• Journal of Physiology & Pathology in Korean Medicine
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• v.29 no.6
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• pp.475-484
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• 2015

In this paper, we studied the anti-oxidative capacities and protective effects of water extract of Gagam-Danguieumja(GDE) against Ultraviolet B(UVB)-induced oxidative damage in human keratinocytes(HaCaT). To evaluate the anti-oxidative activities of GDE, we measured scavenging activities on DPPH radical, hydroxyl radical, hydrogen peroxide, superoxide anion, lipid peroxidation and reducing power of GDE. To detect the protective effects of GDE against UVB, we irradiated with 40 mJ/㎠`s UVB to HaCaT cells then we measured reactive oxygen species(ROS) generation, apoptotic bodies and cell viability using DCFH-DA assay, Hoechst 33342 staining and MTT assay. GDE showed the anti-oxidative activities by scavenging DPPH radical, hydroxyl radical, hydrogen peroxide, superoxide anion, lipid peroxidation. Also GDE showed high reducing values. GDE reduced oxidative stress conditions by inhibition of ROS expression. Also the cell apoptosis by UVB-induced oxidative conditions was decreased by GDE treatment. These results could suggest that GDE had anti-oxidative activities and exhibited protective effects against UVB on HaCaT cells. GDE would be useful for the development of cosmetics treating UVB-induced skin aging.

• The Korea Journal of Herbology
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• v.29 no.6
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• pp.63-71
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• 2014

Objectives : Lonicerae japonicae Flos(LJF) has been reported to exhibit anti-oxidant, anti-inflammatory, anti-viral, anti-rheumatoid properties. However, it is still largely unknown whether LJF inhibits the ultraviolet(UV)B-induced oxidative damage in human HaCaT keratinocytes. Therefore in this paper, we investigated the anti-oxidative capacity and protective effect of LJF against UVB-induced oxidative demage in human HaCaT keratinocytes. Methods : To evaluate the anti-oxidative activity of LJF extracts, we measured total phenolic contents, total flavonoid contents, antioxidant capacity, and superoxide scavenging activity. To give an oxidative stress to HaCaT cells, UVB was irradiated with $200mJ/cm^2$ to HaCaT cells. To detect the protective effect of LJF against UVB, we measured cell viability, DNA fragmentation and reactive oxygen species (ROS) production. In addition, we performed high-performance liquid chromatography (HPLC) analysis to find a major component of LJF. Results : LJF contained phenolic and flavonoid contents, and showed the anti-oxidant and superoxide scavenging activity. The UVB-induced oxidative conditions led to the cell death, DNA fragmentation and reactive oxygen species (ROS) production. However, pretreatment with LJF reduced oxidative conditions, including inhibition of cell death, DNA fragmentation and ROS production. In addition, we found out chlorogenic acid as major component of LJF. Conclusions : These results could suggest that LJF contained anti-oxidative contents and exhibited protective effects against UVB on human HaCaT keratinocytes. And the effective compound of LJF which could show protective activities against UVB is chlorogenic acid. Thus, LJF and chlorogenic acid would be useful for the development of drug or cosmetics treating skin troubles.

• Toxicological Research
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• v.22 no.3
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• pp.283-291
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• 2006

The survival and growth of epithelial cells depends on adhesion to the extracellular matrix. An adhesion signal may regulate the initiation of differentiation, since epidermal keratinocytes differentiate as they leave the basement membrane. A metabolically dead cornified cell envelope is the end point of epidermal differentiation so that this process may be viewed as a specialized form of programmed cell death. In order to investigate the precise cellular signaling events loading to terminal differentiation of keratinocytes, we have utilized HaCaT cells to monitor the biological consequences of $Ca^{2+}$ stimulation and numerous downstream signaling pathways, including activation of the extracellular signal-regulated protein kinase(ERK) pathway and activation of phosphatidylinositol 3-kinase(PI3K). The results presented in this study show that $Ca^{2+}$ function as potent agents for the differentiation of HaCaT keratinocytes, and this differentiation depends or the activation of ERK, Protein kinase B(PKB) and p70 ribosomal protein S6 kinase(p70S6K). Finally, the results show that the expression of Activator protein 1(AP-1; c-Jun and c-Fos) increased following $Ca^{2+}$-mediated differentiation of HaCaT cells, suggesting that ERK-mediated AP-1 expression is critical for initiating the terminal differentiation of keratinocytes.

