-2119 4 g/day and EPA-E 2.7 g/day., Conclusion: Non-inferiority of MND-2119 2 g/day to EPA-E 1.8 g/day for efficacy, and superiority of MND-2119 4 g/day over MND-2119 2 g/day for efficacy were verified. MND-2119 was safe and well tolerated., Competing Interests: Declaration of Competing Interest Takuya Mori, Koichi Hayashi, and Yuichi Yokoyama are employed by Mochida Pharmaceutical Co., Ltd., (Copyright © 2022. Published by Elsevier Inc.)

Background: In Japan, eicosapentaenoic acid ethyl ester (EPA-E) is administered twice-daily or three-times-daily for dyslipidemia. We have developed MND-2119, a novel self-emulsifying formulation of highly purified EPA-E which can be administered once-daily., Objective: The objective of this study was to investigate the safety and efficacy of long-term administration of MND-2119 in hypertriglyceridemia patients., Methods: In this multicenter, 52-week, open-label study, patients with high triglyceride (TG) (TG levels between ≥ 150 and < 500 mg/dL) undergoing lifestyle modification were randomized to MND-2119 2 g/day (n=61) or MND-2119 4 g/day (n=61)., Results: The incidence of adverse events in MND-2119 2 g/day and MND-2119 4 g/day was 70.5% and 62.3%, respectively, and the incidence of adverse drug reactions was 9.8% and 8.2%, respectively. There were no notable problems in the safety assessments of both treatment groups. By Week 4, TG levels had decreased from baseline in both groups, and the TG reducing effect continued up to Week 52 (mean percentage change from baseline in TG at Week 52 [two-sided 95% confidence interval]: MND-2119 2 g/day: -16.71% [-26.61, -6.81], MND-2119 4 g/day: -21.01% [-27.86, -14.16]). In both groups, plasma EPA concentration at Week 4 was maintained up until Week 52 and the plasma EPA concentration at Week 52 was 200.5 ± 54.7 μg/mL in MND-2119 2 g/day and 308.6 ± 98.6 μg/mL in MND-2119 4 g/day., Conclusion: Long-term administration of MND-2119 was not associated with any safety-related problems. TG levels decreased by Week 4, and the TG reducing effect continued up to Week 52., Competing Interests: Declarations of interest T.M and Y.Y are employed by Mochida Pharmaceutical Co., Ltd. This study was supported by Mochida Pharmaceutical Co., Ltd. K.M has acted as the medical expert of this study and has received research/grant support from Mochida Pharmaceutical Co., Ltd., (Copyright © 2022. Published by Elsevier Inc.)

The snailfish Osteodiscus abyssicola sp. nov. is described from a single specimen collected off the Pacific coast of Hokkaido, northern Japan, at a depth of 4,6714,744 m. It is distinguished from all currently recognized congeners by the following combination of characters: vertebrae 49; dorsal-fin rays 44; anal-fin rays 39; principal caudal-fin rays 8; pyloric caeca 5; mouth horizontal; teeth on both jaws simple and sharp, without cusps; upper and lower jaw symphyses without diastema; cephalic pore sizes moderate, similar to or slightly larger than nostril; gill slit extending ventrally to 2nd pectoral-fin ray base; pectoral fin notched; mandibular symphysis to center of anus 101.6% head length (HL); posterior edge of pelvic disk to center of anus 15.5% HL; epural 1, reduced; epipleural ribs absent. An emended diagnosis and key to the species of Osteodiscus are provided.

