Pyrazinamide Safety, Efficacy, and Dosing for Treating Drug-Susceptible Pulmonary Tuberculosis: A Phase 3, Randomized, Controlled Clinical Trial.

RATIONALE: Optimizing pyrazinamide dosing is critical to improve treatment efficacy while minimizing toxicity during tuberculosis treatment. Study 31/ACTG A5349 represents the largest Phase 3 randomized controlled therapeutic trial to date for such investigation. OBJECTIVES: We sought to report pyrazinamide pharmacokinetic parameters, risk factors for lower pyrazinamide exposure, and relationships between pyrazinamide exposure with efficacy and safety outcomes. We aimed to determine pyrazinamide dosing strategies that optimize risks and benefits. METHODS: We analyzed pyrazinamide steady-state pharmacokinetic data using population nonlinear mixed-effects models. We evaluated the contribution of pyrazinamide exposure to long-term efficacy using parametric time-to-event models and safety outcomes using logistic regression. We evaluated optimal dosing with therapeutic windows targeting ≥95% durable cure and safety within the observed proportion of the primary safety outcome. MEASUREMENTS AND MAIN RESULTS: Among 2255 participants with 6978 plasma samples, pyrazinamide displayed 7-fold exposure variability (151-1053 mg·h/L). Body weight was not a clinically relevant predictor of drug clearance and thus did not justify the need for weight-banded dosing. Both clinical and safety outcomes were associated with pyrazinamide exposure, resulting in a therapeutic window of 231-355 mg·h/L for the control and 226-349 mg·h/L for the rifapentine-moxifloxacin regimen. Flat dosing of pyrazinamide at 1000 mg would have permitted an additional 13.1% (n=96) participants allocated to the control and 9.2% (n=70) to the rifapentine-moxifloxacin regimen dosed within the therapeutic window, compared to the current weight-banded dosing. CONCLUSIONS: Flat dosing of pyrazinamide at 1000 mg daily would be readily implementable and could optimize treatment outcomes in drug-susceptible tuberculosis. Clinical trial registration available at www. CLINICALTRIALS: gov, ID: NCT02410772.

Pharmacokinetic-Pharmacodynamic Evidence From a Phase 3 Trial to Support Flat-Dosing of Rifampicin for Tuberculosis.

BACKGROUND: The optimal dosing strategy for rifampicin in treating drug-susceptible tuberculosis (TB) is still highly debated. In the phase 3 clinical trial Study 31/ACTG 5349 (NCT02410772), all participants in the control regimen arm received 600 mg rifampicin daily as a flat dose. Here, we evaluated relationships between rifampicin exposure and efficacy and safety outcomes. METHODS: We analyzed rifampicin concentration time profiles using population nonlinear mixed-effects models. We compared simulated rifampicin exposure from flat- and weight-banded dosing. We evaluated the effect of rifampicin exposure on stable culture conversion at 6 months; TB-related unfavorable outcomes at 9, 12, and 18 months using Cox proportional hazard models; and all trial-defined safety outcomes using logistic regression. RESULTS: Our model-derived rifampicin exposure ranged from 4.57 mg · h/L to 140.0 mg · h/L with a median of 41.8 mg · h/L. Pharmacokinetic simulations demonstrated that flat-dosed rifampicin provided exposure coverage similar to the weight-banded dose. Exposure-efficacy analysis (n = 680) showed that participants with rifampicin exposure below the median experienced similar hazards of stable culture conversion and TB-related unfavorable outcomes compared with those with exposure above the median. Exposure-safety analysis (n = 722) showed that increased rifampicin exposure was not associated with increased grade 3 or higher adverse events or serious adverse events. CONCLUSIONS: Flat-dosing of rifampicin at 600 mg daily may be a reasonable alternative to the incumbent weight-banded dosing strategy for the standard-of-care 6-month regimen. Future research should assess the optimal dosing strategy for rifampicin, at doses higher than the current recommendation.

