Expression In Human Dermal Fibroblasts Research Articles

Alnus Sibirica Extracts Suppress the Expression of Inflammatory Cytokines Induced by Lipopolysaccharides, Tumor Necrosis Factor-α, and Interferon-γ in Human Dermal Fibroblasts.

The effects of Alnus sibirica (AS) extracts on cytokine expression induced by inflammatory stimulants were examined in human dermal fibroblasts (HDFs) and RAW264.7 cells. The anti-oxidative effect and effect on cell viability of AS extracts were evaluated, and four extracts with the highest anti-oxidative effects were selected. HDFs and RAW264.7 cells were treated with inflammatory stimulants, and the expression of cytokines involved in acute (IL-6 and IL-10) and chronic (IL-18) inflammation, the initiation of the immune response (IL-33), and non-specific immune responses (IL-1β, IL-8, and TNF-α) were determined using a reverse-transcription polymerase chain reaction. LPS increased the expression of all the cytokines, except for IL-18; however, AS extracts, particularly AS2 and AS4, reduced this increase, and TNF-α treatment markedly increased the expression of cytokines related to non-specific immune responses. IFN-γ treatment induced no significant changes, except for increased IL-33 expression in HDFs. AS extracts inhibited the increase in the expression of IL-33 and other cytokines in HDFs. Thus, the exposure of HDFs and RAW264.7 cells to inflammatory stimulants increased the expression of cytokines related to all the inflammatory processes. HDFs are involved not only in simple tissue regeneration but also in inflammatory reactions in the skin. AS2 and AS4 may offer effective therapy for related conditions.

Photoprotective Effects of Ice Plant (Mesembryanthemum crystallinum) Callus Extract on Gene Expression of Human Dermal Fibroblast Against UV Exposure

UV exposure is the principal cause of extrinsic photoaging. Antioxidant-related genes (SOD2 and CAT) and collagen-related genes (COL1A1 and TIMP1) were selected for analysis of the mRNA expression in human dermal fibroblasts (CCD-966SK) using qPCR. In this study, UVA-exposed (15 J/cm2 cells showed decrease in SOD2 , CAT (p < 0 001) and COL1A1 (p < 0 05) gene expression, indicating the decline of antioxidant ability and collagen formation. However, treatment with ice plant callus extract (2 mg/mL, 24 h) before UVA exposure significantly up-regulated SOD2 , CAT , COL1A1 and TIMP1 genes (p < 0 01) by 9.5, 2.7, 1.7 and 3.8 times, respectively, compared with those of the control group, and by 10.2, 4.3 (p < 0 001), 2.1 and 3.8 times, respectively, compared with those of the UVA (only) group. These results demonstrated that ice plant extracts affect both antioxidant- and collagen-related gene expressions, and show positive effects on endogenous antioxidant activity and skin collagen preservation.

Yin/Yang expression of CCN family members: Transforming growth factor beta 1, via ALK5/FAK/MEK, induces CCN1 and CCN2, yet suppresses CCN3, expression in human dermal fibroblasts

