The antimicrobial property of JY-1, a complex mixture of Traditional Chinese Medicine, is linked to it abilities to suppress biofilm formation and disrupt membrane permeability.

The substantial increase of infections, caused by novel, sudden, and drug-resistant pathogens, poses a significant threat to human health. While numerous studies have demonstrated the antibacterial and antiviral effects of Traditional Chinese Medicine, the potential of a complex mixture of traditional Chinese Medicine with a broad-spectrum antimicrobial property remains underexplored. This study aimed to develop a complex mixture of Traditional Chinese Medicine (TCM), JY-1, and investigate its antimicrobial properties, along with its potential mechanism of action against pathogenic microorganisms. Antimicrobial activity was assessed using a zone of inhibition assay and the drop plate method. Hyphal induction of Candida albicans was conducted using RPMI1640 medium containing 10% FBS, followed by microscopic visualization. Quantitative real-time PCR (RT-qPCR) was employed to quantify the transcript levels of hyphal-specific genes such as HWP1 and ALS3. The impact of JY-1 on biofilm formation was evaluated using both the XTT reduction assay and scanning electron microscopy (SEM). Furthermore, the cell membrane integrity was assessed by protein and nucleic acid leakage assays. Our results clearly showed that JY-1 significantly inhibits the vegetative growth of Candida spp. and Cryptococcus spp. In addition, this complex mixture is effectively against a wide range of pathogenic bacteria, including Staphylococcus aureus, Vancomycin-resistant enterococci, Escherichia coli, Klebsiella pneumoniae and Enterobacter cloacae. More interestingly, JY-1 plays a direct anti-viral role against the mammalian viral pathogen vesicular stomatitis virus (VSV). Further mechanistic studies indicate that JY-1 acts to reduce the expression of hyphal specific genes HWP1 and ALS3, resulting in the suppression of the hyphal formation of C. albicans. The antimicrobial property of JY-1 could be attributed to its ability to reduce biofilm formation and disrupt the cell membrane permeability, a process resulting in microbial cell death and the release of cellular contents. Taken together, our work identified a potent broad-spectrum antimicrobial agent, a complex mixture of TCM which might be developed as a potential antimicrobial drug.

Shengjihuayu formula ameliorates the oxidative injury in human keratinocytes via blocking JNK/c-Jun/MMPs signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The reactive oxygen species (ROS) surge in the chronic wound tissue of diabetic ulcers (DUs) aggravates the inflammatory response. The oxidative stress state during inflammation will exacerbate inflammation and cause tissue damage, resulting in prolonged wound healing. Shengjihuayu Formula (SJHYF) is a renowned Chinese medicine prescription for treating chronic wounds in diabetic ulcers. Growing clinical evidence has demonstrated that SJHYF exhibits superior therapeutic efficacy and has a favorable safety profile. However, the underlying mechanisms by which SJHYF ameliorates oxidative damage under pathological conditions of DUs remain unclear. OBJECTIVE: To investigate the cytoprotective properties of SJHYF on hydrogen peroxide (H2O2)-induced cell damage in human HaCaT keratinocytes and to explore its potential targets and molecular pathways in treating DUs using RNA-seq. METHODS: HaCaT cells were incubated with H2O2 for 24 h to construct an oxidative stress cell model. Cell viability and proliferation were measured using the MTT and EdU assays, respectively. Cell migration was assessed using the scratch assay, and the fluorescence intensity of ROS was measured using the DCFH-DA probe. The chemical components of SJHYF were analyzed by UPLC-Q-TOF/MS, while the therapeutic effects of SJHYF on H2O2-induced HaCaT cells were analyzed using RNA-Seq. The potential target genes were validated using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). At the same time, the pathway phenotype expression of SJHYF on the protection of H2O2-induced HaCaT cells was explored using Western Blot. RESULTS: The application of SJHY at a concentration of 0.25 mg/mL promoted cell proliferation, cell migration, and reduced ROS production. In addition, SJHYF was detected to have a total of 93 active compounds, including key components such as Galloyl-beta-D-glucose, Danshensu, Procyanidin B2, Catechin, and Alkannin. The RNA-seq analysis identified several core targets namely KRT17, TGM1, JUNB, PRDX5, TXNIP, PRDX1, HSP90AA1, HSP90AB1, HSPA8, and TNF-α. Western blot revealed the presence of the JNK/c-Jun/MMPs pathway and its related transcription factors. CONCLUSION: SJHYF displays significant protective effects on H2O2-induced oxidative cell damage in HaCaT cells via blocking the JNK/c-Jun/MMPs pathway.

