The immune-evasive proline-283 substitution in influenza nucleoprotein increases aggregation propensity without altering the native structure.

Nucleoprotein (NP) is a key structural protein of influenza ribonucleoprotein complexes and is central to viral RNA packing and trafficking. NP also determines the sensitivity of influenza to myxovirus resistance protein 1 (MxA), an innate immunity factor that restricts influenza replication. A few critical MxA-resistant mutations have been identified in NP, including the highly conserved proline-283 substitution. This essential proline-283 substitution impairs influenza growth, a fitness defect that becomes particularly prominent at febrile temperature (39°C) when host chaperones are depleted. Here, we biophysically characterize proline-283 NP and serine-283 NP to test whether the fitness defect is caused by the proline-283 substitution introducing folding defects. We show that the proline-283 substitution changes the folding pathway of NP, making NP more aggregation prone during folding, but does not alter the native structure of the protein. These findings suggest that influenza has evolved to hijack host chaperones to promote the folding of otherwise biophysically incompetent viral proteins that enable innate immune system escape.

Mitochondrial nucleic acids in innate immunity and beyond.

Mitochondria participate in a wide range of cellular processes. One essential function of mitochondria is to be a platform for antiviral signaling proteins during the innate immune response to viral infection. Recently, studies have revealed that mitochondrion-derived DNAs and RNAs are recognized as non-self molecules and act as immunogenic ligands. More importantly, the cytosolic release of these mitochondrial nucleic acids (mt-NAs) is closely associated with the pathogenesis of human diseases accompanying aberrant immune activation. The release of mitochondrial DNAs (mtDNAs) via BAX/BAK activation and/or VDAC1 oligomerization activates the innate immune response and inflammasome assembly. In addition, mitochondrial double-stranded RNAs (mt-dsRNAs) are sensed by pattern recognition receptors in the cytosol to induce type I interferon expression and initiate apoptotic programs. Notably, these cytosolic mt-NAs also mediate adipocyte differentiation and contribute to mitogenesis and mitochondrial thermogenesis. In this review, we summarize recent studies of innate immune signaling pathways regulated by mt-NAs, human diseases associated with mt-NAs, and the emerging physiological roles of mt-NAs.

The Immune-Evasive Proline 283 Substitution in Influenza Nucleoprotein Increases Aggregation Propensity Without Altering the Native Structure.

Nucleoprotein (NP) is a key structural protein of influenza ribonucleoprotein complexes and is central to viral RNA packing and trafficking. In human cells, the interferon induced Myxovirus resistance protein 1 (MxA) binds to NP and restricts influenza replication. This selection pressure has caused NP to evolve a few critical MxA-resistant mutations, particularly the highly conserved Pro283 substitution. Previous work showed that this essential Pro283 substitution impairs influenza growth, and the fitness defect becomes particularly prominent at febrile temperature (39 °C) when host chaperones are depleted. Here, we biophysically characterize Pro283 NP and Ser283 NP to test if the fitness defect is owing to Pro283 substitution introducing folding defects. We show that the Pro283 substitution changes the folding pathway of NP without altering the native structure, making NP more aggregation prone during folding. These findings suggest that influenza has evolved to hijack host chaperones to promote the folding of otherwise biophysically incompetent viral proteins that enable innate immune system escape. Teaser: Pro283 substitution in flu nucleoprotein introduces folding defects, and makes influenza uniquely dependent on host chaperones.

Resveratrol Attenuates the Mitochondrial RNA-Mediated Cellular Response to Immunogenic Stress.

Human mitochondria contain a circular genome that encodes 13 subunits of the oxidative phosphorylation system. In addition to their role as powerhouses of the cells, mitochondria are also involved in innate immunity as the mitochondrial genome generates long double-stranded RNAs (dsRNAs) that can activate the dsRNA-sensing pattern recognition receptors. Recent evidence shows that these mitochondrial dsRNAs (mt-dsRNAs) are closely associated with the pathogenesis of human diseases that accompany inflammation and aberrant immune activation, such as Huntington's disease, osteoarthritis, and autoimmune Sjögren's syndrome. Yet, small chemicals that can protect cells from a mt-dsRNA-mediated immune response remain largely unexplored. Here, we investigate the potential of resveratrol (RES), a plant-derived polyphenol with antioxidant properties, on suppressing mt-dsRNA-mediated immune activation. We show that RES can revert the downstream response to immunogenic stressors that elevate mitochondrial RNA expressions, such as stimulation by exogenous dsRNAs or inhibition of ATP synthase. Through high-throughput sequencing, we find that RES can regulate mt-dsRNA expression, interferon response, and other cellular responses induced by these stressors. Notably, RES treatment fails to counter the effect of an endoplasmic reticulum stressor that does not affect the expression of mitochondrial RNAs. Overall, our study demonstrates the potential usage of RES to alleviate the mt-dsRNA-mediated immunogenic stress response.