Mass Spectrometry-Compatible Elution Technique Enables an Improved Mucin-Selective Enrichment Strategy to Probe the Mucinome.

Mucin-domain glycoproteins are densely O-glycosylated and play critical roles in a host of healthy and disease-driven biological functions. Previously, we developed a mucin-selective enrichment strategy by employing a catalytically inactive mucinase (StcE) conjugated to a solid support. While this method was effective, it suffered from low throughput and high sample requirements. Further, the elution step required boiling in SDS, thus necessitating an in-gel digest with trypsin. Here, we introduce innovative elution conditions amenable to mucinase digestion and downstream analysis using mass spectrometry. This increased throughput and lowered sample input while maintaining mucin selectivity and enhancing the glycopeptide signal. We then benchmarked this technique against different O-glycan binding moieties for their ability to enrich mucins from various cell lines and human serum. Overall, the new method outperformed our previous procedure and all of the other enrichment techniques tested. This allowed for the effective isolation of more mucin-domain glycoproteins, resulting in a high number of O-glycopeptides, thus enhancing our ability to analyze the mucinome.

Analysis of Cell Glycogen with Quantitation and Determination of Branching Using Liquid Chromatography-Mass Spectrometry.

Glycogen is a highly branched biomacromolecule that functions as a glucose buffer. It is involved in multiple diseases such as glycogen storage disorders, diabetes, and even liver cancer, where the imbalance between biosynthetic and catabolic enzymes results in structural alterations and abnormal accumulation of glycogen that can be toxic to cells. Accurate and sensitive glycogen quantification and structural determination are prerequisites for understanding the phenotypes and biological functions of glycogen under these conditions. In this research, we furthered cell glycogen characterization by presenting a highly sensitive method to measure the glycogen content and degree of branching. The method employed a novel fructose density gradient as an alternative to the traditional sucrose gradient to fractionate glycogen from cell mixtures using ultracentrifugation. Fructose was used to avoid the large glucose background, allowing the method to be highly quantitative. The glycogen content was determined by quantifying 1-phenyl-3-methyl-5-pyrazolone (PMP)-derivatized glucose residues obtained from acid-hydrolyzed glycogen using ultra-high-performance liquid chromatography/triple quadrupole mass spectrometry (UHPLC/QqQ-MS). The degree of branching was determined through linkage analysis where the glycogen underwent permethylation, hydrolysis, PMP derivatization, and UHPLC/QqQ-MS analysis. The new approach was used to study the effect of insulin on the glycogen phenotypes of human hepatocellular carcinoma (Hep G2) cells. We observed that cells produced greater amounts of glycogen with less branching under increasing insulin levels before reaching the cell's insulin-resistant state, where the trend reversed and the cells produced less but higher-branched glycogen. The advantage of this method lies in its high sensitivity in characterizing both the glycogen level and the structure of biological samples.

Prothionamide Dose Optimization Using Population Pharmacokinetics for Multidrug-Resistant Tuberculosis Patients.

Prothionamide, a second-line drug for multidrug-resistant tuberculosis (MDR-TB), has been in use for a few decades. However, its pharmacokinetic (PK) profile remains unclear. This study aimed to develop a population PK model for prothionamide and then apply the model to determine the optimal dosing regimen for MDR-TB patients. Multiple plasma samples were collected from 27 MDR-TB patients who had been treated with prothionamide at 2 different study hospitals. Prothionamide was administered according to the weight-band dose regimen (500 mg/day for weight <50 kg and 750 mg/day for weight >50 kg) recommended by the World Health Organization. The population PK model was developed using nonlinear mixed-effects modeling. The probability of target attainment, based on systemic exposure and MIC, was used as a response target. Fixed-dose regimens (500 or 750 mg/day) were simulated to compare the efficacies of various dosing regimens. PK profiles adequately described the two-compartment model with first-order elimination and the transit absorption compartment model with allometric scaling on clearance. All dosing regimens had effectiveness >90% for MIC values <0.4 µg/mL in 1.0-log kill target. However, a fixed dose of 750 mg/day was the only regimen that achieved the target resistance suppression of ≥90% for MIC values of <0.2 µg/mL. In conclusion, fixed-dose prothionamide (750 mg/day), regardless of weight-band, was appropriate for adult MDR-TB patients with weights of 40 to 67 kg.