Ash1L ameliorates psoriasis via limiting neuronal activity-dependent release of miR-let-7b.

BACKGROUND AND PURPOSE: Psoriasis is a common autoimmune skin disease that significantly diminishes patients' quality of life. Interactions between primary afferents of the somatosensory system and the cutaneous immune system mediate the pathogenesis of psoriasis. This study aims to elucidate the molecular mechanisms of how primary sensory neurons regulate psoriasis formation. EXPERIMENTAL APPROACH: Skin and total RNA were extracted from wild-type (WT) and ASH1-like histone lysine methyltransferase (Ash1l+/- ) mice in both naive and imiquimod (IMQ)-induced psoriasis models. Immunohistochemistry, quantitative real-time polymerase chain reaction (qRT-PCR) and fluorescence-activated cell sorting (FACS) were then performed. Microfluidic chamber coculture was used to investigate the interaction between somatosensory neurons and bone marrow dendritic cells (BMDCs) ex vivo. Whole-cell patch clamp recordings were used to evaluate neuronal excitability after Ash1L haploinsufficiency in primary sensory neurons. KEY RESULTS: The haploinsufficiency of ASH1L, a histone methyltransferase, in primary sensory neurons causes both neurite hyperinnervation and increased neuronal excitability, which promote miR-let-7b release from primary afferents in the skin in a neuronal activity-dependent manner. With a 'GUUGUGU' core sequence, miR-let-7b functions as an endogenous ligand of toll-like receptor 7 (TLR7) and stimulates the activation of dermal dendritic cells (DCs) and interleukin (IL)-23/IL-17 axis, ultimately exacerbating the symptoms of psoriasis. Thus, by limiting miR-let-7b release from primary afferents, ASH1L prevents dermal DC activation and ameliorates psoriasis. CONCLUSION AND IMPLICATIONS: Somatosensory neuron ASH1L modulates the cutaneous immune system by limiting neuronal activity-dependent release of miR-let-7b, which can directly activate dermal DCs via TLR7 and ultimately lead to aggravated psoriatic lesion.

METTL14 promotes IL-6-induced viability, glycolysis and inflammation in HaCaT cells via the m6A modification of TRIM27.

Interleukin-6 (IL-6) is a cytokine generated by healthy constituents of the skin, but is also up-regulated by a wide range of skin lesions and inflammatory conditions to trigger cytopathy of skin cells. TRIM27 was identified to contribute to the functional effects of IL-6 on skin cells. However, the underlying mechanism was not clear. Lentivirus infection was used for gene overexpression or silencing. RT-PCR and Western blot were used to respectively assess mRNA and protein levels. Cell viability was assessed by CCK-8 assay. Extracellular flux analysis was used to assess the levels of oxygen consumption rate and extracellular acidification rate. Mouse back skin was treated with imiquimod to produce psoriasis-like inflammation in vivo. Histological assessment and immunohistochemistry staining were respectively applied to analyse lesioned mouse and human skin samples. IL-6-induced increased viability, glycolysis and inflammation in keratinocytes was inhibited both by a chemical methylation inhibitor and by METTL14 knockdown. Further investigation found that METTL14 induces m6A methylation of TRIM27, which is recognized by a m6A reader, IGF2BP2. Elevation of TRIM27 level and activation of IL-6/STAT3 signalling pathway were found in an in vivo psoriasis-like inflammation model, whereas inhibition m6A methylation strongly alleviated the inflammation. Finally, METTL14, TRIM27, STAT3, p-STAT3 and IL-6 expressions were all found to be increased in clinical skin samples of psoriatic patients. Our results unravelled METTL14/TRIM27/IGF2BP2 signalling axis in keratinocyte cytopathy, which plays a critical role in facilitating the activation of IL-6/STAT3 signalling pathway. Our findings should provide inspirations for the design of new therapeutics for skin inflammatory diseases including psoriasis.

Research on the resistance of isoviolanthin to hydrogen peroxide-triggered injury of skin keratinocytes based on Transcriptome sequencing and molecular docking.