Background: Recent research has led to the development of many information technology-supported systems for health care control, including systems estimating nutrition from images of meals. Systems that capture data about eating and exercise are useful for people with diabetes as well as for people who are simply on a diet. Continuous monitoring is key to effective dietary control, requiring systems that are simple to use and motivate users to pay attention to their meals. Unfortunately, most current systems are complex or fail to motivate. Such systems require some manual inputs such as selection of an icon or image, or by inputting the category of the user's food. The nutrition information fed back to users is not especially helpful, as only the estimated detailed nutritional values contained in the meal are typically provided., Objective: In this paper, we introduce healthiness of meals as a more useful and meaningful general standard, and present a novel algorithm that can estimate healthiness from meal images without requiring manual inputs., Methods: We propose a system that estimates meal healthiness using a deep neural network that extracts features and a ranking network that learns the relationship between the degrees of healthiness of a meal using a dataset prepared by a human dietary expert. First, we examined whether a registered dietitian can judge the healthiness of meals solely by viewing meal images using a small dataset (100 meals). We then generated ranking data based on comparisons of sets of meal images (850 meals) by a registered dietitian's viewing meal images and trained a ranking network. Finally, we estimated each meal's healthiness score to detect unhealthy meals., Results: The ranking estimated by the proposed network and the ranking of healthiness based on the dietitian's judgment were correlated (correlation coefficient 0.72). In addition, extracting network features through pretraining with a publicly available large meal dataset enabled overcoming the limited availability of specific healthiness data., Conclusions: We have presented an image-based system that can rank meals in terms of the overall healthiness of the dishes constituting the meal. The ranking obtained by the proposed method showed a good correlation to nutritional value-based ranking by a dietitian. We then proposed a network that allows conditions that are important for judging the meal image, extracting features that eliminate background information and are independent of location. Under these conditions, the experimental results showed that our network achieves higher accuracy of healthiness ranking estimation than the conventional image ranking method. The results of this experiment in detecting unhealthy meals suggest that our system can be used to assist health care workers in establishing meal plans for patients with diabetes who need advice in choosing healthy meals., (©Kyoko Sudo, Kazuhiko Murasaki, Tetsuya Kinebuchi, Shigeko Kimura, Kayo Waki. Originally published in JMIR Formative Research (http://formative.jmir.org), 26.10.2020.)

A new snailfish, Paraliparis flammeus , is described on the basis of 18 specimens collected off the Pacific coast of Tohoku District, northern Japan at depths of 422-890 m. The new species is distinguished from 28 species of Paraliparis described from the North Pacific by the following combination of characters: mouth oblique; uppermost pectoral-fin base below horizontal through posterior margin of maxillary; 60-63 vertebrae, 54-58 dorsal-fin rays, 50 or 51 anal-fin rays, six principal caudal-fin rays, and 17-20 pectoral-fin rays. A maximum likelihood tree based on 106 COI gene sequences (492 bp) of Paraliparis recovered a monophyletic group comprising P. flammeus , Paraliparis cephalus , and Paraliparis dipterus . Paraliparis cephalus is similar to P. flammeus in having an oblique mouth, but it has four caudal-fin rays (vs six rays) and the uppermost pectoral-fin base above a horizontal through the maxillary posterior margin. Paraliparis dipterus differs from P. flammeus in having a horizontal mouth, 12-14 pectoral-fin rays, and lacking pyloric caeca (present in P. flammeus ). Paraliparis flammeus is most similar to the eastern North Pacific Paraliparis mento in having an oblique mouth and the uppermost pectoral-fin base below a horizontal through the posterior margin of the maxillary. However, P. flammeus differs from P. mento in having six caudal-fin rays (vs five rays) and greater preanal length (29.9-35.3% SL vs 26.7-28.5% SL). A poorly known species, Paraliparis mandibularis , previously known from only two specimens collected from Tosa Bay, southern Japan, is redescribed based on the holotype and seven newly collected specimens. It is also similar to the new species but has 27-30 pectoral-fin rays and a shorter pectoral-fin lower lobe (13.8-15.9% SL in P. mandibularis vs 16.7-23.4% SL in P. flammeus )., (Yoshiaki Kai, Kenta Murasaki, Ryo Misawa, Atsushi Fukui, Eisuke Morikawa, Yoji Narimatsu.)