Analyzing polymeric matrix for fabrication of a biodegradable microneedle array to enhance transdermal delivery.

Traditional drug delivery systems, using invasive, transdermal, and oral routes, are limited by various factors, such as the digestive system environment, skin protection, and sensory nerve stimulation. To improve the drug delivery system, we fabricated a polysaccharide-based, dissolvable microneedle-based array, which combines the advantages of both invasive and transdermal delivery systems, and promises to be an innovative solution for minimally invasive drug delivery. In this study, we designed a reusable aluminum mold that greatly improved the efficiency and convenience of microneedle fabrication. Physical characterization of the polysaccharides, individual or mixed at different ratios, was performed to identify a suitable molecule to fabricate the dissolvable microneedle. We used a vacuum deposition-based micro-molding method at low temperature to fabricate the model. Using a series of checkpoints from material into product, a systematic feedback mechanism was built into the "all-in-one" fabrication step, which helped to improve production yields. The physical properties of the fabricated microneedle were assessed. The cytotoxicity analysis and animal testing of the microneedle demonstrated the safety and compatibility of the microneedle, and the successful penetration and effective release of a model protein.

Mega-Cecum: An Unrecognized Cause of Symptoms in Some Female Patients with Uro-Gynecological Symptoms and Severe Slow Transit Constipation.

BACKGROUND: A subset of female patients with severe constipation report overlapping uro-gynecological symptoms which have been attributed to visceral hypersensitivity. AIMS: To study colon morphology and motor function in female patients with medically refractory chronic constipation with or without uro-gynecological symptoms and to assess clinical outcomes following laparoscopic ileo-proctostomy. METHODS: Colon anatomy and cecal emptying time were assessed with plain films and fluoroscopy following a standardized test meal mixed with barium. Transit time was determined with radiopaque markers. IBS-QOL and urinary incontinence questionnaires were employed to assess post-colectomy clinical response. RESULTS: In 21 consecutive patients, mean colon transit time (h) was 211.1 ± 11.3, which was significantly greater than 58.9 ± 5.1 of 10 normal subjects (P < 0.001). Mega-cecum was found in 15 (Group 1) with mean cecal volume of 587 ± 27.9 cm3, significantly greater (P < 0.001) than 169.5 ± 10.4 cm3 of six without mega-cecum (Group 2). Mean cecal empting time (days) of barium-mixed feces in Group 1, 4.0 ± 0.6 was significantly greater than 1.33 ± 0.21 in Group 2 (P < 0.001). Eighteen patients (Groups 1 and 2) who had laparoscopic ileo-proctostomy experienced significantly improved quality of life (P < 0.001). In particular, Group 1 patients benefited significantly from improved uro-gynecological symptoms. CONCLUSIONS: Hitherto an unrecognized mega-cecum with markedly impaired emptying function was found in patients with severe slow transit constipation and uro-gynecological symptoms. Subtotal colectomy relieved constipation and improved significantly uro-gynecological symptoms, suggesting strongly that mega-cecum is causally related to these symptoms.

CDK2 phosphorylation regulates the protein stability of KLF10 by interfering with binding of the E3 ligase SIAH1.