Apoptosis of skin keratinocytes is closely associated with skin problems in humans and natural flavonoids have shown excellent biological activity. Hence, the study of flavonoids against human keratinocyte apoptosis has aroused the interest of numerous researchers. In this study, methyl thiazolyl tetrazolium (MTT) assay and Western blots were used to investigate the skin-protective effect of isoviolanthin, a di-C-glycoside derived from Dendrobium officinale, on hydrogen peroxide (H2O2)-triggered apoptosis of skin keratinocytes. Transcriptome sequencing (RNA-Seq) was used to detect the altered expression genes between the model and treatment group and qRT-PCR was used to verify the accuracy of transcriptome sequencing results. Finally, molecular docking was used to observe the binding ability of isoviolanthin to the selected differential genes screened by transcriptome sequencing. Our results found isoviolanthin could probably increase skin keratinocyte viability, by resisting against apoptosis of skin keratinocytes through downregulating the level of p53 for the first time. By comparing transcriptome differences between the model and drug administration groups, a total of 2953 differential expression genes (DEGs) were identified. Enrichment analysis showed that isoviolanthin may regulate these pathways, such as DNA replication, Mismatch repair, RNA polymerase, Fanconi anemia pathway, Cell cycle, p53 signaling pathway. Last, our results found isoviolanthin has a strong affinity for binding to KDM6B, CHAC2, ESCO2, and IPO4, which may be the potential target for treating skin injuries induced by reactive oxide species. The current study confirms isoviolanthin potential as a skin protectant. The findings may serve as a starting point for further research into the mechanism of isoviolanthin protection against skin damage caused by reactive oxide species (e.g., hydrogen peroxide).

Nano drug delivery systems: a promising approach to scar prevention and treatment.

Scar formation is a common physiological process that occurs after injury, but in some cases, pathological scars can develop, leading to serious physiological and psychological effects. Unfortunately, there are currently no effective means to intervene in scar formation, and the structural features of scars and their unclear mechanisms make prevention and treatment even more challenging. However, the emergence of nanotechnology in drug delivery systems offers a promising avenue for the prevention and treatment of scars. Nanomaterials possess unique properties that make them well suited for addressing issues related to transdermal drug delivery, drug solubility, and controlled release. Herein, we summarize the recent progress made in the use of nanotechnology for the prevention and treatment of scars. We examine the mechanisms involved and the advantages offered by various types of nanomaterials. We also highlight the outstanding challenges and questions that need to be addressed to maximize the potential of nanotechnology in scar intervention. Overall, with further development, nanotechnology could significantly improve the prevention and treatment of pathological scars, providing a brighter outlook for those affected by this condition.

Clinical-mediated discovery of pyroptosis in CD8+ T cell and NK cell reveals melanoma heterogeneity by single-cell and bulk sequence.

Histologically, melanoma tissues had fewer positive cells percentage of pyroptosis-related genes (PRGs), GZMA, GSDMB, NLRP1, IL18, and CHMP4A in epidermal than in normal skin. Pyroptosis, a new frontier in cancer, affects the tumor microenvironment and tumor immunotherapy. Nevertheless, the role of pyroptosis remains controversial, which reason is partly due to the heterogeneity of the cellular composition in melanoma. In this study, we present a comprehensive analysis of the single-cell transcriptome landscape of pyroptosis in melanoma specimens. Our findings reveal dysregulation in the expression of PRGs, particularly in immune cells, such as CD8+ cells (representing CD8+ T cells) and CD57+ cells (representing NK cells). Additionally, the immunohistochemical and multiplex immunofluorescence staining experiments results further confirmed GZMA+ cells and GSDMB+ cells were predominantly expressed in immune cells, especially in CD8 + T cells and NK cells. Melanoma specimens secreted a minimal presence of GZMA+ merged CD8+ T cells (0.11%) and GSDMB+ merged CD57+ cells (0.08%), compared to the control groups exhibiting proportions of 4.02% and 0.62%, respectively. The aforementioned findings indicate that a reduced presence of immune cells within tumors may play a role in diminishing the ability of pyroptosis, consequently posing a potential risk to the anti-melanoma properties. To quantify clinical relevance, we constructed a prognostic risk model and an individualized nomogram (C-index=0.58, P = 0.002), suggesting a potential role of PRGs in malignant melanoma prevention. In conclusion, our integrated single-cell and bulk RNA-seq analysis identified immune cell clusters and immune gene modules with experiment validation, contributing to our better understanding of pyroptosis in melanoma.

