PL-ReliefTMplus Alleviates Atopic Dermatitis and Regulates Inflammatory Responses via Inhibiting NF-κB Signaling Pathway.

BACKGROUND: Atopic dermatitis (AD) has various detrimental effects on individuals with limited drug cure rates which necessitate the development of new treatment methods. PL-ReliefTMplus (PLR) is composed of SupraOlive, Crocus Sativus extracts and Citrus reticulata extracts. The effect of PLR on AD remains to be explored. METHODS: 2,4-dinitrofluorobenzene-induced AD model mice were involved and the histopathology of the skin lesions was observed along with the levels of inflammatory chemokines levels were measured. To further validate the molecular mechanism of PLR, RNA-seq was performed in HaCaT cells. Western blotting and immunofluorescence were performed to investigate NF-κB signaling pathways response in AD. RESULTS: Due to PLR treatment, the thickening of the epidermis and dermis was inhibited and the number of eosinophils, mast cells, and CD4+ T cells in the skin lesion was decreased. In addition, the levels of inflammatory cytokines were decreased in dorsal skin tissues and LPS-stimulated HaCat cells. Furthermore, KEGG pathway analysis suggested that most identified downstream biological functions were associated with inflammatory response. PLR inhibited NF-κB signaling in AD mice and HaCaT cells. CONCLUSIONS: These results indicate that PLR is a potent therapeutic agent for attenuating symptoms of AD.

Measurement of Ca2+ Uptake Through Connexin Hemichannels.

Increasing evidence points to deregulated flux of ionized calcium (Ca2+) mediated by hyperactive mutant connexin (Cx) hemichannels (HCs) as a common gain-of-function etiopathogenetic mechanism for several diseases, ranging from skin disorders to nervous system defects. Furthermore, the opening of nonmutated Cx HCs is associated with an impressive list of widespread diseases including, but not limited to, ischemia/stroke, Alzheimer's disease, and epilepsy. HC inhibitors are attracting a growing attention due to their therapeutic potential for numerous pathologies. This chapter describes a quantitative method to measure Ca2+ uptake though HCs expressed in cultured cells. The assay we developed can be used to probe HC activity as wells as to test HC inhibitors. Furthermore, with minor changes it can be easily adapted to high-throughput high-content platforms and/or primary cells and microtissues.

Anti-Inflammatory Properties of Cannabidiol and Beta-Caryophyllene Alone or Combined in an In Vitro Inflammation Model.

Cannabis contains over 500 different compounds, including cannabinoids, terpenoids, and flavonoids. Cannabidiol (CBD) is a non-psychoactive constituent, whereas beta-caryophyllene (BCP) is one of most the well-known terpenoids of Cannabis sativa. In recent years, there has been an emerging idea that the beneficial activities of these compounds are greater when they are combined. The aim of this study was to evaluate the anti-inflammatory effect of CBD and BCP using the in vitro model of lipopolysaccharide (LPS)-stimulated human keratinocytes (HaCaT) cells. The vitality of the cells was quantified using LDH and MTT assays. The levels of the following pro-inflammatory proteins and genes were quantified: IL-1ß, COX-2, and phospho-NF-κB p65 (p-p65) through Western blotting (WB) and interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and tumor necrosis factor-α (TNFα) through quantitative real-time polymerase chain reaction (RT-qPCR). When present in the incubation medium, CBD and BCP reduced the increased levels of pro-inflammatory proteins (IL-1ß, COX-2, and p-NF-kB) induced by LPS. The anti-inflammatory effects of CBD were blocked by a PPARγ antagonist, whereas a CB2 antagonist was able to revert the effects of BCP. Selected concentrations of CBD and BCP were able to revert the increases in the expression of pro-inflammatory genes (IL-1ß, IL-6, and TNFα), and these effects were significant when the drugs were used in combination. Our results suggest that CBD and BCP work in concert to produce a major anti-inflammatory effect with good safety profiles.

Deciphering Staphylococcus aureus-host dynamics using dual activity-based protein profiling of ATP-interacting proteins.