The role of the microenvironment in driving connective tissue disease is being increasingly appreciated. Matricellular proteins of the CCN family are signaling modifiers that are secreted by cells into the extracellular matrix microenvironment where they have profound, context-dependent effects on organ development, homeostasis and disease. Indeed, CCN proteins are emergent targets for therapeutic intervention. Recent evidence suggests that, in vivo, CCN3 has effects opposing CCN2. Moreover, when CCN3 expression is high, CCN2 expression is low. That is, they appear to be regulated in a yin/yang fashion, leading to the hypothesis that the CCN2:CCN3 ratio is important to control tissue homeostasis. To begin to test the hypothesis that alterations in CCN2:CCN3 expression might be important in skin biology in vivo, we evaluated the relative ex vivo effects of the profibrotic protein TGFbeta1 on dermal fibroblasts on protein and RNA expression of CCN3 and CCN2, as well as the related protein CCN1. We also used signal transduction inhibitors to begin to identify the signal transduction pathways controlling the ability of fibroblasts to respond to TGFbeta1. As anticipated, CCN1 and CCN2 protein and mRNA were induced by TGFbeta1 in human dermal fibroblasts. This induction was blocked by TAK1, FAK, YAP1 and MEK inhibition. Conversely, TGFbeta1 suppressed CCN3 mRNA expression in a fashion insensitive to FAK, MEK, TAK1 or YAP1 inhibition. Unexpectedly, CCN3 protein was not detected in human dermal fibroblasts basally. These data suggest that, in dermal fibroblasts, the profibrotic protein TGFbeta1 has a divergent effect on CCN3 relative to CCN2 and CCN1, both at the mRNA and protein level. Given that the major source in skin in vivo of CCN proteins are fibroblasts, our data are consistent that alterations in CCN2/CCN1: CCN3 ratios in response to profibrotic agents such as TGFbeta1 may play a role in connective tissue pathologies including fibrosis.

Inhibitory role of silk cocoon extract against elastase, hyaluronidase and UV radiation-induced matrix metalloproteinase expression in human dermal fibroblasts and keratinocytes.

Topical delivery of potent antioxidants maintain the redox balance of the skin, which leads to the downregulation of matrix metalloproteinase (MMP) expression and prevents UV radiation-induced photoaging. In this study, we aimed at investigating the inhibitory role of silk cocoon extract (SCE) isolated from the Antheraea assamensis (AA), Bombyx mori (BM), and Philosamia ricini (PR) silk varieties against UV radiation-induced MMP expression. Incubation of elastase and hyaluronidase with Antheraea assamensis silk cocoon extract (AASCE) caused 50% inhibition of activity. The assessment of total collagen content using the Sirius red assay showed that AASCE (10 μg mL-1) and Philosamia ricini silk cocoon extract (PRSCE at 100 μg mL-1 concentration) post-treatment significantly enhanced the total collagen content in UVA1 and UVB irradiated HDF cells, whereas BM silk cocoon extract (BMSCE at 100 μg mL-1 concentration) post-treatment significantly enhanced the total collagen content in UVA1-irradiated HDF cells. Gene expression studies revealed AASCE and PRSCE post-treatment downregulated the expression of interleukin (IL)-6, MMP-1 and upregulated procollagen genes in UV irradiated HDF cells. Gelatin zymography studies with AASCE post-treatment downregulated the release of MMP-2 and MMP-9 by HaCaT cells. The overall results validate AASCE efficiently shielding UV radiation-induced collagen and elastin degradation by downregulation of MMP expression, substantiating its further use as a potent antioxidant complement in skin care formulations.

Antioxidative and antiphotoaging activities of neferine upon UV-A irradiation in human dermal fibroblasts.

Our daily exposure to ultraviolet radiation (UVR) results in the production of reactive oxygen species (ROS), lipids, proteins and DNA damage and alteration in fibroblast structure, thus contributing to skin photoaging. For this reason, the use of natural bioactive compounds with antioxidant activity could be a strategic tool to overcome ultraviolet A (UV-A) induced deleterious effect. Neferine is an alkaloid extract from the seed embryos of lotus (Nelumbo nucifera Gaertn). In the present study, we report the protective effect of neferine against UV-A induced oxidative stress and photoaging in human dermal fibroblasts (HDFs). HDFs subjected to UV-A irradiation showed increased production of ROS and malondialdehyde (MDA). Furthermore, it depleted the cellular enzymatic antioxidant superoxide dismutase (SOD) and non-enzymatic antioxidant glutathione peroxidase (GPx). On the other hand, HDFs treated with neferine followed by UV-A irradiation reversed the process, reduced the ROS and lipid peroxidation and restored the antioxidants pool. Moreover, neferine treatment significantly inhibited UV-A induced matrix metalloproteinase-1 (MMP-1) expression in HDFs. Remarkable morphological and ultrastructural alterations observed in HDFs upon UV-A irradiation, were also reduced with neferine treatment. Taken together, our results suggest that neferine has strong antioxidative and photoprotective properties and thus may be a potential agent for the prevention and treatment of UV-A mediated skin photoaging.