Viral Evolution Shaped by Host Proteostasis Networks.

Understanding the factors that shape viral evolution is critical for developing effective antiviral strategies, accurately predicting viral evolution, and preventing pandemics. One fundamental determinant of viral evolution is the interplay between viral protein biophysics and the host machineries that regulate protein folding and quality control. Most adaptive mutations in viruses are biophysically deleterious, resulting in a viral protein product with folding defects. In cells, protein folding is assisted by a dynamic system of chaperones and quality control processes known as the proteostasis network. Host proteostasis networks can determine the fates of viral proteins with biophysical defects, either by assisting with folding or by targeting them for degradation. In this review, we discuss and analyze new discoveries revealing that host proteostasis factors can profoundly shape the sequence space accessible to evolving viral proteins. We also discuss the many opportunities for research progress proffered by the proteostasis perspective on viral evolution and adaptation.

Analysis of mitochondrial double-stranded RNAs in human cells.

Human mitochondrial genome is transcribed bidirectionally, generating long complementary RNAs that can form double-stranded RNAs (mt-dsRNAs). When released to the cytosol, these mt-dsRNAs can activate antiviral signaling. Here, we present a detailed protocol for the analysis of mt-dsRNA expression. The protocol provides three approaches that can complement one another in examining mt-dsRNAs. While the described protocol is optimized for human cells, this approach can be adapted for use in other animal cell lines and tissue samples. For complete details on the use and execution of this protocol, please refer to Kim et al. (2022).1.

Mitochondrial double-stranded RNAs as a pivotal mediator in the pathogenesis of SjÓ§gren's syndrome.

SjÓ§gren's syndrome (SS) is a systemic autoimmune disease that targets the exocrine glands, resulting in impaired saliva and tear secretion. To date, type I interferons (I-IFNs) are increasingly recognized as pivotal mediators in SS, but their endogenous drivers have not been elucidated. Here, we investigate the role of mitochondrial double-stranded RNAs (mt-dsRNAs) in regulating I-IFNs and other glandular phenotypes of SS. We find that mt-dsRNAs are elevated in the saliva and tears of SS patients (n = 73 for saliva and n = 16 for tears) and in salivary glands of non-obese diabetic mice with salivary dysfunction. Using the in-house-developed 3D culture of immortalized human salivary gland cells, we show that stimulation by exogenous dsRNAs increase mt-dsRNAs, activate the innate immune system, trigger I-IFNs, and promote glandular phenotypes. These responses are mediated via the Janus kinase 1 (JAK1)/signal transducer and activator of transcription (STAT) pathway. Indeed, a small chemical inhibitor of JAK1 attenuates mtRNA elevation and immune activation. We further show that muscarinic receptor ligand acetylcholine ameliorates autoimmune characteristics by preventing mt-dsRNA-mediated immune activation. Last, direct suppression of mt-dsRNAs reverses the glandular phenotypes of SS. Altogether, our study underscores the significance of mt-dsRNA upregulation in the pathogenesis of SS and suggests mt-dsRNAs as propagators of a pseudo-viral signal in the SS target tissue.

Double-stranded RNA induction asa potential dynamic biomarkerfor DNA-demethylating agents.