The utilization of ATP within cells plays a fundamental role in cellular processes that are essential for the regulation of host-pathogen dynamics and the subsequent immune response. This study focuses on ATP-binding proteins to dissect the complex interplay between Staphylococcus aureus and human cells, particularly macrophages (THP-1) and keratinocytes (HaCaT), during an intracellular infection. A snapshot of the various protein activity and function is provided using a desthiobiotin-ATP probe, which targets ATP-interacting proteins. In S. aureus, we observe enrichment in pathways required for nutrient acquisition, biosynthesis and metabolism of amino acids, and energy metabolism when located inside human cells. Additionally, the direct profiling of the protein activity revealed specific adaptations of S. aureus to the keratinocytes and macrophages. Mapping the differentially activated proteins to biochemical pathways in the human cells with intracellular bacteria revealed cell-type-specific adaptations to bacterial challenges where THP-1 cells prioritized immune defenses, autophagic cell death, and inflammation. In contrast, HaCaT cells emphasized barrier integrity and immune activation. We also observe bacterial modulation of host processes and metabolic shifts. These findings offer valuable insights into the dynamics of S. aureus-host cell interactions, shedding light on modulating host immune responses to S. aureus, which could involve developing immunomodulatory therapies. IMPORTANCE: This study uses a chemoproteomic approach to target active ATP-interacting proteins and examines the dynamic proteomic interactions between Staphylococcus aureus and human cell lines THP-1 and HaCaT. It uncovers the distinct responses of macrophages and keratinocytes during bacterial infection. S. aureus demonstrated a tailored response to the intracellular environment of each cell type and adaptation during exposure to professional and non-professional phagocytes. It also highlights strategies employed by S. aureus to persist within host cells. This study offers significant insights into the human cell response to S. aureus infection, illuminating the complex proteomic shifts that underlie the defense mechanisms of macrophages and keratinocytes. Notably, the study underscores the nuanced interplay between the host's metabolic reprogramming and immune strategy, suggesting potential therapeutic targets for enhancing host defense and inhibiting bacterial survival. The findings enhance our understanding of host-pathogen interactions and can inform the development of targeted therapies against S. aureus infections.

miR-26a-5p inhibits the proliferation of psoriasis-like keratinocytes in vitro and in vivo by dual interference with the CDC6/CCNE1 axis.

Psoriasis is a chronic inflammatory proliferative dermatological ailment that currently lacks a definitive cure. Employing data mining techniques, this study identified a collection of substantially downregulated miRNAs (top 10). Notably, 32 targets were implicated in both the activation of the IL-17 signaling pathway and cell cycle dysregulation. In silico analysis revealed that one of these miRNAs, miR-26a-5p, is a highly conserved cross-species miRNA. Strikingly, the miR-26a-5p sequences in humans and mice are identical, and mmu-miR-26a-5p was found to target the same 7 cell cycle targets as its human counterpart, hsa-miR-26a-5p. Among these targets, CDC6 and CCNE1 were the most effective targets of miR-26a-5p, which was further validated in vitro using a dual luciferase reporter system and qPCR assay. The therapeutic assessment of miR-26a-5p revealed its remarkable efficacy in inhibiting the proliferation and G1/S transition of keratinocytes (HaCaT and HEKs) in vitro. In vivo experiments corroborated these findings, demonstrating that miR-26a-5p effectively suppressed imiquimod (IMQ)-induced psoriasis-like skin lesions in mice over an 8-day treatment period. Histological analysis via H&E staining revealed that miR-26a-5p treatment resulted in reduced keratinocyte thickness and immune cell infiltration into the spleens of IMQ-treated mice. Mechanistic investigations revealed that miR-26a-5p induced a cascade of downregulated genes associated with the IL-23/IL-17A axis, which is known to be critical in psoriasis pathogenesis, while concomitantly suppressing CDC6 and CCNE1 expression. These findings were corroborated by qPCR and Western blot analyses. Collectively, our study provides compelling evidence supporting the therapeutic potential of miR-26a-5p as a safe and reliable endogenous small nucleic acid for the treatment of psoriasis.

Correlational analysis of PLIN1 with inflammation in diabetic foot ulcer wounds.