Qinzhu Liangxue inhibits IL-6-induced hyperproliferation and inflammation in HaCaT cells by regulating METTL14/SOCS3/STAT3 axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Psoriasis, an immune-mediated chronic inflammatory skin condition, is treatable with Qinzhu Liangxue (QZLX), a therapeutic medicinal plant formula used in clinical practice. However, further investigation is needed to clarify its molecular mechanisms of action. AIM OF THE STUDY: The potential biological mechanisms of QZLX to alleviate psoriasis involving IL-6-induced hyperproliferation and inflammation by regulating METTL14/SOCS3/STAT3 axis. MATERIALS AND METHODS: HaCaT cell model was induced by IL-6, and dealt with serum containing QZLX. In addition, shRNAs and siRNAs were used for gene silencing, viruses were collected 48 h post-transfection and infected HaCaT cells. Cell viability was detected by CCK-8 assay, cell cycle was determined by flow cytometry. Finally, psoriasis mice model was induced by IMQ cream, then back skin tissue was used for hematoxylin and eosin (H&E). The content of IL-1ß, IL-6, and IL-8 in cell supernatants were analyzed using ELISA kits. Analysis of SOCS3 was used by quantitative RT-PCR, the expression level of SOCS3, METTL3, METTL14, WTAP, SOCS3, YTHDF2, p-STAT3 and STAT3 in HaCaT cells transduced with METTL14 overexpression was detected by Western blot. RESULTS: All results indicated that QZLX could significantly alleviate IL-6-induced HaCaT cell viability, cell cycle progression, and inhibit the level of IL-1ß, IL-6, and IL-8. The m6A levels and level of METTL14 in HaCaT cells treated with IL-6 were enhanced, while it was reversed by QZLX. METTL14 silencing could inhibit IL-6-induced HaCaT cell viability, cell cycle progression and inflammation response, while SOCS3 overexpression also suppressed METTL14-induced HaCaT cell viability, cell cycle progression and inflammation. QZLX could significantly enhance the expression level of SOCS3, while inhibit the level of METTL14, and p-STAT3/STAT3. In addition, QZLX inhibits METTL14-induced HaCaT cell viability, cell cycle progression, and inhibits the level of IL-1ß, IL-6, and IL-8. CONCLUSIONS: Our finding suggested that QZLX ameliorated the inflammation response of psoriasis and performed the potential anti-psoriasis effect by regulating METTL14/SOCS3/STAT3 axis in both mice and HaCaT cells psoriasis model. Therefore, our study demonstrated a significant strategy for inhibiting psoriasis inflammation via targeting METTL14/SOCS3/STAT3 axis.

World Health Organization survey on the level of integration of traditional Chinese medicine in Chinese health system rehabilitation services.

Background: To meet the growing global demand for rehabilitation services, the World Health Organization (WHO) launched Rehabilitation 2030. This study was commissioned by the WHO to investigate the integration degree of traditional Chinese medicine (TCM) in Chinese health system rehabilitation services and the demand for TCM rehabilitation in China. Methods: Twenty TCM rehabilitation experts and relevant government administrators were invited to complete the questionnaire between September 2019 and January 2022. The development of traditional, complementary, and integrative medicine (TCI) rehabilitation in China was assessed primarily based on six different health system components. Results: 26 policies, regulations, and national strategic plans related to TCI rehabilitation were issued by relevant government departments since 2002; notably, 14 policies related to TCI rehabilitation development were intensively introduced from 2016 to 2021. These policies cover the three main areas of financing, infrastructure development, and service delivery. The National Administration of Traditional Chinese Medicine's investment in TCM clinical capacity infrastructure and scientific research in 2019-2021 increased by 66% compared to 2010-2012, and the average number of TCM hospitals with rehabilitation departments in 2020 increased by 6.5% compared to 2018. The proportion of community health service centers providing TCM services in primary medical and health institutions has increased by 30.8% over the past 10 years. Conclusion: Long-term continuous policies, substantial financial investment, and expansion of the scope of TCI rehabilitation services in primary care institutions have effectively contributed to the rapid development of TCI rehabilitation. However, human resources and financing mechanisms for TCI rehabilitation need further improvement.

Classification and biomarker gene selection of pyroptosis-related gene expression in psoriasis using a random forest algorithm.

