Fuchs endothelial corneal dystrophy is one of the most common indications for corneal transplantation, and impaired anti-oxidative function is observed in corneal endothelial cells (CECs). Curcumin ...is well-known for its anti-oxidative property; but, no study has examined the effect of curcumin on anti-oxidative therapeutic roles in corneal endothelial disease. In our experiments, oxidative stress 0.25 mM tert-butyl hydroperoxide for 2 h was induced in immortalized human CECs pretreated with curcumin. Cell behavior and viability, reactive oxygen species production, and the protein expression of the kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2(Nrf2)/antioxidant response element (ARE) pathway were examined; the Keap1/Nrf2/ARE pathway is crucial anti-oxidative pathway of curcumin. The results showed that pretreatment with 12.5 μM curcumin significantly reduced the ROS production and improved the survival of CECs under oxidative stress. In addition, curcumin pretreatment significantly increased the expression of nuclear Nrf2, and the productions of superoxide dismutase 1 and heme oxygenase-1, which were the target anti-oxidative enzymes of the Keap1/Nrf2/ARE pathway. Our findings showed that curcumin enhanced the growth and differentiation of CECs under oxidative stress. The activation of Keap1/Nrf2/ARE pathway by curcumin was crucial for CECs to improve their anti-oxidative capacity.
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•Curcumin benefits the survival of corneal endothelial cells under oxidative stress.•The nuclear translocation of Nrf2 is observed with the addition of curcumin in corneal endothelial cells.•The modulation of Keap1/Nrf2/ARE pathway by curcumin plays an important role for anti-oxidative response.
Antibody‒drug conjugates (ADCs) are innovative biopharmaceutics consisting of a monoclonal antibody, linkers, and cytotoxic payloads. Monitoring circulating payload concentrations has the potential ...to identify ADC toxicity; however, accurate quantification faces challenges, including low plasma concentrations, severe matrix effects, and the absence of stable isotope-labeled internal standards (SIL-IS) for payloads and their derivatives. Previous studies used structural analogs as internal standards, but different retention times between structural analogs and target analytes may hinder effective matrix correction. Therefore, a more flexible approach is required for precise payload quantification.
We developed an LC‒MS/MS method incorporating a postcolumn-infused internal standard (PCI-IS) strategy for quantifying payloads and their derivatives of trastuzumab emtansine, trastuzumab deruxtecan, and sacituzumab govitecan, including DM1, MCC-DM1, DXd, SN-38, and SN-38G. Structural analogs (maytansine, Lys-MCC-DM1, and exatecan) were selected as PCI-IS candidates, and their accuracy performance was evaluated based on the percentage of samples within 80%–120% quantification accuracy. Compared to the approach without PCI-IS correction, exatecan enhanced the accuracy performance from 30-40%–100% for SN-38 and DXd, while maytansine and Lys-MCC-DM1 showed comparable accuracy for DM1 and MCC-DM1. This validated PCI-IS analytical method showed superior normalization of matrix effect in all analytes compared to the conventional internal standard approach. The clinical application of this approach showed pronounced differences in DXd and SN-38 concentrations before and after PCI-IS correction. Moreover, only DXd concentrations after PCI-IS correction were significantly higher in patients with thrombocytopenia (p = 0.037).
This approach effectively addressed the issue of unavailability of SIL-IS for novel ADC payloads and provided more accurate quantification, potentially yielding more robust statistical outcomes for understanding the exposure-toxicity relationship in ADCs. It is anticipated that this PCI-IS strategy may be extrapolated to quantify payloads and derivatives in diverse ADCs, thereby providing invaluable insights into drug toxicity and fortifying patient safety in ADC usage.
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•Developed LC‒MS/MS method with PCI-IS for accurate quantification of ADC payloads.•PCI-IS offers a flexible strategy to quantify payload under the lack of SIL-IS.•Better matrix effect normalization found with PCI-IS than conventional IS approach.•Exposure-toxicity relationship of DXd observed after applying PCI-IS approach.
The aim of this study was to explore the relationships between depressive symptoms and health-related quality of life (HRQOL) measurements for inpatients with major depressive disorder (MDD) before ...and after 6-week fluoxetine treatment, and to elucidate the factors related to the HRQOL changes.
