Pharmacokinetics of the amidine prodrug of a novel oral anticoagulant factor VIIa inhibitor (AS1924269-00) in rats.

AS1924269-00 is a promising orally applicable anticoagulant that inhibits FVIIa but has very low oral absorption. Therefore, in this study, we aimed to develop a prodrug of AS1924269-00, which possesses a carbamate-added amidine functional group, with high membrane permeability. We investigated the pharmacokinetics of the carbamate-type prodrug of AS1924269-00 in rats. The Caco-2 cell monolayer was used as an in vitro model and in parallel an artificial membrane permeability assay (PAMPA) was performed to examine the transport mechanisms of the prodrug. The bioavailability of the active form was determined to be only 0.3% in rats, but the oral absorption of the prodrug was markedly improved, and its bioavailability was 36%. Our in vivo result was consistent with the finding that compared to AS1924269-00, the prodrug showed favorable permeability in Caco-2 cells and PAMPA. We introduced carbamate into the amidine functional group of the FVIIa inhibitor, which possesses the amidine backbone, and converted it to a prodrug using carboxylic acid ethyl ester. This novel prodrug had favorable absorption and membrane permeability in vivo and in vitro. Thus, we suggest a clinical application of the carbamate-added amidine prodrug of the FVIIa inhibitor.

Characterization of in vitro biotransformation of the new oral anticoagulants, the factor VIIa inhibitors AS1927819-00 and AS1932804-00.

We recently developed a prodrug (AS1932804-00, CMP) of the novel FVIIa inhibitor AS1924269-00, which possesses a carbamate amidine backbone. In addition, we developed another type of prodrug (AS1927819-00, OXP) with an oxime amidine backbone. In this study, we investigated the efficiency of conversion of these novel FVIIa prodrugs to their active forms by evaluating the production of the active form in vitro by using microsomes, mitochondria, and cryopreserved hepatocytes, and compared it with the in vivo conversion mechanisms of the prodrugs (oxime amidine vs. carbamate amidine). We observed that OXP and CMP showed improved oral absorption, and the efficiency of conversion of CMP to the active form was higher than that of OXP. The in vivo rate of conversion of OXP to its active form was low in rats, and compared to liver microsomes and mitochondria, cryopreserved hepatocytes supplemented with serum and coenzymes were an appropriate metabolic test tool. On the other hand, the efficiency of conversion of CMP to its active from could be appropriately evaluated using small intestinal microsomes. The development of a prodrug can be optimized when information about the stability of carboxylic acid esters in the presence of serum esterases, membrane permeability of intermediate forms, and differential tissue specificity to metabolic activities for carbamate and oxime backbones of amidine can be obtained.

Impact of the COVID-19 pandemic on the surveillance of antimicrobial resistance.

BACKGROUND: The impact of the coronavirus disease (COVID-19) pandemic on antimicrobial resistance (AMR) is a major concern. AIM: To compare the number of patients and isolation rate of antimicrobial-resistant bacteria before and after the beginning of the COVID-19 pandemic using the comprehensive national surveillance data. METHODS: We utilized comprehensive surveillance data, collected in the Japan Nosocomial Infections Surveillance programme, which included a total of 16.7 million samples of 5.9 million tested patients from >1300 hospitals. We compared the number of patients and isolation rate of five bacteria between 2019 and 2020, including antimicrobial-susceptible and -resistant bacteria of Staphylococcus aureus, Streptococcus pneumoniae, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. FINDINGS: The number of patients and isolation rate of S. aureus and meticillin-resistant S. aureus decreased slightly; those of S. pneumoniae and penicillin-resistant S. pneumoniae decreased by 60%; and those of third-generation cephalosporin-resistant K. pneumoniae increased. The isolation rate of the remaining bacteria apparently increased, although the number of patients decreased. This was due to a substantial decrease in the total number of tested patients (the denominator of the isolation rate), which was larger than that of the number of patients (the numerator of the isolation rate). Consistent results were obtained when the same data were re-aggregated using the procedure of the World Health Organization Global Antimicrobial Resistance Surveillance System, demonstrating the general importance of this problem. CONCLUSION: Surveillance data during the COVID-19 pandemic must be carefully interpreted based on examination of the numerator, denominator and background factors that affect the denominator.