Background: Psoriasis is a chronic and immune-mediated skin disorder that currently has no cure. Pyroptosis has been proved to be involved in the pathogenesis and progression of psoriasis. However, the role pyroptosis plays in psoriasis remains elusive. Methods: RNA-sequencing data of psoriasis patients were obtained from the Gene Expression Omnibus (GEO) database, and differentially expressed pyroptosis-related genes (PRGs) between psoriasis patients and normal individuals were obtained. A principal component analysis (PCA) was conducted to determine whether PRGs could be used to distinguish the samples. PRG and immune cell correlation was also investigated. Subsequently, a novel diagnostic model comprising PRGs for psoriasis was constructed using a random forest algorithm (ntree = 400). A receiver operating characteristic (ROC) analysis was used to evaluate the classification performance through both internal and external validation. Consensus clustering analysis was used to investigate whether there was a difference in biological functions within PRG-based subtypes. Finally, the expression of the kernel PRGs were validated in vivo by qRT-PCR. Results: We identified a total of 39 PRGs, which could distinguish psoriasis samples from normal samples. The process of T cell CD4 memory activated and mast cells resting were correlated with PRGs. Ten PRGs, IL-1ß, AIM2, CASP5, DHX9, CASP4, CYCS, CASP1, GZMB, CHMP2B, and CASP8, were subsequently screened using a random forest diagnostic model. ROC analysis revealed that our model has good diagnostic performance in both internal validation (area under the curve [AUC] = 0.930 [95% CI 0.877-0.984]) and external validation (mean AUC = 0.852). PRG subtypes indicated differences in metabolic processes and the MAPK signaling pathway. Finally, the qRT-PCR results demonstrated the apparent dysregulation of PRGs in psoriasis, especially AIM2 and GZMB. Conclusion: Pyroptosis may play a crucial role in psoriasis and could provide new insights into the diagnosis and underlying mechanisms of psoriasis.

PD-L1 Triggered by Binding eIF3I Contributes to the Amelioration of Diabetes-Associated Wound Healing Defects by Regulating IRS4.

Persistent chronic inflammation and delayed epithelialization lead to stalled healing in diabetic ulcers (DUs). PD-L1 shows anti-inflammatory and proliferative activities in healing defects, whereas its function in DU pathogenesis remains unknown. Lower levels of PD-L1 were found in DU tissues, and exogenous PD-L1 has therapeutic effects in the healing process by accelerating re-epithelialization and attenuating prolonged inflammation, which contributed to the delayed wound closure. We detected the downstream effectors of PD-L1 using transcriptional profiles and screened the interacting proteins using immunoprecipitation in combination with mass spectrometry and coimmunoprecipitation assays. The biological functions of eIF3I‒PD-L1‒IRS4 axis were tested both in vivo and in vitro. Finally, we validated the expression levels of eIF3I, PD-L1, and IRS4 in DU tissues from human clinical samples by immunohistochemistry staining. Mechanistically, PD-L1 binds to eIF3I and promotes cutaneous diabetic wound healing by downregulating IRS4. These findings identify that the eIF3I‒PD-L1‒IRS4 axis contributes to wound healing defects, which can serve as a potential therapeutic target in DUs.

Gene set enrichment analysis and ingenuity pathway analysis to identify biomarkers in Sheng-ji Hua-yu formula treated diabetic ulcers.

ETHNOPHARMACOLOGICAL RELEVANCE: Sheng-ji Hua-yu (SJHY) formula is a Chinese herbal prescription for diabetic ulcers (DUs) treatment, which can accelerate wound reconstruction and shorten the healing time. However, its mechanism role maintains unclear. AIM OF THE STUDY: To elucidate the molecular mechanisms of SJHY application on DUs. MATERIALS AND METHODS: To begin with, transcriptome sequencing was adopted to identified differentially expression mRNAs among normal ulcers, DUs, and DUs + SJHY treatment in vivo. Liquid chromatography-tandem mass spectrometry was applied for the quality control of SJHY formula. GO and KEGG enrichment analysis were used to identify the mechanisms underlying the therapeutic effect of SJHY formula, and then gene set enrichment analysis and ingenuity pathway analysis were conducted for functional analysis. Further, qPCR detection was performed in vivo for validation. RESULTS: SJHY administration could regulate the glucose metabolic process, AMPK and HIF-1 pathway to accelerate healing processes of DUs. Besides, CRHR1, SHH, and GAL were identified as the critical targets, and SLC6A3, GRP, FGF23, and CYP27B1 were considered as the upstream genes of SJHY treatment. Combined with animal experiments, the prediction results were validated in DUs mice model. CONCLUSIONS: This study used modular pharmacology analysis to identify the biomarkers of SJHY formula and provide the potential therapeutic targets for DUs treatment as well.

