Expression In Human Dermal Fibroblasts Research Articles

Antioxidative and Anti-photooxidative Potential of Interruptins from the Edible Fern Cyclosorus terminans in Human Skin Cells.

<p>Background: Human skin is exposed daily to oxidative stress factors such as UV light, chemical pollutants, and invading organisms. Reactive oxygen species (ROS) are intermediate molecules that cause cellular oxidative stress. In order to survive in an oxygen-rich environment, all aerobic organisms, including mammals, have evolved enzymatic and non-enzymatic defence systems. The interruptins from an edible fern Cyclosorus terminans possess antioxidative properties and can scavenge intracellular ROS in adipose-derived stem cells. <p> Objectives: This study aimed to evaluate the antioxidative efficacy of interruptins A, B, and C in cultured human dermal fibroblasts (HDFs) and epidermal keratinocytes (HEKs). Moreover, the anti-photooxidative activity of interruptins in ultraviolet (UV)-exposed skin cells was investigated. <p> Methods: The intracellular ROS scavenging capacity of interruptins in skin cells was measured by flow cytometry. Their induction effects on gene expression of the endogenous antioxidant enzymes was monitored using real-time polymerase chain reaction. <p> Results: Interruptins A and B, but not interruptin C, were highly effective in ROS scavenging, particularly in HDFs. Interruptins A and B upregulated gene expression of superoxide dismutase (SOD)1, SOD2, catalase (CAT), and glutathione peroxidase (GPx) in HEKs, but they only induced SOD1, SOD2, and GPx gene expression in HDFs. Additionally, interruptins A and B efficiently suppressed UVA- and UVB-induced ROS generation in both HEKs and HDFs. <p> Conclusion: The results suggest that these naturally occurring interruptins A and B are potent natural antioxidants and therefore may have the potential in the future of inclusion in antiaging cosmeceutical products.</p>.

Enzymatic upcycling of wild-simulated ginseng leaves for enhancing biological activities and compound K

Compound K (CK), a ginsenoside with high bioavailability, is present at low levels in wild-simulated ginseng leaves (WSGL). WSGL contains the CK precursors, Rd and F2, in amounts up to 26.4 ± 0.4 and 24.1 ± 1.9 mg/g extract, respectively. In this study, CK production in WGSL reached 25.9 ± 1.0 mg/g extract following treatment with Viscozyme, Celluclast 1.5 L, Pectinex Ultra SP-L, and their combination. The antioxidant activities indicated by oxygen radical absorbance capacity, ferric reducing antioxidant power, and ABTS- and DPPH radical scavenging activity of enzyme-treated WSGL were enhanced 1.69-, 2.51-, 2.88-, and 1.80-fold, respectively, compared to non-treated WSGL. Furthermore, the CK-enriched WSGL demonstrated a 1.94-fold decrease in SA-β-galactosidase expression in human dermal fibroblasts and a 3.8-fold enhancement of inhibition of nitric oxide release in lipopolysaccharide-induced RAW 264.7 cells relative to non-treated WSGL. Consequently, WSGL subjected to enzymatic upcycling has potential as a functional material in the food and pharmaceutical industries.

Effects of pulsed electrical stimulation on α-smooth muscle actin and type I collagen expression in human dermal fibroblasts.

Pulsed electrical stimulation (PES) is known to affect cellular activities. We previously found PES to human dermal fibroblasts (HFs) promoted platelet-derived growth factor subunit A (PDGFA) gene expression, which enhanced proliferation. In this study, we investigated PES effects on fibroblast collagen production and differentiation into myofibroblasts. HFs were electrically stimulated at 4800Hz and 5V for 60min. Imatinib, a specific inhibitor of PDGF receptors, was treated before PES. After 6h of PES, PDGFA, α-smooth muscle actin (α-SMA), and collagen type I α1 chain gene expressions were upregulated in PES group. Imatinib suppressed the promoted expression except for PDGFA. Immunofluorescence staining and enzyme-linked immunosorbent assay showed the production of α-SMA and collagen I was enhanced in PES group but suppressed in PES+imatinib group at 48h after PES. Therefore, PES promotes the production of α-SMA and collagen I in fibroblasts, which is triggered by PDGFA that is upregulated early after PES.

Tamoxifen Upregulates Collagenase Gene Expression in Human Dermal Fibroblasts.