Five-year point prevalence survey of healthcare-associated infections and antimicrobial use in a Japanese university hospital.

BACKGROUND: Periodic point prevalence surveys (PPSs) provide a method for assessing changes in healthcare-associated infections (HAIs) and antimicrobial use over time. Following the introduction of an antimicrobial stewardship programme at Nagoya University Hospital (Aichi, Japan) a five-year PPS study was performed to highlight any epidemiological changes. METHODS: One-day PPSs were performed annually in July at Nagoya University Hospital. Data on patient characteristics, medical devices, active HAIs and antimicrobial use were collected using a standard data-collection form. RESULTS: A total of 4339 patients were included. Over the five-year study period the median patient age was 62 years, median duration of hospital admission was nine days, 9% of patients had an HAI and 35.2% received at least one antimicrobial. Overall there were 406 HAIs (95% confidence interval, 369-447) with surgical site infection, pneumonia and febrile neutropenia occurring most frequently. Enterobacterales were the most common pathogens (N = 78, 28.6%) and 32.1% were third-generation cephalosporin-resistant. Meropenem was the most frequently prescribed antimicrobial for HAIs. Surgical antimicrobial prophylaxis changed drastically, with shorter durations and a marked reduction in oral cephalosporin use. However, antimicrobials for medical prophylaxis gradually increased. CONCLUSIONS: This five-year PPS study shows consistent data for patient background, HAIs and causative pathogens and highlights changes in antimicrobial use during the era of the National Action Plan on Antimicrobial Resistance. To describe the epidemiology of Japanese hospitals by PPS, multicentre PPSs including in community hospitals should be performed annually.

A multimicrospray nebulizer for microwave-induced plasma mass spectrometry

We have developed a nebulizer, called a multimicrospray nebulizer (MMSN), that efficiently introduces analytes for plasma mass spectrometry and plasma emission spectrometry. In this nebulizer, both the sample solution and the nebulizer gas are divided into several streams to produce a multispray. That is, the MMSN is a nebulizer that contains several micronebulization units, each unit including an orifice for passing the nebulizer gas and a capillary for introducing a sample solution. The microspray from each micronebulization unit can be operated at a microliter per minute sample uptake rate to achieve high nebulization efficiency. The multimicrospray nebulizer is capable of introducing more analyte to the plasma compared with a single-orifice micronebulizer, which has a very low sample uptake rate. In this work, an MMSN with three orifices was found to be suitable for microwave-induced plasma mass spectrometry (MIP-MS). The sample uptake rate can be varied within a range of 5-250 microL/min. Therefore, the nebulizer is unique in its ability to deal with various sample volumes and provide high nebulization efficiency. The sensitivity for all elements obtained with the MMSN was higher than that obtained with a conventional concentric nebulizer (CCN), which is difficult to achieve with other types of microintroduction nebulizers. For most elements, the MIP-MS sensitivity was improved about 2-fold at a sample uptake rate of 150 microL/min, a much lower rate than that for the CCN (usually 0.5-1.5 mL/min). The sensitivity for arsenic was improved by a factor of 5. The relative standard deviation was found to be less than 2.0%.

Discovery of novel N-phenylglycine derivatives as potent and selective beta(3)-adrenoceptor agonists for the treatment of frequent urination and urinary incontinence.

