Role of caspase-8 and/or -9 as biomarkers that can distinguish the potential to cause toxic- and immune related-adverse event, for the progress of acetaminophen-induced liver injury.

AIMS: Acetaminophen (APAP) overdose can cause acute liver failure. Although it is well known that APAP-induced liver injury (AILI) is caused by toxic mechanism, recently it is also reported to be immune related. However, the detail of the mechanism has been unclear. Therefore, elucidation of the pathophysiology is required. MAIN METHODS: In AILI model rats (800 mg/kg), the levels of AST, ALT and Caspase (C)-3/-8/-9 levels were measured. In in vitro study using human hepatocyte cells (FLC-4) and THP-1 cells, APAP (0.03-1.0 mM) were added to FLC-4 and the cell viability, C-9, cytochrome c, mitochondria membrane potential, and glutathione levels of FLC-4 and inflammasome activation of THP-1 were evaluated. KEY FINDINGS: In AILI model rats, the levels of AST and ALT were increased only at 12-24 h. C-3/-9 levels rose at 6-9 h, whereas C-8 level rose hours later, moreover, 24 h after; C-3/-8/-9 levels re-rose. In FLC-4 cells, cytochrome c was released from the mitochondria which is promoted by oxidative stress due to drug metabolism and C-9 was activated. Thus, AILI was caused mitochondrial damage by NAPQI as early reaction (first stage). In the next stage, inflammasomes of human antigen presenting cells, which released inflammatory cytokines were activated by damage-associated molecular patterns (DAMPs) released from damaged hepatocyte by APAP. SIGNIFICANCE: It is confirmed that AILI includes immune related mechanism. Thereby, in case of N-acetylcysteine refractory, additional administration of steroid hormones should be effective and recommended as a novel strategy for AILI with immune related adverse event (irAE).

Giant Hepatic Cyst: A Possible Cause of Inferior Vena Cava Syndrome.

An 80-year-old man was transferred to our institution with lower limb edema and worsening dyspnea following the administration of diuretic medication. Transthoracic echocardiography and computed tomography revealed a giant hepatic cyst (176×190 mm) compressing his right atrium and inferior vena cava (IVC). Laparoscopic cyst deroofing combined with omental packing and subsequent tube drainage immediately alleviated all his symptoms. The procedure was uneventful, and he was discharged without any complications on postoperative day 9; he had no recurrent symptoms or hepatic cysts at the postoperative 2-month follow-up. Therefore, a giant hepatic cyst can cause IVC syndrome, and laparoscopic deroofing is a beneficial approach for the treatment of accessible cysts.

Remote Management of Pacemaker Patients With Biennial In-Clinic Evaluation: Continuous Home Monitoring in the Japanese At-Home Study: A Randomized Clinical Trial.

BACKGROUND: Current expert consensus recommends remote monitoring for cardiac implantable electronic devices, with at least annual in-office follow-up. We studied safety and resource consumption of exclusive remote follow-up (RFU) in pacemaker patients for 2 years. METHODS: In Japan, consecutive pacemaker patients committed to remote monitoring were randomized to either RFU or conventional in-office follow-up (conventional follow-up) at twice yearly intervals. RFU patients were only seen if indicated by remote monitoring. All returned to hospital after 2 years. The primary end point was a composite of death, stroke, or cardiovascular events requiring surgery, and the primary hypothesis was noninferiority with 5% margin. RESULTS: Of 1274 randomized patients (50.4% female, age 77±10 years), 558 (RFU) and 550 (Conventional follow-up) patients reached either the primary end point or 24 months follow-up. The primary end point occurred in 10.9% and 11.8%, respectively (P=0.0012 for noninferiority). The median (interquartile range) number of in-office follow-ups was 0.50 (0.50-0.63) in RFU and 2.01 (1.93-2.05) in conventional follow-up per patient-year (P<0.001). Insurance claims for follow-ups and directly related diagnostic procedures were 18 800 Yen (16 500-20 700 Yen) in RFU and 21 400 Yen (16 700-25 900 Yen) in conventional follow-up (P<0.001). Only 1.4% of remote follow-ups triggered an unscheduled in-office follow-up, and only 1.5% of scheduled in-office follow-ups were considered actionable. CONCLUSIONS: Replacing periodic in-office follow-ups with remote follow-ups for 2 years in pacemaker patients committed to remote monitoring does not increase the occurrence of major cardiovascular events and reduces resource consumption. Registration: URL: https://clinicaltrials.gov; Unique identifier: NCT01523704.

