HaCaT Research Articles

Betulin-NLC-hydrogel for the Treatment of Psoriasis-like Skin Inflammation: Optimization, Characterisation, and In vitro and In vivo Evaluation.

Psoriasis is a chronic inflammatory skin disorder that poses significant challenges regarding effective and targeted drug delivery. Bioactive substances like betulin have shown tremendous utility in treating these conditions; however, they pose limited utility owing to their physicochemical characteristics. Here, we aimed to develop a novel topical dosage form for treating psoriasis, utilising betulin-loaded solid lipid nanoparticles (NLCs) incorporated into a hydrogel matrix. The optimization of the formulation was meticulously conducted using a design of experiments methodology, and its diverse physicochemical attributes were thoroughly examined. Evaluating betulin's in vitro release pattern from the NLC-hydrogel demonstrated consistent and regulated drug release properties. Additionally, the formulation demonstrated improved skin penetration abilities as determined by in vitro skin permeation experiments employing Franz diffusion cells- furthermore, the therapeutic effectiveness of the betulin-NLC-hydrogel was assessed by an in vivo experiment carried out using an imiquimod-induced psoriasis-like skin inflammation model in BALB/c female mice. The NLCs exhibited a pH of 5.67±0.86, particle size of 148.16±12.66 nm, and zeta potential of -22.84±2.37 mV, ensuring stability and suitability for topical use. The gel, with a pH of 6.05±0.43 and viscosity of 17550±120 cPs, showed enhanced skin hydration and lipid restoration. Drug release studies indicated a slower release from NLCs and gel, improving skin retention. Stability tests revealed that the formulations were stable at room temperature but not at elevated temperatures. The in vitro safety profile of the formulation revealed no significant adverse effects on HaCaT cell lines. The NLC gel demonstrated significant anti-psoriatic activity, reducing inflammation and cytokine levels. The betulin-NLC-hydrogel formulation exhibited promising characteristics for the topical treatment of psoriasis, showcasing optimised drug delivery, sustained release, and notable therapeutic efficacy. The findings from this study provide a foundation for the potential clinical translation of this innovative topical dosage form for improved psoriasis management.

FUNDC1-mediated mitophagy regulates photodamage independently of the PINK1/Parkin-dependent pathway

BackgroundUltraviolet B(UVB) triggers a pro-survival response through mitophagy, but the role of FUNDC1-mediated mitophagy in photodamaged skin remains unexplored. ObjectivesTo clarify the function of mitophagy in UVB-induced photodamaged skin. MethodsTo investigate the role of FUNDC1-mediated mitophagy in UVB-induced mitochondrial damage and cell apoptosis, FUNDC1 knockdown in C57BL/6 mice was performed using adeno-associated virus. Additionally, FUNDC1 overexpression and knockdown in HaCaT cells were conducted using lentivirus. A comprehensive analysis was conducted on a panel of human sun-exposed skin samples, alongside control samples, to assess the expression levels of FUNDC1. ResultsIn UVB-induced C57BL/6 mice, the dorsal skin showed photodamage including erythema, scaling, erosion, and scabs. The expression levels of PINK1, Parkin, and BNIP3 did not show significant changes, while FUNDC1 expression consistently declined along with LC3B. Cytochrome C, Bax, and cleaved-caspase3 were upregulated, while Bcl2 was downregulated. UVB-induced HaCaT cells showed mitochondrial damage, accompanied by FUNDC1 downregulation and BNIP3 upregulation, while PINK1 and Parkin showed no significant changes. FUNDC1 overexpression led to an increase in mtROS and a decrease in mitochondrial membrane potential and ATP levels, indicating complete mitochondrial clearance and exacerbated cell death. FUNDC1 knockdown protected against UVB-induced photodamage in mice and mitigated mitochondrial damage and apoptosis in HaCaT cells by activating compensatory PINK1/Parkin-dependent mitophagy, which was evidenced by upregulation of PINK1 and Bcl2 and downregulation of Bax. In human sun-exposed skin samples, there was a decrease in the number of FUNDC1+ cells compared with non-sun-exposed controls. ConclusionsFUNDC1-mediated mitophagy regulates skin photodamage and provides a novel mechanism for resisting photodamage, presenting a potential target for future therapeutic interventions.

CAPE derivatives: Multifaceted agents for chronic wound healing.