With a novel assay using isolated ferret detrusor to estimate beta(3)-adrenoceptor agonistic activity, we found that a series of glycine derivatives of ritodrine, a beta(2)-adrenoceptor agonist, are potent beta(3)-adrenoceptor agonists, with excellent selectivity versus beta(1) and beta(2) subtypes. Substitution of halogens in the phenyl ring increased potency and selectivity for the beta(3)-adrenoceptor, and this was dependent upon the position of the halogens. The chlorine-substituted derivatives 3f-i exhibited potent beta(3)-adrenoceptor-mediated relaxation of ferret detrusor (EC(50) = 0.93, 11, 14, and 160 nM) and higher potency at beta(3)-adrenoceptors than at beta(1) or beta(2). The intravenous administration of 3h significantly reduced the urinary bladder pressure in anesthetized male rats (ED(50) = 48 microg/kg) without cardiovascular side effects. This article is the first report of structure-activity relationships (SAR) concerning beta(3)-adrenoceptor agonists as agents for the treatment of urinary frequency and incontinence.

Role of neuropeptide Y and its receptor subtypes in neurogenic pulmonary edema.

The effect of neuropeptide Y on the number of perivascular carbon deposits, assessed as a measure of lung vascular permeability, was examined in isolated perfused lung preparations of rats. The number of carbon particle deposits after bronchial application of neuropeptide Y was increased in a dose-dependent manner. In the presence of a beta-adrenoceptor antagonist, norepinephrine augmented the effects of neuropeptide Y. Peptide YY, an analog of neuropeptide Y, demonstrated a much lower potency for increasing the number of carbon deposits, and neuropeptide Y-(18-36), which elicits a weak antagonist action on the neuropeptide Y Y3 receptor, significantly decreased the neuropeptide Y-induced increase. Furthermore, examination of the influence of neuropeptide Y-(18-36) pretreatment on fibrin-induced neurogenic pulmonary edema, in rats, revealed a reduction of the protein concentration ratio of tracheal fluid to serum. Therefore, we conclude that neuropeptide Y may elevate vascular permeability in the pulmonary circulation, conceivably through the neuropeptide Y Y3 receptor, and that neuropeptide Y may in fact play a physiological role even in the in-situ pulmonary circulation.

Effects of neuropeptide Y on lung vascular permeability in the pulmonary circulation of rats.

The effects of neuropeptides on the capillary filtration coefficient in the vessels of the pulmonary circulation were examined, using isolated lung perfusion preparations from rats. Neuropeptide Y and neurokinin A elevated the filtration coefficient, and calcitonin gene related peptide diminished it. Neurotensin and substance P did not affect the value at concentrations less than 10(-7) M. The number of extravasated carbon particle deposits subsequent to tracheal application of neuropeptide Y during spontaneous respiration increased in a dose-dependent manner. From these results, we conclude that neuropeptide Y may increase vascular permeability in the pulmonary circulation.

Repeated 131I treatment of a residual ovarian teratoma containing malignant thyroid tissue.

A 49-year-old woman with ovarian teratoma received 131I treatment three times for an unresectable mass containing malignant thyroid tissue after surgery. Repeated 131I treatment effectively reduced serum thyroglobulin (Tg) level and tumour uptake of 131I, despite absence of any change in size of the treated tumour. Treatment did not inhibit the increase of serum CA-125 and tumour 201Tl uptake, associated with progression of a radioresistant intratumoral hyper-perfused tissue component, detected by colour Doppler ultrasound. Serum CA-125 level and tumour 201Tl uptake were not significantly changed despite temporary increases in serum Tg level after each 131I treatment. These observations indicate the importance of diagnostic measures using combined functional imaging and tumour markers in managing this rare tumour.

Error correcting memorization learning for noisy training examples.

In order to avoid overfitting, we propose error correcting memorization learning. This method is derived from minimization of error between outputs of a trained neural network and correct values for noisy training examples, although the correct values are unknown. We show that noise is adequately suppressed by error correcting memorization learning. The noise suppression mechanism is theoretically clarified. It is found that redundancy plays an essential role for noise suppression and depends on a set of training inputs. We give the condition for the training inputs to provide the redundancy. Moreover, by clarifying the relationships between the proposed method and the weighted least squares estimation with the Mahalanobis norm, we reveal effectiveness of the weighted least squares estimation on noise suppression.

Abnormal brain perfusion demonstrated by Tc-99m MAA total-body scan in two children with complex congenital heart disease.

