In vitro metabolism of fenthion and fenthion sulfoxide by liver preparations of sea bream, goldfish, and rats.

The in vitro metabolism of fenthion and its sulfoxide (fenthion sulfoxide) in sea bream (Pagrus major) and goldfish (Carassius auratus) was investigated and compared with that in rats. Fenthion was oxidized to fenthion sulfoxide and the oxon derivative, but not to its sulfone, in the presence of NADPH by liver microsomes of sea bream, goldfish, and rats. These liver microsomal activities of the fish were lower than those of rats but were of the same order of magnitude. The NADPH-linked oxon- and sulfoxide-forming activities of liver microsomes of the fish and rats were inhibited by SKF 525-A, metyrapone, alpha-naphthoflavone, and carbon monoxide. The oxidizing activity to fenthion sulfoxide was also inhibited by alpha-naphthylthiourea. Several cytochrome P450 isoforms and flavin-containing monooxygenase 1 exhibited these oxidase activities. Fenthion sulfoxide was reduced to fenthion with liver cytosol of the fish and rats upon addition of 2-hydroxypyrimidine, N(1)-methylnicotinamide, or butyraldehyde, each of which is an electron donor of aldehyde oxidase, under anaerobic conditions. The activity was inhibited by menadione, beta-estradiol, and chlorpromazine, which are inhibitors of aldehyde oxidase. The activities in the fish livers were similar to those of rat liver. Aldehyde oxidase purified from the livers of sea bream and rats exhibited the reducing activity. Thus, fenthion and fenthion sulfoxide are interconvertible in fish and rats through the activities of cytochrome P450, flavin-containing monooxygenase, and aldehyde oxidase.     

Impaired stem cell function of CD34+ cells selected by two different immunomagnetic beads systems.

We have been investigating the hematopoietic stem cell (HSC) activity of peripheral blood-derived CD34(+) cells selected by two different laboratory immunomagnetic beads systems (MiniMACS and Isolex 50). In this study, the quality of purified CD34(+) cells was directly compared using clonal cell culture, a cobblestone area-forming cell (CAFC) assay, and an in vivo severe combined immunodeficiency (SCID)-repopulating cell (SRC) assay. It was found that CD34(+) cells selected by these two immunomagnetic methods showed a reduced yield of colony-forming cells and CAFCs compared with cells enriched by the StemSep device (a negative selection method). However, these CD34(+) cells still showed significant SRC activity, including multilineage lymphomyeloid reconstitution. The percentage of human CD45(+) cells in murine bone marrow after transplanting 5 x 10(5) CD34(+) cells selected by the Isolex 50 was significantly lower than after transplanting cells selected by the MiniMACS or the StemSep. Our findings clearly demonstrated that CD34(+) cells selected by the MiniMACS system had superior HSC functions, including SRC activity, compared with cells separated by the Isolex 50 system. More detailed functional analysis of immunomagnetically separated CD34(+) cells may provide useful knowledge for basic research on HSCs as well as for clinical HSC transplantation.     

Prediction of nodal metastasis by comparative genomic hybridization in biopsy specimens from patients with superficial esophageal squamous cell carcinoma.

PURPOSE: Selection of appropriate protocols for treatment of superficial esophageal squamous cell carcinoma (SESCC) is dependent on lymph node metastasis status. Therefore, it is important to know whether lymph node metastasis is present before treatment. EXPERIMENTAL DESIGN: In this study, we examined the relation between DNA sequence copy number aberrations detected by comparative genomic hybridization and lymph node metastasis in 26 surgically resected SESCCs (training samples). We then assessed whether the genetic information is predictive for nodal status in biopsy specimens from eight newly enrolled patients with SESCC (blinded samples). RESULTS: Pathological examination revealed that 17 of 26 training samples (65.4%) did not have associated lymph node metastasis. Gains of 8q24 and/or 20q12-qter were observed in 12, including all (nine of nine) with nodal metastasis. Fourteen training samples did not have gain of either 8q24 or 20q12-qter. Of the blinded samples, two showed no gain of 8q24 or 20q12-qter, and as anticipated the postoperative pathological examination revealed no nodal metastasis. The remaining six blinded samples had gains of 8q24 and/or 20q12-qter, and lymph node metastasis was detected by postoperative examination in four of these tumors. CONCLUSIONS: Absence of gains of 8q24 and/or 20q12-qter appears to be associated with absence of lymph node metastasis in patients with SESCC; therefore, less invasive surgery can be chosen.     

Ischial hypoplasia, tibial hypoplasia and facial abnormalities: a new syndrome?