Chronic wounds significantly impact the patients' quality of life, creating an urgent interdisciplinary clinical challenge. The development of novel agents capable of accelerating the healing process is essential. Caffeic acid phenethyl ester (CAPE) has demonstrated positive effects on skin regeneration. However, its susceptibility to degradation limits its pharmaceutical application. Chemical modification of the structure improves the pharmacokinetics of this bioactive phenol. Hence, two novel series of CAPE hybrids were designed, synthesized, and investigated as potential skin regenerative agents. To enhance the stability and therapeutic efficacy, a caffeic acid frame was combined with quinolines or isoquinolines by an ester (1a-f) or an amide linkage (2a-f). The effects on cell viability of human gingival fibroblasts (HGFs) and HaCaT cells were evaluated at different concentrations; they are not cytotoxic, and some proved to stimulate cell proliferation. The most promising compounds underwent a wound-healing assay in HGFs and HaCaT at the lowest concentrations. Antimicrobial antioxidant properties were also explored. The chemical and thermal stabilities of the best compounds were assessed. In silico predictions were employed to anticipate skin penetration capabilities. Our findings highlight the therapeutic potential of caffeic acid phenethyl ester (CAPE) derivatives 1a and 1d as skin regenerative agents, being able to stimulate cell proliferation, control bacterial growth, regulate ROS levels, and being thermally and chemically stable. An interesting structure-activity relationship was discussed to suggest a promising multitargeted approach for enhanced wound healing.

Human IL-22 receptor-targeted small protein antagonist suppress murine DSS-induced colitis

BackgroundHuman interleukin-22 (IL-22) is known as a “dual function” cytokine that acts as a master regulator to maintain homeostasis, structural integrity of the intestinal epithelial barrier, and shielding against bacterial pathogens. On the other hand, the overexpression of IL-22 is associated with hyper-proliferation and recruitment of pathologic effector cells, leading to tissue damage and chronic inflammation in specific diseases including inflammatory bowel disease (IBD). To study a role of IL-22-mediated signaling axis during intestinal inflammation, we generated a set of small protein blockers of IL-22R1 and verified their inhibitory potential on murine model of colitis.MethodsWe used directed evolution of proteins to identify binders of human IL-22 receptor alpha (IL-22R1), designated as ABR ligands. This approach combines the assembly of a highly complex combinatorial protein library derived from small albumin-binding domain scaffold and selection of promising protein variants using ribosome display followed by large-scale ELISA screening. The binding affinity and specificity of ABR variants were analyzed on transfected HEK293T cells by flow cytometry and LigandTracer. Inhibitory function was further verified by competition ELISA, HEK-Blue IL-22 reporter cells, and murine dextran sulfate sodium (DSS)-induced colitis.ResultsWe demonstrate that ABR specifically recognizes transgenic IL-22R1 expressed on HEK293T cells and IL-22R1 on TNFα/IFNγ-activated HaCaT cells. Moreover, some ABR binders compete with the IL-22 cytokine and function as IL-22R1 antagonists in HEK-Blue IL22 reporter cells. In a murine model of DSS-induced acute intestinal inflammation, daily intraperitoneal administration of the best IL-22R1 antagonist, ABR167, suppressed the development of clinical and histological markers of colitis including prevention of mucosal inflammation and architecture deterioration. In addition, ABR167 reduces the DSS-induced increase in mRNA transcript levels of inflammatory cytokines such as IL-1β, IL-6, IL-10, and IL-17A.ConclusionsWe developed small anti-human IL-22R1 blockers with antagonistic properties that ascertain a substantial role of IL-22-mediated signaling in the development of intestinal inflammation. The developed ABR blockers can be useful as a molecular clue for further IBD drug development.

Functional Properties and Components of Koenigia alpina Extract.