A total of 131 inpatients with MDD were enrolled to receive 20 mg of fluoxetine for 6 weeks. Symptom severity and adverse events were assessed at weeks 0, 1, 2, 3, 4, and 6 using the 17-item Hamilton Depression Rating Scale (HAMD-17) and UKU Side Effect Rating Scale, respectively. HRQOL was measured using the Short Form 36 (SF-36), including 8 subscales, physical component summary (PCS) and mental component summary (MCS), at baseline and week 6. Spearman's coefficient, Cohen's d, and multiple linear regression model were used for statistical analysis.
One hundred and six patients completing all measures at weeks 0 and 6 entered the analysis. HAMD-17 negatively correlated with SF-36 at baseline and week 6. The HAMD-17 had a larger effect size than SF-36. MCS, rather than PCS, showed statistically significant improvement. After using multiple linear regression analysis, age at onset, HAMD-17 score change, and number of adverse events reported during the trial period were related to MCS change after adjusting for confounding variables.
Fluoxetine treatment was associated with an improvement in depressive symptomology and HRQOL. Depressive symptoms had a greater extent of change than HRQOL. Clinicians must consider the negative effects of adverse events caused by antidepressants on the improvement of HRQOL.
http://clinicaltrials.gov , NCT01075529 , retrospectively registered 24/2/2010.
Obesity, dyslipidemia, insulin resistance, oxidative stress, and inflammation are key clinical risk factors for the progression of non-alcoholic fatty liver disease (NAFLD). Currently, there is no ...comprehensive metabolic profile of a well-established animal model that effectively mimics the etiology and pathogenesis of NAFLD in humans. Here, we report the pathophysiological and metabolomic changes associated with NAFLD development in a C57BL/6J mouse model in which NAFLD was induced by feeding a high-fat diet (HFD) for 4, 8, 12, and 16 weeks. Serum metabolomic analysis was conducted using ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC–QTOF–MS) and gas chromatography–mass spectrometry (GC–MS) to establish a metabolomic profile. Analysis of the metabolomic profile in combination with principal component analysis revealed marked differences in metabolites between the control and HFD group depending upon NAFLD severity. A total of 30 potential biomarkers were strongly associated with the development of NAFLD. Among these, 11 metabolites were mainly related to carbohydrate metabolism, hepatic biotransformation, collagen synthesis, and gut microbial metabolism, which are characteristics of obesity, as well as significantly increased serum glucose, total cholesterol, and hepatic triglyceride levels during the onset of NAFLD (4 weeks). At 8 weeks, 5 additional metabolites that are chiefly involved in perturbation of lipid metabolism and insulin secretion were found to be associated with hyperinsulinemia, hyperlipidemia, and hepatic steatosis in the mid-term of NAFLD progression. At the end of 12 and 16 weeks, 14 additional metabolites were predominantly correlated to abnormal bile acid synthesis, oxidative stress, and inflammation, representing hepatic inflammatory infiltration during NAFLD development. These results provide potential biomarkers for early risk assessment of NAFLD and further insights into NAFLD development.
Neurofilaments (Nf) are major structural proteins that occur exclusively in neurons. In spinal cord injury (SCI), the severity of disease is quantified by clinical measures that have limited ...sensitivity and reliability, and no blood-based biomarker has been established to further stratify the degree of injury. We aimed to examine a serum-based NfL immunoassay as predictor of the clinical outcome in SCI.
Longitudinal measurement of serum NfL was performed in patients with central cord syndrome (CCS, n=4), motor-incomplete SCI (iSCI, n=10), motor-complete SCI (cSCI, n=13) and healthy controls (HC, n=67), and correlated with clinical severity, neurological outcome, and neuroprotective effect of the drug minocycline.
Baseline NfL levels were higher in iSCI (21 pg/mL) and cSCI (70 pg/mL) than in HC (5 pg/mL, p=0.006 and p<0.001) and CCS (6 pg/mL, p=0.025 and p=0.010). Levels increased over time (p<0.001) and remained higher in cSCI versus iSCI (p=0.011) and than in CCS (p<0.001). NfL levels correlated with American Spinal Injury Association (ASIA) motor score at baseline (r=-0.53, p=0.004) and after 24 h (r=-0.69, p<0.001) and 3-12-month motor outcome (baseline NfL: r=-0.43, p=0.026 and 24 h NfL: r=-0.72, p<0.001). Minocycline treatment showed decreased NfL levels in the subgroup of cSCI patients.