Interaction of platinum agents, cisplatin, carboplatin and oxaliplatin against albumin in vivo rats and in vitro study using inductively coupled plasma-mass spectrometory.

The protein binding rates (PBR) of platinum-containing agents cisplatin (CDDP), carboplatin (CBDCA) and oxaliplatin (L-OHP) have been reported as 98%, 25-50% and 98%, respectively. To investigate the protein-binding properties of albumin with cisplatin, carboplatin and oxaliplatin, inductively coupled plasma mass spectrometry (ICP-MS) was used to measure their plasma concentration in rats over time. The study also examined the effects of cisplatin, carboplatin and oxaliplatin-binding on albumin in vitro, using CD spectrometry and native-polyacrylamide gel electrophoresis (native PAGE). The ratios of PBR to irreversible PBR, of cisplatin and oxaliplatin were 98%:98% and 90%:87%, respectively, indicating a higher affinity for irreversible binding with albumin. That of carboplatin was 25%:10%, indicating 60-70% reversible binding with albumin. The plasma protein binding rate concentrations of cisplatin, carboplatin and oxaliplatin after in vivo administration were 96%, 15% and 80%, respectively. The CD spectrometry of albumin was unaffected by cisplatin, carboplatin and oxaliplatin binding. Though similar protein binding rates were observed with oxaliplatin and cisplatin, oxaliplatin had a higher mobility rate during PAGE. It was confirmed that the binding of cisplatin and oxaliplatin with albumin affected its electric charge but not the structure. In conclusion, cisplatin and oxaliplatin bind irreversibly with albumin in plasma and may irreversibly interact with tissue protein and/or DNA. The difficulties involved with predicting the tissue concentrations of cisplatin and oxaliplatin from their plasma concentration inhibits their therapeutic drug monitoring. On the contrary, carboplatin, like some generic drugs, reversibly binds to plasma proteins. It is, therefore, possible to conduct therapeutic drug monitoring for carboplatin.

Contributions of caspase-8 and -9 to liver injury from CYP2E1-produced metabolites of halogenated hydrocarbons.

1. Drug-induced liver injury is difficult to predict at the pre-clinical stage. This study aimed to clarify the roles of caspase-8 and -9 in CYP2E1 metabolite-induced liver injury in both rats and cell cultures in vitro treated with carbon tetrachloride (CCl4), halothane or sevoflurane. The human hepatocarcinoma functional liver cell line was maintained in 3-dimensional culture alone or in co-culture with human acute monocytic leukemia cells. 2. In vivo, laboratory indices of liver dysfunction and histology were normal after administration of sevoflurane. CCl4 treatment increased blood AST/ALT levels, liver caspase-3 and -9 activities and liver malondialdehyde, accompanied by centrilobular hepatocyte necrosis. Halothane increased AST/ALT levels, caspase-3 and -8 activities (but not malondialdehyde) concomitant with widespread hepatotoxicity. In vitro, CCl4 treatment increased caspase-9 activity and decreased both mitochondrial membrane potential (MMP) and cell viability. In co-culture, halothane increased caspase-8 activity and decreased MMP and cellular viability. There were no toxic responses in CYP2E1 knockdown in monoculture and co-culture. 3. CYP2E1-inducing compounds play a pivotal role in halogenated hydrocarbon toxicity. 4. Changes in hepatocyte caspase-8 and -9 activities could be novel biomarkers of metabolites causing DILI, and in pre-clinical development of new pharmaceuticals can predict nascent DILI in the clinical stage.

A Novel Analytical Method of Cisplatin Using the HPLC with a Naphthylethyl Group Bonded with Silica Gel (πNAP) Column.

