Inhibition of lysyl oxidase by pharmacological intervention and genetic manipulation alleviates epilepsy-associated cognitive disorder.

Epilepsy-associated cognitive disorder (ECD), a prevalent comorbidity in epilepsy patients, has so far uncharacterized etiological origins. Our prior work revealed that lysyl oxidase (Lox) acted as a novel contributor of ferroptosis, a recently discovered cell death mode in the regulation of brain function. However, the role of Lox-mediated ferroptosis in ECD remains unknown. ECD mouse model was established 2 months later following a single injection of kainic acid (KA) for. After chronic treatment with KA, mice were treated with different doses (30 mg/kg, 100 mg/kg and 300 mg/kg) of Lox inhibitor BAPN. Additionally, hippocampal-specific Lox knockout mice was also constructed and employed to validate the role of Lox in ECD. Cognitive functions were assessed using novel object recognition test (NOR) and Morris water maze test (MWM). Protein expression of phosphorylated cAMP-response element binding (CREB), a well-known molecular marker for evaluation of cognitive performance, was also detected by Western blot. The protein distribution of Lox was analyzed by immunofluorescence. In KA-induced ECD mouse model, ferroptosis process was activated according to upregulation of 4-HNE protein and a previously discovered ferroptosis in our group, namely, Lox was remarkably increased. Pharmacological inhibition of Lox by BAPN at the dose of 100 mg/kg significantly increased the discrimination index following NOR test and decreased escape latency as well as augmented passing times within 60 s following MWM test in ECD mouse model. Additionally, deficiency of Lox in hippocampus also led to pronounced improvement of deficits in ECD model. These findings indicate that the ferroptosis regulatory factor, Lox, is activated in ECD. Ablation of Lox by either pharmacological intervention or genetic manipulation ameliorates the impairment in ECD mouse model, which suggest that Lox serves as a promising therapeutic target for treating ECD in clinic.

Hydrogen attenuates postoperative pain through Trx1/ASK1/MMP9 signaling pathway.

BACKGROUND: Postoperative pain is a serious clinical problem with a poorly understood mechanism, and lacks effective treatment. Hydrogen (H2) can reduce neuroinflammation; therefore, we hypothesize that H2 may alleviate postoperative pain, and aimed to investigate the underlying mechanism. METHODS: Mice were used to establish a postoperative pain model using plantar incision surgery. Mechanical allodynia was measured using the von Frey test. Cell signaling was assayed using gelatin zymography, western blotting, immunohistochemistry, and immunofluorescence staining. Animals or BV-2 cells were received with/without ASK1 and Trx1 inhibitors to investigate the effects of H2 on microglia. RESULTS: Plantar incision surgery increased MMP-9 activity and ASK1 phosphorylation in the spinal cord of mice. MMP-9 knockout and the ASK1 inhibitor, NQDI-1, attenuated postoperative pain. H2 increased the expression of Trx1 in the spinal cord and in BV-2 cells. H2 treatment mimicked NQDI1 in decreasing the phosphorylation of ASK1, p38 and JNK. It also reduced MMP-9 activity, downregulated pro-IL-1ß maturation and IBA-1 expression in the spinal cord of mice, and ameliorated postoperative pain. The protective effects of H2 were abolished by the Trx1 inhibitor, PX12. In vitro, in BV-2 cells, H2 also mimicked NQDI1 in inhibiting the phosphorylation of ASK1, p38, and JNK, and also reduced MMP-9 activity and decreased IBA-1 expression induced by LPS. The Trx1 inhibitor, PX12, abolished the protective effects of H2 in BV-2 cells. CONCLUSIONS: For the first time, the results of our study confirm that H2 can be used as a therapeutic agent to alleviate postoperative pain through the Trx1/ASK1/MMP9 signaling pathway. MMP-9 and ASK1 may be the target molecules for relieving postoperative pain.

Corrigendum: The m6A-methylated mRNA pattern and the activation of the Wnt signaling pathway under the hyper-m6A-modifying condition in the keloid.

[This corrects the article DOI: 10.3389/fcell.2022.947337.].