This paper describes abnormal brain perfusion unexpectedly demonstrated by Tc-99m MAA total-body imaging in two children with intracardiac right-to-left shunt (RLS) associated with complex congenital heart disease. One child was a 12-year-old girl with asplenia cardiac syndrome and multiple cerebral infarctions caused by thromboembolism in the internal carotid artery, and the other child was a 6-month-old boy who developed focal cerebral infarction following shunt operation. In both children, the total-body imaging depicted the brain due to RLS, where radioactivity decreased unilaterally in the cerebral hemisphere. In the first patient, radioactivity also decreased in the contralateral cerebellum, suggesting the crossed cerebellar diaschisis phenomenon. These abnormalities in brain perfusion were confirmed by Tc-99m HMPAO brain SPECT. Careful review of the distribution of the radiotracer in the depicted brain on Tc-99m MAA total-body imaging may provide important information regarding brain perfusion in some patients with a high risk of stroke complication associated with RLS.

Matrix effect on the analysis of oligonucleotides by using a mass spectrometer with a sonic spray ionization source.

Matrix or impurities remaining in a DNA sample solution after various sample treatment procedures may influence a subsequent DNA analysis. In this work, several matrices were investigated concerning their effects on the analysis of oligonucleotide by using an ion-trap mass spectrometer equipped with a sonic spray ionization source. Inorganic salts of sodium chloride and magnesium chloride depressed the signal intensity by about 50% when the content of the salts was about 10 microM. dNTPs and Taq showed more severe depression on the oligonucleotide. However, Tris, or (hydroxymethyl)aminomethane, intensified the signal intensity, if its content was within an appropriate range. When the content of Tris was about 500 microM, the signal intensity was enhanced by factors of 3 and 5 for the 6-mer and the 20-mer oligonucleotides, respectively. With the existence of Tris, matrix effects from the inorganic salts, dNTPs and Taq were reduced.

Hyperlipoproteinemia in rats fed polychlorinated biphenyls.

Hypercholesterolemia due to feeding of polychlorinated biphenyls (PCB) and due to feeding of high cholesterol was compared. Rats in control group were fed a 25% casein diet, and those in PCB group or cholesterol group were fed the 25% casein diet supplemented with 0.03% PCB or 1% cholesterol and 0.25% cholic acid, respectively. Lipoprotein mass (d less than 1.21 g/ml) was higher 1.9- and 1.3-fold in rats fed PCB and cholesterol, respectively, as compared with controls. In rats fed PCB, protein, cholesterol, and phospholipid in the lipoprotein fraction markedly increased. Cholesterol-feeding resulted in the increase in pre-beta-lipoprotein cholesterol, while PCB-feeding increased alpha-lipoprotein cholesterol and slightly slow migrated pre-beta-lipoprotein cholesterol. PCB-treated rats had more apolipoprotein A-I in the lipoprotein fraction than control and cholesterol-fed rats. The data demonstrated that hyperlipoproteinemia induced by PCB is a novel alpha-lipoproteinemia and a useful model for investigating metabolism of high density lipoprotein.

Tc-99m DTPA used as reference imaging to evaluate the distribution of other tumor-seeking tracers in tumors associated with neurofibromatosis.

Two patients with multiple benign and malignant tumors associated with neurofibromatosis underwent radionuclide imaging with Tc-99m DTPA, Tl-201, and Ga-67. In these patients, Tc-99m DTPA accumulated intensively in both the benign and malignant tumors and localized and defined the extent of every tumor. In contrast, Ga-67 and Tl-201 uptake was seen only in focal areas of tumor where there was malignant transformation or at sites that showed progressive tumor growth. Tc-99m DTPA imaging accurately demonstrated areas of neoplastic involvement and identified the areas that would be seen with the other two tracers in individual tumors. Tc-99m DTPA may not always be used for the differential diagnosis of malignant and benign tumors of neurofibromatosis, but it can provide a reference pattern for imaging to evaluate accurately the distribution of Tl-201 and Ga-67 by mapping out the anatomic extent of these tumors.