Tamoxifen is a known inhibitor of fibroblast transforming growth factor beta biosynthesis and wound scar formation. Tamoxifen is also known to be an estrogen antagonist and protein kinase C (PKC) inhibitor. Cells treated with tamoxifen and other PKC/calmodulin inhibitors depolymerize their membrane focal adhesion complexes and cytoskeletal protein structures. These effects result in substrate detachment, cell shape rounding, and upregulation of collagenase synthesis and extracellular matrix degradation. The purpose of our study was to test the hypothesis that tamoxifen treatment of human foreskin fibroblasts results in alteration of cytoskeletal protein organization, cell detachment and rounding, and increased collagenase synthesis similar to known PKC/calmodulin inhibitors such as H-7. We characterized the effects of PKC/calmodulin inhibitors tamoxifen and H-7 on human dermal fibroblast morphology, cytoskeletal protein organization, and collagenase gene expression in monolayer culture and within collagen gels. We found that fibroblasts responded to tamoxifen by initiation of actin filament depolymerization followed by alteration from spindle to spheroidal shapes. This change in cell shape led to increased collagenase synthesis in cells treated with either tamoxifen or H-7 compared with controls. There was also a 23% increase of hydroxyproline release from tamoxifen-treated fibroblast-populated collagen matrices. Tamoxifen may reduce scarring by inhibiting fibroblast PKC/calmodulin activity, which down-regulates pro-fibrotic transforming growth factor beta signaling and upregulates collagenase production. These effects mimic those of the known PKC/calmodulin inhibitor H-7. Overall, these findings suggest that tamoxifen and its analogues are promising agents for clinical investigation as small molecule regulators of fibrosis and scarring disorders.

Development of multifunctional pyrrole-imidazole polyamides that increase hepatocyte growth factor and suppress transforming growth factor-β1

PurposeThe DNA recognition peptide compounds pyrrole-imidazole (PI) polyamides bind to the minor groove and can block the binding of transcription factors to target sequences. To develop more PI polyamides as potential treatments for fibrotic diseases, including chronic renal failure, we developed multifunctional PI polyamides that increase hepatocyte growth factor (HGF) and decrease transforming growth factor (TGF)-β1. MethodsWe designed seven PI polyamides (HGF-1 to HGF-7) that bind to the chicken ovalbumin upstream promoter transcription factor-1 (COUP-TF1) binding site of the HGF promoter sequence. We selected PI polyamides that increase HGF and suppress TGF-β1 in human dermal fibroblasts (HDFs). FindingsGel shift assays showed that HGF-2 and HGF-4 bound the appropriate dsDNAs. HGF-2 and HGF-4 significantly inhibited the TGF-β1 mRNA expression in HDFs stimulated by phorbol 12-myristate 13-acetate. HGF-2 and HGF-4 significantly inhibited the TGF-β1 protein expression in HDFs with siRNA targeting HGF, indicating that HGF-2 and HGF-4 directly inhibited the expression of TGF-β1. ConclusionThe designed and synthetic HGF PI polyamides targeting the HGF promoter, which increased the expression of HGF and suppressed the expression of TGF-β, will be a potential practical medicine for fibrotic diseases, including progressive renal diseases.

Pulsed electrical stimulation and amino acid derivatives promote collagen gene expression in human dermal fibroblasts.

Several collagen types are important for maintaining skin structure and function. Previous reports show that l-hydroxyproline (Hyp), N-acetyl-l-hydroxyproline (AHyp), and l-alanyl-l-glutamine (Aln-Gln) are biological active substances with collagen synthesis-promoting effects. In this study, we combined the promotive effects of pulsed electrical stimulation (PES) with three amino acid derivatives in human dermal fibroblasts. Fibroblasts were exposed to PES with a 4,800Hz pulse frequency and a voltage at 1 or 5V for 15min. The gene expression of type I and III collagen (fibrillar collagen), type IV and VII collagen (basement membrane collagen and anchoring fibril collagen) were measured by RT-PCR 48h after PES. PES alone promoted the expression of COL1A1 and COL3A1 at 5V but did not alter that of COL4A1 and COL7A1. Each AAD and the AAD mixture promoted the expression of COL4A1 and COL7A1 but either repressed, or did not alter, that of COL1A1 and COL3A1. Compared to treatment with each AAD, PES at 5V with Hyp promoted the expression of COL1A1 and COL3A1, enhanced COL3A1 expression with AHyp, and stimulated COL3A1 expression with Aln-Gln, while COL4A1 and COL7A1 expressions were not affected. PES and the AAD mixture significantly promoted COL4A1 expression in a voltage-dependent manner, and COL1A1 and COL3A1 demonstrated a similar but nonsignificant trend, whereas COL7A1 expression was not affected. The combination of PES with each AAD or the AAD mixture may improve skin structure and function by increasing the expression of basement membrane collagen and dermal fibrillar collagen.

