[Construction of TRAF6 ubiquitin site 331 mutant colorectal cancer cell stable line and its effect on biological behavior of colorectal cancer cells].

Objective: To investigate the effect of ubiquitin mutation at position 331 of tumor necrosis factor receptor related factor 6 (TRAF6) on the biological characteristics of colorectal cancer cells and its mechanism. Methods: lentivirus wild type (pCDH-3×FLAG-TRAF6) and mutation (pCDH-3×FLAG-TRAF6-331mut) of TRAF6 gene expression plasmid with green fluorescent protein tag were used to infect colorectal cancer cells SW480 and HCT116, respectively. The infection was observed by fluorescence microscope, and the expressions of TRAF6 and TRAF6-331mut in cells was detected by western blot. Cell counting kit-8 (CCK-8) and plate cloning test were used to detect the proliferation ability of colorectal cancer cells in TRAF6 group and TRAF6-331mut group, cell scratch test to detect cell migration, Transwell chamber test to detect cell migration and invasion, immunoprecipitation to detect the ubiquitination of TRAF6 and TRAF6-331mut with ubiquitinof lysine binding sites K48 and K63. Western blot was used to detect the effects of TRAF6 and TRAF6-331mut over expression on the nuclear factor kappa-B (NF-κB) and mitogen activated protein kinase mitogen-activated protein kinase (MAPK)/activating protein-1(AP-1) signal pathway. Results: The successful infection of colorectal cancer cells was observed under fluorescence microscope. Western blot detection showed that TRAF6 and TRAF6-331mut were successfully expressed in colorectal cancer cells. The results of CCK-8 assay showed that on the fourth day, the absorbance values of HCT116 and SW480 cells in TRAF6-331mut group were 1.89±0.39 and 1.88±0.24 respectively, which were lower than those in TRAF6 group (2.09±0.12 and 2.17±0.45, P=0.036 and P=0.011, respectively). The results of plate colony formation assay showed that the number of clones of HCT116 and SW480 cells in TRAF6-331mut group was 120±14 and 85±14 respectively, which was lower than those in TRAF6 group (190±21 and 125±13, P=0.001 and P=0.002, respectively). The results of cell scratch test showed that after 48 hours, the percentage of wound healing distance of HCT116 and SW480 cells in TRAF6-331mut group was (31±12)% and (33±14)%, respectively, which was lower than those in TRAF6 group [(43±13)% and (43±7)%, P=0.005 and 0.009, respectively]. The results of Transwell migration assay showed that the migration numbers of HCT116 and SW480 cells in TRAF6-331mut group were significantly lower than those in TRAF6 group (P<0.001 and P<0.002, respectively). The results of Transwell invasion assay showed that the number of membrane penetration of HCT116 and SW480 cells in TRAF6-331mut group was significantly lower than those in TRAF6 group (P=0.008 and P=0.009, respectively). The results of immunoprecipitation detection showed that the ubiquitin protein of K48 chain pulled by TRAF6-331mut was lower than that of wild type TRAF6 in 293T cells co-transfected with K48 (0.57±0.19), and the ubiquitin protein of K63 chain pulled down by TRAF6-331mut in 293T cells co-transfected with K63 was lower than that of wild type TRAF6 (0.89±0.08, P<0.001). Western blot assay showed that the protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-HCT116 cells were 0.63±0.08, 0.42±0.08 and 0.60±0.07 respectively, which were lower than those in TRAF6-HCT116 cells (P=0.002, P<0.001 and P<0.001, respectively). The expression level of AP-1 protein in TRAF6-HCT116 cells was 0.89±0.06, compared with that in TRAF6-HCT116 cells. The difference was not statistically significant (P>0.05). The protein expression levels of NF-κB, p-NF-κB and p-AP-1 in TRAF6-331mut-SW480 cells were 0.50±0.06, 0.51±0.04, 0.48±0.02, respectively, which were lower than those in TRAF6-SW480 cells (all P<0.001). There was no significant difference in AP-1 protein expression between TRAF6-331mut-SW480 cells and TRAF6-SW480 cells. Conclusion: The ubiquitin site mutation of TRAF6 gene at 331 may prevent the binding of TRAF6 and ubiquitin lysine sites K48 and K63, and then affect the expressions of proteins related to downstream NF-κB and MAPK/AP-1 signal pathways, and inhibit the proliferation, migration and invasion of colorectal cancer cells.