Hypomethylating agents (HMAs), such as azacitidine and decitabine, induce cancer cell death by demethylating DNAs to promote the expression of tumor-suppressor genes. HMAs also reactivate the transcription of endogenous double-stranded RNAs (dsRNAs) that trigger the innate immune response and subsequent apoptosis via viral mimicry. However, the expression patterns of endogenous dsRNAs and their relevance in the efficacy of HMAs remain largely uninvestigated. Here, we employ amidine-conjugated spiropyran (Am-SP) to examine the dynamic expression pattern of total dsRNAs regulated by HMAs. By analyzing the bone-marrow aspirates of myelodysplastic syndrome or acute myeloid leukemia patients who received the HMAs, we find a dramatic increase in total dsRNA levels upon treatment only in patients who later benefited from the therapy. We further apply our approach in solid tumor cell lines and show that the degree of dsRNA induction correlates with the effectiveness of decitabine in most cases. Notably, when dsRNA induction is accompanied by increased expression of nc886 RNA, decitabine becomes ineffective. Collectively, our study establishes the potential application of monitoring the total dsRNA levels by a small molecule as an analytical method and a dynamic marker to predict the clinical outcome of the HMA therapy.

Mitochondrial double-stranded RNAs govern the stress response in chondrocytes to promote osteoarthritis development.

Protein kinase R (PKR) is an immune response protein that becomes activated by double-stranded RNAs (dsRNAs). PKR overactivation is associated with degenerative diseases with inflammation, including osteoarthritis (OA), but the dsRNA activator remains largely unknown. Here, we find that mitochondrial dsRNA (mt-dsRNA) expression and its cytosolic efflux are facilitated in chondrocytes under OA-eliciting conditions, leading to innate immune activation. Moreover, mt-dsRNAs are released to the extracellular space and activate Toll-like receptor 3 at the plasma membrane. Elevated levels of mt-dsRNAs in the synovial fluids and damaged cartilage of OA patients and in the cartilage of surgery-induced OA mice further support our data. Importantly, autophagy prevents PKR activation and protects chondrocytes from mitochondrial stress partly by removing cytosolic mtRNAs. Our study provides a comprehensive understanding of innate immune activation by mt-dsRNAs during stress responses that underlie the development of OA and suggests mt-dsRNAs as a potential target for chondroprotective intervention.

Image Quality of High-Resolution 3-Dimensional Neck MRI Using CAIPIRINHA-VIBE and GRASP-VIBE: An Intraindividual Comparative Study.

OBJECTIVES: Acquiring high-quality magnetic resonance imaging (MRI) of the head and neck region is often challenging due to motion and susceptibility artifacts. This study aimed to compare image quality of 2 high-resolution three-dimensional (3D) MRI sequences of the neck, controlled aliasing in parallel imaging results in higher acceleration (CAIPIRINHA)-volumetric interpolated breath-hold examination (VIBE), and golden-angle radial sparse parallel imaging (GRASP)-VIBE. MATERIALS AND METHODS: One hundred seventy-three patients indicated for contrast-enhanced neck MRI examination were scanned using 3 T scanners and both CAIPIRINHA-VIBE and GRASP-VIBE with nearly isotropic 3D acquisitions (<1 mm in-plane resolution with analogous acquisition times). Patients' MRI scans were independently rated by 2 radiologists using a 5-grade Likert scale for overall image quality, artifact level, mucosal and lesion conspicuity, and fat suppression degree at separate anatomical regions. Interobserver agreement was calculated using the Cohen κ coefficient. The quality ratings of both sequences were compared using the Mann-Whitney U test. Nonuniformity and contrast-to-noise ratio values were measured in all subjects. Separate MRI scans were performed twice for each sequence in a phantom and healthy volunteer without contrast injection to calculate the signal-to-noise ratio (SNR). RESULTS: The scores of overall image quality, overall artifact level, motion artifact level, and conspicuity of the nasopharynx, oropharynx, oral cavity, hypopharynx, and larynx were all significantly higher in GRASP-VIBE than in CAIPIRINHA-VIBE (all P 's < 0.001). Moderate to substantial interobserver agreement was observed in overall image quality (GRASP-VIBE κ = 0.43; CAIPIRINHA-VIBE κ = 0.59) and motion artifact level (GRASP-VIBE κ = 0.51; CAIPIRINHA-VIBE κ = 0.65). Lesion conspicuity was significantly higher in GRASP-VIBE than in CAIPIRINHA-VIBE ( P = 0.005). The degree of fat suppression was weaker in the lower neck regions in GRASP-VIBE (3.90 ± 0.72) than in CAIPIRINHA-VIBE (4.97 ± 0.21) ( P < 0.001). The contrast-to-noise ratio at hypopharyngeal level was significantly higher in GRASP-VIBE (6.28 ± 4.77) than in CAIPIRINHA-VIBE (3.14 ± 9.95) ( P < 0.001). In the phantom study, the SNR of GRASP-VIBE was 12 times greater than that of CAIPIRINHA-VIBE. The in vivo SNR of the volunteer MRI scan was 13.6 in CAIPIRINHA-VIBE and 20.7 in GRASP-VIBE. CONCLUSIONS: Both sequences rendered excellent images for head and neck MRI scans. GRASP-VIBE provided better image quality, as well as mucosal and lesion conspicuities, with less motion artifacts, whereas CAIPIRINHA-VIBE provided better fat suppression in the lower neck regions.