Gelatin–chitosan–cellulose nanocrystals as an acellular scaffold for wound healing application: fabrication, characterisation and cytocompatibility towards primary human skin cells

Biopolymers that mimic the extracellular matrix are favourable in tissue engineering. However, the rapid degradation and the lack of mechanical and enzymatic stabilities of these biopolymers prompt researchers to composite different biopolymers. In this study, we aim to develop an acellular gelatin-chitosan-cellulose nanocrystal (GCCNC) scaffold as a potential wound dressing. The GCCNC mixture was homogenised via ultrasonication and the genipin crosslinking was performed by magnetic stirring. The mixture was then frozen at − 80 °C for 6 h and freeze-dried. The effects of different ratios of gelatin and chitosan with cellulose nanocrystals on the physiochemical properties, mechanical properties, and cellular biocompatibility were studied. Our results herein showed that G3C7CNC demonstrated a homogenous interconnected porous structure with a good porosity (67.37 ± 9.09%) and pore size (148.46 ± 48.68 µm), acceptable swelling ratio (1071.11 ± 140.26%), adequate water vapour transmission rate (315.59 ± 25.27 g/m2/day), low contact angle (70.21 ± 6.79°), and sufficient mechanical strength (modulus of 64.67 ± 12.42 MPa). The lower biodegradation rate in the G3C7CNC (0.06 ± 0.01 mg/hr) compared to G10CNC (0.48 ± 0.07 mg/hr) together with the absence of glass transition phenomenon indicated an increase in both enzymatic and thermal stabilities. Furthermore, G3C7CNC was non-cytotoxic and biocompatible with human epidermal keratinocytes (HEKs) and human dermal fibroblasts (HDFs). The presence of collagen type I and α-smooth muscle actin expression in HDFs, together with the expression of cytokeratin-14 in HEKs, demonstrated our scaffold’s ability to maintain normal skin physiological functions. Therefore, this study proposes that the fabricated GCCNC scaffold could serve as a potential acellular skin substitute in managing chronic wounds.

1173 Dual role of enhancer of zeste homolog 2 in the regulation of ultraviolet radiation-induced matrix metalloproteinase-1 and type I procollagen expression in human dermal fibroblasts

1173 Dual role of enhancer of zeste homolog 2 in the regulation of ultraviolet radiation-induced matrix metalloproteinase-1 and type I procollagen expression in human dermal fibroblasts

Dual role of enhancer of zeste homolog 2 in the regulation of ultraviolet radiation-induced matrix metalloproteinase-1 and type I procollagen expression in human dermal fibroblasts

Abnormalities in the extracellular matrix (ECM) caused by ultraviolet (UV) radiation are mediated by epigenetic mechanisms. Enhancer of zeste homolog 2 (EZH2) is a histone methyltransferase that is implicated in inflammation, immune regulation, and senescence. However, its role in controlling UV-induced ECM alterations in the skin remains elusive. Here, we investigated the role of EZH2 in UV-induced expression of matrix metalloproteinase (MMP)-1 and type I procollagen. We found that UV induced EZH2 expression in human skin in vivo and in human dermal fibroblasts (HDFs). EZH2 knockdown reduced the expression and promoter activity of MMP-1 and increased those of type I procollagen, whereas EZH2 overexpression had the opposite effects. Mechanistically, EZH2 increased NF-κB activity, and p65 and p50 expression and promoter activity. Intriguingly, chromatin immunoprecipitation assays revealed that the EZH2/p65/p50 complex was recruited and bound to the MMP-1 promoter after UV irradiation, independent of its histone methyltransferase activity. In contrast, EZH2-induced DNA methyltransferase 1 (DNMT1) formed a complex with EZH2 and enhanced the enrichment of H3K27me3 on the COL1A2 promoter following UV irradiation. These findings indicate that EZH2 plays a dual role in regulating MMP-1 and type I procollagen expression and improve our understanding of how this epigenetic mechanism contributes to UV-induced skin responses and photoaging. This study shows that inhibiting EZH2 is a potential anti-aging strategy for preventing UV-induced skin aging by reducing MMP-1 expression and inducing type I procollagen expression.