Molecular Additives Significantly Enhance Glycopolymer-Mediated Transfection of Large Plasmids and Functional CRISPR-Cas9 Transcription Activation Ex Vivo in Primary Human Fibroblasts and Induced Pluripotent Stem Cells.

Fast, efficient, and inexpensive methods for delivering functional nucleic acids to primary human cell types are needed to advance regenerative medicine and cell therapies. Plasmid-based gene editing (such as with CRISPR-Cas9) can require the delivery of plasmids that are large (∼9.5-13 kbp) in comparison to common reporter plasmids (∼5-8 kbp). To develop more efficient plasmid delivery vehicles, we investigated the effect of plasmid size on the transfection of primary human dermal fibroblasts (HDFs) and induced pluripotent stem cells (iPSCs) using a heparin-treated trehalose-containing polycation (Tr4-heparin). Transfections with 4.7 kbp to 10 kbp plasmids exhibited high rates of polyplex internalization with both plasmid sizes. However, transfection with the large plasmid was nearly eliminated in HDFs and significantly reduced in iPSCs. Molecular additives were used to probe intracellular barriers to transfection. Chloroquine treatments were used to destabilize endosomes, and dexamethasone and thymidine were used to destabilize the nuclear envelope. Destabilizing the nuclear envelope resulted in significantly increased large-plasmid-transfection, indicating that nuclear localization may be more difficult for large plasmids. To demonstrate the potential clinical utility of this formulation, HDFs and iPSCs were treated with to dexamethasone-Tr4-heparin polyplexes encoding dCas9-VP64, synthetic transcription activator, targeted to collagen type VII. These transfections enhanced collagen expression in HDFs and iPSCs by 5- and 20-fold, respectively, compared to an untransfected control and were the more effective than the Lipofectamine 2000 control. Functional plasmid transfection efficiency can be significantly improved by nuclear destabilization, which could lead to improved development of nonviral vehicles for ex vivo CRISPR-Cas9 gene editing.

Identification of Matrix Metalloproteinase-1-Suppressive Peptides in Feather Keratin Hydrolysate.

Inhibition of matrix metalloproteinases (MMPs), which degrade collagen and elastin in the dermis of normal skin, is a key strategy for anti-skin aging. In this study, we identified five low-molecular-weight (LMW, <1 kDa) MMP-1-suppressive peptides in feather keratin hydrolysate (FKH) obtained by anaerobic digestion with an extremophilic bacterium. FKH was first subjected to ultrafiltration, followed by size-exclusion chromatography and liquid chromatography/electrospray ionization tandem mass spectrometry analysis. Chemically synthesized peptides identical to the sequences identified suppressed MMP expression in human dermal fibroblasts (HDFs). To investigate the impact of the MMP-1-suppressive peptides on the signaling pathway, we performed antibody array phosphorylation profiling of HDFs. The results suggested that the peptide GGFDL regulates ultraviolet-B-induced MMP-1 expression by inhibiting mitogen-activated protein kinases and nuclear factor κB signaling pathways as well as histone modification. Thus, LMW feather keratin peptides could serve as novel bioactive compounds to protect the skin against intrinsic and extrinsic factors.

Caffeic acid prevents UVB radiation induced photocarcinogenesis through regulation of PTEN signaling in human dermal fibroblasts and mouse skin

Previously, we proved that caffeic acid (CA), a major dietary phenolic acid, prevents skin carcinogenesis by modulating inflammatory signaling in mouse skin. However, the actual mechanisms of CA against UVB (280–320 nm) induced photocarcinogenesis remains unclear. The present results confirms that CA significantly inhibits single UVB-induced CPDs formation, oxidative DNA damage, ROS generation and frequency of apoptotic cell death in human dermal fibroblasts (HDFa). Furthermore, CA prevents UVB-induced expression of PI3K and AKT kinases through activation of PTEN which subsequently promotes XPC dependant NER proteins such as XPC, XPE, TFIIH (p44) and ERCC1 in HDFa cells and mouse skin tissue. Further, CA directly activates PTEN through hydrogen bond and hydrophobic interactions. Taken together, these findings suggest that CA prevents UVB-induced photodamage through the activation of PTEN expression in human dermal fibroblasts and mouse skin.