Downregulation of multiple cell cycle-regulatory molecules is a dominant event in TGF-ß1-mediated growth inhibition of human carcinoma cells. It is known that KLF10 mimics the anti-proliferative and apoptotic effects that TGF-ß1 has on epithelial cell growth and the growth of various tumor cells; based on these findings it is considered as a tumor suppressor. KLF10 protein expression is tightly associated with cell cycle-dependent events. However, the regulatory mechanism and its biological meaning have not been identified. In this study, we have demonstrated that KLF10 is a substrate of CDK2/cyclin E and can be phosphorylated. We also have shown that KLF10 efficiently binds to CDK2, while binding much less to CDK4, and displaying no binding to Cdk6. Using mass spectrometry, site direct mutagenesis, in vitro kinase assays and depletion assays, we have established that CDK2 phosphorylates Ser206, which subsequently affects the steady state level of KLF10 in cells. Our studies have also proved that CDK2 up-regulates the protein level of KLF10 through reducing its association with SIAH1, a KLF10 E3-ubiqutin ligase involved in proteasomal degradation. Taken all together, these findings indicate that CDK2-dependent phosphorylation regulates KLF10 stability and that this affects the role of KLF10 in cell.

Sprouty4 mediates amphiregulin-induced down-regulation of E-cadherin and cell invasion in human ovarian cancer cells.

Sprouty (SPRY) proteins are well-characterized factors that inhibit receptor tyrosine kinase (RTK)-mediated activation of cellular signaling pathways. The down-regulation of SPRY4 expression has been reported in human ovarian cancer. However, the specific roles and mechanisms by which SPRY4 affects ovarian cancer progression are completely unknown. Amphiregulin (AREG) binds exclusively to the epidermal growth factor receptor (EGFR) and has been considered to be a dominant autocrine/paracrine EGFR ligand in ovarian cancer. In the present study, we first examined the effects of AREG on SPRY4 expression and the possible underlying molecular mechanisms involved in this process in two human ovarian cancer cell lines. Our results demonstrated that treatment with AREG up-regulated SPRY4 expression by activating the ERK1/2 signaling pathway. In addition, we showed that small interfering RNA (siRNA)-mediated knockdown of SPRY4 attenuated the AREG-induced down-regulation of E-cadherin by inhibiting the expression of SNAIL but not SLUG. In contrast, overexpression of SPRY4 enhanced AREG-induced down-regulation of E-cadherin by increasing the expression of SNAIL. Moreover, SPRY4 knockdown attenuated AREG-induced cell migration and invasion. Overexpression of SPRY4 enhanced AREG-induced cell invasion. This study reveals that SPRY4 is involved in EGFR-mediated human ovarian cancer progression.

In vitro anti-inflammatory properties of fermented pepino (Solanum muricatum) milk by γ-aminobutyric acid-producing Lactobacillus brevis and an in vivo animal model for evaluating its effects on hypertension.

BACKGROUND: The objectives of this study were to determine the in vitro anti-inflammatory and in vivo antihypertensive effects of fermented pepino (Solanum muricatum) milk by Lactobacillus brevis with the goal of developing functional healthy products. The inflammatory factors of fermented pepino milk with L. brevis were assessed in RAW 264.7 macrophages, including nitric oxide (NO) production. Inflammatory factor genes of cyclooxygenase (COX)-1 and -2, and tumor necrosis factor (TNF)-α were also assayed by a reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Results showed that fermented PE inhibited NO production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells with 150 mg mL(-1) fermented PE completely blocking LPS-induced NO production. The mRNA expressions of COX-1, COX-2, and iNOS were attenuated by treatment with higher concentrations of fermented PE (150 mg/ml). Cells treated with fermented pepino extract (PE) (100 ng mL(-1)) exhibited strikingly decreased LPS-induced expression of TNF-α mRNA. During the feeding trial, rats treated with 10% fermented pepino milk (100 µg 2.5 mL(-1)) and 100% fermented pepino milk (1000 µg 2.5 mL(-1)) exhibited significant decreases in the systolic blood pressure. CONCLUSION: Our results showed that fermented pepino milk has wide potential applications for development as a health food.