A child with facial abnormalities, short stature and a variety of skeletal alterations is reported. The facial abnormalities comprised low-set ears, short nose with a long philtrum, micrognathia and cleft palate. The skeletal alterations included ischial hypoplasia, malformations of the cervical spine, hypoplasia of the lesser trochanters, tibial hypoplasia with bowing of the lower legs, tibio-fibular diastasis with malformed distal tibial epiphyses, clubfeet and brachymesophalangy. The constellation of clinical and radiological findings in the present patient do not fit any known malformation syndrome. Received: 14 February 1998 Accepted: 15 June 1998     

Increased Sensitivity to Long-term 5-Fluorouracil Exposure of Human Colon Cancer HT-29 Cells Resistant to Short-term Exposure

A 5-fluorouracil (5-FU)-resistant subline of human colon cancer HT-29 cells was developed by repeated 1-h exposure in vitro to 5-FU. This subline (HT-29/5-FU/S) had 8-fold resistance to 5-FU in a 1-h exposure assay. However, it had rather increased sensitivity to 5-FU when assayed after a continuous 96-h exposure to it. Significantly less 5-fluorouridine-5'-triphosphate was produced in the resistant cells, leading to a lower level of 5-FU incorporation into the cellular RNA. The reduced activity of orotate phosphoribosyltransferase might explain these results. In contrast, the HT-29/5-FU/S cells were more sensitive to the inhibition of in situ thymidylate synthase (TS) by 5-FU than were the parent cells. The lower in situ TS activity may have made HT-29/5-FU/S cells more sensitive to TS inhibition by 5-FU as compared with the parent cells. The fact that HT-29/5-FU/S was more resistant to short-term 5-FU exposure but more sensitive to long-term exposure than the parent line confirmed the existence of different modes of action of 5-FU, depending on the exposure time.     

Pharmacokinetic study of low- versus high-dose medroxyprogesterone acetate (MPA) in women

The present study was conducted to compare the pharmacokinetics (PK) of low-dose versus high-dose medroxyprogesterone (MPA) as a once-daily oral administration. Of 32 patients, all women, enrolled in this PK study, 18 received 600 mg MPA daily and 14 received 1200 mg daily. Detailed PK data were obtained on day 1 and after more than 4 weeks of MPA treatment. In addition, multiple data for the minimum steady-state concentration (Css min) were analyzed. The MPA serum concentrations were measured by high-performance liquid chromatography. Wide interpatient variability was found in the PK parameters obtained both on day 1 and after more than 4 weeks. There were no clear relationships between the oral dose and the MPA peak concentration (Cmax), area under the time versus concentration curve (AUC), or mean Css min. Weight gains of 10% or more were demonstrated more frequently in the high-dose group (P<0.01). Liver dysfunction (n=5) did not influence the PK of MPA. Five patients demonstrated extremely low AUC and Cmax (<10 ng/ml) values on day 1. Phenobarbital, dexamethasone and betamethasone were being taken concomitantly with the MPA each by one patient. The serum MPA concentrations were markedly increased after the discontinuation of phenobarbital in that patient, suggesting a drug interaction. At present we cannot recommend the high dose of MPA, except in clinical studies, from a PK or a pharmacodynamic points of view. Received: 2 May 1997 / Accepted: 13 October 1997     

Analgesic mechanisms of ketamine in the presence and absence of peripheral inflammation

BACKGROUND: The studies on the mechanisms of ketamine antinociception have led to conflicting results. In this study, the authors investigated the contribution of supraspinal monoaminergic descending inhibitory system to ketamine analgesia for acute nociception and inflammation-induced hyperalgesia. METHODS: Male Sprague-Dawley rats were used. The paw withdrawal latencies to radiant heat stimuli were measured to assess the thermal nociceptive threshold. The analgesic effects of intrathecal or intraperitoneal ketamine were examined in the rats that received unilateral intraplantar carrageenan and in those that were untreated. In addition, it was examined whether pretreatment with intrathecal yohimbine or methysergide inhibited the analgesic effects of ketamine. Using an intrathecal microdialysis method, noradrenaline and 5-hydroxytryptamine concentrations in lumbar cerebrospinal fluid were measured after intraperitoneal ketamine in both saline- and carrageenan-treated rats. RESULTS: In the untreated rats, intraperitoneal but not intrathecal ketamine produced antinociceptive effects in a dose-dependent manner. Pretreatment with intrathecal yohimbine or methysergide inhibited these antinociceptive effects. Intraplantar carrageenan significantly reduced paw withdrawal latencies on the injected paw but not on the contralateral paw. Both intraperitoneal and intrathecal ketamine reversed the shortened paw withdrawal latencies on the injected side in a dose-dependent manner without any effects on the contralateral side. Neither yohimbine nor methysergide inhibited these antihyperalgesic effects. In analyses of monoamines, the magnitude of increase in monoamines after intraperitoneal ketamine was significantly smaller in the carrageenan-treated rats than in the saline-treated rats. CONCLUSION: These results demonstrated that ketamine produced antinociceptive effects through an activation of the monoaminergic descending inhibitory system, whereas, in a unilateral peripheral inflammation-induced hyperalgesic state, the monoaminergic system did not contribute to the antihyperalgesic effects of ketamine. The mechanisms of the antinociceptive and antihyperalgesic properties of ketamine are different.     

