Proliferation Of Human Skin Fibroblasts Research Articles

Synthesis and characterization of electrically conducting polymers for regenerative engineering applications: sulfonated ionic membranes

Conductive polymers have found extensive application in fuel cells, sensors and more recently as scaffolds for tissueand organ regeneration. Scaffolds that can transmit electrical impulses have been shown to be beneficial inregeneration of tissues like muscle and nerve that are electroactive in nature. Most cellular events and cellfunctions are regulated by ion movement, and their imbalance is the cause of several diseases. We report synthesisand characterization of sulfonated polymers of poly(methyl vinyl ether-alt-maleic anhydride) (PMVEMA), poly(etherether ketone) (PEEK), poly(ether sulfone) (PES) and poly(phenylene oxide) (PPO) and evaluate their potential for tis-sue regeneration. The ionic conductive property stems from the presence of sulfonic groups on the polymer back-bone. The structure of the polymer was confirmed using Fourier Transform Infrared Spectroscopy and membranehydrophicity was determined by water contact angle measurement. The electrical conductivity of these sulfonatedmembranes was found to be 53.55, 35.39 and 29.51mS/cm for SPPO, SPEEK and SPMVEMA, respectively. Theconductivity was directly proportional to the sulfonic acid content on the polymer backbone. The ionic membranesnamely SPPO, SPEEK and SPMVEMA demonstrated superior cell adhesion properties (~7–10 fold higher) than cellsseeded onto tissue culture polystyrene. The sulfonated membranes exhibited static water contact angle in the rangeof 70–76°. The membranes supported the proliferation of human skin fibroblasts over 14days in culture asevidencedbyconfocaland electronmicroscopyimaging.Theionicmaterials reportedinthisstudymayserveasscaf-folds for a variety of tissue healing and drug delivery applications. Copyright © 2014 John Wiley & Sons, Ltd.Keywords: Electrically conducting polymers; Ionic conductance; Nerve regeneration; Muscle Regeneration; Biocompatibility

The performance of an orthosilicic acid-releasing silica gel fiber fleece in wound healing

In the present work, we have examined the impact of an inorganic orthosilicic acid-releasing spun fiber fleece (SIFIB) on wound closure in a porcine wound model in vivo as well as on wound healing-relevant parameters in vitro. In vivo SIFIB was completely bio-degradable and had no negative effects on wound closure or the wound healing process. In the in vitro experiments, SIFIB had no negative effects on proliferation of human skin fibroblast (FB) and endothelial cell (EC) cultures but strongly retarded the growth of the human monocyte cell line THP-1, and effectively inhibited human skin keratinocyte (KC) proliferation, which based on significantly enhanced KC differentiation. Furthermore, SIFIB exhibited strong anti-inflammatory properties, which based on SIFIB-dependent inhibition of expression and activity of NF-кB and/or concomitant enhanced expression of IкB, a NF-кB-inhibiting protein. Additionally, SIFIB significantly inhibited TGFβ-induced fibroblast differentiation and collagen synthesis as well as effectively reduced TGF-β synthesis of activated fibroblasts. We have demonstrated wound healing-relevant biological activities of a silica-based bio-degradable inorganic material, which might represent a new therapeutic tool in the treatment of chronic wounds.

Synthesis of strong polycations with improved biological properties

Poly(allylamine hydrochloride) (PAH) has found many applications both in biotechnology and biomedical fields. However, its high toxicity toward various mammalian cells significantly limits its effective usage. This study focuses on improving the biological properties of PAH by its modification to strong polyelectrolytes. The strong polycations were prepared by the direct quaternization of PAH amino groups or by the attachment of glycidyltrimethylammonium chloride to these groups. The biological properties, such as cytotoxicity toward human skin fibroblasts (HSFs), proliferation and migration of the cells on a polymeric surface, and antibacterial activities against two pathogenic bacteria, Staphylococcus aureus and Escherichia coli, were determined. All the modified polyelectrolytes are considerably less toxic to HSFs as compared to PAH. Moreover, the directly quaternized polycations are stronger biocides against S. aureus than the parent polymer. Contrary to PAH, thin films of the modified polyelectrolytes improve or do not affect HSFs proliferation and can stimulate cell migration into the wound, as was demonstrated using an in vitro model. The relationship between the structure of the modified polymers (amount and localization of the quaternary ammonium groups) and the biological activity is discussed. Due to the improved biological properties, the obtained polycations may be potentially useful for a variety of biotechnological and biomedical applications.