Perfil hematológico e bioquímico sérico de capivaras (Hydrochoerus hydrochaeris) de vida livre nos biomas Mata Atlântica e Caatinga / Hematological and serum biochemical profile of free-living capybaras (Hydrochoerus hydrochaeris) in Atlantic Forest and Caatinga biomes

Devido à ausência de estudos sobre capivaras na região Nordeste do Brasil, o objetivo deste estudo foi avaliar a sanidade desses roedores de vida livre em três áreas dos biomas Mata Atlântica (2) e Caatinga (1) do estado de Pernambuco, por meio da determinação de parâmetros da hematologia e bioquímica sérica. De novembro de 2016 a dezembro de 2017, foram capturados 21 animais, dos quais foram coletadas amostras de sangue para avaliação hematológica (eritrograma, leucograma e plaquetometria) e bioquímica sérica (atividade enzimática, perfil proteico, energético e mineral). A maioria dos parâmetros esteve dentro dos valores de normalidade para a espécie, embora alguns apresentassem diferenças estatisticamente significativas de acordo com a área de estudo (hemoglobina, hematócrito, VCM, CHCM, eosinófilos, fosfatase alcalina, proteína total, albumina, ácido úrico, creatinina, lactato, sódio e magnésio) e o sexo dos animais (ácido úrico). Os parâmetros obtidos são apresentados como referência e atestam a sanidade e o bom estado nutricional de populações de capivaras nos biomas Mata Atlântica e Caatinga da região Nordeste do Brasil. As informações aportadas neste estudo pioneiro na região Nordeste contribuem para aumentar o conhecimento sobre a ecofisiologia e a conservação in situ de capivaras.(AU)

Due to the lack of studies about capybaras in the northeast region of Brazil, the objective of this study was to evaluate the health status of free-ranging capybaras in three areas of Atlantic Forest (2) and Caatinga (1) biomes in Pernambuco state, through the determination of hematological and serum biochemical parameters. From November 2016 to December 2017, 21 animals were captured and blood samples were collected for the hematological (erythrogram, leukogram and platelet counts) and serum biochemistry (enzymatic activity, protein, energy and mineral profile) evaluation. Hematological and serum biochemical parameters were within the normal range for the species, but some presented statistically significant variations according to the study area (hemoglobin, hematocrit, MCV, MCHC, eosinophils count, alkaline phosphatase, total proteins, albumin, uric acid, creatinine, lactate, sodium and magnesium) and sex of the animals (uric acid). The parameters obtained are presented as reference and attest to the health and good nutritional status of populations of capybaras in the Atlantic Forest and Caatinga biomes of northeastern Brazil. The information provided in this pioneering study in the northeast region contributes to increased knowledge about the ecophysiology and in situ conservation of capybaras.(AU)

MiR-592 functions as a tumor suppressor in acute myeloid leukemia by targeting ROCK1 and predicts patients' prognosis.

OBJECTIVE: Dysregulation of miR-592 has been reported in several tumors. However, its role in acute myeloid leukemia (AML) remains unknown. The present study aimed at investigating the expression pattern and biological function of miR-592 in AML and to elucidate the mechanism involved. PATIENTS AND METHODS: qRT-PCR was used to analyze the expression of miR-592 in bone marrow and serum obtained from AML patients and healthy controls. The associations between serum miR-592 expression and clinical features and prognosis of AML patients were statistically analyzed. Then we detected the effect of miR-592 on proliferation, metastasis and apoptosis by CCK-8 assay, Transwell assays and flow cytometry, respectively. Dual-luciferase reporter assays were performed to validate the regulation of a putative target of miR-592. Rescue experiments were performed to confirm whether ROCK1 was a direct and functional target of miR-592 in AML. RESULTS: We found that the expression level of miR-592 was significantly lower in AML patients and AML cell lines. Low expression of serum miR-592 was associated with advanced French-American-British classification, cytogenetics and poor prognosis. Multivariate analysis confirmed that serum miR-592 expression was an independent prognostic factor for AML patients. Functionally, overexpression of miR-592 suppressed AML cell growth and metastasis, and promoted apoptosis. Further mechanistic investigation showed ROCK1 was a direct target gene of miR-592. Finally, ROCK1 overexpression rescued the effect of miR-592-mediated AML cell proliferation and metastasis. CONCLUSIONS: These findings suggest that miR-592 acted as a tumor suppressor by targeting ROCK1 and may serve as a potential biomarker in AML.