Effects of ‘NEOGEN V.BIOME LIPOSOME™’ on the Increased Expression of COL1A1 and HAS2 mRNA in Human Dermal Fibroblasts

Purpose: This study investigated functional cosmetic agent for anti-wrinkle effects of 'NEOGEN V.BIOME LIPOSOME™', which is a liposome in the form of a proprietary technology including a vitamin C active ingredient, on human dermal fibroblasts (HDFs).Methods: The cytotoxicity of 'NEOGEN V.BIOME LIPOSOME™' was measured by performing a water-soluble tetrazolium salt (WST-1) assay. The relative mRNA expression of collagen (COL1A1 ) and hyaluronan synthase (HAS2) was measured by quantitative real-time PCR (qRT-PCR) assay.Results: HDF cell viability of 'NEOGEN V.BIOME LIPOSOME™' was no cytotoxic effect at concentrations below 0.25%. The expression of COL1A1 and HAS2 mRNA was increased in a concentration-dependent manner compared with the negative control group.Conclusion: “NEOGEN V.BIOME LIPOSOME™“ helps to increase collagen and hyaluronan synthase production by promoting COL1A1 and HAS2 mRNA expression in HDFs; therefore, it is expected to function as an anti-wrinkle cosmetic material.

Depletion of growth differentiation factor 15 (GDF15) leads to mitochondrial dysfunction and premature senescence in human dermal fibroblasts.

Growth differentiation factor 15 (GDF15) is a stress-responsive cytokine also known as a mitokine; however, its role in mitochondrial homeostasis and cellular senescence remained elusive. We show here that knocking down GDF15 expression in human dermal fibroblasts induced mitochondrial dysfunction and premature senescence, associated with a distinct senescence-associated secretory phenotype. Fibroblast-specific loss of GDF15 expression in a model of 3D reconstructed human skin induced epidermal thinning, a hallmark of skin aging. Our results suggest GDF15 to play a so far undisclosed role in mitochondrial homeostasis to delay both the onset of cellular senescence and the appearance of age-related changes in a 3D human skin model.

Involvement of Aquaporin 1 in the Motility and in the Production of Fibrillin 1 and Type I Collagen of Cultured Human Dermal Fibroblasts

Aminocarbonyl proteins increase with age in the dermal layer. Gene Chip analysis of mRNA expression in human dermal fibroblasts cultured on collagen gels treated with glyceraldehyde as an aminocarbonyl protein and on untreated collagen gels showed a decrease in the amount of aquaporin 1 (AQP1) mRNA. In this study, we clarified the involvement of AQP1 in collagen gel contraction and the production of fibrillin 1 and type I collagen in cultured human dermal fibroblasts. In the experiment, AQP1 siRNA was transfected into cultured human dermal fibroblasts to deplete AQP1, and the cell motility and contractile activity of the collagen gel were assessed. The production of fibrillin 1 and type I collagen was also examined by RT–qPCR and antibody staining. AQP1 depletion decreased the collagen gel contractile activity, and both the amount of mRNA and the antibody staining of fibrillin 1 and type I collagen decreased. Furthermore, the depletion of AQP1 reduced the levels of F-actin and phosphorylated myosin light chain 2, suggesting their involvement in reductions of the motility and collagen gel contractile activity of fibroblasts. These findings suggest that AQP1 is an important biomolecule for cell motility in human dermal fibroblasts and that decreased motility results in decreased expression of extracellular matrix proteins such as fibrillin 1 and type I collagen.