• Korean Journal of Pharmacognosy
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• v.46 no.2
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• pp.116-122
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• 2015

Keratinocytes are first barrier against outer challenges on skin. However, it is still largely unknown about effective protectors against ultraviolet B (UVB), and oxidative stress in human keratinocyte, HaCaT cells. Inducible heme oxygenase (HO)-1 acts against oxidants that are thought to play a role in the pathogenesis of skin disorders. Therefore, the purpose of this study was to evaluate the effect of Portulaca oleracea 70% EtOH extracts against hydrogen peroxide (H₂O₂)-induced oxidative stress in human keratinocytes, HaCaT cells. P. oleracea 70% EtOH extracts showed the potent protective effects on H₂O₂-induced toxicity by induced the expression of HO-1 in human keratinocyte, HaCaT cells. Furthermore, P. oleracea 70 % EtOH extracts caused the nuclear accumulation of nuclear factor E2-related factor 2 (Nrf2) in human keratinocytes, HaCaT cells. In addition, we found that treatment with c-Jun N-terminal kinase (JNK) inhibitor (SP600125) reduced P. oleracea 70% EtOH extracts-induced HO-1 expression, and JNK inhibitor (SP600125) also inhibited protective effects by P. oleracea 70% EtOH extracts. Therefore, these results suggest that P. oleracea 70 % EtOH extracts increases cellular resistance to H₂O₂-induced oxidative injury in human keratinocyte, HaCaT cells, presumably through JNK pathway-Nrf2-dependent HO-1 expression.

• Korean Journal of Microbiology
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• v.47 no.4
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• pp.348-355
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• 2011

Staphylococcus aureus is a major human pathogen that is associated with various types of local and systemic infection. Staphylococcal protin A (SPA), a highly expressed surface component of S. aureus, may have a role in virulence such as activating inflammation and interfering with immune clearance. We examined the effect of recombinant SPA on inflammatory response in human HaCaT keratinocytes. The recombinant SPA protein was prepared using the pET-28a Vector System in Escherichia coli. The expression of pro-inflammatory related adhesion molecules and cytokines in HaCaT cells incubated for 6, 12, and 24 h with SPA (2 ${\mu}g$/ml) was analyzed by comparative RT-PCR or ELISA. The expression of E-selectin, ICAM-1, MCP-1, IL-6 and IL-8 was significantly increased in HaCaT from 6 to 24 h after treatment with SPA. SPA showed the effect on the adhesion-promoting ability of U937 monocytes to HaCaT cells. Our data demonstrate that SPA stimulates inflammatory response of HaCaT cells, implicating an important factor for exacerbation of skin inflammation of immunologic disease.

• Proceedings of the PSK Conference
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• 2003.04a
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• pp.231.1-231.1
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• 2003

Ultraviolet (UV) irradiation is well known to cause human skin aging and skin cancer through activation of matrix metalloproteinases (MMPs) which are responsible for the degradation of collagen, an extracellular matrix component. However, the molecular mechanisms of UV-induced MMP expression are yet to be defined. In this study, we investigated signaling molecules involved in UV-induced MMP expression in HaCaT human keratinocytes. (omitted)

• The Korean Journal of Food And Nutrition
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• v.29 no.3
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• pp.335-340
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• 2016

The objective of the present investigation was to obtain vitamin, mineral, flavonoid, and polyphenol profiles of Rudbeckia laciniata (RL), and to examine the effects of extract of RL (RLE) on various physiological activities of HaCaT keratinocyte for the utilization of RL as natural raw materials to develop functional food. To accomplish this purpose, we checked the contents of the general nutrients of RL. The contents of vitamin A, vitamin $B_1$ and vitamin $B_2$ were $7.49{\mu}g/g$, $51.96{\mu}g/g$, and $132{\mu}g/g$ respectively, while vitamin C and vitamin $D_3$ were not detected. The contents of mineral such as Ca, K and Fe were 2.01 mg/g, 6.06 mg/g and 0.03 mg/g respectively. Total flavonoid contents of RLE were 0.25 mg/g, and total polyphenol were estimated as 1.43 mg/g. Because RL contains high levels of vitamin A which is associated with skin aging, we investigated the effect of RLE on physiological function of keratinocytes with respect to skin aging. We found that RLE significantly increased the growth rate of HaCaT cells and reduced ultraviolet radiation B (UVB)-induced cellular toxicity. Also, the extract of Rudbeckia laciniata attenuated the UVB-induced reactive oxygen species (ROS) generation in a dose-dependent manner in HaCaT cells. In addition, treatment with the extract dose-dependently increased migration activity of HaCaT cells. Thus, these findings indicated that RLE could regulate the physiological activity of keratinocytes, and may be used to develop functional foods.