Flavonoids in Ampelopsis grossedentata as covalent inhibitors of SARS-CoV-2 3CLpro: Inhibition potentials, covalent binding sites and inhibitory mechanisms.

Coronavirus 3C-like protease (3CLpro) is a crucial target for treating coronavirus diseases including COVID-19. Our preliminary screening showed that Ampelopsis grossedentata extract (AGE) displayed potent SARS-CoV-2-3CLpro inhibitory activity, but the key constituents with SARS-CoV-2-3CLpro inhibitory effect and their mechanisms were unrevealed. Herein, a practical strategy via integrating bioactivity-guided fractionation and purification, mass spectrometry-based peptide profiling and time-dependent biochemical assay, was applied to identify the crucial constituents in AGE and to uncover their inhibitory mechanisms. The results demonstrated that the flavonoid-rich fractions (10-17.5 min) displayed strong SARS-CoV-2-3CLpro inhibitory activities, while the constituents in these fractions were isolated and their SARS-CoV-2-3CLpro inhibitory activities were investigated. Among all isolated flavonoids, dihydromyricetin, isodihydromyricetin and myricetin strongly inhibited SARS-CoV-2 3CLpro in a time-dependent manner. Further investigations demonstrated that myricetin could covalently bind on SARS-CoV-2 3CLpro at Cys300 and Cys44, while dihydromyricetin and isodihydromyricetin covalently bound at Cys300. Covalent docking coupling with molecular dynamics simulations showed the detailed interactions between the orthoquinone form of myricetin and two covalent binding sites (surrounding Cys300 and Cys44) of SARS-CoV-2 3CLpro. Collectively, the flavonoids in AGE strongly and time-dependently inhibit SARS-CoV-2 3CLpro, while the newly identified SARS-CoV-2 3CLpro inhibitors in AGE offer promising lead compounds for developing novel antiviral agents.

Spectrophotometric Assays for Sensing Tyrosinase Activity and Their Applications.

Tyrosinase (TYR, E.C. 1.14.18.1), a critical enzyme participating in melanogenesis, catalyzes the first two steps in melanin biosynthesis including the ortho-hydroxylation of L-tyrosine and the oxidation of L-DOPA. Previous pharmacological investigations have revealed that an abnormal level of TYR is tightly associated with various dermatoses, including albinism, age spots, and malignant melanoma. TYR inhibitors can partially block the formation of pigment, which are always used for improving skin tone and treating dermatoses. The practical and reliable assays for monitoring TYR activity levels are very useful for both disease diagnosis and drug discovery. This review comprehensively summarizes structural and enzymatic characteristics, catalytic mechanism and substrate preference of TYR, as well as the recent advances in biochemical assays for sensing TYR activity and their biomedical applications. The design strategies of various TYR substrates, alongside with several lists of all reported biochemical assays for sensing TYR including analytical conditions and kinetic parameters, are presented for the first time. Additionally, the biomedical applications and future perspectives of these optical assays are also highlighted. The information and knowledge presented in this review offer a group of practical and reliable assays and imaging tools for sensing TYR activities in complex biological systems, which strongly facilitates high-throughput screening TYR inhibitors and further investigations on the relevance of TYR to human diseases.

Discovery and characterization of flavonoids in vine tea as catechol-O-methyltransferase inhibitors.

Vine tea has been used as a traditionally functional herbal tea in China for centuries, which exhibits paramount potential for chronic metabolic diseases. Herein, the inhibitory potential of vine tea toward human catechol-O-methyltransferase (hCOMT) was investigated. A practical bioactivity-guided fractionation combined with chemical profiling strategy was developed to identify the naturally occurring hCOMT inhibitors. Five flavonoids in vine tea displayed moderate to strong inhibition on hCOMT with IC50 values ranging from 0.96 µM to 42.47 µM, in which myricetin was the critically potent constituent against hCOMT. Inhibition kinetics assays and molecular docking simulations showed that myricetin could bind to the active site of COMT and inhibited COMT-catalyzed 3-BTD methylation in a mixed manner. Collectively, our findings not only suggested that the strong hCOMT inhibition of vine tea has guiding significance in the drug exposure of catechol drugs, but also identified a promising lead compound for developing more efficacious hCOMT inhibitors.