Koenigia alpina (All.) T.M.Schust. & Reveal (alpine knotweed) is a perennial herb belonging to the Polygonaceae family. Several studies have examined Polygonaceae species' potential applications as cosmeceutical materials; however, the potential of K. alpina as a cosmeceutical has not yet been studied. Hydrogen peroxide (H2O2) and lipopolysaccharide were used to induce an inflammatory response in RAW 264.7 cells. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) radicals and H2O2 were used to evaluate the free-radical scavenging activity of K. alpina extract and its protective effect against reactive oxygen species (ROS)-induced cell damage. The whitening, antiaging, and cell proliferation/migration effects of the extracts were evaluated via tyrosinase inhibition, collagenase/elastase inhibition, and wound healing assays, respectively. The anti-inflammatory effect was confirmed by evaluating nitric oxide (NO) production in RAW 264.7 cells. High-performance liquid chromatography (HPLC), UV, and MS/MS were used to determine the main components of the extract and fractions. The ethyl acetate (EA) fraction and its aglycone fraction showed very high free-radical scavenging activities (47.5 and 47.1µg/mL, respectively). The extract/fractions also showed significant tyrosinase inhibition (IC50 = 0.38mg/mL in EA fraction), collagenase inhibition (IC50 = 0.21mg/mL in EA fraction), and elastase inhibition (IC50 = 0.57mg/mL in aglycone fraction). NO production in lipopolysaccharide-induced RAW 264.7 cells was inhibited by the extract/fractions. The extract also promoted the closure of scratch wounds in HaCaT cells. The K. alpina extract/fractions contained cardamonin, quercetin, and quercitrin. K. alpina extracts/fractions showed antioxidant, antiaging, whitening, and anti-inflammatory activities, suggesting they may have potential as antiaging cosmeceuticals.

New quinoxaline-oxadiazole hybrids as tubulin inhibitors: Synthesis, cytotoxicity, and molecular dynamics simulations

This study describes the synthesis and biological evaluation of a series of novel quinoxaline-oxadiazole hybrid compounds. These compounds were designed to enhance anticancer activity through the incorporation of specific pharmacophoric groups. Structural characterization using NMR and HRMS spectroscopy confirmed the successful synthesis of the hybrids. Cytotoxicity assays revealed that several compounds exhibited significant activity against various cancer cell lines. Compounds 8a and 8f demonstrated potent activity against A549 cells, while 8a, 8b, and 8c were effective against HaCaT cells. Additionally, compounds 8i and 8a showed activity against HepG2 cells. Molecular docking and dynamics simulations suggested that compounds 8c and 8d bind strongly to the tubulin protein, a target for anticancer drugs. These compounds exhibited binding affinities comparable to known anticancer compounds. The results of this study highlight the potential of quinoxaline-oxadiazole hybrids, especially those with halogen substitutions, as promising candidates for the development of novel anticancer therapeutics.

Antibacterial and Antibiofilm potential of Thuja orientalis L. extract targeting cariogenic Enterococcus faecalis ATCC 29212: A combined in-vitro, in-silico study, and cytotoxicity screening

ObjectivesIn this study, we explored the efficacy of methanolic extract of Thuja orientalis (TOME) as a novel antibacterial and antibiofilm agent against a cariogenic bacterium, Enterococcus faecalis ATCC 29212. DesignAntibacterial susceptibility studies were conducted and surface morphology analysis was performed using field emission scanning electron microscopy (FESEM). Antibiofilm activity was evaluated through both qualitative and quantitative biofilm inhibition assays and validated by microscopic analysis. In-silico molecular docking studies were conducted using the EDock server. The effectiveness of TOME was substantiated by biofilm model on dentin discs and cytotoxicity towards the HaCaT cell line was assessed using the MTT assay. ResultsTOME exhibited significant bactericidal activity with minimum inhibitory concentration of 12.5mg/mL and additionally, it effectively compromised bacterial cell wall integrity. Qualitative, quantitative and microscopic studies depicted the inhibition of biofilm formation. TOME significantly impacted the production of extracellular polymeric substance and extracellular DNA. Molecular docking studies identified beta-caryophyllene as a potent inhibitor of the Enterococcal surface protein (Esp). Biofilm model depicted the reduction of bacterial load on dentin discs. Additionally, TOME showed reduced cytotoxicity on HaCaT cells, indicating its potential as a safe therapeutic agent. ConclusionThese findings highlight TOME's promise for developing novel treatments for dental infections and biofilm-associated diseases. Further research should focus on isolating and characterizing the active compounds within TOME, particularly beta-caryophyllene, to elucidate their precise mechanisms of action.

CH25H Promotes Autophagy and Regulates the Malignant Progression of Laryngeal Squamous Cell Carcinoma Through the PI3K-AKT Pathway.