The endoplasmic reticulum proteostasis network profoundly shapes the protein sequence space accessible to HIV envelope.

The sequence space accessible to evolving proteins can be enhanced by cellular chaperones that assist biophysically defective clients in navigating complex folding landscapes. It is also possible, at least in theory, for proteostasis mechanisms that promote strict quality control to greatly constrain accessible protein sequence space. Unfortunately, most efforts to understand how proteostasis mechanisms influence evolution rely on artificial inhibition or genetic knockdown of specific chaperones. The few experiments that perturb quality control pathways also generally modulate the levels of only individual quality control factors. Here, we use chemical genetic strategies to tune proteostasis networks via natural stress response pathways that regulate the levels of entire suites of chaperones and quality control mechanisms. Specifically, we upregulate the unfolded protein response (UPR) to test the hypothesis that the host endoplasmic reticulum (ER) proteostasis network shapes the sequence space accessible to human immunodeficiency virus-1 (HIV-1) envelope (Env) protein. Elucidating factors that enhance or constrain Env sequence space is critical because Env evolves extremely rapidly, yielding HIV strains with antibody- and drug-escape mutations. We find that UPR-mediated upregulation of ER proteostasis factors, particularly those controlled by the IRE1-XBP1s UPR arm, globally reduces Env mutational tolerance. Conserved, functionally important Env regions exhibit the largest decreases in mutational tolerance upon XBP1s induction. Our data indicate that this phenomenon likely reflects strict quality control endowed by XBP1s-mediated remodeling of the ER proteostasis environment. Intriguingly, and in contrast, specific regions of Env, including regions targeted by broadly neutralizing antibodies, display enhanced mutational tolerance when XBP1s is induced, hinting at a role for host proteostasis network hijacking in potentiating antibody escape. These observations reveal a key function for proteostasis networks in decreasing instead of expanding the sequence space accessible to client proteins, while also demonstrating that the host ER proteostasis network profoundly shapes the mutational tolerance of Env in ways that could have important consequences for HIV adaptation.

Characterization of Medication Errors in a Medical Intensive Care Unit of a University Teaching Hospital in South Korea.

OBJECTIVES: The objective of this study was to characterize the current status of medication errors (MEs) throughout the medication therapy process from prescribing to use and monitoring in a medical intensive care unit (MICU) in Korea. METHODS: Four trained research pharmacists collected data through retrospectively reviewing electronic medical records for adults hospitalized in the MICU in 2017. The occurrence of MEs was determined through interprofessional team discussion led by an academic faculty pharmacist and a medical intensivist based on the medication administration records (MARs). The type of MEs and the consequent ME-related outcome severity were categorized according to the Pharmaceutical Care Network Europe and the National Coordinating Council for Medication Error Reporting and Prevention, respectively. RESULTS: Overall, electronic medical records for 293 patients with 78,761 MARs were reviewed in this study. At least one type of ME occurred in 271 patients (92.5%) in association with 16,203 MARs (21%), primarily caused by inappropriate dose (35.5%), drug (27.8%), and treatment duration (25.1%). Clinically significant harmful events occurred in 24 patients (8%), including life-threatening (n = 5) and death (n = 2) cases. The 2 patients died of enoxaparin-induced fatal hemorrhage and neutropenia associated with ganciclovir and cefepime. Antibiotics were the most common culprit medications leading to clinically significant harmful events. CONCLUSIONS: In conclusion, MEs are prevalent in the MICU in Korea, most commonly prescribing errors. Although mostly benign, harmful events including deaths may occur due to MEs, mainly associated with antibiotics. Systematic strategies to minimize these potentially fatal MEs are urgently needed.

