Effectiveness and mechanism of the Chinese medicine Weiren Xiaoyou formula in improving palmoplantar warts.

Background: Palmoplantar warts (PWs) are a usual skin disease associated with human papillomavirus (HPV) that can affect patients' quality of life. The traditional Chinese medicine (TCM) Weiren Xiaoyou formula (WRXYF) is a relatively gentle and effective therapy that has achieved good therapeutic effects in clinical practice, but its mechanism has not yet been studied. Methods: A meta-analysis was carried out to identify the potential advantages of topical TCM for PW treatment. Clinical cases suggested that WRXYF was an effective therapeutic agent against PWs. Network pharmacology was utilized to predict potential targets for the main bioactive compound, tanshinone IIA (Tan IIA), in WRXYF. High-performance liquid chromatography with electrospray mass spectrometry (HPLC/ESI-MS) was applied to detect major components. The bioactivity of Tan IIA against PWs was then validated with quantitative polymerase chain reaction (q-PCR), fluorescence in situ hybridization (FISH), electron microscopy and Western blotting. Results: A meta-analysis was conducted on 10 randomized clinical trials (RCTs) involving 2260 participants suggested that topical TCM could more effectively treat PWs than conventional medications. Network pharmacology identified Tan IIA as a candidate agent from 17 major compounds assessed by HPLC/ESI-MS because of its stable binding with 10 PW targets. HPV2, HPV27, and HPV57 were the main infectious strains in tissues obtained from PW patients and in HPV-infected HaCaT cells. Tan IIA treatment effectively destroyed viral particles and reduced the viral copy numbers of the three HPV subtypes. The results shown that Tan IIA has the ability to halt the cell cycle of HPV-infected HaCaT cells specifically in the G0/G1 phase. A total of 6 cell cycle-related proteins were regulated after Tan IIA treatment, demonstrating the role of Tan IIA in inhibiting the cell cycle. Conclusion: Tan IIA, the primary bioactive constituent in WRXYF, enhances PWs by halting the cell cycle in the G0/G1 phase via modulation of the p53 signaling pathway.

DNA methylation of microRNA-365-1 induces apoptosis of hair follicle stem cells by targeting DAP3.

Background: DNA methylation is a crucial epigenetic alteration involved in diverse biological processes and diseases. Nevertheless, the precise role of DNA methylation in chemotherapeutic drug-induced alopecia remains unclear. This study examined the role and novel processes of DNA methylation in regulating of chemotherapeutic drug-induced alopecia. Methods: A mouse model of cyclophosphamide (CTX)-induced alopecia was established. Hematoxylin-eosin staining and immunohistochemical staining for the Ki67 proportion and a mitochondrial membrane potential assay (JC-1) were performed to assess the structural integrity and proliferative efficiency of the hair follicle stem cells (HFSCs). Immunofluorescence staining and real-time fluorescence quantitative PCR (RT-qPCR) were performed to determine the expression levels of key HFSC markers, namely Lgr5, CD49f, Sox9, CD200, and FZD10. Differential DNA methylation levels between the normal and CTX-induced model groups were determined through simple methylation sequencing and analyzed using bioinformatics tools. The expression levels of miR-365-1, apoptosis markers, and DAP3 were detected through RT-qPCR and western blotting. In parallel, primary mouse HFSCs were extracted and used as a cell model, which was constructed using 4-hydroperoxycyclophosphamide. The luciferase reporter gene assay was conducted to confirm miR-365-1 binding to DAP3. To measure the expression of relevant indicators, superoxide dismutase (SOD) and malondialdehyde (MDA) kits were used. Methylation-specific PCR (MS-PCR) was performed to determine DNA methylation levels. The regulatory relationship within HFSCs was confirmed through plasmid overexpression of miR-365-1 and DAP3. Result: In the alopecia areata model, a substantial number of apoptotic cells were observed within the hair follicles on the mouse backs. Immunofluorescence staining revealed that the expression of HFSC markers significantly reduced in the CTX group. Both RT-qPCR and western blotting demonstrated a noteworthy difference in DNA methyltransferase expression. Simple methylation sequencing unveiled that DNA methylation substantially increased within the dorsal skin of the CTX group. Subsequent screening identified miR-365-1 as the most differentially expressed miRNA. miR-365-1 was predicted and confirmed to bind to the target gene DAP3. In the CTX group, SOD and ATP expression markedly reduced, whereas MDA levels were significantly elevated. Cellular investigations revealed 4-HC-induced cell cycle arrest and decreased expression of HFSC markers. MS-PCR indicated hypermethylation modification of miR-365-1 in the 4-HC-induced HFSCs. The luciferase reporter gene experiment confirmed the binding of miR-365-1 to the DAP3 promoter region. miR-365-1 overexpression dramatically reduced apoptotic protein expression in the HFSCs. However, this effect was slightly reversed after DAP3 overexpression in lentivirus. Conclusion: This study explored the occurrence of miR-365-1 DNA methylation in chemotherapeutic drug-induced alopecia. The results unveiled that miR-365-1 reduces cell apoptosis by targeting DAP3 in HFSCs, thereby revealing the role of DNA methylation of the miR-365-1 promoter in chemotherapeutic drug-induced alopecia.