Toll-like receptor signalling induces the expression of serum amyloid A in epidermal keratinocytes and dermal fibroblasts.

Toll-like receptors (TLRs) play critical roles in innate immune response by sensing pathogen- or damage-associated molecular patterns. Epidermal keratinocytes and dermal fibroblasts also produce proinflammatory cytokines and chemokines under stimulation with TLR ligands. Serum amyloid A (SAA) is an essential factor in the pathogenesis of secondary amyloidosis, and also has immunomodulatory functions. SAA are produced mainly by hepatocytes but also by a variety of cells, including immune cells, endothelial cells, synoviocytes, and epidermal keratinocytes. However, SAA expression in human dermal fibroblasts has not been shown to date. To investigate the effect of TLR ligands on SAA expression in epidermal keratinocytes and dermal fibroblasts. We investigated whether TLR ligands induce the expression of SAA in normal human epidermal keratinocytes (NHEKs) and normal human dermal fibroblasts (NHDFs) by real-time quantitative PCR and ELISA. The effect of SAA on its own expression in NHDFs was also studied. SAA expression was induced via nuclear factor-κB by TLR1/2, 3, 5 and 2/6 ligands in NHEKs. In NHDFs, TLR1/2 and TLR2/6 ligands increased SAA expression. SAA further induced its own expression via TLR1/2 and NF-κB in NHDFs, as previously reported for NHEKs. Our results provide new evidence that the skin's innate immune response contributes to the production of SAA, which might lead to an increased risk of systemic complications such as secondary amyloidosis of recessive dystrophic epidermolysis bullosa.

1149 UV affects histone deacetylases expression in human dermal fibroblasts

1149 UV affects histone deacetylases expression in human dermal fibroblasts

Artesunate Antagonizes Myofibroblast Markers and Fibrosis‐Associated Extracellular Matrix Protein Expression in Human Dermal Fibroblasts

Artemisia annua, a plant species native to eastern Asia, is the source of a compound known as artemenisin that has been shown to have numerous medicinal effects. Artesunate, artemether and other chemical analogs of artemisinin, initially studied as antimalarial compounds, have also demonstrated cytostatic and cytotoxic effects in bacterial and cancer cells. Recently, artemisinin‐derivative compounds have also been demonstrated to attenuate fibrotic pathologies in various preclinical animal models, including those of hepatic and pulmonary fibrosis, but the mechanisms by which this inhibition of fibrosis occurs is not well‐understood. Given the ability of artemisinin derivatives, artesunate in particular, to antagonize various fibrotic pathologies, we decided to investigate the effects of artesunate on a process common to all fibrotic pathologies: the emergence of the highly secretory, contractile myofibroblast. We demonstrated by Western blot analysis that treatment of human dermal fibroblasts with artesunate results in a dose‐dependent decrease in expression of alpha smooth muscle actin (ɑ‐SMA), the de facto marker of fibroblast activation. Treatment of human dermal fibroblasts with artesunate also inhibited activation of fibroblasts mediated by the profibrotic cytokine TGF‐β1, which is largely responsible for differentiation of fibroblasts into myofibroblasts in vivo, and this antagonistic effect of artesunate towards fibroblast activation was consistent among multiple lines of primary human dermal fibroblasts. Further, immunofluorescent analysis for collagen I and collagen III demonstrated a marked reduction of both basal and TGF‐β1‐induced expression of extracellular matrix‐associated proteins. Understanding that fibrosis is the result of excessive differentiation of fibroblasts to myofibroblasts and accumulation of extracellular matrix proteins secreted by myofibroblasts, our data suggest that the antifibrotic mechanisms of artesunate in vivo may be mediated by their antagonistic effects towards myofibroblast phenotypes. We next plan to determine the direct mechanisms underlying the antagonism of artesunate towards fibroblast activation, paying particular attention to the hypotheses regarding the induction of myofibroblast apoptosis or cell cycle arrest, and the dependence of these phenotypes on the generation and activity of peroxide. With mechanistic data underpinning the effects of artesunate as an antagonist to profibrotic fibroblast activation, we hope that our work will enable progression towards the use of artesunate and other artemenisin‐derived compounds as therapies for fibrotic pathologies.Support or Funding InformationThis research was supported by a National Institutes of Health award to Tanja Dominko (grant # R01GM85456) and a National Science Foundation Integrative Graduate Education and Research Traineeship (grant number DGE 1144804) awarded to David Dolivo.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Inhibition Effect of Cnidium officinale Makino Extracts on MMP1 Expression in Human Dermal Fibroblasts