The m6A-methylated mRNA pattern and the activation of the Wnt signaling pathway under the hyper-m6A-modifying condition in the keloid.

Purpose: The present study was carried out to investigate the global m6A-modified RNA pattern and possible mechanisms underlying the pathogenesis of keloid. Method: In total, 14 normal skin and 14 keloid tissue samples were first collected on clinics. Then, three samples from each group were randomly selected to be verified with the Western blotting to determine the level of methyltransferase and demethylase. The total RNA of all samples in each group was isolated and subjected to the analysis of MeRIP sequencing and RNA sequencing. Using software of MeTDiff and htseq-count, the m6A peaks and differentially expressed genes (DEGs) were determined within the fold change >2 and p-value < 0.05. The top 10 pathways of m6A-modified genes in each group and the differentially expressed genes were enriched by the Kyoto Encyclopedia of Genes and Genomes signaling pathways. Finally, the closely associated pathway was determined using the Western blotting and immunofluorescence staining. Results: There was a higher protein level of WTAP and Mettl3 in the keloid than in the normal tissue. In the keloid samples, 21,020 unique m6A peaks with 6,573 unique m6A-associated genetic transcripts appeared. In the normal tissue, 4,028 unique m6A peaks with 779 m6A-associated modified genes appeared. In the RNA sequencing, there were 847 genes significantly changed between these groups, transcriptionally. The genes with m6A-methylated modification and the upregulated differentially expressed genes between two tissues were both mainly related to the Wnt signaling pathway. Moreover, the hyper-m6A-modified Wnt/ß-catenin pathway in keloid was verified with Western blotting. From the immunofluorescence staining results, we found that the accumulated fibroblasts were under a hyper-m6A condition in the keloid, and the Wnt/ß-Catenin signaling pathway was mainly activated in the fibroblasts. Conclusion: The fibroblasts in the keloid were under a cellular hyper-m6A-methylated condition, and the hyper-m6A-modified highly expressed Wnt/ß-catenin pathway in the dermal fibroblasts might promote the pathogenesis of keloid.

Establishment and Application of an Index System for the Risk of Drug Shortages in China: Based on Delphi Method and Analytic Hierarchy Process.

BACKGROUND: At present, the avoidance of drug shortages mainly relies on expert experience. This study aimed to establish an evaluation index system for the risk of drug shortages in medical institutions in China and to apply the system to guide the graded management of drugs in short supply. METHODS: A two-round Delphi process was conducted to determine the indicators in the index system. The weight value of each indicator was calculated using analytic hierarchy process (AHP) methods. The data of drugs in short supply from January 1 to December 31, 2020 in Hunan province were collected and evaluated using this index system. The evaluation scores, which ranged from 0 to 100, were calculated. RESULTS: A three-level index system with four first-level indicators, 11 second-level indicators, and 36 third-level indicators was constructed by the two rounds of the Delphi process. The expert authority coefficient (Cr) of the first and second rounds of consultation were 0.88 and 0.90, respectively. The Kendall's coefficients of concordance (Kendall's W) for the two rounds of consultation were 0.44 and 0.50, respectively (P<.05). For the first-level indicators 'supply stability,' 'causes of shortage,' 'medicine availability in medical institution' and 'pharmaceutical properties,' the weight values were 0.3253, 0.2489, 0.2398, and 0.1860, respectively. Based on the risk evaluation score, drugs (dosage strength) at high risk of shortage included sodium thiosulfate (0.64 g), posterior pituitary lobe hormones (1 mL:6 IU), protamine sulfate (5 mL:50 mg), thrombin (500 U), urokinase (10 WU), and rotundine sulfate (2 mL:60 mg). CONCLUSION: An indexed system for the risk assessment of drug shortages in China was established to guide the graded response to drug shortages in medical institutions and the implementation of differential management strategies to address these shortages.

Distinct skin morphological and transcriptomic profiles between wild and albino Oscar Astronotus ocellatus.

