Carbon monoxide may enhance bile secretion by increasing glutathione excretion and Mrp2 expression in rats

AbstractBackgroundNitric oxide (NO) donors have been reported to induce choleresis via an increased excretion of glutathione. The effects of another gas molecule, carbon monoxide (CO), on bile formation are, however, inconsistent among previous reports. We investigated the sequential changes of bile output and the biliary contents in rats with or without CO supplementation to elucidate the mechanism of CO on bile excretion.MethodsDichloromethane (DCM) was gastrically fed to male Sprague–Dawley rats to yield CO by liver biotransformation. The rats were divided into DCM-treated (n = 7), DCM plus L-NAME-treated (n = 6), and corn oil-treated-(n = 8) groups. Bile samples were collected hourly to examine the flow rate and bile content. Serum levels of nitrite and nitrate 4 hours after DCM supplementation with or without NO synthase (NOS) inhibition were measured by capillary electrophoresis. The expression of hepatic inducible NOS was evaluated by Western blotting 6 hours after DCM administration.ResultsLevels of carboxyhemoglobin rose to around 10% at 4 hours after DCM supplementation and were maintained until the end of the experiments. Bile flow increased after DCM supplementation and was associated with a concomitant increase of biliary glutathione and higher hepatic multidrug resistance-associated protein 2 (Mrp2) expression. Hepatic inducible NOS expression and serum nitrate/nitrite levels were also increased. Treatment with an NOS inhibitor (L-NAME) abolished the CO-induced glutathione excretion and choleresis, but not Mrp2 expression.ConclusionThe present study demonstrated that CO enhanced biliary output in conjunction with NO by increasing the biliary excretion of glutathione. The increment in biliary glutathione was associated with an increased expression of hepatic Mrp2

Serum total antioxidant capacity reflects severity of illness in patients with severe sepsis

INTRODUCTION: We conducted the present study to evaluate the changes in serum total antioxidant capacity (TAC) in patients with severe sepsis and to investigate the association between serum TAC and clinical severity. METHOD: This was a prospective observational study involving a sample of patients who met established criteria for severe sepsis and were admitted to the emergency department of a university teaching hospital. Serum TAC was determined using the total radical-trapping antioxidant parameter method. The levels of TAC, uric acid, albumin, and bilirubin in sera were obtained in the emergency department and evaluated to determine whether there were any correlations between the major antioxidant biomarkers and clinical severity of sepsis. The Acute Physiology and Chronic Health Evaluation (APACHE) II score was used for clinical evaluation of the severity of sepsis. RESULTS: A total of 73 patients with sepsis, with a mean (± standard deviation) APACHE II score of 23.2 ± 8.2 and a mortality rate of 26.0%, were included. Seventy-six healthy individuals served as control individuals. Among the patients, serum TAC levels correlated significantly with APACHE II scores. Patients who died also had higher TAC than did those who survived. Serum uric acid levels correlated significantly with serum TAC and APACHE II scores in patients with severe sepsis. CONCLUSION: Elevated serum TAC level may reflect clinical severity of sepsis. In addition, serum uric acid levels appear to contribute importantly to the higher TAC levels observed in patients with severe sepsis

Two-step magnetic bead-based (2MBB) techniques for immunocapture of extracellular vesicles and quantification of microRNAs for cardiovascular diseases: A pilot study.

Extracellular vesicles (EVs) have attracted increasing attention because of their potential roles in various biological processes and medical applications. However, isolation of EVs is technically challenging mainly due to their small and heterogeneous size and contaminants that are often co-isolated. We have thus designed a two-step magnetic bead-based (2MBB) method for isolation a subset of EVs as well as their microRNAs from samples of a limited amount. The process involves utilizing magnetic beads coated with capture molecules that recognize EV surface markers, such as CD63. Captured EVs could be eluted from beads or lyzed directly for subsequent analysis. In this study, we used a second set of magnetic beads coated with complementary oligonucleotides to isolate EV-associated microRNAs (EV-miRNAs). The efficiencies of 2MBB processes were assessed by reverse transcription-polymerase chain reaction (RT-PCR) with spiked-in exogenous cel-miR-238 molecules. Experimental results demonstrated the high efficiency in EV enrichment (74 ± 7%, n = 4) and miRNA extraction (91 ± 4%, n = 4). Transmission electron micrographs (TEM) and nanoparticle tracking analysis (NTA) show that captured EVs enriched by 2MBB method could be released and achieved a higher purity than the differential ultracentrifugation (DUC) method (p < 0.001, n = 3). As a pilot study, EV-miR126-3p and total circulating cell-free miR126-3p (cf-miR126-3p) in eight clinical plasma samples were measured and compared with the level of protein markers. Compared to cf-miR126-3p, a significant increase in correlations between EV-miR126-3p and cardiac troponin I (cTnI) and N-terminal propeptide of B-type natriuretic peptide (NT-proBNP) was detected. Furthermore, EV-miR126-3p levels in plasma samples from healthy volunteers (n = 18) and high-risk cardiovascular disease (CVD) patients (n = 10) were significantly different (p = 0.006), suggesting EV-miR126 may be a potential biomarker for cardiovascular diseases. 2MBB technique is easy, versatile, and provides an efficient means for enriching EVs and EV-associated nucleic acid molecules