Involvement of bacterial antigens in immunoglobulin A nephropathy.

To investigate the involvement of bacterial antigens in Immunoglobulin A (IgA) nephropathy, we measured IgA, IgG and IgM antibodies to gram-negative Escherichia coli (E.coli) and Haemophilus influenzae (H.influenzae) by ELISA in 24 patients (11 males and 13 females) with IgA nephropathy and 22 normal controls (11 males and 11 females). The titers of IgA and IgM antibodies for E.coli and H.influenzae were significantly higher in the IgA nephropathy group than in the controls. In addition, IgA and IgM antibody titers for E.coli and H.influenzae showed a significant positive correlation with serum IgA and IgM levels. These findings suggest that subclinical infection by these bacteria stimulates IgA production and that this may be a factor in the development and progression of IgA nephropathy.

Serum soluble interleukin 2 receptor in patients with IgA nephropathy.

Serum soluble interleukin 2 receptor (IL-2 R) was determined by the ELISA method in 29 cases of IgA nephropathy and 50 healthy controls. The results showed that the value in IgA nephropathy cases was significantly higher than that in healthy controls. Furthermore, among the cases of IgA nephropathy, the value was significantly higher in the groups with hypertension, elevated serum IgA and depressed creatinine clearance than in that of the corresponding controls. These findings suggest that serum soluble IL-2 R can serve as a prognostic index of IgA nephropathy.

An immunohistochemical study of extracellular matrix components and integrins in human glomerular diseases.

The localization of extracellular matrix components and their cell surface receptors (integrins) was studied in 130 subjects in order to clarify their participation in the progression and aggravation of various types of nephritis. Included in the study were 2 normal subjects, 14 patients with minimal change disease, 2 patients with minimal change nephrotic syndrome, 65 patients with IgA nephropathy, 18 patients with mesangial proliferative glomerulonephritis, 15 patients with membranous glomerulonephritis, 5 patients with membranoproliferative glomerulonephritis and 9 patients with systemic lupus erythematosus (SLE). The distribution of fibronectin (FN), vitronectin (VN), laminin (LN), heparan sulfate proteoglycan (HSPG), type III, IV, V, VI collagen, fibronectin receptor (FNR) and vitronectin receptor (VNR) in the glomerulus was studied employing the indirect immunoperoxidase method. FN, LN, type IV, V and VI collagen, FNR and VNR were found to be distributed in the expanded mesangial region in IgA nephropathy, mesangial proliferative glomerulonephritis and membranoproliferative glomerulonephritis. Deposition of VN was observed in some of the patients. In membranous glomerulonephritis and membranoproliferative glomerulonephritis, the distribution of FN, LN, type IV collagen, FNR and VNR was increased in the thickened loop wall and VN deposition was also observed. Quantitative and functional changes in the extracellular matrix and integrins, therefore, appear to participate in the progression and aggravation of glomerulonephritis.

In vitro immunoglobulin production by peripheral blood mononuclear cells as a prognostic factor in IgA nephropathy.

To determine whether immune system disorders are involved in the exacerbation of IgA nephropathy, the immunoglobulin production of peripheral blood mononuclear cells obtained from 45 IgA nephropathy patients was measured and then compared with that of healthy individuals. The level of IgA production was classified into an elevated group and a non-elevated group and comparisons were made with various clinical factors considered to be related to exacerbation of this disease. The results indicated that although there was no significant difference in immunoglobulin production of the peripheral mononuclear cells between IgA nephropathy cases and healthy individuals in the group not stimulated with pokeweed mitogen (PWM), the group stimulated with PWM revealed a production of IgA, IgG, and IgM which was significantly elevated (P less than 0.01). Also, within the group stimulated with PWM, hypertension, severe proteinuria and microscopic hematuria, elevated BUN and serum creatinine values, decreased 15-min PSP and creatinine clearance values, severe histological damage, and severe IgA deposition were observed more in the elevated IgA production group than in the non-elevated group. These findings suggest that an elevated IgA production plays an important role in the excerbation of this disease.