Oscar Astronotus ocellatus is an important ornamental fish, including albino and wild varieties. Albino individuals attract aquarium hobbyists due to their unique body color, but studies on the species' albinism mechanism are currently scarce. Here, we investigated the morphological and transcriptomic profiles of the skin of albino and wild Oscar. The results showed that the albino type had fewer oval-shaped melanophores and immature melanosomes but that the wild type contained more stellate-shaped melanophores and mature melanosomes. Albino Oscar had a degenerative pigment layer without obvious melanin deposition and content, while the wild type contained more concentrated melanin within the pigment layer. A total of 272,392 unigenes were detected, 109 of which were identified as differentially expressed genes (DEGs) between albino and wild Oscar. Pathways of DEGs, including those involved in complement and coagulation cascades, novobiocin biosynthesis, Th1 and Th2 cell differentiation, and tropane, piperidine and pyridine alkaloid biosynthesis, were significantly enriched. DEGs, including upregulated Sfrp5 and Tat, and downregulated Wnt-10a, Ppp3c, Notch1 and Trim27 involved in the Wnt signaling pathway, Notch signaling pathway, tyrosine metabolism, MAPK signaling pathway and melanogenesis, might be associated with the albinism of Oscar. This study characterized the difference in melanophore morphology between wild and albino Oscar and identified some albinism-related candidate genes and signaling pathways, helping to understand the genetic mechanism of fish albinism.

Lidocaine Alleviates Sepsis-Induced Acute Lung Injury in Mice by Suppressing Tissue Factor and Matrix Metalloproteinase-2/9.

Acute lung injury (ALI) is one of the fatal symptoms of sepsis. However, there were no effective clinical treatments. TF accumulation-induced fibrin deposit formations and coagulation abnormalities in pulmonary vessels contribute to the lethality of ALI. Suppressor of cytokine signaling 3 (SOCS3) acts as an endogenous negative regulator of the TLR4/TF pathway. We hypothesized that inducing SOCS3 expression using lidocaine to suppress the TLR4/TF pathway may alleviate ALI. Hematoxylin and eosin (H&E), B-mode ultrasound, and flow cytometry were used to measure the pathological damage of mice. Gelatin zymography was used to measure matrix metalloproteinase-2/9 (MMP-2/9) activities. Western blot was used to assay the expression of protein levels. Here, we show that lidocaine could increase the survival rate of ALI mice and ameliorate the lung injury of ALI mice including reducing the edema, neutrophil infiltration, and pulmonary thrombosis formation and increasing blood flow velocity. Moreover, in vitro and in vivo, lidocaine could increase the expression of p-AMPK and SOCS3 and subsequently decrease the expression of p-ASK1, p-p38, TF, and the activity of MMP-2/9. Taken together, our study demonstrated that lidocaine could inhibit the TLR4/ASK1/TF pathway to alleviate ALI via activating AMPK-SOCS3 axis.

Evaluation of Factors Impacting the Efficacy of Single or Combination Therapies of Valproic Acid, Carbamazepine, and Oxcarbazepine: A Longitudinal Observation Study.