Downregulation of type I collagen expression in silibinin-treated human skin fibroblasts by blocking the activation of Smad2/3-dependent signaling pathways: Potential therapeutic use in the chemoprevention of keloids

The inhibition of the Smad2/3 pathway is a key step involved in the downregulation of type I collagen synthesis, thus preventing keloid formation in tissue. In this study, we investigated the effect of silibinin on the proliferation of human skin fibroblasts (HSFs), as well as its effect on the expression of type I collagen, matrix metalloproteinase (MMP)-1, Smad2 and Smad3. Our results showed that the proliferation rates of the fibroblasts were not markedly decreased in a dose- and time-dependent manner following treatment with silibinin. Even though silibinin did not exert any cytotoxic effects on HSFs, the expression of type I collagen was markedly decreased in a dose- and time-dependent manner in the silibinin-treated HSFs. Consistent with this finding, the decreased promoter activity of type I collagen was observed in the HSFs following treatment with silibinin. The MMP-1 and MMP-2 expression levels were increased in the silibinin-treated HSFs. Moreover, the silibinin-induced downregulation of type I collagen was associated with the inhibition of Smad2/3 activation in the transforming growth factor‑β1 (TGF-β1)-treated HSFs. We further demonstrated that silibinin attenuated the translocation of Smad2 and Smad3 to the nucleus in the TGF-β1-treated HSFs. Taken together, our data indicate that silibinin has the potential to prevent fibrotic skin changes by inducing the downregulation of type I collagen expression; this effect was partly mediated by the inhibition of the Smad2/3-dependent signaling pathway in HSFs.

The effects of low-level laser irradiation on cellular viability and proliferation of human skin fibroblasts cultured in high glucose mediums

Delayed wound healing is one of the most challenging complications of diabetes mellitus (DM) in clinical medicine. This study has aimed to evaluate the effects of low-level laser therapy (LLLT) on human skin fibroblasts (HSFs) cultured in a high glucose concentration. HSFs were cultured either in a concentration of physiologic glucose (5.5 mM/l) or high glucose media (11.1 and 15 mM/l) for either 1 or 2 weeks after which they were subsequently cultured in either the physiologic glucose or high concentration glucose media during laser irradiation. LLLT was carried out with a helium-neon (He-Ne) laser unit at energy densities of 0.5, 1, and 2 J/cm(2), and power density of 0.66 mW/cm(2) on 3 consecutive days. HSFs' viability and proliferation rate were evaluated with the dimethylthiazol-diphenyltetrazolium bromide (MTT) assay. The LLLT at densities of 0.5 and 1 J/cm(2) had stimulatory effects on the viability and proliferation rate of HSFs cultured in physiologic glucose (5.5 mM/l) medium compared to their control cultures (p = 0.002 and p = 0.046, respectively). All three doses of 0.5, 1, and 2 J/cm(2) had stimulatory effects on the proliferation rate of HSFs cultured in high glucose concentrations when compared to their control cultures (p = 0.042, p = 0.000, and p = 0.000, respectively). This study showed that HSFs originally cultured for 2 weeks in high glucose concentration followed by culture in physiologic glucose during laser irradiation showed enhanced cell viability and proliferation. Thus, LLLT had a stimulatory effect on these HSFs.

Antioxidant activity, total phenolic content and skin care properties of 35 selected plants from Galicia (NW Spain)

In order to contribute to the knowledge of plants from Galician (northwest of the Iberian Peninsula), in the present study, 35 Galician endemic and subendemic plants were examined for their antioxidant activity, total phenolic composition, UV absorption, and skin care properties. The radical scavenging capacity, ferric-reducing antioxidant power, protection against oxidation of the β-carotene/linoleic acid emulsion, antiradical protection of irradiated human skin fibroblast were used to evaluate the antioxidant activity. In several cases, the results of this study strongly indicate in vitro antioxidant activity, which may be due to the presence of a high total phenolic content. Antiradical protection of irradiated human skin fibroblast, stimulation of human skin fibroblast proliferation, and significant absorption of UV radiation were also observed. In order to identify active principles, some extracts were submitted to fractionation and genistein was isolated from Pterospartum tridentatum subsp. tridentatum. The results suggest that some Galician plants extracts have promising skin care properties and could be used as natural antioxidants in cosmetics, pharmaceutical, and food materials.

Influence of RNA interference-induced repression of p53 expression on senescence in human skin fibroblasts

Objective To establish a cell line with repressed expression of p53 by transfecting a plasmid construct expressing short hairpin RNA(shRNA)targeting p53 into human skin fibroblasts(HSFs),and to evaluate the effect of repression of p53 expression on the senescence in HSFs.Methods The eukaryotic expressing plasmid pGCsi-p53 containing shRNA targeting p53 gene was transfected into HSFs with lipofectamine.Subsequently,the cells were selected by G418,and resistant cell clones were chosen and expanded.Reverse transcription-PCR and real time fluorescence-based quanitative PCR were performed to determine the expression of p53 gene,and Western blot to detect the expression of p53 protein in HSFs.The senescence in HSFs was evaluated by SA β-gal staining,and cell proliferation by methyl thiazolyl tetrazolium(MTT)assay.Results A HSF clone with repressed expression of p53 was established successfully.The expressions of p53 mRNA and protein were downregulated in transfected HSFs compared with untransfected HSFs(0.09 ± 0.03 vs.0.32 ± 0.04,0.11 ± 0.04 vs.0.84 ± 0.05,both P ＜ 0.01).The percentage of senescent cells was 13.47％ ± 1.01％ in the transfected HSFs,significantly lower than that in untransfected HSFs(18.10％ ± 0.66％,P ＜ 0.05).As MTT assay showed,the proliferation was accelerated in transfected HSFs compared with untransfected HSFs(P ＜ 0.05).Conclusions The repression of p53 expression decelerates the senescence in HSFs,but promotes the proliferation of HSFs. Key words: Fibroblasts; Cell aging; Genes,p53; RNA interference

ChemInform Abstract: Constituents from the Roots of Taraxacum platycarpum and Their Effect on Proliferation of Human Skin Fibroblasts.