Cisplatin is the most widely used anticancer drug in the world. Mono-chloro and none-chloro complexes of cisplatin may be believed to be the activated compounds. The separation of these compounds using octa decyl silyl column or aminopropylsilyl silica gel column is difficult because of high-reactivity and structural similarity. In this study, cisplatin, hydroxo complexes, and OH-dimer were determined by HPLC using a naphthylethyl group bonded with silica gel (πNAP) column. The analytical conditions of HPLC were as follows: analytical column, πNAP column; wave length, 225 nm; column temperature, 40°C; mobile phase, 0.1 M sodium perchlorate, acetonitrile, and perchloric acid (290 : 10 : 3), flow rate, 1.0 mL/min. Sample (20 µL) was injected onto the HPLC system. Retention time of cisplatin, mono-chloride, OH-dimer, and none-chloride was 3.2, 3.4, 3.6, and, 4.3-6.6 min, respectively. Measurable ranges with this method were 1×10-5 to 4×10-3 M for cisplatin. Correlation coefficient of the calibration curves of cisplatin was 0.999 (p<0.01). The within- and between-day variations of coefficient of variation (CV) were 5% or lower. In this study, injectable formulations in physiological saline solution, water for injection, 5% glucose solution, and 7% sodium bicarbonate precisely were measured the stability and compositional changes upon mixing by πNAP column rather than C18 column. We successfully determined cisplatin, hydroxo complexes, and OH-dimer by HPLC using a πNAP column. Thus the measurement of cisplatin (cis-diamminedichloro-platinum(II), cis-[PtCl2(NH3)2]) (CDDP) should be done using a πNAP column rather than a C18 column or aminopropylsilyl silica gel column.

Hydrogen gas attenuates embryonic gene expression and prevents left ventricular remodeling induced by intermittent hypoxia in cardiomyopathic hamsters.

The prevalence of sleep apnea is very high in patients with heart failure (HF). The aims of this study were to investigate the influence of intermittent hypoxia (IH) on the failing heart and to evaluate the antioxidant effect of hydrogen gas. Normal male Syrian hamsters (n = 22) and cardiomyopathic (CM) hamsters (n = 33) were exposed to IH (repeated cycles of 1.5 min of 5% oxygen and 5 min of 21% oxygen for 8 h during the daytime) or normoxia for 14 days. Hydrogen gas (3.05 vol/100 vol) was inhaled by some CM hamsters during hypoxia. IH increased the ratio of early diastolic mitral inflow velocity to mitral annulus velocity (E/e', 21.8 vs. 16.9) but did not affect the LV ejection fraction (EF) in normal Syrian hamsters. However, IH increased E/e' (29.4 vs. 21.5) and significantly decreased the EF (37.2 vs. 47.2%) in CM hamsters. IH also increased the cardiomyocyte cross-sectional area (672 vs. 443 µm(2)) and interstitial fibrosis (29.9 vs. 9.6%), along with elevation of oxidative stress and superoxide production in the left ventricular (LV) myocardium. Furthermore, IH significantly increased the expression of brain natriuretic peptide, ß-myosin heavy chain, c-fos, and c-jun mRNA in CM hamsters. Hydrogen gas inhalation significantly decreased both oxidative stress and embryonic gene expression, thus preserving cardiac function in CM hamsters. In conclusion, IH accelerated LV remodeling in CM hamsters, at least partly by increasing oxidative stress in the failing heart. These findings might explain the poor prognosis of patients with HF and sleep apnea.

Chronological changes in circulating levels of soluble tumor necrosis factor receptors 1 and 2 in rats with carbon tetrachloride-induced liver injury.

Carbon tetrachloride (CCl4) facilitates the generation of hepatotoxins that can result in morphologic abnormalities, and these abnormalities are reasonably characteristic and reproducible for each particular toxin. It is also known that tumor necrosis factor-alpha (TNF-α) may participate in CCl4-induced liver injury (CILI). In this study, we observed the chronological changes in circulating soluble tumor necrosis factor receptors 1 and 2 (sTNF-R1 and -R2) in rats with CILI. Laboratory data; circulating levels of TNF-α, sTNF-R1, and sTNF-R2; and TNF-α levels in liver tissues were measured at various time-points. In the CCl4 group, the plasma aspartate aminotransferase (AST, 7694±3041IU/l)/alanine aminotransferase (ALT, 3241±2159 IU/l) levels peaked at 48 h after CCl4 administration, but the other laboratory data did not differ significantly from the corresponding data in the controls. Centrilobular hepatocyte necrosis and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells near the central vein area were observed via hematoxylin eosin (HE) and TUNEL staining, respectively, at 24 and 48 h after CCl4 administration. Compared to the control group, the CCl4 group did not show significantly the increased circulating TNF-α levels. But TNF-α levels in the liver tissues first peaked at 1h (5261±2253 pg/g liver), and a second peak was observed at 12h (3806±533 pg/g liver) after CCl4 administration. Compared to the control group, the CCl4 group showed significantly increased circulating levels of both sTNF-R1 (797±121pg/ml) and sTNF-R2 (5696±626 pg/ml) 1h after CCl4 administration. Since the hepatocyte apoptosis may be resulted from binding of TNF-α with TNF-R1 at 24h after administration, and consequently the circulating TNF-R2 level might be approximately 10-fold higher than the circulating TNF-R1 level. In conclusion, increased circulating levels of sTNF-R1 and -R2 potentially contribute to drug-induced liver injury, together with AST/ALT.

