Chronic Treatment with Metformin Has No Disrupting Effect on the Hepatic Circadian Clock in Mice.

Background and Objectives: The antidiabetic agent metformin is known to activate AMP-activated protein kinase (AMPK) in various tissues. Because AMPK can modulate intracellular circadian clocks through regulating the stability of clock components, a single dose of metformin has been reported to affect circadian clocks in the peripheral tissues. In this study, therefore, we investigated whether chronic treatment with metformin causes the impairment of circadian clocks, especially if given at an inappropriate time. Materials and Methods: Non-diabetic C57BL/6J mice were allowed access to food only during 4 h at the beginning of the dark period, and repeatedly i.p. injected with a nearly maximum non-toxic dose of metformin, once daily either at 4 h after the beginning of the dark period or at the beginning of the light period. Diabetic ob/ob mice were given free access to food and treated with metformin in drinking water. Results: Under the controlled feeding regimen, 8-day treatment with metformin did not alter the mRNA expression rhythms of clock genes in both liver and adipose tissue of C57BL/6J mice, regardless of dosing time. In addition, chronic treatment with metformin for 2 weeks affected hepatic AMPK activation rhythm but did not disrupt the circadian clocks in the liver and adipose tissues of the ob/ob mice. Conclusions: These results mitigate concerns that treatment with metformin impairs peripheral circadian clocks, although confirmation is needed in humans.

Influence of the aldehyde dehydrogenase 2 polymorphism on the vasodilatory effect of nitroglycerin in infants with congenital heart disease and pulmonary arterial hypertension.

PURPOSE: The influence of the aldehyde dehydrogenase 2 (ALDH2) gene polymorphism on the pharmacokinetics and haemodynamics of nitroglycerin (GTN) was determined in human subjects. METHODS: Eighteen infants (nine each with and without ALDH2 gene polymorphism) with congenital heart disease and pulmonary arterial hypertension participated in this study. GTN treatment started at a dose of 2 µg/kg/min, and the dose was escalated by 1-2 µg/kg/min until pulmonary vascular resistance (PVR) was reduced by more than 30%. The plasma GTN concentration and PVR were measured at the end of each infusion period. RESULTS: Plasma GTN concentrations were significantly higher in patients with the ALDH2 gene polymorphism than in those without the polymorphism. Conversely, the reduction in PVR was smaller in patients with the ALDH2 gene polymorphism than in those without. CONCLUSIONS: These data suggest that the ALDH2 gene polymorphism influences the pharmacokinetics and haemodynamics of GTN in human subjects.

Influence of renal ischaemia-reperfusion injury on renal neutrophil gelatinase-associated lipocalin receptor (24p3R) in rats.

The neutrophil gelatinase-associated lipocalin (NGAL) receptor (24p3R) is expressed in distal nephron and contributes to the endocytosis of NGAL in urine. This study was undertaken to evaluate an influence of renal ischaemia-reperfusion injury on 24p3R. Unilateral renal pedicle was clamped for 0, 10, 20, 30, or 45 minutes in male Wistar rats. Urine was collected for 24 hours after reperfusion, and ischaemic kidney and blood sample were obtained. Apparent histological injury in the ischaemic kidney was detected in the 30 and 45 minutes-treated groups. Urinary NGAL excretion elevated in rats with renal ischaemia for more than 20 minutes, while serum creatinine increased in rats for more than 30 minutes of ischaemia. Renal protein expression of NGAL did not significantly change. Renal mRNA expressions of megalin and cubilin, which are expressed at renal proximal tubules and uptake NGAL, decreased in animals with renal ischaemia for more than 20 minutes. Renal protein expression of 24p3R, which is expressed at renal distal tubules and uptake NGAL, decreased in rats with renal ischaemia for 45 min. This study showed for the first time that renal 24p3R decreased in response to renal ischaemia. As relatively longer renal ischaemia (45 minutes) decreased renal 24p3R protein and increased urinary NGAL excretion, the down-regulation of 24p3R protein might contribute to the elevated urinary excretion of NGAL in rats with unilateral ischaemia-reperfusion injury.

Increased tacrolimus blood concentration by Beni-Madonna - a new hybrid citrus cultivar categorized as 'Tangor', in a liver transplant patient: likely furanocoumarin-mediated inhibition of CYP3A4 or P-glycoprotein.

