Circular RNA KIAA0564 Serves as a Competitive Endogenous RNA for MicroRNA-424-5p, Mediating the Expression of Lysine Demethylase 4a, Thereby Facilitating Intervertebral Disc Degeneration.

A growing body of research has confirmed the involvement of circular RNAs (circRNAs) in the regulation of intervertebral disc degeneration (IDD) progression. However, the underlying molecular networks remain largely elusive. This study aimed to explore whether a novel circRNA, named circKIAA0564, affects nucleus pulposus (NP) cell injury and to elucidate its molecular mechanism. Both in vivo and in vitro IDD models were established, and the expression patterns of circKIAA0564/miR-424-5p/lysine demethylase 4a (KDM4A) were evaluated through quantitative reverse transcription PCR and Western blot analysis. Actinomycin D, RNase R, and Northern blotting were utilized to assess the circular structure of circKIAA0564. The Cell Counting Kit-8, flow cytometry, enzyme-linked immunosorbent assay, commercial assay kits, Western blotting, and reactive oxygen species (ROS) probes were employed to assess the inflammatory and oxidative stress status in NP cells and tissues. Hematoxylin and eosin and TUNEL staining were used to evaluate pathological damage in mouse NP tissues. RNA immunoprecipitation and dual-luciferase reporter assays were conducted to assess the direct targeting relationships among circKIAA0564, miR-424-5p, and KDM4A. CircKIAA0564 was found to be abnormally overexpressed in IDD, functioning as a novel circRNA. Knockdown of circKIAA0564 ameliorated interleukin-1 beta (IL-1ß)-induced inflammation and oxidative stress in NP cells. The therapeutic effect of circKIAA0564 knockdown on NP cells was reversed by the silencing of miR-424-5p. Overexpression of circKIAA0564 exacerbated IL-1ß-induced NP cell injury, a process that was reversed by knockdown of KDM4A. CircKIAA0564 activated the toll-like receptor 4 (TLR4)/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)/NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) signaling pathway by regulating the miR-424-5p/KDM4A axis. CircKIAA0564 exacerbates IL-1ß-induced inflammation and oxidative stress in NP cells by competitively binding miR-424-5p, thereby mediating KDM4A and activating the TLR4/NF-κB/NLRP3 signaling pathway. These findings provide robust data support for targeted therapy of IDD and the development of future pharmaceuticals.

Thermodynamic Modeling and Exergy Analysis of A Combined High-Temperature Proton Exchange Membrane Fuel Cell and ORC System for Automotive Applications.

A combined system consisting of a high-temperature proton exchange membrane fuel cell (HT-PEMFC) and an organic Rankine cycle (ORC) is provided for automotive applications in this paper. The combined system uses HT-PEMFC stack cathode exhaust gas to preheat the inlet gas and the ORC to recover the waste heat from the stack. The model of the combined system was developed and the feasibility of the model was verified. In addition, the evaluation index of the proposed system was derived through an energy and exergy analysis. The numerical simulation results show that the HT-PEMFC stack, cathode heat exchanger, and evaporator contributed the most to the total exergy loss of the system. These components should be optimized as a focus of future research to improve system performance. The lower current density increased the ecological function and the system efficiency, but reduced the system's net out-power. A higher inlet temperature and higher hydrogen pressures of the stack and the lower oxygen pressure helped improve the system performance. Compared to the HT-PEFC system without an ORC subsystem, the output power of the combined system was increased by 12.95%.

Performance Analysis of a HT-PEMFC System with 6FPBI Membranes Doped with Cross-Linkable Polymeric Ionic Liquid.

In this paper, a high-temperature proton-exchange membrane fuel cell (HT-PEMFC) system using fluorine-containing polybenzimidazole (6FPBI) composite membranes doped with cross-linkable polymer ionic liquid (cPIL) is developed and studied. The reliability of the model is verified by a comparison with the experimental data. The performance of the HT-PEMFC system using 6FPBI membranes with different levels of cPIL is analyzed. The results show that when the HT-PEMFC uses 6FPBI membranes with a cPIL content of 20 wt % (6FPBI-cPIL 20 membranes), the single cell power density is 4952.3 W·m-2. The excessive cPIL content will lead to HT-PEMFC performance degradation. The HT-PEMFC system using the 6FPBI-cPIL 20 membranes shows a higher performance, even at higher temperatures and pressures, than the systems using 6FPBI membranes. In addition, the parametric study results suggest that the HT-PEMFC system should be operated at a higher inlet temperature and hydrogen pressure to increase system output power and efficiency. The oxygen inlet pressure should be reduced to decrease the power consumption of the ancillary equipment and improve system efficiency. The proposed model can provide a prediction for the performance of HT-PEMFC systems with the application of phosphoric-acid-doped polybenzimidazole (PA-PBI) membranes.

Simultaneous determination of TPN729 and its five metabolites in human plasma and urine by liquid chromatography coupled to tandem mass spectrometry.