BACKGROUND: The persistent presence of inflammation is a recognized pathogenic mechanisms of diabetic foot ulcers (DFUs). We aimed to investigate the expression of PLIN1 in tissues from DFU patients and assess its potential association with inflammation-induced damage. METHODS: We performed transcriptome sequencing and correlation analysis of the foot skin from patients with or without DFUs. Additionally, we examined the correlation between PLIN1 and related inflammatory indicators by analyzing PLIN1 expression in tissue and serum samples and through high-glucose stimulation of keratinocytes (HaCaT cells). RESULTS: PLIN1 is upregulated in the tissue and serum from DFU patients. Additionally, PLIN1 shows a positive correlation with leukocytes, neutrophils, monocytes, C-reactive protein, and procalcitonin in the serum, as well as IL-1ß and TNF-α in the tissues. Experiments with Cells demonstrated that reduced expression of PLIN1 leads to significantly decreased expression of iNOS, IL-1ß, IL-6, IL-18, and TNF-α. PLIN1 may mediate wound inflammatory damage through the NF-κB signaling pathway. CONCLUSION: Our findings suggest that PLIN1 mediates the inflammatory damage in DFU, offering new prospects for the treatment of DFU.

Protective autophagy enhances antistress ability through AMPK/ULK1 signaling pathway in human immortalized keratinocytes.

Keratinocytes, located in the outermost layer of human skin, are pivotal cells to resist environmental damage. Cellular autophagy plays a critical role in eliminating damaged organelles and maintaining skin cell homeostasis. Low-dose 5-Aminolevulinic acid photodynamic therapy (ALA-PDT) has been demonstrated to enhance skin's antistress ability; however, the regulatory mechanisms of autophagy in keratinocytes remain unclear. In this study, we treated immortalized human keratinocytes (HaCaT cells) with low-dose ALA-PDT (0.5 mmol/L, 3 J/cm2). Through RNA-sequencing analysis, we identified that low-dose ALA-PDT modulated autophagy-related pathways in keratinocytes and pinpointed Unc-51-like kinase 1 (ULK1) as a key gene involved. Western blot results revealed that low-dose ALA-PDT treatment upregulated the expression of autophagy-related proteins Beclin-1 and LC3-II/LC3-I ratio. Notably, low-dose ALA-PDT regulated autophagy by inducing an appropriate level of reactive oxygen species (ROS), transiently reducing mitochondrial membrane potential, and decreasing adenosine triphosphate production; all these processes functioned on the AMP-activated protein kinase (AMPK)/ULK1 pathway to activate autophagy. Finally, we simulated external environmental damage using ultraviolet B (UVB) at a dose of 60 mJ/cm2 and observed that low-dose ALA-PDT mitigated UVB-induced cell apoptosis; however, this protective effect was reversed when using the autophagy inhibitor 3-methyladenine. Overall, these findings highlight how low-dose ALA-PDT enhances antistress ability in HaCaT cells through controlling ROS generation and activating the AMPK/ULK1 pathway to arouse cellular autophagy.

Punicalagin from pomegranate ameliorates TNF-α/IFN-γ-induced inflammatory responses in HaCaT cells via regulation of SIRT1/STAT3 axis and Nrf2/HO-1 signaling pathway.

Punicalagin (PUN) was isolated from the peel of pomegranate (Punica granatum L.), is a polyphenol with anti-inflammatory, hepatoprotective, and antioxidant activities. However, it remains unclear whether PUN alleviates the inflammation and anti-inflammatory mechanisms in pro-inflammatory cytokines-induced human keratinocyte HaCaT cells. Here, we investigated that tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) mixture-stimulated HaCaT cells were treated with various concentrations of PUN, followed by analyzed the expression of inflammation-related mediators and evaluate anti-inflammatory-related pathways. Our results demonstrated that PUN ≤ 100 µM did not reduce HaCaT cell viability, and PUN ≥ 3 µM was sufficient to decrease interleukin-6 (IL-6), IL-8, monocyte chemoattractant protein-1 (MCP-1), chemokine ligand 5 (CCL5), CCL17 and CCL20 concentrations. We found that PUN ≥ 10 µM and ≥ 3 µM significantly increased sirtuin 1 (SIRT1) expression and inhibited signal transducer and activator of transcription 3 (STAT3) phosphorylation, respectively. PUN downregulated inflammation-related proteins cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), enhanced nuclear factor erythroid-2-related factor-2 (Nrf2) and heme oxygenase-1 (HO-1) expression. Moreover, PUN decreased intercellular adhesion molecule-1 (ICAM-1) expression and inhibited monocyte adhesion to inflamed HaCaT cells. PUN also suppressed inflammatory-related pathways, including mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signaling pathways in TNF-α/IFN-γ- stimulated HaCat cells. Collectively, there is significant evidence that PUN has effective protective defenses against TNF-α/IFN-γ-induced skin inflammation by enhancing SIRT1 to mediate STAT3 and Nrf2/HO-1 signaling pathway.