Serum NfL concentrations in SCI patients show a close correlation with acute severity and neurological outcome. Our data provide evidence that serum NfL is of prognostic value in SCI patients for the first time. Further, blood NfL levels may qualify as drug response markers in SCI.
ABSTRACT
Introduction: The negative results in trials of vitamin C in Charcot–Marie–Tooth disease (CMT) type 1A have highlighted the lack of sensitive outcome measures. Neurofilaments are abundant ...neuronal cytoskeletal proteins, and their concentration in blood is likely to reflect axonal breakdown. We therefore examined plasma neurofilament heavy‐chain (NfH) concentration as a potential biomarker in CMT. Methods: Blood samples were collected from healthy controls and patients with CMT over a 2‐year period. Disease severity was measured using the CMT Examination Score. An in‐house enzyme‐linked immunoabsorbent assay was used to measure plasma NfH levels. Results: There was no significant difference in plasma NfH concentrations between CMT patients and controls (P = 0.449). There was also no significant difference in plasma NfH levels in the CMT group over 1 year (mean difference = –0.02, SEM = 4.44, P = 0.98). Conclusions: Plasma NfH levels are not altered in patients with CMT and are not a suitable biomarker of disease activity. Muscle Nerve 53: 972–975, 2016
Exosomes secreted by adipose-derived stem cells (ADSC-exo) reportedly improve nerve regeneration after peripheral nerve injury. Herein, we investigated whether pretreatment of ADSCs with FK506, an ...immunosuppressive drug that enhances nerve regeneration, could secret exosomes (ADSC-F-exo) that further augment nerve regeneration. Designed exosomes were topically applied to injured nerve in a mouse model of sciatic nerve crush injury to assess the nerve regeneration efficacy. Outcomes were determined by histomorphometric analysis of semi-thin nerve sections stained with toluidine blue, mouse neurogenesis PCR array, and neurotrophin expression in distal nerve segments. Isobaric tags for relative and absolute quantitation (iTRAQ) were used to profile potential exosomal proteins facilitating nerve regeneration. We observed that locally applied ADSC-exo and ADSC-F-exo significantly enhanced nerve regeneration after nerve crush injury. Pretreatment of ADSCs with FK506 failed to produce exosomes possessing more potent molecules for enhanced nerve regeneration. Proteomic analysis revealed that of 192 exosomal proteins detected in both ADSC-exo and ADSC-F-exo, histone deacetylases (HDACs), amyloid-beta A4 protein (APP), and integrin beta-1 (ITGB1) might be involved in enhancing nerve regeneration.
Recently, there has been significant interest in the influences of the human gut microbiota on many diseases, such as cardiovascular disease (CVD) and metabolic disorders. Trimethylamine N-oxide ...(TMAO) is one of the most frequently discussed gut-derived metabolites. Dried blood spot (DBS) sampling has been regarded as an attractive alternative sampling strategy for clinical studies and offers many advantages. For DBS sample processing, whole-spot analysis could minimize hematocrit-related bias, but it requires blood volume calibration. This study developed a method combining matrix-induced ion suppression (MIIS) with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) to estimate blood volume and quantify TMAO and its precursors and derivatives, including choline, carnitine and acetylcarnitine, in DBSs. The MIIS method used an ion suppression indicator (ISI) to measure the extent of ion suppression caused by the blood matrix, which was related to the blood volume. The results showed that the volume estimation accuracy of the MIIS method was within 91.7–109.7%. The combined MIIS and LC-MS/MS method for quantifying TMAO, choline, carnitine and acetylcarnitine was validated in terms of linearity, precision and accuracy. The quantification accuracy was within 91.2–113.2% (with LLOQ <119%), and the imprecision was below 8.0% for all analytes. A stability study showed that the analytes in DBSs were stable at all evaluated temperatures for at least 30 days. The validated method was applied to quantify DBS samples (n = 56). Successful application of the new method demonstrated the potential of this method for real-world DBS samples and to facilitate our understanding of the gut microbiota in human health.
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•We developed a method to quantify TMAO and its precursors and derivatives in DBSs.•The MIIS method can be used to correct blood volume for polar metabolites in DBSs.•Validation results indicated that the quantitation was precise and accurate.•The developed method was applied to analyze 56 real paired plasma/DBS samples.
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