Konjac Ceramide (kCer)-Mediated Signal Transduction of the Sema3A Pathway Promotes HaCaT Keratinocyte Differentiation

Histamines suppress epidermal keratinocyte differentiation. Previously, we reported that konjac ceramide (kCer) suppresses histamine-stimulated cell migration of HaCaT keratinocytes. kCer specifically binds to Nrp1 and does not interact with histamine receptors. The signaling mechanism of kCer in HaCaT cells is also controlled by an intracellular signaling cascade activated by the Sema3A-Nrp1 pathway. In the present study, we demonstrated that kCer treatment induced HaCaT keratinocyte differentiation after migration of immature cells. kCer-induced HaCaT cell differentiation was accompanied by some features of keratinocyte differentiation markers. kCer induced activating phosphorylation of p38MAPK and c-Fos, which increased the protein levels of involucrin that was the latter differentiation marker. In addition, we demonstrated that the effects of both kCer and histamines are regulated by an intracellular mechanism of Rac1 activation/RhoA inhibition downstream of the Sema3A/Nrp1 receptor and histamine/GPCR pathways. In summary, the effects of kCer on cell migration and cell differentiation are regulated by cascade crosstalk between downstream Nrp1 and histamine-GPCR pathways in HaCaT cells.

Carotenoids from Persimmon (Diospyros kaki Thunb.) Byproducts Exert Photoprotective, Antioxidative and Microbial Anti-Adhesive Effects on HaCaT

Persimmon (Diospyros kaki Thunb.) fruits are a remarkable source of carotenoids, which have shown protective effects against UV radiation in bacteria, fungi, algae, and plants. The aim of this study was to analyze the photoprotection provided by an acetone extract, rich in carotenoids and obtained from byproducts derived from the persimmon juice industry, against UV-induced cell death in the keratinocyte HaCaT cell line. For this purpose, the cytotoxicity and phototoxicity of carotenoid extract, as well as its intracellular reactive oxygen species (ROS) scavenging and anti-adhesive activities towards HaCaT cells, were evaluated. The in vitro permeation test provided information about the permeability of the carotenoid extract. Persimmon extracts, rich in carotenoids (PEC), were absorbed by HaCaT keratinocyte cells, which reduced the UV-induced intracellular ROS production in treated cells. Thus, PEC exerted a photoprotective and regenerative effect on UV-irradiated HaCaT cells, and this protection was UV dose-dependent. No cytotoxic effect was observed in HaCaT cultures at the concentration tested. PEC treatment also stimulated the adhesion capacity of skin microbiome to HaCaT cells, while exhibiting a significant anti-adhesive activity against all tested pathogens. In conclusion, PEC showed potential for use as a functional ingredient in skin-care products.

A Cornflower Extract Containing N-Feruloylserotonin Reduces Inflammation in Human Skin by Neutralizing CCL17 and CCL22 and Inhibiting COX-2 and 5-LOX

Thymus and Activation-Regulated Chemokine (TARC/CCL17) and Macrophage-Derived Chemokine (MDC/CCL22) are two key chemokines exerting their biological effect via binding and activating a common receptor CCR4, expressed at the surface of type 2 helper T (Th2) cells. By recruiting Th2 cells in the dermis, CCL17 and CCL22 promote the development of inflammation in atopic skin. The aim of this research was to develop a plant extract whose biological properties, when applied topically, could be beneficial for people with atopic-prone skin. The strategy which was followed consisted in identifying ligands able to neutralize the biological activity of CCL17 and CCL22. Thus, an in silico molecular modeling and a generic screening assay were developed to screen natural molecules binding and blocking these two chemokines. N-Feruloylserotonin was identified as a neutraligand of CCL22 in these experiments. A cornflower extract containing N-feruloylserotonin was selected for further in vitro tests: the gene expression modulation of inflammation biomarkers induced by CCL17 or CCL22 in the presence or absence of this extract was assessed in the HaCaT keratinocyte cell line. Additionally, the same cornflower extract in another vehicle was evaluated in parallel with N-feruloylserotonin for cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX) enzymatic cellular inhibition. The cornflower extract was shown to neutralize the two chemokines in vitro, inhibited COX-2 and 5-LOX, and demonstrated anti-inflammatory activities due mainly to the presence of N-feruloylserotonin. Although these findings would need to be confirmed in an in vivo study, the in vitro studies lay the foundation to explain the benefits of the cornflower extract when applied topically to individuals with atopic-prone skin.

Anti-Inflammatory Effects of Weigela subsessilis Callus Extract via Suppression of MAPK and NF-κB Signaling

Weigela subsessilis is used in folk medicine to treat pain and allergic syndromes in Korea. However, the antibacterial and anti-inflammatory activities of W. subsessilis callus extract remain unexplored. In this study, we aimed to evaluate the W. subsessilis callus of pharmacological activity. Therefore, we first established in vitro calluses of W.subsessilis via plant tissue culture methods. We then evaluated the antioxidant and anti-inflammatory effects of W. subsessilis callus extract in lipopolysaccharide (LPS)-treated RAW264.7 macrophage cells. The W. subsessilis callus extract showed antioxidant and anti-inflammatory effects. These effects were regulated via suppression of mitogen-activated protein kinase signaling through LPS-induced translocation of nuclear factor kappa B (NF-κB) p65 from the cytoplasm to the nucleus. W. subsessilis callus extract also showed antibacterial and anti-inflammatory activities in Propionibacterium acnes-treated HaCaT keratinocyte cells. These results indicate that W. subsessilis callus extract has antioxidant, antibacterial and anti-inflammatory activities, suggesting its possible application in the treatment of inflammatory disorders.