1-Kestose Blocks UVB-Induced Skin Inflammation and Promotes Type I Procollagen Synthesis via Regulating MAPK/AP-1, NF-κB and TGF-ß/Smad Pathway.

Solar UVB irradiation cause skin photoaging by inducing the high expression of matrix metalloproteinase (MMPs) to inhibit the expression of Type1 procollagen synthesis. 1-Kestose, a natural trisaccharide, has been indicated to show a cytoprotective role in UVB radiation-induced-HaCaT cells. However, few studies have confirmed the anti-aging effects. In the present study, we evaluated the anti-photoaging and pathological mechanism of 1-kestose using Human keratinocytes (HaCaT) cells. The results found that 1-kestose pretreatment remarkably reduced UVB-generated reactive oxygen species (ROS) accumulation in HaCaT cells. 1-Kestose suppressed UVB radiation-induced MMPs expressions by blocking MAPK/AP-1 and NF-κB p65 translocation. 1-Kestose pretreatment increased Type 1 procollagen gene expression levels by activating TGF-ß/Smad signaling pathway. Taken together, our results demonstrate that 1-kestose may serve as a potent natural trisaccharide for inflammation and photoaging prevention.

In Vitro Profiling of Toxicity Effects of Different Environmental Factors on Skin Cells.

The skin is constantly exposed to a variety of environmental threats. Therefore, the influence of environmental factors on skin damage has always been a matter of concern. This study aimed to investigate the cytotoxic effects of different environmental factors, including cooking oil fumes (COFs), haze (PM2.5), and cigarette smoke (CS), on epidermal HaCaT cells and dermal fibroblast (FB) cells. Cell viability, intracellular reactive oxygen species (ROS) generation, inflammatory cytokine levels, and collagen mRNA expression were used as toxicity endpoints. Additionally, the effects of ozone (O3) on cell viability and release of inflammatory cytokines in 3D epidermal cells were also examined. The results showed that the organic extracts of CS, COFs, and PM2.5 significantly inhibited the viability of HaCaT and FB cells at higher exposure concentrations. These extracts also increased intracellular ROS levels in FB cells. Furthermore, they significantly promoted the release of inflammatory cytokines, such as IL-1α and TNF-α, in HaCaT cells and down-regulated the mRNA expression of collagen I, III, IV, and VII in FB cells. Comparatively, SC organic extracts exhibited stronger cytotoxicity to skin cells compared to PM2.5 and COFs. Additionally, O3 at all test concentrations significantly inhibited the viability of 3D epidermal cells in a concentration-dependent manner and markedly increased the levels of TNF-α and IL-1α in 3D epidermal cells. These findings emphasize the potential cytotoxicity of COFs, PM2.5, CS, and O3 to skin cells, which may lead to skin damage; therefore, we should pay attention to these environmental factors and take appropriate measures to protect the skin from their harmful effects.

Induction of ferroptosis in human keratinocyte HaCaT cells by squalene hydroperoxide: Possible prevention of skin ferroptosis by botanical extracts.

Ever since the proposal of ferroptosis, it has been studied as a nonapoptotic cell death caused by iron ion-dependent phospholipid (PL) peroxidation. We previously showed that treatment of human hepatoma cell line HepG2 with prepared PL hydroperoxide (PLOOH) resulted in ferroptosis. However, in human sebum, the major hydroperoxide is not PLOOH but squalene hydroperoxide (SQOOH), and to our knowledge, it is not established yet whether SQOOH induces ferroptosis in the skin. In this study, we synthesized SQOOH and treated human keratinocyte HaCaT cells with SQOOH. The results showed that SQOOH induces ferroptosis in HaCaT cells in the same way that PLOOH causes ferroptosis in HepG2 cells. Some natural antioxidants (botanical extracts) could inhibit the ferroptosis in both the cell types. Consequently, future research focus would revolve around the involvement of SQOOH-induced ferroptosis in skin pathologies as well as the prevention and treatment of skin diseases through inhibition of ferroptosis by botanical extracts.

Collagen I protects human keratinocytes HaCaT against UVB injury via restoring PINK1/parkin-mediated mitophagy.