Protective effects of Evodiae Fructus extract against ultraviolet-induced MMP-1 and MMP-3 expression in human dermal fibroblasts

Evodiae Fructus (EF) has been used as a traditional medicine for abdominal pain, headache, bleeding, and menorrhea in Asian countries. However, the effect of EF extract (EFE) on ultraviolet (UV)B-induced photoaging remains unknown. This study aimed to investigate the effects and molecular mechanisms of UVB-induced matrix metalloproteinase (MMP)−1 and MMP-3 expression in primary human dermal fibroblasts (HDFs). In HDFs, the expression of MMP-1, MMP-3 and pro-collagenA2 were analyzed using western blotting and real-time polymerase chain reaction. The phosphorylation levels of mitogen-activated protein kinase (MAPK) and activation of nuclear factor (NF)-κB and activator protein (AP)−1 were assessed using western blotting. Results showed that EFE reduced UVB-induced MMP-1 and MMP-3 expression, and this effect was mediated by the inhibition of MAPK/NF-κB and AP-1 activation. Moreover, EFE prevented UVB-induced inhibition of collagen synthesis. Taken together, these results suggest that EFE may have a preventive potential against photoaging.

Acetylated Resveratrol and Oxyresveratrol Suppress UVB-Induced MMP-1 Expression in Human Dermal Fibroblasts.

Resveratrol (RES) and oxyresveratrol (OXYRES) are considered and utilized as active ingredients of anti-aging skin cosmetics. However, these compounds are susceptible to oxidative discoloration and unpleasant odor in solutions, limiting their use in cosmetics. Accordingly, RES and OXYRES were chemically modified to acetylated derivatives with enhanced stability, and their anti-aging effect on the skin and detailed molecular mechanism of their acetylated derivatives were investigated. Acetylated RES and OXYRES lost their acetyl group and exerted an inhibitory effect on H2O2-induced ROS levels in human dermal fibroblast (HDF) cells. In addition, RES, OXYRES, and their acetylated derivatives suppressed UVB-induced matrix metalloproteinase (MMP)-1 expression via inhibition of mitogen-activated protein kinases (MAPKs) and Akt/mTOR signaling pathways. Furthermore, RES, OXYRES, and their acetylated derivatives suppressed type I collagen in TPA-treated HDF cells. Collectively, these results suggest the beneficial effects and underlying molecular mechanisms of RES, OXYRES, and their acetylated derivatives for anti- skin aging applications.

Atomoxetine and circadian gene expression in human dermal fibroblasts from study participants with a diagnosis of attention-deficit hyperactivity disorder

Atomoxetine (ATO) is a second line medication for attention-deficit hyperactivity disorder (ADHD). We proposed that part of the therapeutic profile of ATO may be through circadian rhythm modulation. Thus, the aim of this study was to investigate the circadian gene expression in primary human-derived dermal fibroblast cultures (HDF) after ATO exposure. We analyzed circadian preference, behavioral circadian and sleep parameters as well as the circadian gene expression in a cohort of healthy controls and participants with a diagnosis of ADHD. Circadian preference was evaluated with German Morningness-Eveningness-Questionnaire (D-MEQ) and rhythms of sleep/wake behavior were assessed via actigraphy. After ex vivo exposure to different ATO concentrations in HDF cultures, the rhythmicity of circadian gene expression was analyzed via qRT-PCR. No statistical significant effect of both groups (healthy controls, ADHD group) for mid-sleep on weekend days, mid-sleep on weekdays, social jetlag, sleep WASO and total number of wake bouts was observed. D-MEQ scores indicated that healthy controls had no evening preference, whereas subjects with ADHD displayed both definitive and moderate evening preferences. ATO induced the rhythmicity of Clock in the ADHD group. This effect, however, was not observed in HDF cultures of healthy controls. Bmal1 and Per2 expression showed a significant ZT × group interaction via mixed ANOVA. Strong positive correlations for chronotype and circadian genes were observed for Bmal1, Cry1 and Per3 among the study participants. Statistical significant different Clock, Bmal1 and Per3 expressions were observed in HDFs exposed to ATO collected from ADHD participants exhibiting neutral and moderate evening preference, as well as healthy participants with morning preferences. The results of the present study illustrate that ATO impacts on circadian function, particularly on Clock, Bmal1 and Per2 gene expression.