Background: Recent clinical trials' findings have revealed the therapeutic noninferiority of direct oral anticoagulant (DOAC) to standard therapy with vitamin K antagonist (VKA) in patients with pulmonary thromboembolism (PTE). However, few studies have quantitatively analyzed thrombus reduction in the pulmonary artery., Methods: This observational study included 38 symptomatic PTE patients with stable hemodynamics. All patients received an intravenous heparin bolus followed by continual heparin injections immediately after the PTE diagnosis. The heparin was discontinued after edoxaban therapy began in the DOAC group (n = 22) or after the therapeutic range for the prothrombin time-international normalized ratio was achieved in the VKA group (n = 16). The thrombus volumes in the pulmonary arteries were quantitatively analyzed using contrast-enhanced computed tomography scans, and they were compared at baseline and at 2 weeks after admission., Results: The pulmonary thrombus volumes declined in the VKA and DOAC groups from 7.9 to 4.2 cm 3 (P = 0.048) and from 7.1 to 3.7 cm 3 (P < 0.01), respectively, and the thrombus reduction rates did not differ significantly between the groups (-34% vs. -64%, respectively; P = 0.38). The fibrinogenolysis parameter changes during the14 days after admission were similar in both groups. Compared with the VKAgroup, the average hospital stay was 9days shorter in the DOAC group. There were no in-hospital deaths, and 1 case experienced major bleeding in the VKA group., Conclusions: In relation to pulmonary artery thrombus volume reduction, DOAC monotherapy for PTE may be comparable with standard therapy involving VKAs., (Copyright © 2020 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.)

Background: In "real-world" practice, anticoagulant therapy is indicated for patients whose clinical profiles are not addressed in randomized clinical trials. We assessed the effectiveness and safety of dabigatran versus warfarin in "real-world" Japanese patients with non-valvular atrial fibrillation (NVAF)., Methods: Among 613 NVAF patients who initiated dabigatran or warfarin therapy during the period between 2011 and 2013, 362 patients were included in the study after propensity score adjustment. The median follow-up period was 1.3 years. The effectiveness and safety outcomes were thromboembolism and major bleeding, respectively., Results: The propensity-matched hazard ratios of thromboembolism and major bleeding with dabigatran were 1.03 (95% CI: 0.12-8.04, p =0.971) and 0.15 (95% CI: 0.01-0.90, p =0.037), respectively., Conclusions: The ability of dabigatran to prevent thromboembolism is comparable to that of warfarin; however, the major bleeding rate is lower with dabigatran in "real-world" NVAF patients.

Background: Non-vitamin K antagonist oral anticoagulants (NOACs) show a favorable balance between efficacy and safety compared with warfarin for patients with non-valvular atrial fibrillation (NVAF). In "real-world" practice, however, NOAC adherence and persistence among patients are not clear. The aim of this study is to evaluate NOAC and warfarin persistence in Japanese patients with NVAF who newly started these drugs., Methods: We retrospectively studied 401 patients with NVAF who had newly started NOACs during the first 18 months after our hospital adopted their use (197 dabigatran, 107 rivaroxaban, 102 apixaban) and 200 patients with NVAF who had newly started warfarin during the same period. The endpoint was drug discontinuation for each drug., Results: During the follow-up period (up to a maximum of 24 months), 113 (28%) patients who had newly started NOACs and 33 (17%) patients who had newly started warfarin discontinued the drug. The persistence rates of patients prescribed NOACs was lower than that of patients prescribed warfarin at 3, 6, and 12 months (85% versus 93%, 79% versus 88%, and 70% versus 82%, respectively). One-tenth of patients who had newly started NOACs discontinued the drug by their own decision. Drug adverse events, worsening renal dysfunction, and patient desire were the major causes of NOAC discontinuation., Conclusions: The rate of persistence of prescribed NOACs was significantly lower than that of warfarin in Japanese patients with NVAF.

The whisker pad area (WP) is innervated by the second branch of the trigeminal nerve and experiences allodynia and hyperalgesia following transection of the mental nerve (MN; the third branch of the trigeminal nerve). However, the mechanisms of this extra-territorial pain remain unclear. The ionotropic P2X(7) ATP receptor (P2X(7)) in microglia is known to potentiate, via cytokines, the perception of noxious stimuli, raising the possibility that P2X(7) and cytokines are involved in this extra-territorial pain. One day after MN transection (MNT), WP allodynia/hyperalgesia developed, which lasted for > 8 wks. Activation of microglia and up-regulation of P2X(7), membrane-bound tumor necrosis factor (TNF)-α (mTNF-α), and soluble TNF-α (sTNF-α) in the trigeminal sensory nuclear complex (TNC) were evident for up to 6 wks after MNT. Allodynia/hyperalgesia after MNT was blocked by intracisternal administration of etanercept, a recombinant TNF-α receptor (p75)-Fc fusion protein. Intracisternal A438079, a P2X(7) antagonist, also attenuated allodynia/hyperalgesia and blocked up-regulation of mTNF-α and sTNF-α in the TNC. We conclude that sTNF-α released by microglia following P2X(7) activation may be important in both the initiation and maintenance of extra-territorial pain after MNT.