A specific and sensitive method was firstly developed using high performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS) to simultaneously quantify TPN729 and its metabolites (TPN729-D1, TPN729-D2, TPN729M15-3 and TPN729M3) in human plasma and (TPN729-D1, TPN729-D2, TPN729M15-3 and TPN729M14) in human urine. Protein precipitation and direct dilution were used to extract TPN729 and its metabolites from plasma and urine, respectively. Ionization of TPN729, TPN729-D1, TPN729-D2, TPN729M15-3, TPN729M3, TPN729M14 and sildenafil (internal standard, IS) was performed using an electrospray ionization (ESI) source in positive mode and detection was carried out with multiple reaction monitoring (MRM) mode. This assay method for TPN729 and its five metabolites has been fully validated in terms of sensitivity, linearity, lower limit of quantification (LLOQ), precision, accuracy, stability, matrix effect and recovery. The LLOQ of TPN729/TPN729-D1/TPN729-D2/TPN729M15-3/TPN729M3 in human plasma and TPN729/TPN729-D1/TPN729-D2/TPN729M15-3/TPN729M14 in human urine were 0.200/0.500/2.00/0.500/1.00 ng/mL and 4.00/2.50/10.0/2.50/1.00 ng/mL, respectively. Inter- and intra-batch precision of TPN729 and its metabolites were less than 15% and the accuracy was within ±15% for both plasma and urine. The extraction recoveries of all analytes at three concentration levels were consistent. In conclusion, the validation results showed that this method was robust, specific, and sensitive and it can successfully fulfill the requirement of clinical pharmacokinetic study of TPN729 in Chinese healthy subjects.

A single and multiple postprandial dose study investigating the pharmacokinetics and pharmacodynamics of edoxaban in healthy Chinese volunteers
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AIMS: This study investigated the pharmacokinetics (PK) and pharmacodynamics (PD) of once-daily postprandial doses of edoxaban 60 mg in healthy Chinese subjects. METHODS: 6 male and 6 female healthy Chinese volunteers, aged 18 - 45 years, were enrolled into this open-label, phase-I trial. Subjects received single oral doses of edoxaban 60 mg after a meal, followed by successive once-daily doses for 7 days. Serial blood samples were taken pre- and postdose to measure plasma concentrations of edoxaban and its major active metabolite D21-2393 as well as prothrombin time (PT) and activated partialprothrombin time (aPTT). Safety was assessed throughout the study. RESULTS: Eoxaban was quickly absorbed after dosing. The resultant maximum and total exposure of edoxaban after single postprandial dose were similar to those after the same dose in fasting condition, but tmax was about half an hour longer. Meanwhile, the exposure of D21-2393 and the metabolite-over-parent ratio were both lower vs. the fasting condition, suggesting involvement of food on D21-2393 formation. Steady state was attained after two successive daily doses. The PK parameters of edoxaban with multiple postprandial doses were comparable to those observed in Caucasian and Japanese volunteers. Similarly, the PD profiles and the concentration-response relationship of edoxaban were not changed with repeated doses. Minor bleeding was the most commonly reported adverse event during the study. CONCLUSION: Once daily postprandial doses of edoxaban 60 mg was safe and well tolerated in healthy Chinese volunteers. The PK and PD characteristics of edoxaban were comparable among Chinese, Caucasian, and Japanese subjects.
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A single-dose study investigating the pharmacokinetics and pharmacodynamics of edoxaban at 30-90 mg in healthy Chinese volunteers.

1. This study investigated the pharmacokinetics and pharmacodynamics of single-dose edoxaban in healthy Chinese subjects. 2. This single-centre, open-label, 3-occasion, cross-over study enrolled six males and six females. Subjects received a single-oral-dose of edoxaban 30-90 mg in each study occasion under fasting condition. Serial blood samples were collected to measure the plasma concentrations of edoxaban and its major active metabolite D21-2393. Meanwhile, PT, INR, aPTT were measured pre- and post-dose. Safety was assessed. 3. After administration, edoxaban was rapidly absorbed (median Tmax = 1-2 h). With rapid transformation, peak concentration of D21-2393 was reached within 2-h post-dose. The elimination half-life of edoxaban was 5-20 h. The dose-exposure relationships were slightly lower-than-dose-proportional for both edoxaban and D21-2393. Although women had higher plasma exposure of edoxaban and D21-2393 than men, it was considered clinically insignificant. 4. The effects of edoxaban on all pharmacodynamic biomarkers were concentration-dependent and linearly-correlated to plasma level of the compound. Minor bleeding was the most commonly reported adverse events. 5. Single oral doses of edoxaban 30-90 mg were safe and well tolerated in healthy Chinese volunteers. 6. The pharmacokinetic and pharmacodynamic profiles of edoxaban in Chinese subjects were comparable to those observed in Caucasian and Japanese populations.

A unified strategy in selection of the best allometric scaling methods to predict human clearance based on drug disposition pathway.