Transcriptional profiling and circRNA-miRNA-mRNA network analysis identify the biomarkers in Sheng-ji Hua-yu formula treated diabetic wound healing.

ETHNOPHARMACOLOGICAL RELEVANCE: Sheng-ji Hua-yu (SJHY) formula is a traditional Chinese herbal which is effective in treating diabetic ulcers. It has been indicated to accelerate re-epithelialization and healing time of cutaneous wounds in a Streptozotocin (STZ)-induced diabetic mouse model. However, its mechanisms remain undetermined. AIM OF THE STUDY: To reveal the molecular mechanisms of SJHY formula in treating diabetic wounds through transcriptional profiling and circRNA-miRNA-mRNA network analysis clues. MATERIALS AND METHODS: Protein expressions of tumor necrosis factor (TNF-α), interleukin (IL)-6, IL-1ß in skin tissues of wounds from SJHY formula-treated and untreated mice were analyzed by Bio-Plex assay. Differentially expressed (DE) genes were detected by whole transcriptome sequencing (RNA-seq). Using predicted miRNA targets, circRNA-miRNA-mRNA networks were constructed. Furthermore, quantitative real-time PCR (qRT-PCR) was utilized to validate the circRNA-miRNA-mRNA networks. RESULTS: Bio-Plex assay illustrated that the protein expressions of TNF-α, IL-1ß, IL-6 were downregulated in SJHY formula-treated diabetic wounds compared with untreated wounds. RNA-seq identified 11 DE circRNAs and 476 DE mRNAs between SJHY formula-treated and diabetic mice, including 4 upregulated and 7 downregulated circRNAs, 311 upregulated and 165 downregulated mRNAs. CircRNA-Krt13/miR-665-3p/Itga3 and circRNA-Krt14/miR-706/Mylk4 pathways were built, which may contribute to the healing of SJHY formula-treated diabetic wounds. CONCLUSIONS: Overall, this study suggests that these 2 circRNA-miRNA-mRNA networks are potential biomarkers for evaluation of SJHY formula efficacy in diabetic wound healing, which provides evidence to support its clinical applications.

Fabrication of Composite Structures of Lysozyme Fibril-Zein using Antisolvent Precipitation: Effects of Blending and pH Adjustment Sequences.

Antisolvent precipitation is a widely used method to fabricate prolamin-based composites. In the present study, composite structures of lysozyme amyloid fibrils with zein proteins were fabricated using the antisolvent precipitation method by applying different blending and pH adjustment sequences. Globular prolamins were bound to the amyloid fibrils to combine their respective advantages. The dynamic light scattering showed that the composites with a characteristic stabilized behavior (43.60 ± 1.75 mV ∼ 35.20 ± 0.65 mV) were formed at pH 4.0-5.0, in which noncovalent interactions between fibril and particles occurred. Two different structures: fruit tree-like structure and beaded-like structure, were presented in AFM and TEM images due to the different pH adjustment sequences, while blending sequences had negligible effect on the morphology of the composites. A fruit tree-like entity was detected for lysozyme fibril-zein composites, where its "branches" bear zein globular particles. A beaded-like structure was observed for lysozyme fibril-zein composites, where lysozyme fibril was the thread and zein aggregates were the beads. The potential mechanism of this phenomenon can be explained as the fruit tree-like structure being primarily formed through electrostatic interactions while the beaded-like structure is mainly caused by hydrophobic interactions. The composites of fruit tree-like structures hold a more promising stability than those with beaded-like structures. The results of this research would give constructive information for the fabrication of amyloid fibril-prolamin protein composites, which may exhibit the combined advantages of each components and have potential applications in encapsulation and protection of bioactive substances and stabilizing emulsions.

Herb-drug interaction between Styrax and warfarin: Molecular basis and mechanism.