Background: The present study directly addresses the roles of the P2X(7) receptor (P2X(7)R), an ionotropic adenosine triphosphate (ATP) receptor, and cytokines in the induction of orofacial pain following chronic constriction injury (CCI) of the infraorbital nerve (IoN)., Methods: Rats were anesthetized, and ligatures of 4-0 chromic gut were tied around the IoN. A438079, a P2X(7)R antagonist or SB203580, a phosphorylated (p)-p38 mitogen-activated protein kinase (MAPK) inhibitor, was infused intrathecally into CCI-treated rats. In another group of rats, 3'-O-(4-benzoylbenzoyl) adenosine 5'-triphosphate (BzATP), a P2X(7) R agonist, was infused intrathecally with A438079, SB203580 or etanercept, a tumor necrosis factor (TNF)-α receptor-binding recombinant drug., Results: CCI of the IoN induced tactile allodynia/hyperalgesia and up-regulation of P2X(7)R, membrane-bound TNF-α (mTNF-α) and soluble TNF-α (sTNF-α) in the trigeminal sensory nuclear complex (TNC). Tactile allodynia/hyperalgesia or up-regulation of mTNF-α and sTNF-α in the TNC following CCI of the IoN was inhibited by A438079. SB203580 also attenuated tactile allodynia/hyperalgesia or up-regulation of mTNF-α, but not the up-regulation of sTNF-α in the TNC. Treatment of rats with BzATP induced tactile allodynia/hyperalgesia and up-regulation of sTNF-α and p-p38 in the TNC. Tactile allodynia/hyperalgesia or up-regulation of sTNF-α following BzATP treatment was inhibited by SB203580 and etanercept., Conclusions: Based on these findings, phosphorylation of p38 MAPK via P2X(7)R may induce tactile allodynia/hyperalgesia, which is most likely mediated by sTNF-α released by microglia., (© 2012 European Federation of International Association for the Study of Pain Chapters.)

Background: Although a lower target prothrombin time-international normalized ratio (PT-INR) with warfarin therapy is recommended in Japan for atrial fibrillation (AF) patients ≥70 years of age, few studies have provided supporting data. The current study aimed to evaluate the clinical outcome in elderly Japanese patients with non-valvular AF who were taking warfarin., Methods: We conducted a cohort study of 845 consecutive non-valvular AF patients ≥70 years of age who were taking warfarin (median age, 74 years; 30.5% women) with a median follow-up period of 27 months (4-69 months). Of these patients, 29.7% had a history of stoke/transient ischemic attack (TIA), and 73.1% of the patients had a CHADS(2) score ≥2. The occurrence of thromboembolic events, including ischemic stroke, TIA and other systemic embolisms, and major bleeding events were validated through a review of medical records., Results: The incidence of thromboembolic and major bleeding events were 3.8 and 2.1% per year, respectively. A higher incidence of both events was observed in patients with a CHADS(2) score ≥3. The multivariate analysis showed that prior stroke/TIA (odds ratio 1.7, 95% CI 1.0-2.7) and diabetes (odds ratio 1.7, 95% CI 1.0-2.8) were independent risks of thromoembolic events. A HAS-BLED score ≥3 represented a risk for major bleeding (hazard ratio 2.8, 95% CI 1.7-4.6). A PT-INR of 1.5-2.5 indicated a low incidence of thromboembolic and major bleeding events in patients with a CHADS(2) score ≥2., Conclusions: Our results demonstrate that a target PT-INR of 2.0 and a range of 1.5-2.5 may be safe for elderly Japanese patients with non-valvular AF., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