Destabilization of KLF10, a tumor suppressor, relies on thr93 phosphorylation and isomerase association.

KLF10 is now classified as a member of the Krüppel-like transcription factor family and acts as a tumor suppressor. Although KLF10 is originally named as TGF-ß-inducible early gene-1 and mimicking the anti-proliferative effect of TGF-ß in various carcinoma cells, the transcriptional upregulatory function of KLF10 has been described for a variety of cytokines and in many diseases. Through in vivo and in vitro phosphorylation assays, we identified that KLF10 is a phosphorylated protein in cells. Using yeast-two hybrid screening and site direct mutagenesis, we also identified PIN1 as a novel KLF10 associated protein. PIN1 is a peptidyl-prolyl isomerase enzyme belonging to the parvulin family, which specifically recognizes phosphorylated Ser/Thr-Pro containing substrates. Through protein-protein interaction assays, we showed that the Pro-directed Ser/Thr-Pro motif at Thr-93 in the KLF10 N-terminal region is essential for the interaction between KLF10 and PIN1. More importantly, PIN1 interacts with KLF10 in a phosphorylation-dependent manner and this interaction promotes KLF10 protein degradation in cells. Therefore, KLF10 shows shorter protein stability compared with mutant KLF10 that lacks PIN1 binding ability after cycloheximide treatments. The reversely correlated expression profile between KLF10 and PIN1 as observed in cell lines was also shown in clinic pancreatic cancer specimen. Using in vitro kinase assays and depletion assays, we were able to show that RAF-1 phosphorylates the Thr-93 of KLF10 and affects the KLF10 expression level in cells. Thus these findings as a whole indicate that RAF-1 phosphorylation and PIN1 isomerization together regulate KLF10 stability and further affect the role of KLF10 in tumor progression.

Dovitinib synergizes with oxaliplatin in suppressing cell proliferation and inducing apoptosis in colorectal cancer cells regardless of RAS-RAF mutation status.

BACKGROUND: Cancer is the result of a multistep process of genomic alterations, including mutations in key regulatory proteins that result in loss of balanced gene expression and subsequent malignant transformation. Throughout the various stages of colorectal carcinoma (CRC), complex genetic alterations occur, of which over-expression of growth factors, such as vascular endothelial growth factor, fibroblast growth factor and platelet-derive growth factor and their corresponding receptor tyrosine kinases, have been shown to correlate with invasiveness, tumor angiogenesis, metastasis, recurrence, and poor prognosis of colorectal cancer. To evaluate the therapeutic effect, we combined Dovitinib, an orally bioavailable, potent inhibitor of class III-V receptor tyrosine kinases with chemotherapeutic drug, oxaliplatin in preclinical models of colon cancer. METHODS: Human colon cancer cells with different RAS-RAF mutation status (HCT-116, HT-29, SW-480, CaCO2 and LS174T) were treated with a combination of Dovitinib and Oxaliplatin at low dosage followed by assays to investigate the effect of the combination on cell proliferation, cell migration, cell apoptosis and signaling pathways involved in molecular mechanism of drug(s). The antitumor effects of either of the drugs were compared to the combination using human colon carcinoma cell line HT-29 xenograft model. Treated vs untreated tumor sections were also compared for proliferation and angiogenesis markers by immunohistochemistry. RESULTS: The combination of dovitinib and oxaliplatin showed higher in vitro cytotoxicity in colon cell lines irrespective of their RAS-RAF status as compared to either of the drugs alone. Simultaneous inhibition of MAP kinase and AKT pathways and induction of apoptosis via activation of caspases 9/caspases 3 contributed to the synergistic effect of this combination therapy. In the xenograft model, the combination showed a significantly higher antitumor activity. Immunohistochemistry of post treatment tumors showed a significant decrease in proliferation and angiogenesis as compared to either of the treatments alone. CONCLUSIONS: This study demonstrates the synergistic antitumor activity of combination of dovitinib and oxaliplatin against colon cancer with different RAS-RAF status. The combination also showed its antitumor efficacy in a multidrug resistant phenotype xenograft model. This provides a basis for further investigation for its potential in clinical setting for colorectal cancer.