Collagen I is a major component of extracellular matrix in human skin, and is also widely used in a variety of skin-care products. In this study, we investigated the modulatory roles of collagen I on human immortalized keratinocytes HaCaT, especially when cells were irradiated with UVB. Interestingly, the cells grown on plates coated by molecular collagen I, but not fibrillar collagen I, acquired certain resistance against UVB damages, as shown by increased survival and reduced apoptosis. The accumulation of dysfunctional mitochondria in UVB-treated cells was attenuated by molecular collagen I-coating. Interestingly, molecular collagen I rescued the loss of mitochondrial biogenesis in cells treated with UVB. Loss of PINK1/parkin-mediated mitophagy was dominant for the accumulation of dysfunctional mitochondria after UVB irradiation. Of note, cells cultured on molecular collagen I-precoated plates exhibited reserved mitophagy after UVB irradiation, as reflected by the enhanced protein level of PINK1/parkin, increased mitochondrial ubiquitin and the co-localization of lysosomes and mitochondria. Moreover, in UVB-treated cells, inhibiting mitophagy by Cyclosporin A, or by silencing PINK1 or parkin, disturbed the resolution of mitochondrial stress and reduced the protective effect of molecular collagen I, indicating that mitophagy is pivotal for the protection of collagen I against UVB damage in keratinocytes HaCaT. Collectively, this study reveals an unexpected protective role of collagen I, which facilitates mitophagy to rescue cells under UVB irradiation, providing a new direction for clinical application of collagen products.

Deoxynivalenol exposure induces oxidative stress and apoptosis in human keratinocytes via PI3K/Akt and MAPK signaling pathway.

Deoxynivalenol (DON) is a mycotoxin frequently occurring in human and animal food worldwide, which raises increasing public health concerns. In the present study, we used human keratinocytes (HaCaT cells) as an in vitro model to explore the cytotoxic effect of DON. The results showed that the cells exhibited varying degrees of damage, including decreased cell number and viability, cell shrinkage and floating, when treated with 0.125, 0.25, and 0.5 µg/mL DON for 6, 12, and 24 h, respectively. Furthermore, exposure to DON for 24 h significantly increased the lactate dehydrogenase (LDH) release and intracellular reactive oxygen species (ROS), and prominently decreased the superoxide dismutase (SOD) and catalase (CAT) activity. Additionally, DON exposure induced mitochondrial damage and cell apoptosis through reducing mitochondrial membrane potential. Then, we performed RNA-sequencing to investigate the molecular changes in HaCaT cells after DON exposure. The RNA-sequencing results revealed that DON exposure altered the gene expression involved in apoptosis, MAPK signaling pathway, and PI3K/Akt signaling pathway. Moreover, DON exposure significantly decreased the mRNA and protein expression of Bcl-2, and increased the mRNA and protein expression of Bax, Caspase 3 and COX-2, the protein expression of PI3K, and the phosphorylation levels of Akt, ERK, p38, and JNK. Taken together, these findings suggest that DON exposure could induce cell damage, oxidative stress, and apoptosis in HaCaT cells through the activation of PI3K/Akt and MAPK pathways.

Chlorogenic acid attenuates deoxynivalenol-induced apoptosis and pyroptosis in human keratinocytes via activating Nrf2/HO-1 and inhibiting MAPK/NF-κB/NLRP3 pathways.

Deoxynivalenol (DON) is a common mycotoxic contaminant, frequently found in food and feed, causing a severe threat to human and animal health. Because of the widespread contamination of DON, humans involved in agricultural practices may be directly exposed to DON through the skin route. Chlorogenic acid (CGA) is a phenolic acid, which has anti-inflammatory and antioxidant properties. However, it is still unclear whether CGA can protect against DON-induced skin damage. Here, the effect of CGA on mitigating damage to human keratinocytes (HaCaT) triggered by DON, as well as its underlying mechanisms were investigated. Results demonstrated that DON exposure significantly decreased cell viability, and induced excessive mitochondrial reactive oxygen species (mtROS) generation, mitochondrial damage, oxidative stress, cell apoptosis and pyroptosis. However, CGA pretreatment for 2 h significantly increased cell viability and reversed DON-induced oxidative stress by improving antioxidant enzyme activities such as superoxide dismutase (SOD), glutathione (GSH), catalase (CAT), reducing mtROS generation and enhancing mitochondrial function through activating Nrf2/HO-1 pathway. Moreover, CGA significantly increased the Bcl-2 protein expression, decreased the protein expressions of Bax and cleaved Caspase-3, and suppressed the phosphorylated of ERK, JNK, NF-κB. Further experiments revealed that CGA could also inhibit the pyroptosis-related protein expressions including NLRP3, cleaved Caspase-1, GSDMD-N, cleaved IL-1ß and IL-18. In conclusion, our results suggest that CGA could attenuate DON-induced oxidative stress, inflammation, and apoptosis by activating the Nrf2/HO-1 pathway and inhibiting MAPK/NF-κB/NLRP3 pathway. CGA might be a novel promising therapeutic agent for alleviating the dermal damage triggered by DON.