It has been reported that the whisker pad (WP) area, which is innervated by the second branch of the trigeminal nerve, shows allodynia/hyperalgesia following transection of the mental nerve (MN: the third branch of the trigeminal nerve). However, the mechanisms of this extra-territorial pain induction still remain unclear. Glia and cytokines are known to facilitate perception of noxious input, raising a possibility that these non-neuronal elements are involved in the induction and spread of allodynia/hyperalgesia at non-injured skin territory. One day after MN transection, tactile allodynia/hyperalgesia developed on the ipsilateral WP area, which is in the non-injured skin territory. The tactile allodynia/hyperalgesia lasted for more than 56 days. In response to MN transection, astrocytes and microglia appeared to be in an activated state, and interleukin (IL)-1beta was up-regulated in astrocytes in the trigeminal subnucleus caudalis (Vc). Allodynia/hyperalgesia at WP area induced by MN transection was attenuated dose-dependently by IL-1 receptor antagonist IL-1ra (i.t., 0.05, 0.5, and 5 pg/rat). Fos-like immunoreactive (Fos-Li) neurons were observed in the Vc after non-noxious mechanical stimulation of the WP area in the rats with MN transection. Administration of IL-1ra also attenuated the number of Fos-Li neurons dose-dependently. Administration of a noncompetitive antagonist of NMDA receptors MK-801 (i.t., 5 μg/rat) reversed allodynia/hyperalgesia. IL-1 receptor type I (IL-1RI) was localized in Fos- and phospho NR1-immunoreactive neurons. These results suggest that IL-1beta in the Vc plays an important role in the development of extra-territorial tactile allodynia/hyperalgesia after MN transection., (Copyright © 2010 European Federation of International Association for the Study of Pain Chapters. Published by Elsevier Ltd. All rights reserved.)

The first total synthesis of (-)-fructigenine A and a novel approach to (-)-5-N-acetylardeemin through a common imine intermediate (+)-3 are described. The key steps include highly enantioselective preparation of (+)-3 via domino olefination/isomerization/Claisen rearrangement (OIC) of 5, reductive cyclization (RC), and regioselective oxidation of (-)-4 and a novel assembly of the pyrazino ring of these alkaloids via Ugi three-component reaction/cyclization of (+)-3 with the corresponding amino acid and isonitrile.

Background: Thromboembolic events account for significant morbidity and mortality after the Fontan procedure, but the underlying mechanisms remain unclear. P-selectin on platelets indicates platelet activation. Thrombomodulin (TM), a receptor for thrombin and a major anticoagulant proteoglycan on the endothelial membrane, reflects the anticoagulant activity of the endothelium. The present study investigated the hypothesis that the balance between platelet activation and endothelial biological function is impaired in Fontan patients., Methods and Results: Platelet P-selectin as a marker of platelet activation, plasma TM levels and protein C activity, as markers of anticoagulant activity of the endothelium, and thrombin-antithrombin complex III (TAT) were examined in 43 Fontan patients. P-selectin levels on platelets (4.5 +/-1.4 vs 3.4 +/-0.4 mean fluorescence intensity, P<0.001) and TAT levels (80.2 +/-322.6 vs 1.9 +/-0.9 ng/ml, P<0.05) were significantly higher in Fontan patients than in control subjects. On the other hand, plasma TM levels (1.5 +/-0.8 vs 2.2 +/-0.3 FU/ml, P<0.01) and protein C activity (71 +/-35 vs 118 +/-25%, P<0.001) were significantly lower in Fontan patients compared with controls. These abnormalities were not seen in patients after other surgical procedures for congenital heart disease., Conclusions: Platelet activation is enhanced and endothelial function is impaired in patients after the Fontan procedure, which may partly explain the thromboembolic complications in Fontan patients.