Prevention by the Natural Artocarpin of Morphological and Biochemical Alterations on UVB-Induced HaCaT Cells

Natural substances have gained considerable attention for skin protection against UV light reactions. Artocarpus altilis plant’s heartwood extract is comprised of artocarpin as a major substance, already known for its interesting biological attributes as an antimicrobial, an anti-inflammatory, an antioxidant, and a melanogenesis inhibitor. The present work clarified the mechanism of natural artocarpin (NAR) with a purity of approximately 99% against the effects of UVB-induced HaCaT keratinocyte apoptosis. The indicated results showed that NAR suppresses free radical production (ROS and nitrite) and apoptosis-related molecule activation (caspase-3, p-p53, p-p38, and NF-κB p65) and secretion (TNF-α). Additionally, NAR prevented structural damages (nuclei condensation and fragmentation, apoptotic body formation, impaired cell adherence and round cell shape, disruption of F-actin filament, and clustering of cell death receptor CD95/Fas) and biophysical changes (plasma membrane rigidification). Thus, NAR acts directly from scavenging free radicals generated by UV and indirectly by suppressing morphological and biochemical UV-induced cell damages. Its biological effects are mainly attributed to antioxidant and antiapoptotic properties. Taken together, NAR could be considered as an effective natural product for photoprotective formulations.

Isolation, characterization and evaluation of anti-proliferative properties of andrographolide isolated from Andrographis paniculata on cultured HaCaT cells

Summary Introduction: Psoriasis is an inflammatory skin disease characterized by hyper-proliferation, abnormal epidermal keratinocytes and inflammatory infiltration. It affects approximately 4% of the population globally. Herbal extracts have better results with less toxic effects than the synthetic drugs in the treatment of psoriasis. Objective: Present study was aimed to access the anti-psoriatic effect of andrographolide extracted from Andrographis paniculate (A. paniculata). Method: We extracted, characterized, and screened the extracted andrographolide for anti-proliferative characteristics using cultured cell model of human HaCaT keratinocyte. Results: Andrographolide at 31.25 µg/mL (90 µM) demonstrated significant inhibitory effect on human HaCaT keratinocytes proliferation in cell culture. To our best knowledge, we reported the anti-proliferative potency of andrographolide extracted from A. paniculata for the first time. Conclusion: The results suggest that the andrographolide extracted from A. paniculata plant may have potential to be used in the management of psoriasis.

Intracellular Ca2+-Mediated AE2 Is Involved in the Vectorial Movement of HaCaT Keratinocyte

Keratinocyte migration is initiated toward the wound skin barrier as a crucial process in wound healing. However, the migratory machinery used by keratinocytes is relatively unknown. Histamine signaling, including an increase in the Ca2+ signal, mediated the enhanced protein expression and chloride/bicarbonate exchange activity of anion exchanger AE2 in keratinocytes. In this study, we applied an agarose spot assay to induce a vectorial motion. The vectorial stimulation of the histamine-containing agarose spot enhanced the HaCaT keratinocyte migration, compared to non-directional stimulation. AE2 is associated with the vectorial movement of HaCaT keratinocytes. Enhanced expression of AE2 was mainly associated with an increase in Ca2+ and was abolished by the treatment with the Ca2+ chelating agent BAPTA-AM. These findings revealed that the directionality of Ca2+-exerted stimulation can play a prominent role in facilitating migration through the involvement of AE2 as a migratory machinery in HaCaT keratinocytes.

The effects of oscillatory temperature on HaCaT keratinocyte behaviors

BACKGROUND: Keratinocytes are exposed to a thermal gradient throughout epidermal layers in human skin depending on environmental temperatures. OBJECTIVE: Here, the effect of cyclic temperature changes (ΔT) on HaCaT cell behaviors was explored. METHODS: HaCaT cells were cultured at constant temperature (37 °C or 25 °C) or under ΔT conditions. The morphology, mechanics, cell cycle progression, proliferation, and lipid synthesis of HaCaT cells were determined. RESULTS: ΔT conditions led to the inhomogeneous arrangement of the cytoskeleton in HaCaT cells, which resulted in enlarged size, rounder shape, and increased stiffness. Accumulation in the G2/M phase in the cell cycle, a decreased proliferation rate, and a delayed lipogenesis were detected in HaCaT cells cultured under ΔT conditions. CONCLUSIONS: ΔT conditions resulted in the re-arrangement of the cytoskeleton in HaCaT cells, which showed similarity to the temperature-induced disassemble and re-assemble of cytoskeletons in keratinocyte in vivo. The altered cytoskeleton arrangement resulted in the cell enlargement and stiffening, which reflected the changes in cellular functions. The application of oscillatory temperature in the in vitro culture of keratinocytes provides a way to gain more insights into the role of skin in response to environmental stimuli and maintaining its homeostasis in vivo.