Chilaiditi's Syndrome Mimicking Crohn's.

Chilaiditi's sign (colonic interposition) is a rare anomaly due to an abnormally located portion of the colon that is interposed in between the liver and the diaphragm. This rare anomaly is often incidentally seen on chest or abdominal radiographs. Chilaiditi's radiographic sign is usually asymptomatic, whereas the medical condition accompanied by clinical symptoms is termed Chilaiditi's syndrome. Possible causes of the syndrome include a long and mobile colon, scarring of the liver (cirrhosis), ascites, long-standing lung disease, as well as laxity of the falciform ligament. The most common clinical signs of Chilaiditi's syndrome include gastrointestinal symptoms; however, clinical presentation can vary. This report describes a case of a 21-year-old male patient who presented with a longstanding history of left upper quadrant epigastric abdominal pain with diarrhea (six to eight loose watery stools). The patient was diagnosed with Crohn's colitis and had tried a myriad of medical therapies with no adequate response. He chose to seek a second opinion and was subsequently discovered to have Chilaiditi's syndrome via computed tomography (CT) and confirmed by barium enema. The patient then elected to undergo a right laparoscopic colectomy to resolve the symptoms. By postoperative day five, all symptoms had resolved including abdominal pain and diarrhea. Therefore, it is important to consider Chilaiditi's syndrome as a differential diagnosis in persons presenting with left upper quadrant pain and symptoms of Crohn's colitis, especially those treated with adequate medical therapy without alleviation of symptoms.

Computational Discovery of Intermolecular Singlet Fission Materials Using Many-Body Perturbation Theory.

Intermolecular singlet fission (SF) is the conversion of a photogenerated singlet exciton into two triplet excitons residing on different molecules. SF has the potential to enhance the conversion efficiency of solar cells by harvesting two charge carriers from one high-energy photon, whose surplus energy would otherwise be lost to heat. The development of commercial SF-augmented modules is hindered by the limited selection of molecular crystals that exhibit intermolecular SF in the solid state. Computational exploration may accelerate the discovery of new SF materials. The GW approximation and Bethe-Salpeter equation (GW+BSE) within the framework of many-body perturbation theory is the current state-of-the-art method for calculating the excited-state properties of molecular crystals with periodic boundary conditions. In this Review, we discuss the usage of GW+BSE to assess candidate SF materials as well as its combination with low-cost physical or machine learned models in materials discovery workflows. We demonstrate three successful strategies for the discovery of new SF materials: (i) functionalization of known materials to tune their properties, (ii) finding potential polymorphs with improved crystal packing, and (iii) exploring new classes of materials. In addition, three new candidate SF materials are proposed here, which have not been published previously.

Xylosyltransferase Bump-and-hole Engineering to Chemically Manipulate Proteoglycans in Mammalian Cells.

Mammalian cells orchestrate signalling through interaction events on their surfaces. Proteoglycans are an intricate part of these interactions, carrying large glycosaminoglycan polysaccharides that recruit signalling molecules. Despite their importance in development, cancer and neurobiology, a relatively small number of proteoglycans have been identified. In addition to the complexity of glycan extension, biosynthetic redundancy in the first protein glycosylation step by two xylosyltransferase isoenzymes XT1 and XT2 complicates annotation of proteoglycans. Here, we develop a chemical genetic strategy that manipulates the glycan attachment site of cellular proteoglycans. By employing a tactic termed bump- and-hole engineering, we engineer the two isoenzymes XT1 and XT2 to specifically transfer a chemically modified xylose analogue to target proteins. The chemical modification contains a bioorthogonal tag, allowing the ability to visualise and profile target proteins modified by both transferases in mammalian cells. The versatility of our approach allows pinpointing glycosylation sites by tandem mass spectrometry, and exploiting the chemical handle to manufacture proteoglycans with defined GAG chains for cellular applications. Engineered XT enzymes permit a view into proteoglycan biology that is orthogonal to conventional techniques in biochemistry.