Identification of IRF1 as a Novel Pyroptosis-Related Prognostic Biomarker of Atopic Dermatitis.

Purpose: The aim of this study was to characterize key biomarkers associated with pyroptosis in atopic dermatitis (AD). Materials and methods: To identify the differentially expressed pyroptosis-related genes (DEPRGs), the gene expression profiles GSE16161 and GSE32924 from the Gene Expression Omnibus (GEO) database were utilized. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted to determine the potential biological functions and involved pathways. Furthermore, protein-protein interaction network analyses were performed to identify hub genes. The types and proportions of infiltrating immune cells were detected by immune filtration analysis using CIBERSORT. A 12-axis competing endogenous RNA (ceRNA) network was constructed utilizing the miRNet database. Immunohistochemistry (IHC) further validated the differential expression of a key gene IRF1 in the skin tissues collected from AD patients. The collection of skin tissue from human subjects in this study were reviewed and approved by the IRB of Yueyang Integrated Chinese and Western Medicine Hospital (KYSKSB2020-125). Results: The study identified a total of 76 DEPRGs, which were enriched in genes associated with the inflammatory response and immune regulation. There was a higher percentage of activated dendritic cells and a lower percentage of resting mast cells in AD samples. PVT1 expression was associated with upregulation of hub genes including CXCL8, IRF1, MKI67, and TP53 in the ceRNA network and was correlated with activated dendritic cells in AD. As a transcription factor, IRF1 could regulate the production of downstream inflammatory factors. The IHC study revealed that IRF1 was overexpressed in the skin tissues of AD patients, which were consistent with the results of the bioinformatic study. Conclusions: IRF1 and its related genes were identified as key pyroptosis-related biomarkers in AD, which is a crucial pathway in the pathogenesis of AD.

Therapeutic effects of the extract of Sancao Formula, a Chinese herbal compound, on imiquimod-induced psoriasis via cysteine-rich protein 61.

OBJECTIVE: Psoriasis is a common chronic inflammatory skin disease that is prone to recurrence, and the proinflammatory factor, cysteine-rich protein 61 (Cyr61), is important in its pathophysiology. Long-term clinical practice has shown that Sancao Formula (SC), a Chinese herbal compound, is effective in the treatment of psoriasis, but the precise mechanism remains unknown. In this study, we investigate the mechanism by which SC extract alleviates imiquimod (IMQ)-induced psoriasis. METHODS: The expression of Cyr61 in psoriatic lesions and normal healthy skin was detected using immunohistochemical analysis to investigate the biological role of Cyr61 in models of psoriatic inflammation. A psoriatic mouse model was established by topical application of IMQ, and the effect of topical application of SC extract was evaluated using the psoriasis area and severity index (PASI) score, hematoxylin-eosin staining, and histopathological features of the skin. Next, a HaCaT cell inflammation model was established using interferon-γ (IFN-γ), and the effect of SC extract on the mRNA and protein levels of Cyr61 and intercellular cell adhesion molecule-1 (ICAM-1) was confirmed using Western blot and quantitative real-time polymerase chain reaction analyses. RESULTS: Immunohistochemical staining showed that the expression of Cyr61 in psoriatic lesions was higher than that in normal skin samples (78.26% vs 41.18%, P < 0.05), and the number of Cyr61-positive cells in psoriatic lesions was also significantly higher than in normal skin (18.66 ± 2.51 vs 4.33 ± 1.52, P < 0.05). Treatment in mice with IMQ-induced psoriasis showed that SC extract could significantly improve the inflammatory phenotype, PASI score (10.875 ± 0.744 vs 3.875 ± 0.582, P < 0.05), and pathological features compared with those in IMQ model group; SC treatment was also associated with decreased levels of Cyr61 and ICAM-1. In the IFN-γ-induced inflammatory cell model, the mRNA and protein levels of Cyr61 and ICAM-1 were upregulated, while the SC extract downregulated the levels of Cyr61 and ICAM-1. CONCLUSION: The results provide a theoretical basis for the involvement of Cyr61 in the pathogenesis of psoriasis, and suggest that SC should be used to target Cyr61 for the prevention of psoriasis recurrence.