Laryngeal squamous cell carcinoma (LSCC) is a type of cancer of the respiratory tract that often presents with subtle symptoms at the early stage and is susceptible to recurrence and metastasis. To find out key regulatory genes involved in LSCC development, we downloaded LSCC-related sequencing datasets for bioinformatics analysis. WGCNA was performed on GSE142083 and differential analysis was conducted on GSE51985 and TCGA-HNSC. Intersectiongenes were taken from the above three datasets. To confirm the function of genes, we overexpressed and knocked down genes in cells and treated them with autophagy agonist Rapamycin and PI3K-AKT pathway inhibitor. At the cellular level, the expression of CH25H, autophagy-related proteins (LC3 I, LC3 II, p62, and Beclin 1), and PI3K-AKT pathway-related proteins (PI3K, AKT, and p-AKT) were assessed via Western blot; the mRNA level of CH25H was evaluated through qRT-PCR; the cell activity was examined by CCK8; the apoptosis was assessed through flow cytometry; and the cell migration and invasion were assessed through wound healing and Transwell assays. Through bioinformatics analysis, we screened 7 genes (CH25H, NELL2, STC2, TMEM158, ZIC2, HOXD11, and HOXD10). Ultimately, CH25H was selected for follow-up experiments. By detecting CH25H expression in human immortalized keratinocytes (HaCaT) and LSCC cells (Tu-686, SNU899, and AMC-HN-8), it was found out that CH25H expression was higher in HaCaT cells than in LSCC cells. To elucidate the role of CH25H in LSCC development, we overexpressed CH25H in Tu-686 cells and downregulated its expression in AMC-HN-8 cells. CH25H was revealed to reduce the proliferation, activity, invasion, and migration of LSCC cells while increasing their apoptosis levels. Significant changes were also observed in the expressions of autophagy- and PI3K-AKT pathway-related proteins. To further investigate the roles of autophagy and the PI3K-AKT pathway in LSCC development, we respectively employed autophagy agonists and inhibitors targeting the PI3K-AKT pathway to intervene the cells, and found that CH25H regulated the PI3K-AKT pathway to promote autophagy, thus enhancing the apoptosis of LSCC cells. We further investigated CH25H's impact on tumor growth, autophagy, and the PI3K-AKT pathway at the animal level and found that CH25H promoted autophagy of LSCC cells and inhibited the PI3K-AKT pathway, and ultimately inhibiting the progression of LSCC. In summary, CH25H promotes autophagy and affects the malignant progression of LSCC through the PI3K-AKT pathway.

P2X7R-primed keratinocytes are susceptible to apoptosis via GPCR-Gβγ-pERK signal pathways

BackgroundCell death constitutes a pivotal biological phenomenon essential for the preservation of homeostasis within living organisms. In the context of maintaining a functional skin barrier, keratinocytes exert positively and negatively control cell death signals. However, in patients with severe drug eruptions, anomalous overexpression of the formyl peptide receptor 1 (FPR1) in keratinocytes elicits a distinctive mode of cell death known as necroptosis, thereby suffering a loss of the skin barrier. The precise molecular mechanisms connecting FPR1 activation to this cell death remain unclear. ObjectiveWe have investigated the intracellular signal transduction cascade governing FPR1-mediated cell death in cultured keratinocytes. MethodsWe used HaCaT cells as a model keratinocyte. The expression of FPR1 was detected with qPCR. The presence of cell death events was monitored through live-cell fluorescent staining and LDH release assays. Furthermore, the phosphorylation of ERK was assessed via Western blot analysis. Intracellular signal pathways were investigated using specific inhibitors. ResultsLigand stimulation of an endogenous ion channel, purinergic receptor P2X7 (P2X7R), increased the FPR1 expression level. This upregulated FPR1 demonstrated functional competence in the phosphorylation of downstream MAP kinase and the initiation of cell death. Notably, this cell death was ameliorated upon the administration of inhibitors targeting Gβγ, ERK, and caspases. ConclusionThe induction and stimulation of FPR1 initiated apoptosis in keratinocytes via the Gβγ-pERK signaling pathway. Our findings postulate that the downstream components of FPR1 represent an alternative therapeutic target for preventing unintended keratinocyte cell death.