• Journal of Microbiology and Biotechnology
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• v.17 no.2
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• pp.271-279
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• 2007

Black rice (Oryza sativa L. var. japonica) has been used in folk medicine in Asia. To understand the effects of black rice hydrolyzed peptides (BRP) from germinated black rice, we assessed the expression levels of about 20,000 transcripts in BRP-treated HaCaT keratinocytes using human 1A oligo microarray analysis. As a result, the BRP treatment showed a differential expression ratio of more than 2-fold: 745 were activated and 1,011 were repressed. One of the most interesting findings was a 2-fold increase in hyaluronan synthase 2 (HAS2) gene expression by BRP. Semiquantitative RT-PCR showed that BRP increased HAS2 mRNA in dose-dependent manners. ELISA showed that BRP effectively increased hyaluronan (HA) production in HaCaT keratinocytes.

• Biomolecules & Therapeutics
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• v.16 no.4
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• pp.394-402
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• 2008

Atopic dermatitis (AD) is an inflammatory skin disease commonly characterized by infiltration of inflammatory cells into skin lesions. Keratinocytes produce many chemokines that are involved in the pathogenesis of skin disorders. In particular, macrophage-derived chemokine (MDC/CCL22) and thymus and activationregulated chemokine (TARC/CCL17) are Th2-type cytokines. Serum MDC and TARC levels are increased in AD patients. In this study, we investigated the anti-inflammatory effect and mechanism of action of the active fraction from Prunus yedoensis bark. We evaluated their inhibitory effects on the AD-like inflammatory markers (MDC and TARC) and JAK-STAT pathway (STAT1) in HaCaT keratinocytes. The EtOAc fraction of the crude extract (80% EtOH) and the E5 sub-fraction potently inhibited the induction of MDC and TARC mRNA and protein at 50 ${\mu}g$/mL in HaCaT cells. In addition, the E5 sub-fraction inhibited the phosphorylation of STAT1 protein associated with IFN-$\gamma$ signaling transduction in a dose-dependent manner. Thus, P. yedoensis may have antiatopic activity by suppressing the inflammatory chemokines (MDC and TARC).

• Biomolecules & Therapeutics
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• v.11 no.4
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• pp.224-231
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• 2003

The aim of this study was to investigate the effect of resveratrol on the oxidative stress-induced inhibition of gap junctional intercellular communication in HaCaT keratinocytes. Anti-oxidative activity of resveratrol was measured by $\alpha,\alpha$-diphenyl-$\beta$-picrylhydrazyl assay and dichlorodihydrofluorescein diacetate oxidation assay. Gap junctional intercellular communication in HaCaT keratinocytes was assessed using the scrape loading/dye transfer technique. Western blots and reverse transcription-polymerase chain reaction were also analyzed for connexin 43 protein and mRNA expression, respectively. Resveratrol scavenged directly the stable $\alpha,\alpha$-diphenyl-$\beta$-picrylhydrazyl radical over a concentration range of 4 mg/ml ($78.2{\pm}2.7$% of control) to 500 mg/ml ($29.9{\pm}4.2$% of control) and decreased the intracellular reactive oxygen species induced by ultraviolet A (UVA) irradiation ($89.3{\pm}1.1$% of UVA group), ultraviolet B (UVB) irradiation ($70.9{\pm}1.7$% of UVB group) and 12-0-tet-radecanoylphorbol-13-acetate (TPA, $48.3{\pm}1.1$% of TPA group), respectively. UVA irradiation and TPA markedly reduced gap junctional intercellular communication, which was restored by resveratrol. There were no significant differences in the level of connexin 43 protein and mRNA expression among any of the experimental groups. Our data suggests that resveratrol has the protective effect on the oxidative stress-induced inhibition of gap junctional intercellular communication in HaCaT keratinocytes, and this protection is likely due to the scavenging of reactive oxygen species.