Recent studies have suggested important roles of inflammation in the pathophysiology of unstable angina (UA). We investigated whether activation of the circulating platelets and neutrophils were implicated in inflammatory reactions associated with unstable angina Expressions of platelet P-selectin and neutrophil CD11b, and neutrophil-platelet aggregates were evaluated by flow cytometry in anticoagulated peripheral venous blood from 71 patients with UA and 22 patients with stable angina (SA). Expressions of platelet P-selectin and neutrophil CD11b, and neutrophil-platelet aggregates on the admission day were all significantly higher in 71 patients with UA than 22 with SA (median, mean fluorescence intensity [MFI]: 7.00 vs 4.51, P < 0.01, 64.68 vs 47.75, P = 0.0007; and % of 10 000 neutrophils: 7.84 vs 3.40, P = 0.0001, respectively). These three parameters in 43 patients with UA were significantly decreased (MFI: 4.23, P = 0.003, 50.82, P = 0.0003; and % of 10 000 neutrophils: 5.04, P = 0.0001, respectively) 7 days after the first measurement. These results indicate that circulating activated platelets and neutrophils are more strongly implicated in the acute phase of UA. These findings also suggest that thrombus formation after rupture of atherosclerotic plaques as well as plaque formation involves inflammatory reactions.

Inflammatory and immunological mechanisms are implicated in the development of idiopathic dilated cardiomyopathy (DCM). Since activated T lymphocytes express surface HLA-DR antigens, an increased level of these cells in the circulation could indicated an ongoing immune response. While the role of activated T lymphocytes in experimental myocarditis has been elucidated, the contribution of T lymphocyte activation in clinical DCM remains unclear. We therefore examined the role of T-cell activation in peripheral blood samples obtained from 10 patients with DCM (mean age, 49 +/- 12 years) and from 10 age-matched healthy controls. Citrated whole blood was mixed with fluorescein isothiocyanate- or phycoerythrin-conjugated specific monoclonal antibodies and analyzed using a fluorescence-activated cell sorter (FACS). The ratio (%) of histocompatibility leukocyte antigen (HLA)-DR positive cells in the FACS gated lymphocyte population was significantly higher in DCM patients than in controls (7.9% +/- 5.3% vs 2.0% +/- 0.9%; P < 0.01). The expression of CD40L on T cells determined as mean fluorescence intensity (MFI) was also significantly higher in DCM patients than in controls (3.6 +/- 2.1 vs 1.8 +/- 0.4 MFI; P < 0.05). Furthermore, the ratios of T cells expressing HLA-DR and serum brain natriuretic peptide (BNP) levels closely correlated (P = 0.0008). We showed that HLA-DR on peripheral T cells significantly correlated with serum BNP levels and that high CD40L expression on T cells was concomitant with increased BNP levels (P < 0.05). Therefore the magnitude of T-cell expression, such as increased expression of HLA-DR and CD40L, contributes to myocardial dysfunction in DCM.

Recent studies have shown that circulating platelets play an important role in the development of restenosis early after coronary stent implantation. We investigated P-selectin expression and CD36 blockade on platelets by flow cytometry in 48 consecutive patients who underwent coronary stenting. P-selectin expression was significantly higher 1 day after stenting in patients who had restenosis (n = 15) than in those who had no restenosis (n = 28), and the odds ratio for restenosis in patients with high P-selectin levels (MFI > 6.5) was 11.67 (P < 0.001) as compared with patients who had intermediate and low P-selectin levels. CD36 blockade was assessed with the use of two anti-CD36 antibodies, OKM5 and GS95 (our new anti-CD36 antibody), the binding of which indicates total CD36 amount and free CD36 unoccupied by lipid-related ligands, respectively. Binding of OKM5 to platelets was similar before and after stenting in both groups. CD36 blockade on platelets was seen 1 day after stenting in the non-restenosis group, and the odds ratio for restenosis in patients without CD36 blockade [GS95 binding ratio >0.8 as compared with binding before stenting] on day 1 was 28.60 (P < 0.001). P-selectin expression and unoccupied CD36 on platelets shortly after stenting may be strong predictors of post-stent restenosis.