Comparison of genetic diversity between wild-caught broodstock and hatchery-produced offspring populations of the vulnerable Korean kelp grouper (Epinephelus bruneus) by microsatellites.

The kelp grouper Epinephelus bruneus (Perciformes: Haemulidae), is one of the most economically important fishery resources in Korea. This fish is regarded as a target for prospective aquaculture diversification; therefore, maintenance of stock quality is important. To investigate the effects of current artificial reproduction in a hatchery facility, genetic variation in wild-caught broodstock and hatchery-produced offspring of kelp grouper was analyzed using eight polymorphic nuclear microsatellite DNA loci; 77 alleles were identified. Allelic variability ranged from 2 to 22 in the broodstock and from 1 to 10 in the offspring. The average observed and expected heterozygosities were 0.620 and 0.623 in the broodstock and 0.600 and 0.513 in the offspring, respectively. The possibility of a recent genetic bottleneck was suggested in both populations of E. bruneus. The minor, but significant, genetic differentiation (FST = 0.047, P < 0.05) observed was mainly due to statistically significant reductions in the number of alleles in the offspring compared with the broodstock, suggesting that these genetic changes could be related to genetic drift. Our results demonstrate the usefulness of microsatellite markers to monitor genetic variation and raise concerns about potential harmful genetic effects of inappropriate hatchery procedures. Therefore, genetic variation between broodstock and offspring in a hatchery should be monitored in both breeding and release programs as a routine hatchery operation, and inbreeding should ideally be controlled to improve kelp grouper hatchery management. Our data provide a useful genetic basis for future planning of sustainable culture and management of E. bruneus in fisheries.

Large magnetoelectric coupling in Co4Nb2O9.

Magnetoelectric materials which simultaneously exhibit electric polarization and magnetism have attracted more and more attention due to their novel physical properties and promising applications for next-generation devices. Exploring new materials with outstanding magnetoelectric performance, especially the manipulation of magnetization by electric field, is of great importance. Here, we demonstrate the cross-coupling between magnetic and electric orders in polycrystalline Co4Nb2O9, in which not only magnetic-field-induced electric polarization but also electric field control of magnetism is observed. These results reveal rich physical phenomenon and potential applications in this compound.

Gender differences in electrophysiological characteristics of idiopathic ventricular tachycardia originating from right ventricular outflow tract.

It has become increasingly apparent in recent years that there are important differences of many cardiovascular disorders including ventricular tachycardias in men and women. Nevertheless, so far just few studies have addressed possible gender differences in electrophysiological characteristics of idiopathic ventricular tachycardia from right ventricular outflow tract (RVOT-VT), other than epidemiological ones. This study explored possible gender differences in electrophysiological characteristics and catheter ablation outcome in RVOT-VT patients. Ninety-three patients (mean age 38.7+/-15.5 years, 30 males) with idiopathic RVOT-VT were enrolled and analyzed in our study. Male patients had longer QRS width (99.9+/-19.4 ms vs. 88.4+/-20.7 ms, p=0.02). Female patients had lower right ventricular mean voltage (3.0+/-0.7 mV vs. 3.7+/-0.9 mV, p=0.03), and more low voltage zone over the right ventricular outflow tract free wall (27.0 % vs. 6.7 %, p=0.02). Eighty-one patients passed catheter ablation (23 males). The acute success rate, repeated catheter ablation rate and VT recurrence rate were similar in both genders. The present study provides evidence of the gender differences in electrophysiological findings in patients with idiopathic RVOT-VT. Studies on gender-specific differences in arrhythmia could lead to a better understanding of its mechanism(s) and provide valuable information for the development of optimal treatment strategies.

Development of microsatellite markers for the kelp grouper Epinephelus bruneus by 454 pyrosequencing and transfer to related species.