Arzanol, a natural phloroglucinol α-pyrone, protects HaCaT keratinocytes against H2O2-induced oxidative stress, counteracting cytotoxicity, reactive oxygen species generation, apoptosis, and mitochondrial depolarization.

Skin oxidative stress results in structural damage, leading to premature senescence, and pathological conditions such as inflammation and cancer. The plant-derived prenylated pyrone-phloroglucinol heterodimer arzanol, isolated from Helichrysum italicum ssp. microphyllum (Willd.) Nyman aerial parts, exhibits anti-inflammatory, anticancer, antimicrobial, and antioxidant activities. This study explored the arzanol protection against hydrogen peroxide (H2O2) induced oxidative damage in HaCaT human keratinocytes in terms of its ability to counteract cytotoxicity, reactive oxygen species (ROS) generation, apoptosis, and mitochondrial membrane depolarization. Arzanol safety on HaCaT cells was preliminarily examined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and microscopic observation. The arzanol pre-incubation (5-100 µM, for 24 h) did not induce cytotoxicity and morphological alterations. The phloroglucinol, at 50 µM, significantly protected keratinocytes against cytotoxicity induced by 2 h-incubation with 2.5 and 5 mM H2O2, decreased cell ROS production induced by 1 h-exposure to all tested H2O2 concentrations (0.5-5 mM), as determined by the 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) assay, and lipid peroxidation (thiobarbituric acid reactive substances [TBARS] method). The 2-h incubation of keratinocytes with H2O2 determined a significant increase of apoptotic cells versus control cells, evaluated by NucView® 488 assay, from the dose of 2.5 mM. Moreover, an evident mitochondrial membrane potential depolarization, monitored by fluorescent mitochondrial dye MitoView™ 633, was assessed at 5 mM H2O2. Arzanol pre-treatment (50 µM) exerted a strong significant protective effect against apoptosis, preserving the mitochondrial membrane potential of HaCaT cells at the highest H2O2 concentrations. Our results validate arzanol as an antioxidant agent for the prevention/treatment of skin oxidative-related disorders, qualifying its potential use for cosmeceutical and pharmaceutical applications.

Identification of potential biomarkers and infiltrating immune cells from scalp psoriasis.

BACKGROUND: Scalp psoriasis seriously affects the appearance and psychological status of patients. The aim of this study was to investigate the effect and potential mechanism of RPL9 and TIFA in scalp psoriasis, so as to provide a precise and effective way for the clinical treatment of scalp psoriasis. METHODS: The Gene Expression Omnibus (GEO) database was employed to download the GSE75343 dataset to search for differentially expressed genes (DEGs) in scalp psoriasis through Sangerbox. Gene set enrichment analysis (GSEA) and gene set variation analysis (GSVA) enrichment analysis, functional enrichment analysis, immune cell infiltration analysis, immune responses and correlation analysis with 12 hub genes were performed. Then, STRING was used to develop a protein-protein interaction (PPI) network, used Cytoscape to locate hub genes, and SVM-RFE and random forest were utilized to identified RPL9 as the targeted gene. TIFA-RPL9 interaction predictions were made viathe Open Targets Platform and Uniprot. Further, the RPL9 and TIFA expression, molecular mechanism, and function were assessed in scalp psoriasis. RESULTS: Immunohistochemistry, qPCR, and western blotting verified that RPL9 and TIFA were highly expressed in lesional tissues of scalp psoriasis and IL17A-stimulated HaCaT cells. RPL9 knockdown effectively suppressed the proliferative capacity of IL17A-stimulated HaCaT cells in the CCK8 assay. The co-immunoprecipitation results revealed that RPL9 could interact with TIFA in IL17A-stimulated HaCaT cells. In qPCR and western blotting, RPL9 knockdown significantly inhibited TIFA at the mRNA and protein levels in IL17A-stimulated HaCaT cells. In ELISA, the secretion of TNF-α was markedly inhibited after downregulating RPL9 in IL17A-stimulated HaCaT cells. CONCLUSION: To our knowledge, we have elucidated the expression and role of RPL9 and TIFA in scalp psoriatic skin and keratinocytes, and our findings confirm that RPL9 might act as a candidate therapeutic target for scalp psoriasis.