Photoprotective properties of four structure propolis from Heterotrigona itama stingless beehive: Fractionation, bioactivity analysis, and chemical profiling

Heterotrigona itama stingless bee propolis extract is known for its diverse bioactive compounds, making it a potential natural shield against UV radiation. This research assesses the photoprotective potential of crude ethanol extract from H. itama propolis collected from four structures (involucrum, pillar, pot, and entrance) of five bee hives (H1-H5), totalling 20 samples. Initially, the samples underwent testing for SPF value and UV absorption spectra. The crude ethanol extract (E) from the involucrum (H4) with the highest SPF value and broadest spectrum was selected for fractionation using hexane and water. Subsequently, the extract (E) and its hexane (H) and water (W) fractions were subjected to SPF analysis, UVA/UVB absorption assessment, determination of total phenolic and flavonoid content, free radical scavenging capacity, anti-collagenase effects, and cytotoxicity assessment. Additionally, LC-MS/MS analysis was performed to identify chemical constituents in the active fraction (W). The extract E demonstrated an SPF of 8.23 ± 0.09 and UV absorption. Notably, its fraction W exhibited the highest SPF (16.55 ± 0.24) at 100 μg/mL, surpassing the H fraction (SPF 5.7 ± 0.45). Phenolic content was highest in the H fraction (388.95 ± 4.54 mg/g GAE DW), followed by the W fraction (286.76 ± 6.48 mg/g GAE DW) and crude E (91.83 ± 4.12 mg/g GAE DW) from the involucrum. Regarding flavonoids, the fraction W led with 79.82 ± 6.21 mg/g QE DW, followed by the H fraction (45.56 ± 0.05 mg/g QE DW) and E (34.57 ± 1.11 mg/g QE DW). The extract E also exhibited modest DPPH scavenging (EC50 = 120 μg/mL), while the H fraction demonstrated stronger activity (EC50 = 4.37 μg/mL), and the W fraction displayed moderate effects (EC50 = 17.55 μg/mL). Notably, the W fraction showed remarkable anti-collagenase activity, outperforming the positive control, EG. HaCaT cell cytotoxicity revealed that the extract E was cytotoxic, whereas the H and W fractions showed no toxicity. LC-MS/MS analysis identified bioactive flavonoids (e.g., pratensein, quercetin) in the W fraction. These findings highlight the superior photoprotective properties of the water fraction from the involucrum of H. itama stingless bee propolis extract, suggesting its potential as a natural and effective ingredient for sunscreen and skincare formulations.

Protective Effects of Sesame Glycoproteins on Ultraviolet-Induced Skin Aging: In Vitro and In Vivo Studies.

Ultraviolet (UV) radiation is a primary factor in skin photoaging, leading to wrinkles, reduced elasticity, and pigmentation changes due to damage to cellular DNA, proteins, and lipids. Glycoproteins from sesame cake (SPE) have potential protective effects against UV-induced skin aging. This study investigated the anti-photoaging effects of SPE on UV-induced damage in human keratinocyte HaCaT cells and SKH-1 hairless mice. SPE was evaluated for its ability to mitigate UV-induced damage in HaCaT cells by assessing MMP-1 protein and mRNA expression levels, as well as the activity of transcription factors AP-1 and NF-κB. The phosphorylation of AKT and MAPK pathways was also analyzed. In vivo, SKH-1 hairless mice were exposed to UV radiation, and the effects of SPE on wrinkle formation and skin structure were assessed by measuring wrinkle length, area, and volume. SPE significantly inhibited UV-induced MMP-1 protein and mRNA expression in HaCaT cells, indicating suppression of AP-1 and NF-κB transcription factors involved in MMP-1 production. Additionally, SPE reduced UV-induced phosphorylation of AKT and MAPK pathways. In SKH-1 hairless mice, SPE treatment led to significant reductions in wrinkle length, area, and volume, preserving skin structure in UV-exposed mice. The findings demonstrate that SPE has protective effects against UV-induced photoaging by inhibiting key molecular pathways associated with skin aging. SPE shows promise as a natural anti-photoaging agent, providing a foundation for future skincare product development. Further studies are warranted to explore the molecular mechanisms in detail and to validate these effects through clinical trials.