The kelp or longtooth grouper (Epinephelus bruneus), which inhabits Eastern Asia, is the most economically important of 11 grouper species that inhabit the Southern Sea near Jeju Island in Korea. This species is listed as vulnerable by the International Union for the Conservation of Nature and Natural Resources because of a rapid decrease in its resources. We developed microsatellite markers for E. bruneus using the pyrosequencing technique for applications in resource management and aquaculture. In addition, we tested the cross-species transferability of the microsatellite markers in four species belonging to the Epinephelus genus. Among 66,452 simple sequence repeats, 64 loci containing more than eight CA or TG repeats were randomly selected for primer synthesis; 45 primer sets (75.0%) produced polymerase chain reaction (PCR) products of 100-300 bp and were selected as candidates. After primary testing with four E. bruneus fish, 28 polymorphic loci were selected as the final microsatellite markers, and 23 sets showing clear amplification of polymorphic loci were used to analyze 71 fish. These loci have allele numbers ranging from 2 to 23. Null alleles were detected at three loci, and three loci showed an excess of homozygotes in the Hardy-Weinberg equilibrium test. Of the three species used for cross-species transfer of these markers, Epinephelus moara showed the highest transferability (92.9%) and polymorphism (67.9%), followed by Epinephelus fuscoguttatus (75.0 and 67.9%, respectively) and Epinephelus septemfasciatus (57.1 and 46.4%, respectively). These results suggested that these microsatellite loci should be valuable tools for population genetic studies of the species Epinephelus.

Estrogen can modulate menopausal women's heart rate variability.

The aim of our study was to compare the responses of heart rate variability (HRV) with two different types of hormonal substitution therapy (HT) in post-menopausal women (cross-sectional study) and to reveal an effect of HT shortly after beginning of its administration (follow-up study). To elucidate the influence of menopause and effects of different protocols of a HT on autonomic control of heart rate, we evaluated the heart rate variability (HRV) in 5 groups: premenopausal women (n=140), postmenopausal women without HT (n=360), women on HT with conjugated estrogen only (n=168), women on continuous combined estrogen-progesterone HT (n=117), and men (n=140). Frequency-domain of short-term stationary R-R intervals was performed to evaluate the total variance, low frequency power (LF; 0.04-0.15 Hz), high frequency power (HF; 0.15-0.40 Hz), portion of low frequency power (LF%) and ratio of LF to HF (LF/HF). Significantly lower portion of the LF was found in premenopausal women [46.9 (+/-2.7) nu] when compared to untreated postmenopausal women [54.3 (+/-2.9) nu] and men [55.2 (+/-3.0) nu]. Treatment by estrogen only was proved to decrease the LF% [40.1 (+/-2.1) nu] while no effect on HRV was observed in women treated with combination of estrogen and progesterone [57.2 (+/-3.1) nu]. Also the HF was lower in postmenopausal women [4.16 (+/-0.16) ms(2)] than in premenopausal women [4.79 (+/-0.22) ms(2)] and women treated with estrogen only [4.98 (+/-0.25) ms(2)] while in women treated with combined hormonal therapy the average value [3.99 (+/-0.21) ms(2)] did not significantly differ from that of untreated postmenopausal women. The follow-up study also proved increase of high frequency power already after two months of estrogen substitution therapy [4.86 (+/-0.14) ms(2) vs. 4.19 (+/-0.15) ms(2)]. These results suggest that higher vagal modulation of heart rate that seems typical for younger women becomes after menopause similar to that of men. We also proved a positive shift of HRV parameters toward more beneficial values as for a cardiovascular risk in postmenopausal women treated with estrogens but not in those treated by combined estrogen - progesterone substitution therapy.

Experimental evaluation of the cardiac rhythm originating in myocardial sleeves of pulmonary veins using a monophasic action potential.

Spontaneous depolarization similar to that from the sinus node was documented from the myocardial sleeves of pulmonary veins (PV) after isolation procedures. It was then hypothesized that sinus node-like tissue is present in the PVs of humans. Based on a number of features, the myocardium of myocardial sleeves (MS) is highly arrhythmogenic. Membrane potentials originating from MS are invariably recordable at the PVs ostia in patients with atrial fibrillation (AF) and delayed conduction around the PVs ostia may play a role in re-entry process responsible for the initiation and maintenance of AF. Diagnostic and therapeutic evidence of premature atrial beats induced in MS of PVs and resulting in launch of AF was detected by 3D electroanatomic method of monophasic action potential (MAP). MAP recording plays an important role in a direct view of human myocardial electrophysiology under both physiological and pathological conditions. Its crucial importance lies in the fact that it enables the study of the action potential of myocardial cell in vivo and, therefore, the study of the dynamic relation of this potential with all the organism variables. The knowledge of pathological MAPs from PV myocardial sleeves can help us to confirm a diagnosis when finding the similar action potential morphology. MAP can be also used to evaluate the therapeutic efficiency of vagal nerves suppression, radiofrequency ablation or other treatment procedures in PVs myocardial sleeves as well as for post-treatment following up.