ADVERSE EVENT: A drug interaction leading to greater exposure to tacrolimus. DRUG IMPLICATED: Tacrolimus and Beni-Madonna (a new cultivar citrus categorized as 'Tangor'). THE PATIENT: A 9-month-old girl with biliary atresia (body weight, 7.5 kg) taking tacrolimus after liver transplantation. EVIDENCE THAT LINKS THE DRUG TO THE EVENT: The time course was consistent with the appearance of the interaction, which was confirmed by an increase in the blood concentration of tacrolimus. Dihydroxybergamottin was detected in peel of Beni-Madonna and in peel and fruit pulp of grapefruit. MANAGEMENT: Avoiding Beni-Madonna intake. MECHANISM: Inhibition of activity of CYP3A4, P-glycoprotein, or both, by Beni-Madonna. IMPLICATION FOR THERAPY: Clinicians should be aware of this potential interaction, and patients taking drugs such as tacrolimus (the kinetics of which are affected by grapefruit juice) should avoid Beni-Madonna intake. HYPOTHESIS TO BE TESTED: Further study is required to determine if other Citrus species categorized as Tangor contain furanocoumarins.

Dosing-time-dependent effect of rivaroxaban on coagulation activity in rats.

The anticoagulant effect of rivaroxaban, a direct inhibitor of activated factor X (FX), might be influenced by its dosing time because the activity of the coagulofibrinolytic system exhibits daily rhythmicity. In rats, FX activity follows a 24-h rhythm with a peak in the middle of the light phase and a trough at the beginning of the dark phase. Consistent with these findings, a single dose of rivaroxaban had a stronger inhibitory effect on FX activity after dosing at the beginning of the light phase than after dosing at the beginning of the dark phase. A similar chronopharmacological effect was seen in a quantitative model of venous stasis thrombosis. In comparison, the dosing time had minimal influence on the pharmacokinetics of rivaroxaban. These data indicate that the anticoagulant effect of rivaroxaban is influenced by the dosing time. Further studies should confirm this finding in a clinical setting.

Effect of a dosing-time on quetiapine-induced acute hyperglycemia in mice.

Although rare, second-generation antipsychotic drugs cause severe hyperglycemia within several days after the initiation of therapy. Because glucose tolerance exhibits circadian rhythmicity, we evaluated an effect of a dosing-time on quetiapine-induced acute hyperglycemia in mice. A single intraperitoneal dose of quetiapine dosing-time-independently induced insulin resistance in fasted C57BL/6J mice. However, acute hyperglycemic effect was detected only after dosing of the drug at the beginning of an active phase. Under the conditions in which hepatic glucose production was stimulated by pyruvate administration, hyperglycemic effect of quetiapine was dosing-time-independently observed. In addition, the dosing-time-dependent hyperglycemic effect of quetiapine disappeared in the liver-specific circadian clock-disrupted mice in which circadian rhythmicity in hepatic glucose production is deranged. Furthermore, the dosing-time had little impact on the pharmacokinetics of quetiapine in normal mice. These results suggest that quetiapine acutely causes hyperglycemia only when hepatic glucose production elevates. Therefore, quetiapine therapy with once daily dosing at a rest phase might be safer than that at an active phase. Further studies are needed to confirm the hypothesis.

Association between ABCG2 and SLCO1B1 polymorphisms and adverse drug reactions to regorafenib: a preliminary study
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OBJECTIVE: Due to the occurrence of severe adverse drug reactions to regorafenib, a drug used in cancer therapy, the identification of a predictive marker(s) is needed to increase the therapeutic applicability of this compound. We therefore investigated whether polymorphisms in the ABCG2 and SLCO1B genes are associated with adverse drug reactions to regorafenib. METHODS: For these analyses, 37 Japanese cancer patients were treated with regorafenib, genotyped for polymorphisms in ABCG2 and SLCO1B, and evaluated for drug-related adverse drug reactions. RESULTS: There was no association between the ABCG2 421C>A variant and adverse drug reactions to regorafenib. After treatment, the incidences of increased aspartate aminotransferase (AST) and alanine aminotransferase (ALT) as well as increased total bilirubin (grade ≥ 2) were 8%, 4%, and 12%, and 42%, 25%, and 25% among SLCO1B1*1b carriers and non-carriers, respectively. There were no significant associations between elevated ALT and bilirubin and the SLCO1B1*1b allele. However, there were significantly lower incidences of increased AST (8% vs. 42%) and anemia (16% vs. 50%) in SLCO1B1*1b carriers than in non-carriers. CONCLUSIONS: The absence of SLCO1B1*1b allele appears to be associated with the development of adverse drug reactions to regorafenib; however, further studies involving larger test groups and other populations are needed to confirm these findings.
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Prediction of the therapeutic response to methotrexate at 24 weeks by methotrexate-polyglutamates concentration in erythrocytes at 8 weeks in patients with rheumatoid arthritis.