Phytochemical profile and biological activities of Bromelia antiacantha extracts / Perfil fitoquímico e atividades biológicas de extratos de Bromelia antiacantha

Reports from popular medicine usually act as a basis for the development of new drugs from natural compounds with therapeutic actions for serious diseases and prevalence such as cancer. Bromelia antiacantha Bertol. is a species of the Bromeliaceae family, considered an unconventional food plant, found in the south and midwest regions of Brazil. Despite the high nutritional content and pharmacological potential of its fruits, few scientific studies report its biological actions. Thus, this study evaluates the phytochemical profile of aqueous and ethanol extracts obtained from B. antiacantha fruits, as well as their possible antioxidant, antitumor, and cytotoxic activities. The aqueous extract exhibited phenolic compounds and flavonoids, while ethanol extracts indicated the presence of flavonoids and coumarin in their composition, regardless of the region of collection. The ethanolic extract demonstrated a more promising antioxidant effect than the aqueous extract and also induced a significant inhibition in the viability of human cervical cancer cells of the SiHa strain. In addition, treatment with both extracts did not alter the viability of non-tumor cells of the immortalized human keratinocyte lineage (HaCaT). These results bring new data about extracts obtained from a native plant, edible and traditionally used in popular medicine, opening new perspectives for its possible therapeutic application.

Relatos da medicina popular costumam atuar como referencial para o desenvolvimento de novos fármacos a partir de moléculas naturais com ações terapêuticas para doenças de alta gravidade e prevalência como o câncer. Bromelia antiacantha Bertol. é uma espécie da família Bromeliaceae, considerada uma planta alimentícia não convencional (PANC), encontrada nas regiões sul e centro-oeste do Brasil. Apesar do alto teor nutritivo e potencial farmacológico de seus frutos, poucos estudos científicos relatam suas ações biológicas. Desta forma, este estudo avalia o perfil fitoquímico de extratos aquoso e etanólico obtidos de frutos de B. antiacantha, bem como a sua possível ação antioxidante, antitumoral e citotóxica. O extrato aquoso apresentou compostos fenólicos e flavonoides, enquanto os extratos etanólicos apontam a presença de flavonóides e cumarina em sua composição, independente da região de coleta. O extrato etanólico demonstrou efeito antioxidante mais promissor do que o extrato aquoso e também induziu uma inibição significativa na viabilidade de células humanas de câncer cervical da linhagem SiHa. Além disso, o tratamento com ambos extratos não alterou a viabilidade de células não tumorais da linhagem de queratinócitos humanos imortalizados (HaCaT). Estes dados trazem novas informações sobre extratos obtidos de uma espécie vegetal nativa, comestível e já utilizada tradicionalmente, mas abrindo novas perspectivas quanto a possíveis aplicações terapêuticas.

Phytochemical profile and biological activities of Bromelia antiacantha extracts

Abstract Reports from popular medicine usually act as a basis for the development of new drugs from natural compounds with therapeutic actions for serious diseases and prevalence such as cancer. Bromelia antiacantha Bertol. is a species of the Bromeliaceae family, considered an unconventional food plant, found in the south and midwest regions of Brazil. Despite the high nutritional content and pharmacological potential of its fruits, few scientific studies report its biological actions. Thus, this study evaluates the phytochemical profile of aqueous and ethanol extracts obtained from B. antiacantha fruits, as well as their possible antioxidant, antitumor, and cytotoxic activities. The aqueous extract exhibited phenolic compounds and flavonoids, while ethanol extracts indicated the presence of flavonoids and coumarin in their composition, regardless of the region of collection. The ethanolic extract demonstrated a more promising antioxidant effect than the aqueous extract and also induced a significant inhibition in the viability of human cervical cancer cells of the SiHa strain. In addition, treatment with both extracts did not alter the viability of non-tumor cells of the immortalized human keratinocyte lineage (HaCaT). These results bring new data about extracts obtained from a native plant, edible and traditionally used in popular medicine, opening new perspectives for its possible therapeutic application.