Trivalent arsenicals induce skin toxicity through thiol depletion

Arsenic, a widespread environmental contaminant, is highly toxic to human health. Arsenic exposure is associated with the occurrence of skin lesions and diseases. This study investigated the dermal toxicity of trivalent arsenicals (AsIII and MMAIII) and its underlying mechanism using human keratinocyte cell line and ex vivo porcine skin. AsIII and MMAIII induced concentration-dependent cell apoptosis and necrosis in HaCaT cells, which was confirmed in ex vivo porcine skin. AsIII and MMAIII increased reactive oxygen species generation and GSH depletion. Interestingly, radical scavenger antioxidants such as Vitamin C failed to mitigate arsenic-induced cytotoxicity, while thiol-containing compounds effectively alleviated it, suggesting a key role of thiol depletion in the trivalent arsenical-induced dermal toxicity. DMSA showed the strongest protective effects against AsIII and MMAIII-induced cytotoxicity in HaCaT cells. Of note, DMSA restored arsenical-induced tissue damage, and reduced the apoptosis in ex vivo porcine skin, highlighting its potential use to alleviate arsenic-induced skin lesions and diseases.

Role of FOXM1 and AURKB in regulating keratinocyte function in psoriasis.

This study investigated the effect of forkhead box M1 (FOXM1) and Aurora kinase B (AURKB) on the epidermal function of keratinocytes. Bioinformatics analysis was used to analyze the co-expression network of FOXM1 and its correlation with AURKB. The expression of FOXM1 and AURKB in tissues and cells was detected by immunofluorescence and real-time quantitative polymerase chain reaction, respectively. HaCaT cells were transfected with si-FOXM1 to knock down FOXM1. Cell proliferation was detected by cell counting kit-8 assay. Cell migration was detected by scratch assay. Cell invasion was detected by the Transwell invasion assay. Cell apoptosis and cell cycle were detected by flow cytometry. FOXM1 and AURKB were positively correlated and highly expressed in psoriatic lesions. After transfection of si-FOXM1, the expression levels of FOXM1 and AURKB genes significantly decreased. The proliferation of HaCaT cells decreased, the apoptosis rate increased significantly, and the proportion of cells in the G1 phase increased significantly, while the proportion of cells in the S phase decreased significantly. The scratch closure of HaCaT cells was reduced, and the number of cell invasions decreased significantly. FOXM1 and AURKB may affect the progression of psoriasis by regulating the proliferation, cell cycle, migration, and invasion of keratinocytes.

New Supramolecular Hydrogels Based on Diastereomeric Dehydrotripeptide Mixtures for Potential Drug Delivery Applications.

Self-assembly of peptide building blocks offers unique opportunities for bottom-up preparation of exquisite nanostructures, nanoarchitectures, and nanostructured bulk materials, namely hydrogels. In this work we describe the synthesis, characterization, gelation, and rheological properties of new dehydrotripeptides, Cbz-L-Lys(Cbz)-L,D-Asp-∆Phe-OH and (2-Naph)-L-Lys(2-Naph)-L,D-Asp-∆Phe-OH, containing a N-terminal lysine residue Nα,ε-bis-capped with carboxybenzyl (Cbz) and 2-Naphthylacetyl (2-Naph) aromatic moieties, an aspartic acid residue (Asp), and a C-terminal dehydrophenylalanine (∆Phe) residue. The dehydrotripeptides were obtained as diastereomeric mixtures (L,L,Z and L,D,Z), presumably via aspartimide chemistry. The dehydrotripeptides afforded hydrogels at exceedingly low concentrations (0.1 and 0.04 wt%). The hydrogels revealed exceptional elasticity (G' = 5.44 × 104 and 3.43 × 106 Pa) and self-healing properties. STEM studies showed that the diastereomers of the Cbz-capped peptide undergo co-assembly, generating a fibrillar 3D network, while the diastereomers of the 2-Naph-capped dehydropeptide seem to undergo self-sorting, originating a fibril network with embedded spheroidal nanostructures. The 2-Naph-capped hydrogel displayed full fast recovery following breakup by a mechanical stimulus. Spheroidal nanostructures are absent in the recovered hydrogel, as seen by STEM, suggesting that the mechanical stimulus triggers rearrangement of the spheroidal nanostructures into fibers. Overall, this study demonstrates that diastereomeric mixtures of peptides can be efficacious gelators. Importantly, these results suggest that the structure (size, aromaticity) of the capping group can have a directing effect on the self-assembly (co-assembly vs. self-sorting) of diastereomers. The cytotoxicity of the newly synthesized gelators was evaluated using human keratinocytes (HaCaT cell line). The results indicated that the two gelators exhibited some cytotoxicity, having a small impact on cell viability. In sustained release experiments, the influence of the charge on model drug compounds was assessed in relation to their release rate from the hydrogel matrix. The hydrogels demonstrated sustained release for methyl orange (anionic), while methylene blue (cationic) was retained within the network.