OBJECTIVES: The objective of this study is to evaluate the pharmacokinetics and pharmacodynamics of methotrexate-polyglutamates (MTX-PGs) in erythrocytes in patients with rheumatoid arthritis and correlate them with the efficacy. METHODS: MTX-PG concentrations in erythrocytes were measured in 42 MTX-naïve patients repeatedly for 24 weeks by high-performance liquid chromatography. In 56 patients receiving stable MTX doses for at least 12 weeks, the correlation between MTX doses and MTX-PG concentrations was examined. The efficacy was measured by the change of DAS28CRP (ΔDAS28CRP). RESULTS: There were moderate correlations between MTX dose and MTX-PG 3, 4, and 5. At 24 weeks, MTX-PG2, 3, 4, and 1-5 were higher in patients with ΔDAS28CRP >1.2 than in those with ≤1.2. The cutoff value of MTX-PG1-5 to discriminate ΔDAS28CRP >1.2 from ≤1.2 at 24 weeks was 68.7 nM. Among 20 patients with MTX-PG1-5 > 50.6 nM at 8 weeks, seven already improved at 8 weeks and additional 11 improved at 24 weeks (p < 0.001). On the contrary, among the nine patients with MTX-PG1-5 ≤ 50.6 nM at 8 weeks, none improved at 8 weeks and only one improved at 24 weeks (p = 0.500). CONCLUSIONS: Erythrocyte MTX-PGs might be a potential indicator and predictor of MTX efficacy.

Effects of telmisartan and olmesartan on insulin sensitivity and renal function in spontaneously hypertensive rats fed a high fat diet.

Although telmisartan, an angiotensin II receptor blocker (ARB), has an agonistic action for proliferator-activated receptor (PPAR)-γ in vitro, it remains to be determined whether telmisartan exerts such an action in vivo using a non-toxic dose (<5 mg/kg in rats). To address the issue, telmisartan (2 mg/kg) and olmesartan (2 mg/kg), another ARB without PPAR-γ agonistic action, were given to spontaneously hypertensive rats (SHR) fed a high fat diet (HFD). HFD decreased plasma adiponectin, and caused insulin resistance, hypertriglyceridemia and renal damage, which were improved by ARBs. Protective effects of telmisartan and olmesartan did not significantly differ. In addition, in vitro study showed that 1 µM of telmisartan did not elevate the mRNA expression of adipose protein 2, which is a PPAR-γ-stimulated adipogenic marker gene, in preadipocytes with 3% albumin. To obtain 1 µM of plasma concentration, oral dose of telmisartan was calculated to be 6 mg/kg, which indicates that PPAR-γ agonistic action is negligible with a non-toxic dose of telmisartan (<5 mg/kg) in rats. This study showed that 2 mg/kg of telmisartan and olmesartan ameliorated insulin resistance, hypertriglyceridemia and renal damage in SHR fed a HFD. As beneficial effects of telmisartan and olmesartan did not significantly differ, these were mediated through the PPAR-γ-independent actions.

Influence of genetic polymorphisms of multidrug and toxin extrusion protein 1 on its mRNA expression in peripheral blood cells.

This study aimed to determine the effect of multidrug and toxin extrusion protein 1 (MATE1) genetic variants on its transcript expression in peripheral blood cells. Consistent with previous in vitro findings, MATE1 mRNA levels were significantly higher in subjects carrying rs2453579, but not rs2252281, compared to those without either of these promoter variants. In addition, the mRNA levels did not differ between subjects with both variants and those with neither allele. Thus, this study reveals that the influence of MATE1 genetic variants on its mRNA expression can be detected in vivo using peripheral blood.