Resumo Relatos da medicina popular costumam atuar como referencial para o desenvolvimento de novos fármacos a partir de moléculas naturais com ações terapêuticas para doenças de alta gravidade e prevalência como o câncer. Bromelia antiacantha Bertol. é uma espécie da família Bromeliaceae, considerada uma planta alimentícia não convencional (PANC), encontrada nas regiões sul e centro-oeste do Brasil. Apesar do alto teor nutritivo e potencial farmacológico de seus frutos, poucos estudos científicos relatam suas ações biológicas. Desta forma, este estudo avalia o perfil fitoquímico de extratos aquoso e etanólico obtidos de frutos de B. antiacantha, bem como a sua possível ação antioxidante, antitumoral e citotóxica. O extrato aquoso apresentou compostos fenólicos e flavonoides, enquanto os extratos etanólicos apontam a presença de flavonóides e cumarina em sua composição, independente da região de coleta. O extrato etanólico demonstrou efeito antioxidante mais promissor do que o extrato aquoso e também induziu uma inibição significativa na viabilidade de células humanas de câncer cervical da linhagem SiHa. Além disso, o tratamento com ambos extratos não alterou a viabilidade de células não tumorais da linhagem de queratinócitos humanos imortalizados (HaCaT). Estes dados trazem novas informações sobre extratos obtidos de uma espécie vegetal nativa, comestível e já utilizada tradicionalmente, mas abrindo novas perspectivas quanto a possíveis aplicações terapêuticas.

Phytochemical and Nutraceutical Screening of Ethanol and Ethyl Acetate Phases of Romanian Galium verum Herba (Rubiaceae).

Galium species are used worldwide for their antioxidant, antibacterial, antifungal, and antiparasitic properties. Although this plant has demonstrated its antitumor properties on various types of cancer, its biological activity on cutaneous melanoma has not been established so far. Therefore, the present study was designed to investigate the phytochemical profile of two extracts of G. verum L. herba (ethanolic and ethyl acetate) as well as the biological profile (antioxidant, antimicrobial, and antitumor effects) on human skin cancer. The extracts showed similar FT-IR phenolic profiles (high chlorogenic acid, isoquercitrin, quercitrin, and rutin), with high antioxidant capacity (EC50 of ethyl acetate phase (0.074 ± 0.01 mg/mL) > ethanol phase (0.136 ± 0.03 mg/mL)). Both extracts showed antimicrobial activity, especially against Gram-positive Streptococcus pyogenes and Staphylococcus aureus bacilli strains, the ethyl acetate phase being more active. Regarding the in vitro antitumor test, the results revealed a dose-dependent cytotoxic effect against A375 melanoma cell lines, more pronounced in the case of the ethyl acetate phase. In addition, the ethyl acetate phase stimulated the proliferation of human keratinocytes (HaCaT), while this effect was not evident in the case of the ethanolic phase at 24 h post-stimulation. Consequently, G. verum l. could be considered a promising phytocompound for the antitumor approach of cutaneous melanoma.

KRT17 from skin cells with high glucose stimulation promotes keratinocytes proliferation and migration.

Impaired diabetic wound healing is an important issue in diabetic complications. Proliferation and migration of keratinocytes are major processes of skin wound repair after injury. However, hyperkeratosis can affect the speed of wound healing. Based on the results of preliminary experiments on increased KRT17 expression after high glucose stimulation of human skin tissue cells, a cell model of human immortalized keratinocyte (HaCaT) stimulation with different concentrations of KRT17 was established in vitro, and the promotion in cell proliferation and migration were discovered. KRT17 silencing promoted diabetic wound healing in the db/db diabetic wound model. Transcriptome sequencing (RNA-seq) was performed on HaCaT cells after KRT17 stimulation, and analysis showed significant enrichment in the PI3K-AKT signaling pathway, in which the regulation of cell c-MYB mRNA, a key molecule regulating cell proliferation and migration, was significantly upregulated. In vitro assays showed increased c-MYB expression and enhanced pAKT activity after HaCaT cell stimulation by KRT17. We speculate that KRT17 is upregulated under high glucose and promotes keratinocyte proliferation and migration caused hyperkeratosis, through the c-MYB/PI3K-AKT pathway, contributing to delayed wound healing.