ëª©ì  천궁은 산형과에 속하는 약초로써 중국이 ì›ì‚°ì§€ì´ê³ , 한국, 중국, 일본에서 많이 사용되어 왔다. 천궁은 진통, 항염증, 월경 ê°œì„ , 비타민 결핍증 ì–µì œ 및 ê³ í˜ˆì••ì— 약으로 사용되어 왔다. 하지만 아직 피부에서의 ì—­í• ì€ ì•Œë ¤ì§€ì§€ 않았다. 때문에 본 연구에서는 천궁의 ì¸ê°„ì§„í”¼ì„¬ìœ ì•„ì„¸í¬ì—ì„œì˜ ì—­í• ì„ í™•ì¸í•˜ê³ ìž 한다. 방법 천궁에 의한 matrix metalloproteinase-1 (MMP1)의 발현 변화는 quantitative real time polymerase chain reaction (qRT-PCR)로 í™•ì¸í•˜ì˜€ê³ , 추가로 activator protein 1 (AP-1)ê³¼ c-Jun N-terminal protein kinase (JNK)의 활성은 각각 luciferase assay와 western blot을 통해서 확인하였다. ê²°ê³¼ ìžì™¸ì„ ì— 의해서 증가된 MMP1의 발현이 천궁추출물에 의해 다시 감소되었다. 또한 ìžì™¸ì„ ì´ 조사된 ì¸ê°„ì§„í”¼ì„¬ìœ ì•„ì„¸í¬ì—ì„œ 천궁추출물은 AP-1의 ì „ì‚¬í™œì„±ì„ ê°ì†Œì‹œí‚¤ê³ , JNK의 인산화를 감소시키는 것으로 확인되었다. ê²°ë¡ ë³¸ ë ¼ë¬¸ì˜ 연구결과를 통해서 천궁추출물은 AP-1ê³¼ JNK 인산화를 ì¡°ì ˆí•¨ìœ¼ë¡œì¨ MMP1의 발현을 감소시킴을 알 수 있다. 때문에 천궁추출물은 피부의 collagen 분해하는 MMP1의 발현을 감소시켜 collagen의 분해를 ì¡°ì ˆí• ìˆ˜ 있는 화장품원료로서 가능성을 ê°€ì§€ê³ ìžˆë‹¤. 핵심용어: MMPs, AP-1, JNK, Cnidium officinale Makino, UV

MicroRNA-378b regulates α-1-type 1 collagen expression via sirtuin 6 interference