The effect of capsaicin on circulating biomarkers, soluble tumor necrosis factor and soluble tumor necrosis factor-receptor-1 and -2 levels in vivo using lipopolysaccharide-treated mice.

The circulating soluble tumor necrosis factor (sTNF) and sTNF-receptor (R) 1 and -R2 have known as septic biomarker. The pungent component of capsicum, capsaicin (Cap), has several associated physiological activities, including anti-oxidant, anti-bacterial and anti-inflammatory effects. The aim of this study was to elucidate the effect of Cap on circulating sTNF and sTNF-R1 and -R2 in vivo using lipopolysaccharide (LPS)-treated mice. LPS (20 mg/kg, ip)-treated group was significantly increased circulating sTNF, sTNF-R1, and -R2 and TNF-α mRNA expression levels compared to the vehicle group. Treatment with LPS (20 mg/kg, ip) + Cap (4 mg/kg, sc)-treated group was significantly decreased both circulating sTNF levels (after 1 h only) and TNF-α mRNA expression (after 6 h) compared to the LPS-treated group. There is an early increase in circulating sTNF, sTNR-R1, and -R2 observed in the LPS-treated mice. Since Cap inhibits this initial increase as biomarkers, circulating sTNF, it is considered a potent treatment option for TNF-α-related diseases, such as septicemia. In conclusion, Cap interferes with TNF-α mRNA transcription and exerts an inhibiting effect on TNF-α release from macrophages in the early phase after LPS stimulation. Thus, Cap is considered a potent agent for the treatment of TNF-α-related diseases, such as septicemia.

Systemic inflammatory response syndrome criteria and vancomycin dose requirement in patients with sepsis.

PURPOSE: Vancomycin has been used in patients with sepsis infected by MRSA and shows large interindividual variability in its dosing. In this observational study the potential influence of sepsis status on the vancomycin dose requirement in relation to systemic inflammatory response syndrome (SIRS) criteria was assessed. METHODS: From about 250 patients receiving serum vancomycin monitoring from May 2006 to April 2011 at the Osaka National Hospital, 105 adult patients who had been assessed using the SIRS criteria were identified. Patients on chemotherapy or intermittent positive pressure ventilation in whom the SIRS criteria could not accurately evaluate inflammatory status were excluded. Using two vancomycin serum concentrations at peak and trough, individual pharmacokinetic parameters were calculated by the Bayesian estimation method using a two-compartment model. Creatinine clearance rate was estimated by the Cockcroft-Gault formula (eCcr). RESULTS: Patients with SIRS had a significantly higher vancomycin clearance than those without SIRS, indicating that SIRS patients had a higher elimination capacity. The vancomycin clearance was positively correlated with the SIRS score defined as the number of positive items in the criteria, and negatively with age, except in patients with renal dysfunction. A linear relationship between the vancomycin clearance and eCcr remained even in the supernormal eCcr phase (more than approximately 120 mL/min). CONCLUSIONS: This study provides a new insight into the need for quick prediction of dose requirement. That is, an increased vancomycin dosage would be needed in patients with a higher SIRS score to maintain the therapeutic target concentration, in particular in those with a high eCcr value.

Application of electrolysis to inactivation of antibacterials in clinical use.