Keratinocyte derived extracellular vesicles mediated crosstalk between epidermis and dermis in UVB-induced skin inflammation.

Ultraviolet-B (UVB) light induces dermal inflammation, although it is mostly absorbed in the epidermis. Recent reports suggest extracellular vesicles (EVs) act as a mediator of photodamage signaling. Melatonin is reported to be a protective factor against UV-induced damage. We hypothesized that EVs derived from UVB-irradiated keratinocytes might trigger proinflammatory responses in dermal cells and tested whether melatonin can ameliorate UVB-induced inflammation. We used UVB-irradiated HaCaT cells, primary keratinocytes and STING knock-out mice to model production of EVs under photodamaging conditions and performed immunoblotting and ELISA to measure their effect on dermal macrophages. UVB-irradiated keratinocytes produce an increased number of EVs that contain higher concentrations of DNA and protein compared with controls. KC-derived EVs (KEVs) induced a STING- and inflammasome-mediated proinflammatory response in macrophages in vitro, and a pronounced inflammatory infiltrate in mouse dermis in vivo. Melatonin ameliorated KEVs inflammatory effect both in vitro and in vivo. This data suggests EVs are mediators in a crosstalk that takes place between keratinocytes and their neighboring cells as a result of photodamage. Further studies exploring EVs induced by damaging doses of UVB, and their impact on other cells will provide insight into photodamage and may help develop targeted therapeutic approaches.

Cold Plasma Ameliorates Imiquimod-Induced Psoriasis-Like Skin Inflammation in Mice.

Cold plasma has shown efficacy in various dermatological applications by reduces inflammatory responses and modulating cytokine expression. Therefore, this study aimed to investigate the therapeutic effects of cold plasma on psoriasis. In psoriasis HaCaT cells with cold plasma, we confirmed the expression of inflammatory cytokines involved in psoriasis formation and MAPK pathway, cell cycle, and apoptosis-related factors. In psoriasis-like BALB/c mice model, the effects of cold plasma treatment on skin were visually assessed. The expression of psoriasis-related factors was confirmed through qPCR, Western blotting, and Immunohistochemistry. Cold plasma led to a reduction in inflammatory cytokines including IL-17A, IL-23A, IL-24, IL-1β, and TNF-α in the psoriasis cell line. It also modulated factors involved in the MAPK pathway and the cell cycle. In the psoriasis-like mice model, cold plasma resulted in improvements in skin thickness, erythema, scaling, and PASI. Additionally, decreases in inflammatory cytokines like INF-γ, IL-23, and S100a7 were observed, along with improvements in MAPK pathway activation, apoptosis, and other psoriasis-related factors. Through in vitro and in vivo studies, our research highlights the potential of cold plasma as a novel therapeutic approach for psoriasis. Furthermore, cold plasma could serve as an adjunctive treatment for skin immunological diseases.

Vanillic acid promotes keratinocyte migration, proliferation, and angiogenesis

Wound healing is a complex, multi-phase process involving the coordinated interaction of various cells and molecules. This study evaluates the effects of vanillic acid (VA) on wound healing using HaCaT cells. The results demonstrate that VA treatment increases the levels of phosphorylated AMPK protein, thereby activating the AMPK pathway. This activation leads to an upregulation of PNCA expression and enhances the proliferation of HaCaT cells. Additionally, VA treatment reduces the expression of Bax and cleaved Caspase-3 while increasing Bcl2 expression, which inhibits apoptosis in HaCaT cells. Furthermore, VA treatment upregulates the expression of MMP-2 and MMP-9, promoting HaCaT cell migration. VA also induces the secretion of FGF, VEGF, and PDGF, which enhances tube formation in HUVECs. In conclusion, VA may have potential therapeutic benefits in wound healing by promoting keratinocyte proliferation, migration, and angiogenesis.

Linoleic Acid Induces Metabolic Reprogramming and Inhibits Oxidative and Inflammatory Effects in Keratinocytes Exposed to UVB Radiation.