Biomechanical characterization of a low density silicone elastomer filled with hollow microspheres for maxillofacial prostheses.

An ideal material for maxillofacial prostheses has not been found. We created a novel material: silicone elastomer filled with hollow microspheres and characterized its biomechanical properties. Expancel hollow microspheres were mixed with MDX4-4210 silicone elastomer using Q7-9180 silicone fluid as diluent. The volume fractions of microspheres were 0, 5, 15, and 30% v/v (volume ratio to the total volume of MDX4-4210 and microspheres). The microspheres dispersed well in the matrix. The physical properties and biocompatibility of the composites were examined. Shock absorption was the greatest by the 5% v/v composite, and decreased with increasing concentrations of microspheres. The density, thermal conductivity, Shore A hardness, tear and tensile strength decreased with increasing concentrations of microspheres, while elongation at break increased. Importantly, the tear strength of all composites was markedly lower than that of pure silicone elastomer. Cell viability assays indicated that the composite was of good biocompatibility. The composite with a volume fraction of 5% exhibited the optimal properties for use as a maxillofacial prosthesis, though its tear strength was markedly lower than that of silicone elastomer. In conclusion, we developed a novel light and soft material with good flexibility and biocompatibility, which holds a promising prospect for clinical application as maxillofacial prosthesis.

Rnh1 promotes differentiation and myelination via RhoA in oligodendrocytes.

Increases in Rattus norvegicus ribonuclease/angiogenin inhibitor 1 (Rnh1) are observed in rat primary neuron injury and/or the regeneration process and in differentiated oligodendrocytes. However, the roles of Rnh1 in the central nervous system are still largely unexplored. RhoA is an important signaling protein that has been implicated in oligodendrocyte differentiation and myelination. We demonstrate enhanced differentiation and myelination of oligodendrocytes mediated by Rnh1 in vitro. We further show that Rnh1 is expressed in oligodendrocyte precursors and oligodendrocytes. Importantly, Rnh1 strongly affects oligodendrocyte differentiation through RhoA-ROCK signaling. Moreover, changes in Rnh1 expression in oligodendrocytes regulates the expression and phosphorylation of Fyn, a regulator of RhoA activity. Finally, Rnh1 promotes myelination in vitro. These results show that Rnh1-mediated RhoA inactivation enhances the differentiation and myelination in oligodendrocytes. Overall, Rnh1 might contribute to oligodendrocyte differentiation and myelination processes in vitro.

SCIRR39 promotes differentiation of oligodendrocyte precursor cells and regulates expression of myelin-associated inhibitory factors.

SCIRR39 is an identified upregulated gene in rat primary neuron injury and/or regeneration process. However, roles of SCIRR39 in the regeneration of central nervous system (CNS) injury are still largely unexplored. Using real-time quantitative PCR and Western blotting, SCIRR39 expression was detected in oligodendrocyte precursor cells (OPCs) and oligodendrocytes. Moreover, the results from cell proliferation and cell cycle indicated that SCIRR39 inhibited OPCs proliferation and induced cell cycle arrest in G0/G1 and G2/M phases. Importantly, SCIRR39 positively regulated OPC differentiation and the expression of myelin basic protein. We also examined the effect of SCIRR39 on expression of myelin-associated inhibitory factors, including myelin-associated glycoprotein (MAG), oligodendrocyte myelin glycoprotein (OMgp), and Nogo A. Nogo A level was markedly regulated by SCIRR39 overexpression or knockdown in oligodendrocytes and cortical neurons co-cultures, while the expression of MAG and OMgp was not obviously changed by SCIRR39 overexpression or knockdown. Taken together, our results indicate the important role of SCIRR39 either in OPC differentiation or in axon myelination, and may provide a new therapeutic target for the treatment of CNS injury.

Interleukin-27 as a negative regulator of human neutrophil function.