Contamination of surface water by antibacterial pharmaceuticals (antibacterials) from clinical settings may affect aquatic organisms, plants growth, and environmental floral bacteria. One of the methods to decrease the contamination is inactivation of antibacterials before being discharged to the sewage system. Recently, we reported the novel method based on electrolysis for detoxifying wastewater containing antineoplastics. In the present study, to clarify whether the electrolysis method is applicable to the inactivation of antibacterials, we electrolyzed solutions of 10 groups of individual antibacterials including amikacin sulfate (AMK) and a mixture (MIX) of some commercial antibacterials commonly prescribed at hospitals, and measured their antibacterial activities. AMK was inactivated in its antibacterial activities and its concentration decreased by electrolysis in a time-dependent manner. Eighty to ninety-nine percent of almost all antibacterials and MIX were inactivated within 6h of electrolysis. Additionally, cytotoxicity was not detected in any of the electrolyzed solutions of antibacterials and MIX by the Molt-4-based cytotoxicity test.

Essential role of macrophages in the initiation of allergic rhinitis in mice sensitized intranasally once with cedar pollen: regulation of class switching of immunoglobulin in B cells by controlling interleukin-4 production in T cells of submandibular lymph nodes.

The production of allergen-specific IgE antibodies (Abs) in allergen-sensitized patients or animals has a mutual relationship with the immunologic response leading to allergic rhinitis. We recently reported that, after an intranasal injection of cedar pollen into mice, an interleukin-4 (IL-4)-dependent increase in serum nonspecific IgE Abs was a prerequisite for the production of serum allergen-specific IgE Abs. Here, we explored which lymphoid organs were responsive to the intranasally injected allergen and how IL-4 and IgE Abs were produced in the lymphocytes. Time-dependent changes in the total cell numbers and in in vitro IgE Ab production in various lymphoid organs revealed that the submandibular lymph nodes were the main responsible organ. After treatment with allergen (for IgE production) or allergen and complete Freund's adjuvant (for IgG production), we separated submandibular lymph node cells into macrophage-, lymphocyte-, and granulocyte-rich populations by discontinuous Percoll density-gradient centrifugation. Unexpectedly, bulk cells, but not the lymphocyte- or macrophage-rich populations, produced significant amounts of IL-4, IgE, and IgG; whereas production was restored by addition of Mac-1(+) cells from the macrophage-rich to the lymphocyte-rich fraction. Furthermore, a combination of the lymphocyte-rich population (for IgG [or IgE]) production) and the macrophage-rich population (for IgE [or IgG]) production) produced a large amount of IgE (or IgG). These results indicate that, in the initiation of allergic rhinitis, macrophages in the submandibular lymph nodes are essential not only for IL-4 or immunoglobulin production, but also for class switching of immunoglobulin in lymphocytes.

Role of cytokines in lavage or drainage fluid after hemithyroidectomy in wound healing: involvement of histamine in the acceleration and delay of wound healing.

Wound healing is a sophisticated biologic process. In the case of hemithyroidectomy, the operation time is relatively short with small tissue damage and without skin excision, and bacterial contamination before, during, and after the operation is uncommon. Here, we explored which cytokine(s) affected the rates of healing of skin wounds after hemithyroidectomy of 29 patients. We assessed the amounts of cytokines (e.g., interleukin-6, platelet-derived growth factor, basic fibroblast growth factor, vascular endothelial growth factor, and tumor necrosis factor-α) in either the preoperative or postoperative lavage fluids, or in the drainage fluids on postoperative days (PODs) 1-8. All of these cytokines showed a similar pattern; after reaching a peak on POD1, the production fell sharply on POD2-8, revealing that wound healing commenced on POD1. The rates of wound healing were inversely related to the levels of histamine in six patients (i.e., those with the three largest and those with the three smallest total volumes of drainage fluid on POD1): high (or low) levels of histamine in the postoperative lavage fluids with low (or high) levels in the drainage fluids on POD1 caused earlier (or the delay of) wound healing, suggesting involvement of histamine in the acceleration and delay of wound healing.

Application of electrolysis for detoxification of an antineoplastic in urine.

Antineoplastics in excreta from patients have been considered to be one of the origins of cytotoxic, carcinogenic, teratogenic, and mutagenic contaminants in surface water. Recent studies have demonstrated that antineoplastics in clinical wastewater can be detoxified by electrolysis. In this study, to develop a method for the detoxification of antineoplastics in excreta, methotrexate solution in the presence of human urine was electrolyzed and evaluated. We found that urine inhibits detoxification by electrolysis; however, this inhibition decreased by diluting urine. In urine samples, the concentrations of active chlorine generated by anodic oxidation from 0.9% NaCl solution for inactivation of antineoplastics increased in dilution-dependent and time-dependent manner. These results indicate that electrolysis with platinum-based iridium oxide composite electrode is a possible method for the detoxification of a certain antineoplastic in urine.