Different chronotherapeutic effects of valsartan and olmesartan in non-dipper hypertensive patients during valsartan treatment at morning.

This study was undertaken to evaluate the differences in chronotherapeutic effects of angiotensin-II receptor blockers, valsartan and olmesartan in hypertensive patients with non-dipper blood pressure (BP) pattern during valsartan at morning. Ninety four patients were enrolled, and 40 patients were judged to be non-dippers. In these patients, same dose of valsartan was changed to evening (Val-E, n = 12), or olmesartan (equivalent dose of valsartan) was given at morning (Olm-M, n = 13) or evening (Olm-E, n = 15) for 4 months. BP decreased during sleep and increased during waking hours in Val-E group. In Olm-M and Olm-E groups, BP decreased during sleep and waking hours. Percent reduction in BP at night-time compared to BP at waking hours significantly increased after changing the dose regimen in each group. Serum creatinine decreased and estimated glomerular filtration rate (eGFR) elevated in Olm-M and Olm-E, but not Val-E groups. Positive correlation between systolic BP (SBP) during sleep and serum creatinine, and negative correlation between SBP during sleep and eGFR were detected. These data suggest that dipper BP pattern could be obtained by chronotherapeutic approach using valsartan and olmesartan in non-dipper patients with valsartan at morning. Morning and evening olmesartan, but not evening valsartan improved renal function in these patients.

Radiofrequency ablation combined with chemolipiodolization in a porcine liver: Comparison of the pharmacokinetic analysis with cisplatin powder and miriplatin.

AIM: To compare the pharmacokinetics of radiofrequency (RF) ablation with chemolipiodolization using cisplatin (CDDP) powder and miriplatin (MPT) in a porcine liver. METHODS: Twelve pigs were divided equally into four groups. After each CDDP powder-lipiodol suspension (n = 6; groups A and B) or MPT-lipiodol suspension (n = 6; groups C and D) was injected into the lateral left artery, one RF ablation was performed at the lateral left lobe of each pig. Six pigs (groups A and C) were killed on the same day as treatment, whereas the other pigs (groups B and D) were killed 7 days after the treatment. The platinum concentrations in venous blood were assayed at 15, 60 and 120 min, and 7 days after treatment. The platinum concentrations in the ablated area and the surrounding liver were also examined. RESULTS: Plasma platinum concentrations of the CDDP group peaked at 15 min, and then gradually diminished over time (µg units), while plasma platinum levels in the MPT group gradually increased over time (ng units). Liver tissue platinum concentrations of the CDDP group were significantly lower in non-ablative areas than in ablated areas at days 0 and 7, while liver concentrations of the MPT group were significantly higher in non-ablative areas than in ablated areas at day 7. CONCLUSION: MPT may be a suitable chemotherapeutic agent to stagnate platinum in the surrounding liver.

Space environment of an asteroid preserved on micrograins returned by the Hayabusa spacecraft.

Records of micrometeorite collisions at down to submicron scales were discovered on dust grains recovered from near-Earth asteroid 25143 (Itokawa). Because the grains were sampled from very near the surface of the asteroid, by the Hayabusa spacecraft, their surfaces reflect the low-gravity space environment influencing the physical nature of the asteroid exterior. The space environment was examined by description of grain surfaces and asteroidal scenes were reconstructed. Chemical and O isotope compositions of five lithic grains, with diameters near 50 µm, indicate that the uppermost layer of the rubble-pile-textured Itokawa is largely composed of equilibrated LL-ordinary-chondrite-like material with superimposed effects of collisions. The surfaces of the grains are dominated by fractures, and the fracture planes contain not only sub-µm-sized craters but also a large number of sub-µm- to several-µm-sized adhered particles, some of the latter composed of glass. The size distribution and chemical compositions of the adhered particles, together with the occurrences of the sub-µm-sized craters, suggest formation by hypervelocity collisions of micrometeorites at down to nm scales, a process expected in the physically hostile environment at an asteroid's surface. We describe impact-related phenomena, ranging in scale from 10(-9) to 10(4) meters, demonstrating the central role played by impact processes in the long-term evolution of planetary bodies. Impact appears to be an important process shaping the exteriors of not only large planetary bodies, such as the moon, but also low-gravity bodies such as asteroids.

[Chronotherapy of hypertension].