Preoperative assessment of cervical lymph node metastases in patients with papillary thyroid carcinoma: Incremental diagnostic value of dual-energy CT combined with ultrasound.

PURPOSE: To determine whether dual-energy CT (DECT) has incremental diagnostic value when combined with ultrasound (US) in the diagnosis of metastatic cervical lymph nodes (LNs) in patients with papillary thyroid carcinoma (PTC). METHODS: This was a single-center retrospective cohort study of patients diagnosed with PTC between October 2019 and August 2020. US features of LNs to include hyperechogenicity, round shape, microcalcification, cystic component, and homogeneous/peripheral vascularity were considered suggestive of metastasis. The HU of arterial phase (HUarterial) and DECT-derived CT images [contrast media (CM) and areas under the 100 keV monoenergetic curve (AUC100keV)] were measured. Effective atomic numbers (Zeff), iodine concentration (mg/mL), and slope of the HU curve (λHU) were also obtained. The values for metastatic and benign LNs were compared using Student's t-test with false-discovery correction. Logistic regression with areas under the receiver operating characteristic curves (AUCs) were performed for predicting metastatic LNs. RESULTS: A total of 102 patients were included (49 metastatic and 53 benign LNs; mean age, 46±15 years). Metastatic LNs showed significantly higher values for HUarterial, CM, Zeff, λHU, AUC100keV, and iodine concentration (all, P = 0.001). In logistic regression, the HUarterial demonstrated the highest AUC (0.824; 95% confidence interval [CI], 0.751-0.897), followed by CM HU (0.762; 95% CI, 0.679-0.846). Combination of DECT parameters with US features improved the AUC from 0.890 to 0.941. CONCLUSION: Compared to US features alone, combination with DECT-derived quantitative parameters improved diagnostic performance in predicting metastatic cervical LNs in patients with PTC.

Standardized hot water extract from the leaves of Hydrangea serrata (Thunb.) Ser. alleviates obesity via the AMPK pathway and modulation of the gut microbiota composition in high fat diet-induced obese mice.

Obesity is an increasing health problem worldwide as it is the major risk factor for metabolic diseases. In the present study, we investigated the anti-obesity effects of WHS by examining its effects on high fat diet (HFD)-induced obese mice. Male C57BL/6 mice were fed either a normal diet (ND) or a high fat diet (HFD) with or without WHS. At the end of the experiment, we observed the changes in their body weight and white adipose tissue (WAT) weight and lipid profiles in plasma. We performed western blot and histological analyses of WAT and liver to elucidate the molecular mechanisms of action. We also conducted fecal 16S rRNA analysis for investigating the gut microbiota. Our results indicated that pre- and post-oral administration of WHS significantly prevented body weight gain and reduced body fat weight in HFD-induced obese mice. In addition, WHS was found to improve adipocyte hypertrophy and liver fat accumulation by regulating the AMPK and AKT/mTOR pathways. WHS ameliorated hyperlipidemia by reducing total cholesterol and low-density lipoprotein (LDL) and decreased the energy metabolism-related hormones, leptin and insulin, in mouse plasma. Furthermore, we found that WHS modulated gut dysbiosis by normalizing HFD-induced changes. Taken together, our in vivo data implicate that WHS can be considered as a potential dietary supplement for alleviating obesity.

Efficacy and safety of thread embedding acupuncture combined with acupuncture for chronic low back pain: A randomized, controlled, assessor-blinded, multicenter clinical trial.