Ultraviolet (UV) light mediates skin aging and induces destruction of the dermis by modulating the expression levels of extracellular matrix‑associated genes, including collagen and matrix metalloproteinases. Sirtuin 6 (SIRT6), a member of the sirtuin family of proteins, regulates collagen metabolism and is an established anti‑aging protein. However, the exact underlying mechanism by which SIRT6 expression is regulated in dermal fibroblasts during the aging process is unclear. The present study demonstrated that expression of microRNA‑378b (miR‑378b) is induced in UVB‑exposed human dermal fibroblasts (HDFs), and this was inversely associated with the mRNA expression levels of α‑1‑type 1 collagen (COL1A1). In addition, knockdown of miR‑378b enhanced the mRNA expression levels of COL1A1 in HDFs. A target analysis for miR‑378b was performed, and the results revealed that SIRT6, a regulator of COL1A1, contains a target sequence for miR‑378b in its 3'untranslated region. Notably, the present study demonstrated that an miR‑378b mimic and inhibitor may directly regulate SIRT6 expression in HDFs. In conclusion, the present study suggested that miR‑378b represses the mRNA expression levels of COL1A1 via interference with SIRT6 in HDFs, and may contribute to the underlying molecular mechanism by which UVB inhibits collagen I in dermal fibroblasts.

Lactoferrin induces tropoelastin expression by activating the lipoprotein receptor-related protein 1-mediated phosphatidylinositol 3-kinase/Akt pathway in human dermal fibroblasts.

Dermal fibroblasts generate the extracellular matrix component elastin, which is synthesized as tropoelastin (TE) and play a critical role in maintaining skin elasticity. Lactoferrin (Lf), an 80-kDa iron-binding glycoprotein, has biological functions such as anti-bacterial, -inflammatory, and -cancer activities. We previously reported that bovine Lf increases TE mRNA expression in human dermal fibroblasts. However, it remains unclear how Lf up-regulates TE expression. Here, we investigated molecular mechanisms underlying this effect. Lf promoted the phosphorylation of Akt1 and extracellular signal-regulated protein kinase (ERK)1/2. As expected, the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 and the MAPK inhibitor U0126 inhibited Lf-induced phosphorylation of Akt1 and ERK1/2, respectively. In contrast, LY294002, but not U0126, inhibited Lf-induced TE expression. Human dermal fibroblasts expressed lipoprotein receptor-related protein 1 (LRP-1) mRNA, and the LRP1 inhibitor receptor-associated protein attenuated Lf-induced increases in TE expression. Furthermore, siRNA-mediated knockdown of LRP-1 significantly suppressed Lf-increased TE expression and Lf-induced Akt1 phosphorylation. Iron-saturated Lf (holo-Lf) increased TE expression and promoted Akt1 phosphorylation, when compared to those parameters in cells treated with iron-free Lf (apo-Lf). Transforming growth factor (TGF)-β1 also increased TE expression. LY294002 inhibited TGF-β1-mediated TE upregulation, whereas TGF-β1 activated Akt2, but not Akt1, phosphorylation. These results indicate that holo-Lf, but not apo-Lf, increases TE expression through LRP-1 in human dermal fibroblasts and suggest that holo-Lf and TGF-β1 enhance TE expression by activating the PI3K/Akt1 and PI3K/Akt2 pathways, respectively.

Integrin α4β1 and TLR4 Cooperate to Induce Fibrotic Gene Expression in Response to Fibronectin’s EDA Domain

Alternative splicing of fibronectin increases expression of the EDA+ isoform of fibronectin (EDA+Fn), a damage-associated molecular pattern molecule, which promotes fibro-inflammatory disease through the activation of toll-like receptors. Our studies indicate that the fibronectin EDA domain drives two waves of gene expression in human dermal fibroblasts. The first wave, seen at 2 hours, consisted of inflammatory genes, VCAM1, and tumor necrosis factor. The second wave, evaluated at 24 hours, was composed of the fibrosis-associated cytokines IL-10 and IL-13 and extracellular matrix genes fibronectin and osteopontin. Gene expression was coordinately regulated by the α4β1 integrin and the innate immune receptor toll-like receptor 4. Additionally, we found a significant toll-like receptor 4/α4β1-dependent enrichment in the ratio of EDA+Fn to total fibronectin in response to EDA, consistent with EDA+Fn initiating further production of EDA+Fn. Our data also suggest that the EDA/α4β1 integrin interaction primes the cell for an enhanced response to toll-like receptor 4 ligands. Our studies provide evidence that remodeling of the fibronectin matrix in injured or diseased tissue elicits an EDA-dependent fibro-inflammatory response in dermal fibroblasts. The data suggest a paradigm of damage-associated molecular pattern-based signaling whereby damage-associated molecular pattern binding integrins cooperate with innate immune receptors to stimulate inflammation and fibrosis.