Interleukin-27 (IL-27) is a novel cytokine of the IL-6/12 family with a broad range of immune regulation properties, which has been considered as a potential therapeutic agent for immune diseases and cancers. However, little is known about the effect of IL-27 on human neutrophils before its clinical administration. In this study, we investigated the effects of IL-27 on human neutrophil functions including adhesion, reactive oxygen species (ROS)/cytotoxic granule components production, inflammatory cytokines production, major histocompatibility complex (MHC) molecules expression and neutrophils' survival. We showed that IL-27 receptor complex, WSX-1/TCCR and gp130, is constitutively expressed on human neutrophils. In vitro, IL-27 suppressed neutrophil adhesion in response to fMLP, which might depend on the down-regulation of Mac-1. IL-27 also suppressed lipopolysaccharide-induced ROS production and attenuated cytotoxic granule components production in the cytoplasm of human neutrophils. In addition, IL-27 enhanced the production of IL-1ß but not TNF-α from neutrophils. However, IL-27 failed to regulate the expression of MHC molecules and the survival of human neutrophils. In conclusion, our data demonstrate that IL-27 mainly down-modulates human neutrophil function, which might extend our understanding of the role of IL-27 in the innate immune response.

New insights into application of cardiac monophasic action potential.

Monophasic action potential (MAP) recording plays an important role in a more direct view of human myocardial electrophysiology under both physiological and pathological conditions. The procedure of MAP measuring can be simply performed using the Seldinger technique, when MAP catheter is inserted through femoral vein into the right ventricle or through femoral artery to the left ventricle. The MAP method represents a very useful tool for electrophysiological research in cardiology. Its crucial importance is based upon the fact that it enables the study of the action potential (AP) of myocardial cell in vivo and, therefore, the study of the dynamic relation of this potential with all the organism variables. This can be particularly helpful in the case of arrhythmias. There are no doubts that physiological MAP recording accuracy is almost the same as transmembrane AP as was recently confirmed by anisotropic bidomain model of the cardiac tissue. MAP recording devices provide precise information not only on the local activation time but also on the entire local repolarization time course. Although the MAP does not reflect the absolute amplitude or upstroke velocity of transmembrane APs, it delivers highly accurate information on AP duration and configuration, including early afterdepolarizations as well as relative changes in transmembrane diastolic and systolic potential changes. Based on available data, the MAP probably reflects the transmembrane voltage of cells within a few millimeters of the exploring electrode. Thus MAP recordings offer the opportunity to study a variety of electrophysiological phenomena in the in situ heart (including effects of cycle length changes and antiarrhythmic drugs on AP duration).

DFT study on the bromination pattern dependence of electronic properties and their validity in quantitative structure-activity relationships of polybrominated diphenyl ethers.

With quantum chemical computation of density functional theory (DFT), the electronic properties including the polarisabilities, polarisability anisotropies and quadrupole moments of a total of 209 congeners of polybrominated diphenyl ethers (PBDEs) were evaluated. The electronic properties were shown to be highly dependent on the bromination pattern, i.e. their values changed sensitively with the number and sites of bromination. Being similar to the 2,3,7,8-, 1,4,6,9-chlorination of dioxins, respectively, 3,3',4,4'-, 2,2',5,5'-bromination of PBDEs can impose relatively greater effects on the electronic properties. Some of electronic properties were found to be potent in explaining the variance of toxicity, and the potency was verified by the development of quantitative structure-activity relationships (QSARs). To further improve the stability and predictability of QSARs for toxicity, two-dimensional topological indices were introduced. In QSARs, polarisability anisotropy was more significant than other polarisability tensors, indicating the implicit occurrence of dispersion interaction between the ligand and aryl hydrocarbon receptor (AhR). For PBDEs, the quadrupole moment was as significant as shown previously for dioxins. As interesting descriptors with encoded information about dispersion and electronics, the electronic properties analysed herein are helpful in obtaining a better understanding of the congener-specific toxicities of PBDEs, and are applicable and may be extended to research into the toxicology of structurally similar compounds, such as halogenated aromatics.

Decreased urinary secretion of belotecan in folic acid-induced acute renal failure rats due to down-regulation of Oat1 and Bcrp.

The effects of folic acid-induced acute renal failure on the renal excretion of belotecan were investigated in rats after intravenous administration. Both glomeruli and renal tubules were seriously damaged by folic acid-induced acute renal failure. The renal excretion clearance, CLr, of belotecan was significantly decreased by folic acid-induced acute renal failure. Furthermore, glomerular filtration rate and secretion clearance of the drug were dramatically decreased by folic acid-induced acute renal failure. In vivo renal uptake of belotecan was inhibited by p-aminohippurate, whereas renal excretion was inhibited by GF120918, but not by verapamil and bromosulphalein. This indicates that Oat1/3 and Bcrp are involved in the renal uptake and urinary excretion of belotecan, respectively. Both mRNA and protein levels of Oat1, Oat3 and Bcrp were significantly decreased in folic acid-induced acute renal failure rats. Based on the finding that belotecan is a substrate of OAT1 but not of OAT3, the decrease in CLr of belotecan in folic acid-induced acute renal failure could, therefore, mainly be attributed to the down-regulation of Oat1 and Bcrp, in addition to the decrease in glomerular filtration rate.