BACKGROUND: Low back pain is a very common disease. Many patients with chronic low back pain (CLBP) have been treated by complementary and alternative medicine such as acupuncture (AT) treatment. A type of AT, thread embedding acupuncture (TEA), consists of a thread that can continually stimulate at the AT points and has mechanical and chemical effects. Although TEA was widely used in clinical practice, there was little evidence of its efficacy and safety for CLBP. METHODS: This clinical trial was randomized, controlled, assessor-blinded, two-armed, parallel, and conducted in multiple centers. Four Korean medical institutions recruited 38 outpatients with CLBP. The participants were randomly allocated to a treatment group (TEA combined with AT) or a control group (only AT) in a 1:1 ratio. All participants received conventional AT twice a week for 8 weeks (16 sessions) at 15 AT points (GV3 and bilateral BL23, BL24, BL25, BL26, BL40, BL60, and EX-B5) and the treatment group participants additionally received TEA once a week for 8 weeks (8 sessions) on 10 AT points in the multifidus, spinal erector, and lumbar quadrate muscles. The primary outcome measure of this study was the change of visual analog scale (VAS) from baseline (0 week) to the end of intervention (8 weeks). Secondary outcome measures included clinically relevant improvement (minimal clinically important difference) and 3% to 50% decrease on VAS, disability level (Korean version of Roland and Morris disability questionnaire), quality of life (Korean version of European quality of life 5dimension), global assessment (patient global impression of change), economic analysis, credibility test, and safety assessment. RESULTS: The treatment group showed a significant reduction in VAS scores when compared with the control group (-33.7 ±â€Š25.1 vs -15.6 ±â€Š17.0, P = .013). As for the secondary outcome measures, the treatment group showed significant difference in 50% decrease on VAS and patient global impression of change. There was no serious adverse event associated with TEA and AT. CONCLUSION: This clinical trial documents the efficacy and safety of TEA combined with AT for the management of CLBP.

Improved Modeling of Thioamide FRET Quenching by Including Conformational Restriction and Coulomb Coupling.

Thioamide-containing amino acids have been shown to quench a wide range of fluorophores through distinct mechanisms. Here, we quantitatively analyze the mechanism through which the thioamide functional group quenches the fluorescence of p-cyanophenylalanine (Cnf), tyrosine (Tyr), and tryptophan (Trp). By comparing PyRosetta simulations to published experiments performed on polyproline ruler peptides, we corroborate previous findings that both Cnf and Tyr quenching occurs via Förster resonance energy transfer (FRET), while Trp quenching occurs through an alternate mechanism such as Dexter transfer. Additionally, optimization of the peptide sampling scheme and comparison of thioamides attached to the peptide backbone and side chain revealed that the significant conformational restriction associated with the thioamide moiety results in a high sensitivity of the apparent FRET efficiency to underlying conformational differences. Moreover, by computing FRET efficiencies from structural models using a variety of approaches, we find that quantitative accuracy in the role of Coulomb coupling is required to explain contributions to the observed quenching efficiency from individual structures on a detailed level. Last, we demonstrate that these additional considerations improve our ability to predict thioamide quenching efficiencies observed during binding of thioamide-labeled peptides to fluorophore-labeled variants of calmodulin.

Quantitation of plasma and biliary cefpiramide concentrations in human samples using high-performance liquid chromatography.

Cefpiramide is frequently used to treat biliary infections. However, no bioanalytical method has been validated to quantitate cefpiramide in human samples, particularly in bile. Therefore, this study was conducted to develop a simple, selective and validated high-performance liquid chromatographic method to determine cefpiramide in human plasma and bile. A protein precipitation procedure was used to extract cefpiramide and cefoperazone (internal standard, IS) from 200 µl of plasma and bile. Utilizing a Capcell Pak C18 column (4.6 × 250 mm), cefpiramide and IS were separated using the timed-gradient mobile phase consisting of 0.1 m sodium acetate (pH 5.2) and acetonitrile at a flow rate of 1 ml/min with photodiode array detector (wavelength set at 273 nm). The calibration curves showed linearity at concentrations ranging from 1 to 150 µg/ml in both plasma and bile (r2 > 0.999). The within- and between-run coefficients of variation (CVs) for plasma samples were 0.570-4.43 and 1.10-2.76%, respectively; for bile samples, the within- and between-day precision (CV) was 0.814-6.34 and 2.05-4.00%, respectively. Our newly developed bioanalytical method was successfully employed to quantify cefpiramide concentrations in both plasma and bile at multiple time points in patients with acute cholangitis.

Prevalence and Seriousness of Analgesic-Induced Adverse Events in Korea: A 10-Year Nationwide Surveillance.