Blunted blood pressure (BP) decline during night-time leads to the higher risk of cardiovascular events in hypertensive patients. Therefore, to improve the prognosis of the patients, it is essential for properly controlling BP during night-time sleep as well as day-time activity. In addition, the dosing-time dependent changes are observed in the pharmacokinetics and pharmacodynamics of some antihypertensive drugs. Thus, the efficacy and toxicity of drugs might be affected by their dosing-time. Chronotherapy is the therapeutic application of chronopharmacology and chronotoxicology to enhance the effectiveness and tolerance of drugs by determining optical dosing-time of drugs from a circadian perspective. In this article, chronotherapy of angiotensin-converting enzyme inhibitor, Ca channel blocker, α1 blocker, diuretics and angiotensin II receptor blocker will be discussed.

Selenoprotein P as a diabetes-associated hepatokine that impairs angiogenesis by inducing VEGF resistance in vascular endothelial cells.

AIMS/HYPOTHESIS: Impaired angiogenesis induced by vascular endothelial growth factor (VEGF) resistance is a hallmark of vascular complications in type 2 diabetes; however, its molecular mechanism is not fully understood. We have previously identified selenoprotein P (SeP, encoded by the SEPP1 gene in humans) as a liver-derived secretory protein that induces insulin resistance. Levels of serum SeP and hepatic expression of SEPP1 are elevated in type 2 diabetes. Here, we investigated the effects of SeP on VEGF signalling and angiogenesis. METHODS: We assessed the action of glucose on Sepp1 expression in cultured hepatocytes. We examined the actions of SeP on VEGF signalling and VEGF-induced angiogenesis in HUVECs. We assessed wound healing in mice with hepatic SeP overexpression or SeP deletion. The blood flow recovery after ischaemia was also examined by using hindlimb ischaemia model with Sepp1-heterozygous-knockout mice. RESULTS: Treatment with glucose increased gene expression and transcriptional activity for Sepp1 in H4IIEC hepatocytes. Physiological concentrations of SeP inhibited VEGF-stimulated cell proliferation, tubule formation and migration in HUVECs. SeP suppressed VEGF-induced reactive oxygen species (ROS) generation and phosphorylation of VEGF receptor 2 (VEGFR2) and extracellular signal-regulated kinase 1/2 (ERK1/2) in HUVECs. Wound closure was impaired in the mice overexpressing Sepp1, whereas it was improved in SeP (-/-)mice. SeP (+/-)mice showed an increase in blood flow recovery and vascular endothelial cells after hindlimb ischaemia. CONCLUSIONS/INTERPRETATION: The hepatokine SeP may be a novel therapeutic target for impaired angiogenesis in type 2 diabetes.

Temsirolimus induces surfactant lipid accumulation and lung inflammation in mice.

Interstitial lung disease (ILD) is a well-known adverse effect of mammalian target of rapamycin (mTOR) inhibitors. However, it remains unknown how lung toxicities are induced by mTOR inhibitors. Here, we constructed a mouse model of mTOR inhibitor-induced ILD using temsirolimus and examined the pathogenesis of the disease. Male ICR mice were treated with an intraperitoneal injection of different doses of temsirolimus (3 or 30 mg·kg(-1)·wk(-1)) or vehicle. Temsirolimus treatment increased capillary-alveolar permeability and induced neutrophil infiltration and fibrinous exudate into the alveolar space, indicating alveolar epithelial and/or endothelial injury. It also induced macrophage depletion and the accumulation of excessive surfactant phospholipids and cholesterols. Alveolar macrophage depletion is thought to cause surfactant lipid accumulation. To further examine whether temsirolimus has cytotoxic and/or cytostatic effects on alveolar macrophages and alveolar epithelial cells, we performed in vitro experiments. Temsirolimus inhibited cell proliferation and viability in both alveolar macrophage and alveolar epithelial cells. Temsirolimus treatment caused some signs of pulmonary inflammation, including upregulated expression of several proinflammatory cytokines in both bronchoalveolar lavage cells and lung homogenates, and an increase in lymphocytes in the bronchoalveolar lavage fluid. These findings indicate that temsirolimus has the potential to induce alveolar epithelial injury and to deplete alveolar macrophages followed by surfactant lipid accumulation, resulting in pulmonary inflammation. This is the first study to focus on the pathogenesis of mTOR inhibitor-induced ILD using an animal model.