Background: The basic mechanisms of thromboembolism in cyanotic congenital heart disease (CCHD) have not been well clarified. P-selectin on the platelets reflects platelet activation. Thrombomodulin is a critical cofactor for thrombin-mediated activation of protein C and reflects the anticoagulant activity of the endothelium. The present study was performed to evaluate whether platelet activation exists in patients with CCHD., Methods and Results: Platelet P-selectin as a marker of platelet activation, plasma thrombomodulin level and protein C activity as markers of anticoagulant activity of the endothelium and thrombin - antithrombin complex III (TAT) were examined in 35 patients with CCHD. Plasma thrombomodulin level (1.1+/-0.9 vs 2.2+/-0.3 FU/ml) and protein C activity (71.1+/-29.8 vs 117.8+/-24.8%) were significantly lower in patients with CCHD as compared with the control subjects. The levels of plasma TAT (255+/-811 vs 1.9+/-0.9 ng/ml) and P-selectin on platelets (6.3 +/-4.5 vs 3.3+/-0.3 mean fluorescence intensity) were significantly higher in the patients with CCHD than in the controls. Four of the CCHD patients who experienced thromboembolic events had elevated levels of platelet P-selectin (p=0.02) compared with CCHD patients without thromboembolic events., Conclusion: Platelet activation exists in patients with CCHD and it may play an important role in the thromboembolic events in CCHD.

Glycoprotein (GP) VI is a major receptor for collagen and belongs to the immunoglobulin super family. Here, we examined the localization of GPVI in resting and activated human platelets by immunogold scanning and transmission electron microscopy and flow cytometry. Ultrastructural observation detected immunolabelling for GPVI that was distributed uniformly over the entire surface of resting platelets, and revealed that GPVI was also localized on both the membranes of the surface-connected open canalicular system (OCS) and alpha-granules. The OCS- and alpha-granule-associated GPVI pools were an estimated 35.4 +/- 3.2% (mean +/- standard deviation) of the total. Little GPVI labelling was observed in any part of GPVI-deficient platelets. A remarkable time-dependent increase in GPVI surface expression was found by flow cytometry when platelets were activated by collagen-related peptide (CRP) and convulxin. The GPVI-mediated activation of platelets by CRP or convulxin resulted in similar ultrastructural changes and an increased GPVI labelling density on the activated platelet surface, which was accompanied by a decreased interior expression. GPVI was also expressed on microparticles generated from activated platelets. Thus, our study demonstrates that platelets have internal pools of GPVI, and that GPVI is increasingly redistributed to the surface membrane and to microparticles during platelet activation.

The thrombotest (TT) technique has been widely used in Japan for monitoring oral anticoagulant therapy (OAT). The therapeutic range was originally recommended to be 10%-25%. However, the International Committee for Standardization in Hematology/International Committee on Thrombosis and Hemostasis (ICSH/ICTH) recommended using the international normalized ratio of prothrombin time (PT-INR) for monitoring OAT. It is necessarv to use a universal standard measure for monitoring OAT in accordance with the ICSH/ISTH recommendation. We simultaneously measured TT and PT in blood samples from 1,157 patients on long-term warfarin therapy, and studied the correlation between TT and PT-INR. An excellent linear correlation was obtained between TT-INR and PT-INR with the regression equation PT-INR = 1.0420 TT-INR - 0.0987 (r = 0.905, P < 0.001). We also examined the correlation between the incidence of thromboembolism in 170 patients receiving warfarin therapy after prosthetic valve replacement; 50.5% received concomitant antiplatelet therapy. Thromboembolism occurred in 9 of 170 patients during a mean follow-up period of 2.44 years. The average TT values in patients with and without thromboembolism were 26.4% (PT-INR: 1.53) and 21.1% (1.73), respectively (P < 0.01). The incidence of thromboembolism did not differ significantly between patients on warfarin alone (average TT: 22.2%) and those on warfarin and antiplatelet agent (average TT: 20.9%). Our results suggest that the incidence of thromboembolism is low in Japan despite a less intensive regimen having been adopted.

A 53-year-old man, who had undergone tricuspid valve replacement with Hancock valve and direct closure of a ventricular septal defect when aged 34 years, was admitted with signs of right heart failure. Two-dimensional echocardiography showed bioprosthetic tricuspid valve malfunction with right atrial thrombus. He was treated by tricuspid valve replacement using a Hancock II valve and removal of the right atrial thrombus with remarkable improvement. Transesophageal echocardiography was the most useful method for recognizing the presence of right atrial thrombus and assessing its actual or potential hemodynamic effects.