18ß-glycyrrhetinic acid derivative promotes proliferation, migration and aquaporin-3 expression in human dermal fibroblasts.

Licorice (Glycyrrhiza) species have been widely used as a traditional medicine and a natural sweetener in foods. The 18β-glycyrrhetinic acid (18β-GA) is a bioactive compound in licorice that exhibits potential anti-cancer, anti-inflammatory, and anti-microbial activities. Many synthesized derivatives of 18β-GA have been reported to be cytotoxic and suggested for the treatment of malignant diseases. In this study, we explored the possible pharmacological roles of an 18β-GA derivative in skin biology using primary human dermal fibroblasts and HaCaT keratinocytes as cell models. We found that this 18β-GA derivative did not cause cell death, but significantly enhanced the proliferation of dermal fibroblasts and HaCaT keratinocytes. A scratch wound healing assay revealed that the 18β-GA derivative promoted the migration of fibroblasts. Due to the important role of aquaporin-3 in cell migration and proliferation, we also investigated the expression of aquaporin-3 and found this compound up-regulated the expression of aquaporin-3 in dermal fibroblasts and HaCaT keratinocytes. In dermal fibroblasts, the 18β-GA derivative induced the phosphorylation of Akt, ERK, and p38. The inhibitor of Akt predominantly suppressed the 18β-GA derivative-induced expression of aquaporin-3. Collectively, this compound had a positive effect on the proliferation, migration, and aquaporin-3 expression of skin cells, implying its potential role in the treatment of skin diseases characterized by impaired wound healing or dermal defects.

Protective effect of Arthrospira platensis extracts against ultraviolet B-induced cellular senescence through inhibition of DNA damage and matrix metalloproteinase-1 expression in human dermal fibroblasts

Ultraviolet (UV) light exposure causes skin photoaging, which is known to be preventable and controllable by application of UV-protective agents. In this study, we demonstrated, for the first time, that the extract of microalgae Arthrospira platensis has a reverse effect on UV-induced photodamage such as loss of cell viability, cellular senescence, DNA damage, and collagen destruction in dermal fibroblasts. Forty-eight extracts were prepared from the cell biomass by controlling culture light conditions, extract solvents, and disruption methods. Then, we analyzed their cytotoxicities using WST-1 assay and separated low and high cytotoxic extracts with normal human dermal fibroblasts (nHDFs). Using the low cytotoxic extracts, we performed UVB protection assay and selected the most effective extract demonstrating protective effect against UVB-induced nHDF damage. Flow cytometric analysis and senescence-associated (SA) β-galactosidase assay showed that pretreatment with the extract reversed UVB-induced G2/M phase cell cycle arrest and senescence in nHDFs. Furthermore, UVB-induced DNA damage in nHDFs, such as cyclobutane pyrimidine dimer formation, was significantly suppressed by the extract. Further, quantitative real-time PCR experiments revealed that the extract significantly inhibited UVB-induced upregulation of matrix metalloproteinase 1 (MMP1) and MMP3 expression in nHDFs. Therefore, we concluded that the microalgae extract can be a potential anti-photoaging agent.

Fli1 Deficiency Induces CXCL6 Expression in Dermal Fibroblasts and Endothelial Cells, Contributing to the Development of Fibrosis and Vasculopathy in Systemic Sclerosis.