OBJECTIVES: The objectives of this study were to describe the prevalence and seriousness of analgesic-induced adverse events (AEs) and to identify factors associated with serious analgesic-related AEs in Korea. METHODS: Voluntarily reported analgesic-induced AEs to the Korea Adverse Event Reporting System from 2007 to 2016 were retrospectively reviewed. Analgesic medications were classified into nonopioids and opioids based on the Anatomical Therapeutic Chemical classification system. All AEs were grouped using System Organ Classes according to the World Health Organization-Adverse Reaction Terminology. Logistic regression was performed to identify factors associated with serious AEs. RESULTS: Overall, 194,566 AEs (32.2% for nonopioids, 67.8% for opioids) were included in this analysis. The most common causative nonopioid and opioid analgesics was ketorolac (n = 10,789) and tramadol (n = 53,727), respectively. The most frequent AEs were skin and appendage disorders for nonopioids (31.8%) and gastrointestinal disorders (59.5%) for opioids. Serious AEs occurred in 6102 (9.7%) and 3326 (2.5%) cases of the nonopioid and opioid groups, respectively. The most common serious AEs were skin and appendage disorders (33.2%) for nonopioids and neurologic disorders (19.3%) for opioids. Serious AEs were significantly associated with male (odds ratio [OR] = 1.423), advanced age (OR = 1.570), certain causality (OR = 2.304), nonopioid analgesics (OR = 4.182), and polypharmacy (OR = 1.009; P <0.001 for all). CONCLUSIONS: In Korea, analgesic-induced AEs are prevalent with opioids more commonly implicated. Tramadol is the most common etiologic medication. Serious AEs are more frequently caused by nonopioids with skin and appendage disorders most common.

Quantification of metronidazole in human bile fluid and plasma by liquid chromatography-tandem mass spectrometry.

This study was conducted to develop a highly selective, sensitive, and validated method for quantifying metronidazole in human plasma and bile fluid. Metronidazole and metronidazole-d4 (internal standard) were extracted from 100 µL of plasma and bile fluid by liquid-liquid extraction. Liquid chromatography with a Hydrosphere C18 column (50 × 2.0 mm) was performed using 10 mM ammonium formate (pH 4.0) and acetonitrile (20:80, v/v) as the mobile phase. Triple quadrupole mass spectrometry was operated with an electrospray ionization interface in multiple reaction monitoring and positive ion modes. The calibration curves were linear for bile and plasma samples over the range of 50-20,000 ng/mL (r2 > 0.999). The intra- and inter-day coefficients of variation (CVs) for plasma ranged from 2.50% to 7.85% and 3.11% to 16.9%, respectively; for bile, the intra-and inter-run precision (CVs) ranged from 2.76% to 13.2% and 3.16% to 11.5%, respectively. The mean extraction recovery for metronidazole ranged from 76.5% to 82.1% in plasma and from 78.8% to 87.8% in bile, respectively. Our proposed analytical method was successfully applied to determine metronidazole concentrations in bile as well as in plasma at multiple time points in a patient with acute cholangitis.

Pharmacokinetics and bioequivalence of two fenofibrate choline formulations in healthy subjects under fed and fasted condition
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OBJECTIVE: The objective of this study was to evaluate the pharmacokinetics and bioequivalence of two formulations (the original capsule ("reference") and the new tablet ("test") formulations) of 135-mg choline fenofibrate under fed and fasted conditions. MATERIALS AND METHODS: This was an open-label, randomized, single-dose, crossover bioequivalence study in healthy Korean males. A total of 40 individuals were separately enrolled in the high-fat fed and the fasting study, respectively, and were randomized in a 1:1 ratio into two sequences. Serial blood samples were collected over 72 hours after drug administration. Plasma concentrations of fenofibric acid were determined by a validated LC-MS/MS method. Pharmacokinetic (PK) parameters were estimated using noncompartmental methods. RESULTS: Overall, 37 and 35 individuals completed the fed and the fasting study, respectively, as planned. The estimated Cmax, AUC0-∞, and AUC0-last were comparable between the test and the reference formulations in both fed and fasting studies (p > 0.05). The 90% confidence intervals for the geometric mean ratios of Cmax, AUC0-∞, and AUC0-last were 0.92 - 1.06, 0.95 - 1.01, and 0.95 - 1.01 in the fed study; and 0.94 - 1.12, 0.94 - 1.00, and 0.94 - 1.00 in the fasting study, respectively. For both formulations, tmax was significantly prolonged under fed condition compared to fasting condition (p < 0.0001); all other PK parameters were comparable between the fed and the fasting studies (p > 0.05). CONCLUSION: The reference and the test formulations of 135 mg choline fenofibrate show comparable pharmacokinetic profiles of fenofibric acid under both fed and fasted conditions and are considered bioequivalent.
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