The curative effect of ALSS on 1-month mortality in AoCLF patients after 72 to 120 hours.

BACKGROUND: Hepatitis B virus (HBV)-related acute-on-chronic liver failure (AoCLF) is associated with a high mortality rate. An artificial liver support system (ALSS) creates a better environment for the self-regeneration of retained hepatocytes. AIM AND PATIENTS: We investigated the curative effect of ALSS on 1-month mortality at 72-120 h post-ALSS in 289 AoCLF patients. METHODS: Of the 289 patients, 117 who survived for at least 1 month post-ALSS comprised the survival group; the remaining cases who died within 1 month served as controls. The improvements in laboratory data and clinical syndromes at 72-120 h post-ALSS were compared with those at 24 h. RESULTS: Total bilirubin, international normalized ratio, and creatinine levels, and encephalopathy were significantly improved at 24 h post-ALSS in both the groups (p<0.05); however, these variables showed deterioration at 72-120 h; a rebound occurred in the nonsurvivors (p>0.05). The improvements in these variables in the nonsurvivors were considerably smaller than those in the survivors (p<0.05), particularly at 72-120 h. One-month mortality was more accurately predicted by the logistic regression model at 72-120 h than at 24 h. CONCLUSIONS: The prognosis of AoCLF patients was highly dependent on the improvement in encephalopathy, total bilirubin, international normalized ratio, and creatinine levels at 72-120 h post-ALSS. These variables are useful, therefore, as disease severity indexes to determine organ allocation priorities for liver transplant.

Stable bioavailability of cyclosporin A, regardless of food intake, from soft gelatin capsules containing a new self-nanoemulsifying formulation.

AIM: We recently succeeded in preparing soft gelatin capsules containing a new self-nanoemulsifying formulation consisting of cyclosporin A (CsA), triacetin, polyoxyl 40 hydrogenated castor oil, polysorbate 20, medium chain triglycerides and medium chain mono- and diglycerides. The soft capsules containing the new formulation exhibited a significantly improved physical stability in terms of the appearance of the gelatin capsule shells and the composition of the fill mass during long-term storage, compared to commercially available soft capsules containing CsA, in which ethanol was employed as a cosolvent of CsA. In the present study, the influence of a fat-rich meal on the bioavailability of CsA from the soft capsule containing the new formulation (test drug) was evaluated and the results compared to those obtained with a representative soft capsule of CsA. VOLUNTEERS AND METHODS: A randomized, open-label, 3-way crossover study was performed in the test capsules and reference soft capsules, in a fasted state or after a fat-rich breakfast. 18 healthy male volunteers received a single dose of the reference formulation (Neoral, Novartis AG, Basel, Switzerland) or test formulation (2 capsules each, 200 mg as CsA) with 240 ml of water with a 1-week washout period between the treatments, after a fat-rich (670 kcal, 45 g fat) breakfast (for the test drug, Treatment A; for the reference drug, Treatment B) or a 12-h fasting (for the test drug, Treatment C). Serial blood samples, collected over a 24-h period after the administration, were assayed for blood CsA concentrations using a specific monoclonal radioimmunoassay. RESULTS: The differences in bioavailability parameters (i.e., AUC(0-24h), AUC(0-infinity) and C(max)) between the treatments were within the range of 80-125% of the reference treatment. An analysis of variance (ANOVA) revealed no significant differences (p > 0.05) between subjects, formulations or periods. The 90% confidence intervals (CI) indicated that the differences between the treatments (Treatments A and B, Treatments A and C) were also within the criteria. CONCLUSION: These results indicate that the bioavailability of CsA from the test drug is equivalent to reference in the fed state, and is likely to be less influenced by a fat-rich meal. Therefore, the new formulation of CsA using triacetin appears to have an advantage over the commercial soft capsules of CsA using a volatile cosolvent such as ethanol.