1. It is critical to develop a unified strategy to select the best allometric scaling (AS) method for a given group of drugs. 2. A total of 446 drugs with known human CLiv, clear disposition pathway and animal (rat, dog, monkey) CLiv were analyzed. All drugs were stratified based on their disposition pathway, liver extraction ratio (ERH) and ratios of unbound clearance to renal glomerular filtration rate (RGFR). Up to 22 AS methods were applied and compared in prediction of human CLiv to each group of drugs. 3. AS methods that give the best prediction of human CLiv, were identified for drugs primarily eliminated through liver with a fraction of renal elimination (frenal) within 0.3-0.5 or ERH > 0.3, where human CLiv of more than 80% or 90% drugs could be accurately (within 2- or 3-fold error) predicted. For drugs with ERH < 0.3, acceptable accuracy could be achieved by a two species method TSR,D resulting more than 60% or 75% drugs were predicted within 2- or 3-fold error. 4. By stratified analysis of drugs, according to their disposition pathway and organ extraction ratio, a unified strategy was developed to select the best AS method in prediction of human CLiv.

Tissue distribution and ontogeny of multidrug resistance protein 2, a phosphatidylcholine translocator, in rats.

Multidrug resistance protein 2 (Mdr2), encoded by ATP-binding cassette b4 (Abcb4), serves as a phospholipid flippase that is indispensable for phosphatidylcholine translocation. However, little was known about the regulation of Mdr2 in Sprague-Dawley rats, although they are commonly used for pre-clinical investigation as well as mechanistic study. Present study aims at determining the tissue distribution, gender difference and ontogeny of Mdr2 in rats on both gene and protein levels. Results showed that Mdr2 was highly expressed in liver, modestly enriched in brain and testis, and less distributed in gastrointestinal tracts. Gender-divergent and male-dominated distribution was observed in the Mdr2 mRNA expression of liver and generative organs. Developmental pattern of rat Mdr2 on protein level was not exactly consistent with that on mRNA level. In conclusion, there was a considerable distribution of rat Mdr2 in the brain, testis and intestine besides liver, and the ontogeny of Mdr2 performed in an age-dependent pattern with the post-transcriptional regulation.

Simultaneous determination of imigliptin and its three metabolites in human plasma and urine by liquid chromatography coupled to tandem mass spectrometry.

A specific and sensitive method was firstly developed using high performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS) to simultaneously quantify imigliptin (KBP-3853) and its three metabolites (KBP-3926, KBP-3902, KBP-5493) in human plasma and urine. Solid-phase extraction (SPE) and direct dilution were used to extract imigliptin and its three metabolites from plasma and urine, respectively. The extracts were injected onto a SymmetryShield RP8 column with a gradient elution of acetonitrile and water containing 5mM ammonium acetate (pH 7). Ionization of KBP-3853, KBP-3926, KBP-3902, KBP-5493, and XZP-3244 (internal standard, IS) was performed using an electrospray ionization (ESI) source in positive mode and detection was carried out with multiple reaction monitoring (MRM) mode. The lower limits of quantitation (LLOQ) of KBP-3853/KBP-3926/KBP-3902/KBP-5493 in human plasma and urine were 0.500/0.500/0.500/0.500ng/mL and 20.0/20.0/10.0/10.0ng/mL, respectively. Inter- and intra-batch precision of imigliptin and its three metabolites were less than 15% and the accuracy was within 85-115% for both plasma and urine. The extraction recoveries of all analytes at three concentration levels were consistent. The specificity, matrix effect, linearity and stabilities under various conditions were validated for imigliptin and its three metabolites in human plasma and urine. In conclusion, the validation results showed that this method was robust, specific, and sensitive and it can successfully fulfill the requirement of clinical pharmacokinetic study of imigliptin hydrochloride in Chinese healthy subjects.

Pharmacokinetics study of calf thymus DNA in rats and beagle dogs with (3)H-labeling and tracing method.

This study developed a radioisotope detection and tracing method to investigate the pharmacokinetic properties of calf thymus DNA (ctDNA) in rats and beagle dogs. The radioactivity labeling result was detected through gel electrophoresis analysis, and pharmacokinetic analytical methods for (3)H-ctDNA in rat and beagle dog plasma were developed, respectively. Full method validation indicated that the established radioisotope method was sensitive, specific, rapid and reliable, and the results were all in accordance with the analysis requirement in biological samples. After intravenous administration of the planned doses of (3)H-ctDNA to the rats and beagle dogs, plasma concentrations from the various dose groups declined rapidly. In addition, the radioactive concentration of (3)H-ctDNA in the plasma from single and multiple dosings decreased in a similar trend. Through comparative analysis of the pharmacokinetic parameters, we inferred that the elimination of ctDNA accorded with the linear pharmacokinetic characteristic. The results demonstrated that ctDNA was rapidly eliminated in rat and beagle dog plasma and would not accumulate, indicating the safe use of ctDNA as an immunoadsorptive material without bringing out potential risk.