Krüppel-like factor 10 expression as a prognostic indicator for pancreatic adenocarcinoma.

Deregulation of transforming growth factor (TGF)-ß function is a common feature of pancreatic cancer, rendering these cancers unresponsive to TGF-ß-stimulated growth inhibition. Recent findings have supported a primary role for Krüppel-like factor 10 (KLF10) as an important transcription factor involved in mediating TGF-ß1 signaling. The aim of this study was to evaluate the correlation between KLF10 expression and the clinical and pathologic features of pancreatic cancer. Tissue specimens from patients with pancreatic adenocarcinoma were retrospectively collected for immunohistochemical analysis. To demonstrate that Klf10 expression was primarily regulated by methylation status, the Klf10 promoter was examined by methylation-specific PCR using a pancreatic cancer cell line (Panc-1). DNA methyltransferase (DNMT) inhibitor and small-interfering RNA depletion of DNMT genes were used to reverse KLF10 expression in the Panc-1 cells. In parallel, DNMT1 expression was evaluated in the pancreatic cancer tissue specimens. In 95 pancreatic cancer tissue specimens, KLF10 expression was inversely correlated with pancreatic cancer stage (P = 0.01). Multivariable analysis revealed that, in addition to the presence of distant metastasis at diagnosis (P = 0.001 and 0.001, respectively), KLF10 was another independent prognostic factor related to progression-free and overall survival (P = 0.018 and 0.037, respectively). The loss of KLF10 expression in advanced pancreatic cancer is correlated with altered methylation status, which seems to be regulated by DNMT1. Our results suggest that KLF10 is a potential clinical predictor for progression of pancreatic cancer.

Translation directionality and the Inhibitory Control Model: a machine learning approach to an eye-tracking study.

Introduction: Based on such physiological data as pupillometry collected in an eye-tracking experiment, the study has further confirmed the effect of directionality on cognitive loads during L1 and L2 textual translations by novice translators, a phenomenon called "translation asymmetry" suggested by the Inhibitory Control Model, while revealing that machine learning-based approaches can be usefully applied to the field of Cognitive Translation and Interpreting Studies. Methods: Directionality was the only factor guiding the eye-tracking experiment where 14 novice translators with the language combination of Chinese and English were recruited to conduct L1 and L2 translations while their pupillometry were recorded. They also filled out a Language and Translation Questionnaire with which categorical data on their demographics were obtained. Results: A nonparametric related-samples Wilcoxon signed rank test on pupillometry verified the effect of directionality, suggested by the model, during bilateral translations, verifying "translation asymmetry" at a textual level. Further, using the pupillometric data, together with the categorical information, the XGBoost machine-learning algorithm yielded a model that could reliably and effectively predict translation directions. Conclusion: The study has shown that translation asymmetry suggested by the model was valid at a textual level, and that machine learning-based approaches can be gainfully applied to Cognitive Translation and Interpreting Studies.

Optimized mucin-selective enrichment strategy to probe the mucinome.

Mucin-domain glycoproteins are densely O-glycosylated and play critical roles in a host of healthy and disease-driven biological functions. Previously, we developed a mucin-selective enrichment strategy by employing a catalytically inactive mucinase (StcE) conjugated to solid support. While this method was effective, it suffered from low throughput and high sample requirements. Further, the elution step required boiling in SDS, thus necessitating an in-gel digest with trypsin. Here, we optimized our previous enrichment method to include elution conditions amenable to mucinase digestion and downstream analysis with mass spectrometry. This increased throughput and lowered sample input while maintaining mucin selectivity and enhancing glycopeptide signal. We then benchmarked this technique against different O-glycan binding moieties for their ability to enrich mucins from various cell lines and human serum. Overall, the new method outperformed our previous procedure and all other enrichment techniques tested. This allowed for effective isolation of more mucin-domain glycoproteins, resulting in a high number of O-glycopeptides, thus enhancing our ability to analyze the mucinome.