Suppression of Matrix Metallopeptidase-3 Expression in Human Dermal Fibroblasts by Decursin from Angelica gigas Nakai Root Extracts Fermented with Jeju Lava Seawater

ëª©ì  최본 연구는 ì œì£¼ 용암해수로 숙성된 참당귀 추출물의 인간 ì§„í”¼ì„¬ìœ ì•„ì„¸í¬ 내 콜라겐분해효소 발현 감소 효과를 ë¶„ì„í•˜ê³ ê·¸ 효과에 대한 지표물질을 íŒŒì• í•˜ëŠ” 것을 ëª©ì ìœ¼ë¡œ 진행되었다. 방법 ì œì£¼ 용암해수를 이용해 ì €ì˜¨ìˆ™ì„±í•˜ì—¬ 추출한 참당귀 추출물의 지표물질 확인을 위해 ê³ ì„±ëŠ¥ 액체 í¬ë¡œë§ˆí† ê·¸ëž˜í”¼(HPLC) 분석을 진행하였다. 참당귀 뿌리 추출물 및 decursin의 인간 ì§„í”¼ì„¬ìœ ì•„ì„¸í¬ 내 세포생존율을 ì¸¡ì •í•˜ê³ ì‹¤ì‹œê°„ ìœ ì „ìžì¦í­ ì •ëŸ‰(qRT-PCR) 분석을 통한 콜라겐분해효소 발현 변화를 분석하였다. Decursin으로 진행한 결과와 참당귀 뿌리 추출물의 콜라겐분해효소 발현 감소 효과를 비교하여 ê·¸ 상관관계를 íŒŒì• í•˜ì˜€ë‹¤. ê²°ê³¼ HPLC 분석 ê²°ê³¼, 참당귀 추출물 내 주요한 지표물질로 decursin이 확인되었다. 인간 ì§„í”¼ì„¬ìœ ì•„ì„¸í¬ì—ì„œ 참당귀 뿌리 추출물과 decursin은 각각 200 μg/mL, 20 μM 미만의 농도에서 세포 생존율에 낮은 영향을 보였다. 또한, MMP3 발현에 대한 참당귀 뿌리 추출물과 decursin의 감소 효과를 qRT-PCR을 통해 확인하였다. ê²°ë¡ ë³¸ 연구결과를 통해 ì œì£¼ 용암해수로 숙성된 참당귀 뿌리 추출물과 이 추출물로부터 순수분리된 지표물질 decursin의 콜라겐분해효소인 MMP3 발현 감소 효능을 확인하였으며, 이를 통해 참당귀 뿌리 추출물 및 decursion은 ì£¼ë¦„ê°œì„ ìš© 기능성화장품 소재로 활용 가능함을 ê³¼í•™ì ìœ¼ë¡œ ì¦ëª í•˜ì˜€ë‹¤. 핵심용어: 참당귀 뿌리, Decursin, 콜라겐분해효소, ì œì£¼ 용암해수, 인간 ì§„í”¼ì„¬ìœ ì•„ì„¸í¬

Artesunate inhibits myofibroblast formation via induction of apoptosis and antagonism of pro-fibrotic gene expression in human dermal fibroblasts.

The anti-malaria drug artesunate and other chemical analogs of artemisinin have demonstrated cytostatic and cytotoxic effects in bacterial and cancer cells. Artemisinin-derived compounds have also been demonstrated to attenuate fibrosis in preclinical animal models, but the mechanisms by which this inhibition occurs are not well-understood. We investigated the effects of artesunate on the emergence of the myofibroblast, which is causally implicated in pro-fibrotic pathologies. CRL-2097 human dermal fibroblasts were analyzed for protein and transcript expression after treatment with artesunate to analyze fibroblast activation. Proliferation and apoptosis were also evaluated following treatment with artesunate in this cell line. Treatment of human dermal fibroblasts with artesunate antagonized fibroblast activation and pro-fibrotic extracellular matrix (ECM) deposition, both at basal culture conditions and when cultured in the presence of exogenous transforming growth factor-β1 (TGF-β1), a major pro-fibrotic cytokine. Artesunate-treated fibroblasts also demonstrated decreased proliferation and increased apoptosis. Transcript analysis by quantitative real-time polymerase chain reaction demonstrated that artesunate downregulated expression of pro-fibrotic genes including canonical myofibroblast markers, ECM genes, and several TGF-β receptors and ligands, and upregulated expression of cell cycle inhibitors and matrix-metalloproteinases. Together, these data demonstrate that artesunate antagonizes fibroblast activation and decreases expression of pro-fibrotic genes, while also promoting myofibroblast apoptosis, suggesting that these mechanisms may be responsible in part for the anti-fibrotic effects of artesunate described previously.

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 &lt; 0 001) and COL1A1 (p &lt; 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 &lt; 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 &lt; 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.