Curcumin and genistein additively potentiate G551D-CFTR.

BACKGROUND: The G551D mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) is a common cause of cystic fibrosis (CF). G551D-CFTR is characterized by an extremely low open probability despite its normal trafficking to the plasma membrane. Numerous small molecules have been shown to increase the activity of G551D-CFTR presumably by binding to the CFTR protein. METHODS: We investigated the effect of curcumin, genistein and their combined application on G551D-CFTR activity using the patch clamp technique. RESULTS: Curcumin increased G551D-CFTR whole-cell and single-channel currents less than genistein did at their maximally effective concentrations. However, curcumin further increased the channel activity of G551D-CFTR that had been already maximally potentiated by genistein, up to ~50% of the WT-CFTR level. In addition, the combined application of genistein and curcumin over a lower concentration range synergistically rescued the gating defect of G551D-CFTR. CONCLUSIONS: The additive effects between curcumin and genistein not only support the hypothesis that multiple mechanisms are involved in the action of CFTR potentiators, but also pose pharmaceutical implications in the development of drugs for CF pharmacotherapy.

Is cyclosporine A transport inhibited by pravastatin via multidrug resistant protein 2?

BACKGROUND AND PURPOSE: Cyclosporine A (CyA) is an immunosuppressant drug used to treat various autoimmune diseases and transplantations. It has been reported that, in humans, CyA is metabolized by cytochrome P450 (CYP) 3A or excreted by P-glycoprotein/multidrug resistant protein (MRP) 2. Pravastatin, a statin, is used to treat hyperlipidemia and has also been reported to be excreted primarily by MRP2. We observed an increased blood CyA level in a patient following pravastatin administration, suggesting the possibility that CyA interacted with the pravastatin via MRP2. The aim of the study reported here was to investigate the effects of pravastation on CyA transport via MRP2 using a human colon adenocarcinoma (Caco-2) monolayer model system. METHODS: Calcein, a substrate of MRP families, was first added to the tissue culture medium of the Caco-2 cells, and CyA (5, 50 microM) and pravastatin (0.1, 1.0 mM) were then added to the apical and basolateral sides. After a 30-min incubation, calcein was effluxed from the Caco-2 cells and the level in the culture medium was assayed. CyA was then added to the tissue culture medium of the Caco-2 cells, and pravastatin (0.1, 0.5, 1.0 mM) was added to the apical and basolateral sides. After a 30-min incubation, CyA was effluxed from the Caco-2 cells, and the level in the culture medium was assayed. RESULTS: The calcein efflux to the apical side was decreased significantly by the addition of pravastatin (1.0 mM) and CyA (5, 50 microM), respectively. The CyA efflux to the apical side was decreased significantly by the addition of pravastatin (1.0 mM). CONCLUSIONS: Based on these results, we suggest that CyA transport may be competitively inhibited by pravastatin via MRP2.

Ion-exclusion/cation-exchange chromatographic determination of common inorganic ions in human saliva by using an eluent containing zwitterionic surfactant.

Ion-exclusion/cation-exchange chromatography with an eluent containing the bile salt-type zwitterionic surfactant CHAPS was performed in order to evaluate variations in anion (SO(4)(2-), NO(3)(-), and SCN(-)) and cation (Na(+), K(+), NH(4)(+), Mg(2+), and Ca(2+)) concentrations in human saliva. CHAPS prevents the adsorption of proteins to the stationary phase, i.e., weakly acidic cation-exchange resin, since it aggregates proteins without denaturing them. Addition of 1mM CHAPS to the eluent comprising 6mM tartaric acid and 7 mM 18-crown-6 yielded reproducible separations of anions and cations in protein-containing saliva. The resolutions of anions and cations were not significantly affected by the addition of CHAPS to the eluent. The concentrations of Na(+) and K(+) varied before and after meals; or that of SCN(-), upon smoking. The relative standard deviations of peak areas ranged from 0.3 to 5.1% in 1 day (n=20) and from 1.4 to 5.8% over 6 days (n=6).

Changes of midazolam pharmacokinetics in Wistar rats treated with lipopolysaccharide: relationship between total CYP and CYP3A2.