BACKGROUND: Styrax, one of the most famous folk medicines, has been frequently used for the treatment of cardiovascular diseases and skin problems in Asia and Africa. It is unclear whether Styrax or Styrax-related herbal medicines may trigger clinically relevant herb-drug interactions. PURPOSE: This study was carried out to investigate the inhibitory effects of Styrax on human cytochrome P450 enzymes (CYPs) and to clarify whether this herb may modulate the pharmacokinetic behavior of the CYP-substrate drug warfarin when co-administered. STUDY DESIGN: The inhibitory effects of Styrax on CYPs were assayed in human liver microsomes (HLM), while the pharmacokinetic interactions between Styrax and warfarin were investigated in rats. The bioactive constituents in Styrax with strong CYP3A inhibitory activity were identified and their inhibitory mechanisms were carefully investigated. METHODS: The inhibitory effects of Styrax on human CYPs were assayed in vitro, while the pharmacokinetic interactions between Styrax and warfarin were studied in rats. Fingerprinting analysis of Styrax coupled with LC-TOF-MS/MS profiling and CYP inhibition assays were used to identify the constituents with strong CYP3A inhibitory activity. The inhibitory mechanism of oleanonic acid (the most potent CYP3A inhibitor occurring in Styrax) against CYP3A4 was investigated by a panel of inhibition kinetics analyses and in silico analysis. RESULTS: In vitro assays demonstrated that Styrax extract strongly inhibited human CYP3A and moderately inhibited six other tested human CYPs, as well as potently inhibited warfarin 10-hydroxylation in liver microsomes from both humans and rats. In vivo assays demonstrated that compared with warfarin given individually in rats, Styrax (100 mg/kg) significantly prolonged the plasma half-life of warfarin by 2.3-fold and increased the AUC(0-inf) of warfarin by 2.7-fold when this herb was co-administrated with warfarin (2 mg/kg) in rats. Two LC fractions were found with strong CYP3A inhibitory activity and the major constituents in these fractions were characterized by LC-TOF-MS/MS. Five pentacyclic triterpenoid acids (including epibetulinic acid, betulinic acid, betulonic acid, oleanonic acid and maslinic acid) present in Styrax were potent CYP3A inhibitors, and oleanonic acid was a competitive inhibitor against CYP3A-mediated testosterone 6ß-hydroxylation. CONCLUSION: Styrax and the pentacyclic triterpenoid acids occurring in this herb strongly modulate the pharmacokinetic behavior of warfarin via inhibition of CYP3A.

TRIM27 promotes IL-6-induced proliferation and inflammation factor production by activating STAT3 signaling in HaCaT cells.

The IL-6/STAT3 signaling pathway is required for the development of psoriatic lesions, and tripartite motif-containing 27 (TRIM27) is a protein inhibitor of activated STAT3 (PIAS3)-interacting protein that could modulate IL-6-induced STAT3 activation. However, whether TRIM27 is associated with the IL-6/STAT3 signaling pathway in psoriasis remains enigmatic. TRIM27 expression and gene set enrichment analysis in patients with psoriasis were determined using bioinformatics. Human keratinocyte HaCaT cells treated with recombinant protein IL-6 (rh-IL-6) were transduced with lentivirus silencing TRIM27 and/or PIAS3 or, otherwise, transduced with lentivirus expressing TRIM27 and/or lentivirus silencing STAT3, or MG132, a proteasome-specific protease inhibitor. Cell proliferation and inflammation factor production were measured using Cell Counting Kit-8 and ELISA, respectively. TRIM27, proliferation marker protein Ki-67 (Ki67), phospho-STAT3 (p-STAT3), STAT3, and PIAS3 expressions were determined using real-time quantitative PCR, immunofluorescence staining, or Western blot analysis. Coimmunoprecipitation combined with ubiquitination analysis was performed to explore the interaction between TRIM27 and PIAS3. In the present study, TRIM27 expression was increased in psoriatic lesions, associated with the IL-6 signaling pathway, and induced by rh-IL-6 in a time-dependent manner. The increased cell proliferation, inflammation factor production, and expression of Ki67 and of p-STAT3 relative to STAT3 induced by rh-IL-6 and TRIM27 overexpression were significantly inhibited by TRIM27 silencing and STAT3 silencing, respectively. More importantly, TRIM27 interacted with PIAS3, and its overexpression promoted PIAS3 ubiquitination in HaCaT cells. PIAS3 silencing also significantly promoted TRIM27-dependent and IL6-induced STAT3 activation, cell proliferation, and inflammation factor production. In conclusion, our results highlight that TRIM27 expression is significantly increased by IL-6 and suggest a TRIM27/STAT3-dependent mechanism for regulation of inflammation and proliferation-associated development of psoriasis.