Objective: This study aimed to determine the efficacy and clinical factors related to the pharmacodynamics of single or combination therapies of valproic acid (VPA), carbamazepine (CBZ), and oxcarbazepine (OXC), three commonly used anti-epileptic drugs (AEDs) in China. Methods: The study evaluated the records of 2027 outpatients in a Changsha hospital, located in China, from December 23, 2015 to October 28, 2019. The baseline seizure frequency was assessed during the first visit. AED efficacy was determined based on the reduction in seizures from baseline at the subsequent visits. Multivariable ordinal regression analysis was used to determine the association between the clinical factors (demographic characteristics, clinical features, and medication situation) and AED efficacy. For validation, the clinical efficacies of AEDs were compared as both single agents and in combinations. Differences in adverse effect (AEs) categories were analyzed by Chi-square between AED groups. Results: Records of patients receiving VPA, CBZ, and OXC were evaluated. Serum concentrations of VPA and CBZ is significantly correlated with efficacy (OR 1.030 [1.024-1.037], p < 0 0.0001; OR 1.250 [1.146-1.63], p < 0.0001, respectively) and OXC efficacy correlated to the serum concentration of the metabolite 10,11-dihydro-10-hydroxy-carbazepine (monohydroxy derivative, MHD) serum concentrations (OR 1.060 [1.031-1.089], p < 0.0001). Significant differences existed between females and males in VPA efficacy (OR 1.318 [1.033-1.682], p = 0.027). After validation, VPA, in combination with OXC (OR 1.93 [1.38-2.70], p<0.001), or with VGB (Vigabatrin) (OR 2.36 [1.38-2.70], p = 0.002), showed significantly better efficacy than as a single agent. OXC efficacy was also affected by the duration of epilepsy (OR 0.965 [0.946-0.984], p < 0.001). Additionally, the efficacies of OXC and VPA were also affected by the seizure type. Seizure reduction improved significantly with an increasing number of pharmacists' educations in the first three visits period. There were no differences in AEs incidence among these 3 AEDs except for Psychiatric (0.02) and nervous system disorders (0.0001). Conclusion: Serum concentrations of VPA and CBZ may positively affect their efficacies, while OXC efficacies are correlated to MHD serum concentrations. The efficacy of VPA was higher in females compared to males. VPA-OXC and VPA-VGB combinations had higher efficacies compared to monotherapy. Besides, OXC efficacy is probably reducing by the duration of epilepsy. Additionally, VPA efficacy for focal or generalized seizures is superior to mixed-type seizures. OXC was more effective for focal seizures compared to mixed-type ones. Education provided by pharmacists improved the seizures to some extent, and there were no significant differences between most categories of adverse effects for the investigated AEDs.

Effect of rifampicin on anticoagulation of warfarin: A case report.

BACKGROUND: The drug interaction between warfarin and rifampicin is widely known, but there are still some difficulties in managing the combination of the two drugs. CASE SUMMARY: A patient with brucellosis received strict monitoring from a Chinese pharmacist team during combination of warfarin and rifampicin. The dose of warfarin was increased to 350% in 3 mo before reaching the lower international normalized ratio treatment window. No obvious adverse reaction occurred during the drug-adjustment period. This is the first case report of long-term combined use of rifampicin and warfarin in patients with brucellosis and valve replacement in China based on the Chinese lower warfarin dose and international normalized ratio range. CONCLUSION: Anticoagulation for valve replacement in Chinese patients differs from that in other races. Establishment of a pharmacist clinic provides vital assistance in warfarin dose adjustment.

Population pharmacokinetics of unbound valproic acid in pediatric epilepsy patients in China: a protein binding model.

PURPOSE: The purpose of this study was to establish a protein binding model of unbound valproic acid (VPA) based on Chinese pediatric patients with epilepsy and provide a reference for clinical medication. METHODS: A total of 313 patients were included and both their total and unbound VPA concentrations (375 pairs of concentrations) were measured. NONMEM software was used for population pharmacokinetic modeling. The stepwise method was used to screen the potential covariates. Goodness-of-fit plot, bootstrap, and visual predictive check were used for model evaluation. In addition, dose recommendations for typical patients aged 0 to 16 years were proposed by Monte Carlo simulations. RESULTS: A one-compartment model of first-order absorption and first-order elimination was used to describe the pharmacokinetic characteristics of unbound VPA, and the linear non-saturable binding equation was introduced to describe the protein binding. Body weight, age-based maturation, and co-medicated with lamotrigine could affect the CL/F of unbound and bound VPA. Model evaluation showed satisfactory robustness of the final model. The dosing regimens for children aged 0 to 16 years were proposed based on the final established model. CONCLUSION: We developed a population pharmacokinetic model of unbound and bound VPA that took account of protein binding. The VPA dosing regimen in pediatric patients with epilepsy needs to be optimized by the body weight, age, and co-medications.

Determination of unbound valproic acid in plasma using centrifugal ultrafiltration and gas chromatography:Application in TDM.