CXCL6, a chemokine with proangiogenic property, is reported to be involved in vasculopathy associated with systemic sclerosis (SSc). We investigated the contribution of CXCL6 to SSc development by focusing on the association of friend leukemia virus integration 1 (Fli1) deficiency, a potential predisposing factor of SSc, with CXCL6 expression and clinical correlation of serum CXCL6 levels. mRNA levels of target genes and the binding of Fli1 to the CXCL6 promoter were evaluated by quantitative reverse transcription-PCR and chromatin immunoprecipitation, respectively. Serum CXCL6 levels were determined by ELISA. FLI1 siRNA significantly enhanced CXCL6 mRNA expression in human dermal fibroblasts and human dermal microvascular endothelial cells, while Fli1 haploinsufficiency significantly suppressed CXCL6 mRNA expression in murine peritoneal macrophages stimulated with lipopolysaccharide. Supporting a critical role of Fli1 deficiency to induce SSc-like phenotypes, CXCL6 mRNA expression was higher in SSc dermal fibroblasts than in normal dermal fibroblasts. Importantly, Fli1 bound to the CXCL6 promoter in dermal fibroblasts, endothelial cells, and THP-1 cells. In patients with SSc, serum CXCL6 levels correlated positively with the severity of dermal and pulmonary fibrosis and were elevated in association with cardiac and pulmonary vascular involvement and cutaneous vascular symptoms, including Raynaud phenomenon, digital ulcers (DU)/pitting scars, and telangiectasia. Especially, serum CXCL6 levels were associated with DU/pitting scars and heart involvement by multiple regression analysis. CXCL6 expression is upregulated by Fli1 deficiency in fibroblasts and endothelial cells, potentially contributing to the development of fibrosis and vasculopathy in the skin, lung, and heart of SSc.

Inhibition of DNA Methylation in the COL1A2 Promoter by Anacardic Acid Prevents UV-Induced Decrease of Type I Procollagen Expression

UV radiation decreases type I procollagen production mainly by inhibiting the transforming growth factor-β/Smad signaling pathway. Because further epigenetic regulatory mechanisms are unclear, we investigated the roles of DNA methylation and histone acetylation in UV-induced regulation of COL1A2 transcription in human dermal fibroblasts. Anacardic acid, a p300 histone acetyltransferase inhibitor, rescued the UV-induced decrease of type I procollagen expression in human dermal fibroblasts. Although UV irradiation induced global histone acetylation, it reduced the local recruitment of histone H3 acetylation as well as p300, and Smad2/3 to the p300 binding site (-1406/-1393), in the COL1A2 promoter as shown by chromatin immunoprecipitation. This effect was reversed by anacardic acid treatment. In contrast, pyrosequencing analysis showed that UV irradiation induced DNA methylation in the same region of the COL1A2 promoter, which was reversed by anacardic acid and a DNA methyltransferase inhibitor (5-AZA-2'-deoxycytidine). Inhibition of UV-induced DNA methylation led to an increase of UV-induced histone acetylation in the COL1A2 promoter and increased the recruitment of transcription factors, leading to up-regulation of type I collagen after UV irradiation. Collectively, our findings indicate that the epigenetic crosstalk between DNA methylation and histone acetylation plays a crucial role in COL1A2 transcription induced by UV irradiation.

Enhanced topical delivery of fish scale collagen employing negatively surface-modified nanoliposome

In this study, we investigated the effect of fish scale collage (FSC) on type I procollagen expression in cultured human dermal fibroblasts using real-time polymerase chain reaction and ELISA, under normal and UV exposure condition. Also, FSC encapsulated negatively surface modified nanoliposome (NLsurf mod) was fabricated, and the physicochemical characteristics of the liposomes and skin permeation properties were evaluated. FSC increased type I procollagen mRNA and protein expression in human dermal fibroblasts. The mean particle size, zeta potential value and encapsulation efficiency of the NLsurf mod were about 175 nm, −44 mV and 11%, respectively. The skin localization effect was enhanced by NLsurf mod. Conclusively, FSC is able to elevate the type I procollagen expression in human dermal fibroblasts and FSC-loaded NLsurf mod would be a good candidate for the topical delivery of FSC.