Branched Chain Amino Acids Promote ATP Production Via Translocation of Glucose Transporters

PurposeWe have previously shown that maintenance of ATP levels is a promising strategy for preventing neuronal cell death, and that branched chain amino acids (BCAAs) enhanced cellular ATP levels in cultured cells and antagonized cell death. BCAAs attenuated photoreceptor degeneration and retinal ganglion cell death in rodent models of retinal degeneration or glaucoma. This study aimed to elucidate the mechanisms through which BCAAs enhance ATP production.MethodsIntracellular ATP concentration was measured in HeLa cells under glycolysis and citric acid cycle inhibited conditions. Next, glucose uptake was quantified in HeLa cells and in 661W retinal photoreceptor-derived cells under glycolysis inhibition, endoplasmic reticulum stress, and glucose transporters (GLUTs) inhibited conditions, by measuring the fluorescence of fluorescently labeled deoxy-glucose analog using flow cytometry. Then, the intracellular behavior of GLUT1 and GLUT3 were observed in HeLa or 661W cells transfected with enhanced green fluorescent protein-GLUTs.ResultsBCAAs recovered intracellular ATP levels during glycolysis inhibition and during citric acid cycle inhibition. BCAAs significantly increased glucose uptake and recovered decreased glucose uptake induced by endoplasmic reticulum stress or glycolysis inhibition. However, BCAAs were unable to increase intracellular ATP levels or glucose uptake when GLUTs were inhibited. Fluorescence microscopy revealed that supplementation of BCAAs enhanced the translocation of GLUTs proteins to the plasma membrane over time.ConclusionsBCAAs increase ATP production by promoting glucose uptake through promotion of glucose transporters translocation to the plasma membrane. These results may help expand the clinical application of BCAAs in retinal neurodegenerative diseases, such as glaucoma and retinal degeneration.     

Constipation Is a Frequent Problem Associated with Vascular Complications in Patients with Type 2 Diabetes: A Cross-sectional Study

Objective Diabetes is recognized as an underlying disease of constipation. However, the prevalence of constipation varies according to the diagnostic criteria applied. We investigated the prevalence of constipation based on the new guideline for constipation in Japanese patients with type 2 diabetes and examined the relationship with the clinical background, including diabetic vascular complications. Methods Questionnaire surveys including items concerning the diagnosis and treatment status of constipation were administered to 410 patients with type 2 diabetes. Results Although 29% of the patients considered that they had experienced constipation (self-judged), only 14% had consulted a physician about constipation. The prevalence of chronic constipation based on the guideline was 26%. After including laxative users, constipation was finally found in 36%. Despite the use of laxatives (n=81), 51% of the patients were still diagnosed with chronic constipation. Patients with constipation (chronic constipation or laxative use) were significantly older and had a longer duration of diabetes than those without constipation. The body mass index (BMI) of patients with constipation (24.9±3.8 kg/m²) was significantly lower than that of those without constipation (26.3±4.6 kg/m²). Diabetic neuropathy (49% vs. 32%) and coronary heart disease (CHD) (27% vs. 13%) were significantly more frequent in the patients with constipation than in those without constipation. A multivariate logistic regression analysis revealed that gender, BMI, diabetic neuropathy, insulin use, and CHD were significantly associated with constipation. Conclusion An accurate diagnosis of constipation is desirable in patients with type 2 diabetes because constipation is independently associated with CHD.     

Living and working environments are important determinants of glycemic control in patients with diabetes during the COVID‐19 pandemic: A retrospective observational study

AimTo investigate (1) the association of lifestyle changes and living and working conditions with glycemic control and (2) whether treatment was intensified appropriately in patients with diabetes under the first COVID‐19 state of emergency in Japan.Materials and MethodsA total of 321 participants were included. Participants completed a questionnaire regarding lifestyle changes, including diet, physical activity, and living and working conditions during the COVID‐19 pandemic. The change in hemoglobin A1c (HbA1c) levels was estimated before (June 1, 2019 to August 31, 2019) and during (June 1, 2020 to August 31, 2020) the pandemic. Factors associated with changes in HbA1c levels were examined by multiple linear regression analysis. The proportion of patients who received treatment intensification for diabetes was compared between before and during the pandemic.ResultsThere was no significant change in HbA1c levels before the pandemic and during the pandemic (7.13 ± 0.98% vs 7.18 ± 1.01%, P = 0.186). Teleworking (estimate 0.206, P = 0.004) and living with a dog (estimate −0.149, P = 0.038) were significantly associated with changes in HbA1c levels after adjusting for covariates. There was no significant difference in the proportion of patients who received treatment intensification for diabetes during the pandemic and before the pandemic in either the elderly or non‐elderly patients.ConclusionsOverall glycemic control did not worsen during the pandemic. Nonetheless, environmental factors, including telework, were found to influence glycemic control in patients with diabetes. Further studies are needed to clarify whether the COVID‐19 pandemic could affect treatment intensification for diabetes.