AIM: In order to monitor the free concentration of VPA in plasma, a simple and rapid method needs to be developed. METHODS: The free fraction of VPA in plasma was obtained by centrifugal ultrafiltration (CF-UF) devices. Cyclohexanecarboxylic acid was used as internal standard. Valproate in plasma was converted to VPA by sulphuric acid acidification, and dichloromethane was used as solvent for extraction. Nitrogen was the carrier gas, the samples were separated by capillary column, and the flame ionization detector was used to detect VPA fragment ions for quantitative analysis. RESULTS: The assay had good specificity and stability. The linear range of the assay was 0.56-28.11 mg/L. The intra-day and inter-day precision (RSDs) of the assay were all within 15%, and the accuracy (RE) was 2.58%. The recoveries of VPA with three different concentrations were 102.03 ±â€¯1.05, 101.45 ±â€¯2.08 and 102.58 ±â€¯3.38. The results of therapeutic drug monitoring (TDM) in pediatric inpatient group and outpatient group showed significant differences between the two groups (P < 0.001). CONCLUSION: This assay has low cost and good analytical performance, so it can be developed into a routine TDM method of unbound VPA. We recommend the monitoring of unbound VPA concentration in pediatric inpatients during clinical use of VPA.

Pharmacist impact on adherence of valproic acid therapy in pediatric patients with epilepsy using active education techniques.

There is limited information on the impact of active education by a pharmacist in the population of pediatric patients with epilepsy (PWE) in China. The objective of this study was to assess the effect of education by pharmacists on medication adherence and percentage of valproic acid (VPA) samples reaching therapeutic reference range in these patients. This study was conducted at two teaching hospitals in Changsha, China. Patients were retrospectively identified from January 2016 to December 2017. Active education by a pharmacist in both oral and written formats was provided at the intervention hospital whereas standard passive pharmacist service (dispensing and answering questions) was provided at the control hospital. Medication adherence was assessed by the simplified medication adherence questionnaire (SMAQ), and serum concentrations of VPA were collected. The correlation between pharmacist education and medication adherence and percentage of VPA samples reaching therapeutic reference range were analyzed. A total of 2165 patients and 4343 serum VPA concentrations were included in the analysis. For the first therapeutic drug monitoring (TDM) measurement, there was no statistical difference between the two hospitals: 41.3% of VPA samples reached therapeutic range at the intervention hospital compared with 45.4% at the control hospital (χ2 = 3.686, P > 0.05). After pharmacist intervention at the intervention hospital, however, there were significant differences in the percentage of therapeutic VPA samples reaching therapeutic range between the first and the second, third, fourth, and fifth TDM measurements (χ2 = 9.756, P < 0.01; χ2 = 22.840, P < 0.01; χ2 = 15.816, P < 0.01; χ2 = 27.613, P < 0.01). Based on the SMAQ adherence assessment, adherence increased from a minimum of 56.0% to a maximum of 73.9% with stabilization during the last six months of follow-up at the intervention hospital. Both the medication adherence rate and the percentage of VPA samples reaching therapeutic range increased as the result of active education by a pharmacist, suggesting that continuous pharmacist intervention had a positive impact in outpatient pediatric PWE.

Progress in Research and Application of HIV-1 TAT-Derived Cell-Penetrating Peptide.

Human immunodeficiency virus type I (HIV-1) transactivator of transcription (TAT) is encoded by HIV-1. It is a peptide rich in basic amino acids and belongs to the protein transduction domain family. It has been found that HIV-1 TAT and its core peptide segment TAT47-57 play an important role in promoting the cellular uptake of coupled bioactive macromolecules, such as peptides, proteins, oligonucleotides, and drug molecules. HIV-1 TAT can also significantly increase the soluble expression of extrinsic proteins. However, the mechanism behind the cellular uptake of HIV-1 TAT-derived cell-penetrating peptide remains unclear. This review focuses on the research into HIV-1 TAT-derived cell-penetrating peptide over the last years. We briefly discuss TAT's structural features, functions and applications, the mechanism of its cellular internalization, current challenges, and their possible solutions. At the end of this review, we provide a summary and predict the future research directions and potential applications of HIV-1 TAT when it is used as a cell-penetrating peptide.