It has been reported that infection interferes with drug metabolism, resulting in changes in pharmacokinetics. In this study, we investigated the effects of lipopolysaccharide (LPS) on hepatic total cytochrome P450 (CYP), CYP3A2, and CYP2C11 contents in a transient, LPS-induced, endotoxemia model of rats. In addition, to assess the effects on CYP3A2 activities, the pharmacokinetics of midazolam (CYP3A2 substrate) and 1-OH-midazolam (metabolite of midazolam) were investigated. Hepatic total CYP contents were significantly low until day 3 (P < 0.05) but returned to the control level on day 5. Hepatic CYP3A2 contents were significantly decreased on day 1 until day 5 (P < 0.05) but returned to the control level on day 7. Hepatic CYP2C11 contents were continuously low until day 7, and lowest on day 3. The AUC of 1-OH-midazolam was significantly decreased on day 1 after LPS administration (P < 0.01). In conclusion, LPS (5 mg/kg) challenge decreased hepatic total CYP, CYP3A2, and CYP2C11 contents and also decreased the activities of hepatic CYP3A2. It took at least 7 days for hepatic total CYP and CYP3A2 to recover to control levels, and it was suggested that the changes of hepatic total CYP contents might correlate with those of hepatic CYP3A2 contents and activities. Additionally, it is shown that their changes might reflect the recovery process from inflammation.

Changes in digoxin pharmacokinetics treated with lipopolysaccharide in Wistar rats.

Lipopolysaccharide (LPS) is a highly bioactive substance that can cause local as well as systemic damage to various organs of both humans and animals, even at very low doses. However, there are a few reports on drug pharmacokinetics during endotoxemia. In this study, we analyzed the pharmacokinetics of digoxin (a therapeutic agent for cardiac insufficiency) as a probe drug for a two-compartment model in a rat model of endotoxemia induced by LPS for 5 d. Digoxin was given to Wistar rats intravenously (i.v.), orally (p.o.), and intra-intestinally using an in situ closed-loop method (loop). The AUCi.v. was significantly increased in the LPS (+) group throughout the experiment (p<0.05). There was significant decrease in V2 (volume of distribution of tissue compartment) on Day 1-3 (p<0.05). On Day 1-2 after LPS administration, the AUCp.o. was significantly increased in the LPS (+) group (p<0.05). The AUCloop was significantly increased throughout the experiment (p<0.05). The elimination rate constant was unchanged. Thus LPS administration affected the absorption but not the excretion of digoxin. The findings of this study suggest that digoxin absorption increased and the volume of distribution of tissue compartment decreased after LPS administration (5 mg/kg, i.p.). It appears that digoxin pharmacokinetics recover over 3 d after LPS administration.

Effects of CpG-DNA from Escherichia coli on digoxin pharmacokinetics.

Deoxyribonucleic acid (DNA) from bacteria or viruses has been reported as one of the pathogen-associated molecular patterns (PAMPs) and a substance that can induce endotoxemia-like inflammation in animals. However, there has been no report on digoxin pharmacokinetics in the inflammation induced by bacterial DNA containing unmethylated CpG motifs (CpG-DNA). In this study, we investigated the effects of CpG-DNA on digoxin pharmacokinetics. We determined the degree of lipopolysaccharide contamination in CpG-DNA solution and examined the changes in digoxin pharmacokinetics in rats after CpG-DNA administration. In addition, plasma concentrations of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and nitrite/nitrate (NOx) were determined after CpG-DNA administration (5 mg/kg, i.p.). The AUC0-24 of digoxin increased significantly on Day 1-3 and CL/F decreased on Day 1 and Day 2 after CpG-DNA administration. On Day 7 after CpG-DNA administration, there were no significant differences in AUC0-24 and CL/F compared with the control group (without CpG-DNA administration). However, Kel remained relatively unchanged throughout the experiment. Plasma TNF-alpha concentrations were significantly increased at 1 h and plasma IL-1beta concentrations were significantly decreased at 6 h after administration of CpG-DNA, while plasma NOx concentrations were significantly increased at 12 h after CpG-DNA administration, compared with the control group. These findings suggest that CpG-DNA (5 mg/kg) induces a transient inflammatory condition, and that AUC0-24 and CL/F of digoxin were altered after CpG-DNA administration. Digoxin pharmacokinetics recovered within 7 d after CpG-DNA exposure.