Early detection of renal injury using urinary vanin-1 in rats with experimental colitis.

Renal complications are often detected in patients with inflammatory bowel disease (IBD). Because conventional markers such as serum creatinine and ß2-microglobulin are not sensitive and/or specific, a new renal biomarker is needed. We have recently identified urinary vanin-1 as an early biomarker for the detection of nephrotoxicant-induced renal injury. In this study, we compared the usefulness of urinary vanin-1 with other newly developed biomarkers [urinary monocyte chemoattractant protein-1 (MCP-1), kidney injury molecule-1 (Kim-1) and N-acetyl-beta-D-glucosaminidase (NAG)] for the detection of renal complications in rats with experimental colitis. On day 2 after intracolonic injection of 2,4,6-trinitrobenzene sulfonic acid (TNBS), male Wistar rats developed not only colitis, but histologically evident renal injury. Urinary vanin-1 started to elevate on day 1, whereas serum creatinine and urinary excretions of glucose, total protein, albumin, Kim-1, MCP-1 and NAG significantly increased only on day 2. The mRNA expressions of vanin-1 and Kim-1 significantly increased in the kidney, but not in the colon. In addition, vanin-1 did not appear in the blood. On the other hand, colonic mRNA expression and the serum concentration of MCP-1 were significantly higher in the TNBS-treated rats than in the control animals. These results suggest that, in contrast to MCP-1, urinary vanin-1 and Kim-1 mainly originated from the kidney rather than the colon in this model. Compared with Kim-1 and MCP-1, vanin-1 might be an earlier biomarker for the detection of renal injury in rats with experimental colitis.

Two nursing mothers treated with zonisamide: Should breast-feeding be avoided?

Zonisamide, an antiepileptic drug, is excreted into breast milk, but information regarding the safety of breast-feeding while using this drug is limited. We present the cases of two nursing mothers, taking 300 and 100 mg/day zonisamide. At 5 days after delivery, the milk concentrations and relative infant doses of the drug were 18.0 and 5.1 µg/mL, and 44 and 36%, respectively. In the first case, the mother fed colostrum and continued partial breast-feeding thus reducing the relative infant dose to 8%. The neonatal serum concentration of zonisamide declined to below the limit of detection at day 34 after birth. In the second case, the mother breast-fed partially until 2 weeks postpartum. No adverse effect was observed in the infants. These findings suggest that mothers taking zonisamide should not breast-feed exclusively, but may not have to avoid partial breast-feeding, with significant caution regarding adverse effects in infants.

Involvement of Wee1 in the circadian rhythm-dependent intestinal damage induced by docetaxel.

Docetaxel, a semisynthetic taxane, is effective for the treatment of some solid cancers; however, docetaxel-induced intestinal damage leads to poor prognosis in some patients. Although such adverse effects have been reported to depend on the dosing-time of docetaxel, the mechanisms involved remain unclear. Wee1 expression is controlled by the clock gene complex, clock/bmal1, and contributes to cell-cycle progression. The present study was undertaken to evaluate the potential role of Wee1 in the circadian rhythm-dependent profile of docetaxel. Male mice were maintained under a 12-hour light/dark cycle. Intestinal damage after repeated dosing of docetaxel (20 mg/kg) for 3 weeks was more severe at 14 hours after light on (HALO) than at 2 HALO. The intestinal protein expressions of Wee1, phosphorylated CDK1, and cleaved Caspase-3 were higher in the 14-HALO group than in the 2-HALO group, whereas that of survivin was lower in the 14-HALO group. Thus, it is speculated that elevated Wee1 expression inhibited CDK1 activity more by phosphorylation, which in turn caused the lower expression of survivin and consequently more activated Caspase-3 in the 14-HALO group. There were no significant differences in plasma docetaxel concentrations between the 2- and 14-HALO groups. Bindings of CLOCK and BMAL1 to the E-box regions at the wee1 gene promoter were not altered by docetaxel treatment at 2 and 14 HALO. These findings suggest that Wee1 is directly or indirectly involved in the mechanism of the circadian rhythm-dependent changes in docetaxel-induced intestinal damage. However, the mechanism for a circadian rhythm-dependent change in intestinal Wee1 expression by docetaxel remains to be determined.