Model-Based Efficacy and Toxicity Comparisons of Moxifloxacin for Multidrug-Resistant Tuberculosis.

BACKGROUND: Moxifloxacin (MOX) is used as a first-choice drug to treat multidrug-resistant tuberculosis (MDR-TB); however, evidence-based dosing optimization should be strengthened by integrative analysis. The primary goal of this study was to evaluate MOX efficacy and toxicity using integrative model-based approaches in MDR-TB patients. METHODS: In total, 113 MDR-TB patients from 5 different clinical trials were analyzed for the development of a population pharmacokinetics (PK) model. A final population PK model was merged with a previously developed lung-lesion distribution and QT prolongation model. Monte Carlo simulation was used to calculate the probability target attainment value based on concentration. An area under the concentration-time curve (AUC)-based target was identified as the minimum inhibitory concentration (MIC) of MOX isolated from MDR-TB patients. RESULTS: The presence of human immunodeficiency virus (HIV) increased clearance by 32.7% and decreased the AUC by 27.4%, compared with HIV-negative MDR-TB patients. A daily dose of 800 mg or a 400-mg, twice-daily dose of MOX is expected to be effective in MDR-TB patients with an MIC of ≤0.25 µg/mL, regardless of PK differences resulting from the presence of HIV. The effect of MOX in HIV-positive MDR-TB patients tended to be decreased dramatically from 0.5 µg/mL, in contrast to the findings in HIV-negative patients. A regimen of twice-daily doses of 400 mg should be considered safer than an 800-mg once-daily dosing regimen, because of the narrow fluctuation of concentrations. CONCLUSIONS: Our results suggest that a 400-mg, twice-daily dose of MOX is an optimal dosing regimen for MDR-TB patients because it provides superior efficacy and safety.

Restoring Prohealing/Remodeling-Associated M2a/c Macrophages Using ON101 Accelerates Diabetic Wound Healing.

Diabetic wounds exhibit chronic inflammation and delayed tissue proliferation or remodeling, mainly owing to prolonged proinflammatory (M1) macrophage activity and defects in transition to prohealing/proremodeling (M2a/M2c; CD206+ and/or CD163+) macrophages. We found that topical treatment with ON101, a plant-based potential therapeutic for diabetic foot ulcers, increased M2c-like (CD163+ and CD206+) cells and suppressed M1-like cells, altering the inflammatory gene profile in a diabetic mouse model compared with that in the controls. An in vitro macrophage-polarizing model revealed that ON101 directly suppressed CD80+ and CD86+ M1-macrophage polarization and M1-associated proinflammatory cytokines at both protein and transcriptional levels. Notably, conditioned medium collected from ON101-treated M1 macrophages reversed the M1-conditioned medium‒mediated suppression of CD206+ macrophages. Furthermore, conditioned medium from ON101-treated adipocyte progenitor cells significantly promoted CD206+ and CD163+ macrophages but strongly inhibited M1-like cells. ON101 treatment also stimulated the expression of GCSF and CXCL3 genes in human adipocyte progenitor cells. Interestingly, treatment with recombinant GCSF protein enhanced both CD206+ and CD163+ M2 markers, whereas CXCL3 treatment only stimulated CD163+ M2 macrophages. Depletion of cutaneous M2 macrophages inhibited ON101-induced diabetic wound healing. Thus, ON101 directly suppressed M1 macrophages and facilitated the GCSF- and CXCL3-mediated transition from M1 to M2 macrophages, lowering inflammation and leading to faster diabetic wound healing.