Linoleic acid (LA), the primary ω-6 polyunsaturated fatty acid (PUFA) found in the epidermis, plays a crucial role in preserving the integrity of the skin's water permeability barrier. Additionally, vegetable oils rich in LA have been shown to notably mitigate ultraviolet (UV) radiation-induced effects, including the production of reactive oxygen species (ROS), cellular damage, and skin photoaging. These beneficial effects are primarily ascribed to the LA in these oils. Nonetheless, the precise mechanisms through which LA confers protection against damage induced by exposure to UVB radiation remain unclear. This study aimed to examine whether LA can restore redox and metabolic equilibria and to assess its influence on the inflammatory response triggered by UVB radiation in keratinocytes. Flow cytometry analysis unveiled the capacity of LA to diminish UVB-induced ROS levels in HaCaT cells. GC/MS-based metabolomics highlighted significant metabolic changes, especially in carbohydrate, amino acid, and glutathione (GSH) metabolism, with LA restoring depleted GSH levels post-UVB exposure. LA also upregulated PI3K/Akt-dependent GCLC and GSS expression while downregulating COX-2 expression. These results suggest that LA induces metabolic reprogramming, protecting against UVB-induced oxidative damage by enhancing GSH biosynthesis via PI3K/Akt signaling. Moreover, it suppresses UVB-induced COX-2 expression in HaCaT cells, making LA treatment a promising strategy against UVB-induced oxidative and inflammatory damage.

Recombinant filaggrin-2 improves skin barrier function and attenuates ultraviolet B (UVB) irradiation-induced epidermal barrier disruption

The integrity of the skin barrier is essential for maintaining skin health, with the stratum corneum and filaggrin 2 (FLG-2) playing a key role. FLG-2 deficiency or mutation has been linked to diseases such as atopic dermatitis, while external stressors such as ultraviolet B (UVB) radiation further damage the epidermal barrier. This study investigated the effects of recombinant filaggrin (rFLG) on skin barrier function and UVB induced epidermal destruction. Cell experiments showed that 10 μg/mL of rFLG could increase the mobility of HaCaT cells from 20 % to 42 %, increase the epithelial resistance (TEER) value by about 2 times, and up-regulate the tight junction associated protein by about 2 times. In mouse models of UVB-induced epidermal barrier destruction, rFLG at concentrations of 0.5, 1, and 2 mg/mL showed effective cell uptake and skin penetration, alleviating erythema, and reducing skin thickness in mice by 1.5–3 times. Among them, 2 mg/mL of rFLG treatment restored the expression of tight junction proteins (LOR, ZO-1, and caspase-14), reduced collagen degradation, and reduced oxidative stress by normalizing serum hydroxyproline and superoxide dismutase levels. In addition, 2 mg/mL of rFLG inhibited UVB-induced upregulation of matrix metalloproteinases (MMP-3 and MMP-9) and reduced pro-inflammatory factors (IL-10, IL-1α, IL-6, and TNF-α) and apoptotic markers (P38, Bax, and Bcl-2) to normal levels. These findings suggested that rFLG effectively enhanced skin barrier integrity and mitigated UVB-induced epidermal barrier destruction, highlighting its potential as a therapeutic agent for diseases associated with skin barrier dysfunction.

Novel Collagen Analogs with Multicopy Mucin-Type Sequences for Multifunctional Enhancement Properties Using SUMO Fusion Tags.

Multifunctional enhanced collagen materials in green biomanufacturing are highly desired yet challenging due to the poor comprehensive performance caused by the adoption of targeting monofunctional peptides. Herein, novel collagen analog design strategy using multicopy tandem of mucin-type sequence (GAPGAPGSQGAPGLQ) derived from human COL1α1 to construct basic building blocks is reported, in which SUMO tag is added to the N-terminal of the protein as a stabilizing core. In particular, novel collagen analogs (named S1506, S1511, S1523, and S1552) with multicopy mucin-type sequences (repeated 6, 11, 23, and 52 times), which were constructed in Escherichia coli, have distinct orientation preferences of functional enhancement (including cell proliferation, differentiation, migration, antioxidant activity, and anti-inflammatory property) compared to COL1α1 in HaCaT and THP-1 cell experiments due to variant three-dimensional structures (the different-length mucin-type polypeptide chains wind around central SUMO tag). Our findings suggest that the innovative protein design and synthesis approaches employed in the construction of these novel S15 proteins have the potential to advance the development of new types of recombinant collagen analogs.