AbstractA variety of triterpenes, sesquiterpene glycosides and a phenylpropyl glycoside, including eight new triterpenes such as (I)—(V) and sequiterpene glycoside (VI) are isolated.

Constituents from the Roots of Taraxacum platycarpum and Their Effect on Proliferation of Human Skin Fibroblasts

A MeOH extract from the roots of Taraxacum platycarpum has shown significant effects on the proliferation of normal human skin fibroblasts. Chemical analysis of the extract resulted in the isolation of 26 compounds, including eight new triterpenes, one new sesquiterpene glycoside, and seventeen known compounds. The structure of each new compound was established using NMR spectroscopy. Some triterpenes had a significant effect on the proliferation of normal human skin fibroblasts.

Effect of PRX-2 gene transferred by lipofectamine on the proliferation of human skin fibroblasts

To explore the effects of PRX-2 gene transferred by lipofectamine on the human skin fibroblasts. Normal human skin fibroblasts were cultured and PRX-2 gene was transferred by lipofectamine. The proliferation of fibroblasts was detected by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) and flow cytometry. The proliferation of PRX-2-transfected fibroblasts was stronger than that of normal counterparts. There were fewer cells during G0-G1 period and more cells during S and G2-M periods. The proliferative index increased. The proliferation of fibroblasts may be modified by transfected PRX-2. Thus PRX-2 plays an important role during the healing of human skin wound.

Protective effect against oxygen reactive species and skin fibroblast stimulation of Couroupita guianensis leaf extracts

In this study, hydroalcoholic leaf extracts of Couroupita guianensis were examined for antioxidant activity, phytochemical and total phenolic composition, stimulation of human skin fibroblast (HSF) proliferation and UV-absorption. The radical scavenging capacity, reducing power and protection against joint oxidation of linoleic acid and β-carotene bleaching oxidation in emulsion were used to evaluate the antioxidant activity. The results of this study strongly indicate in vitro antioxidant activity, which may be due to the presence of a high total phenolic content. In order to identify active principles, the extracts were submitted to fractionation and the compounds isolated were the flavonoids 2′,4′-dihydroxy-6′-methoxy-3′,5′-dimethylchalcone (1), 7-hydroxy-5-methoxy-6,8-dimethylflavanone (2) and the phenolic acid 4-hydroxybenzoic acid (3). In addition, a high level of stimulation of HSF proliferation and significant absorption of UV radiation were also observed. The results suggest that the hydroalcoholic leaf extracts of C. guianensis have promising skin care properties.

8,12;8,20-Diepoxy-8,14-secopregnane glycosides from roots of Asclepias tuberosa and their effect on proliferation of human skin fibroblasts

A pregnane glycoside fraction from the roots of Asclepias tuberosa L. caused normal human skin fibroblasts to proliferate. This fraction contained 21 pregnane glycosides whose structures were established using NMR spectroscopic analysis and chemical evidence. The aglycones of most of these compounds were identified as 8,12;8,20-diepoxy-8,14-secopregnanes, such as tuberogenin or 5,6-didehydrotuberogenin, the same aglycones as constituents of the aerial parts of this plant. Some of these compounds also caused proliferation of skin fibroblasts.

Elastin peptides prepared from piscine and mammalian elastic tissues inhibit collagen‐induced platelet aggregation and stimulate migration and proliferation of human skin fibroblasts

We obtained pure elastin peptides from bovine ligamentum nuchae, porcine aorta, and bonito bulbus arteriosus. The inhibitory activity of these elastin peptides on platelet aggregation induced by collagen and the migratory and proliferative responsivenesses of human skin fibroblasts to these elastin peptides were examined. All of bonito, bovine, and porcine elastin peptides found to inhibit platelet aggregation, but bonito elastin peptides showed a higher inhibitory activity than bovine and porcine elastin peptides did. All elastin peptides enhanced the proliferation of fibroblasts 3.5- to 4.5-fold at a concentration of 10 µg/ml. Bovine and porcine elastin peptides stimulated the migration of fibroblasts, with the optimal response occurring at 10(-1) µg/ml, while maximal response was at 10(2) µg/ml for bonito elastin peptides. Furthermore, pretreatment of fibroblasts by lactose depressed their ability to migrate in response to all elastin peptides, suggesting the involvement of elastin receptor in cell response. These results suggest that both mammalian and piscine elastin peptides can be applied as useful biomaterials in which elasticity, antithrombotic property, and the enhancement of cell migration and proliferation are required.

Hyaluronan Used for Intradermal Injections Is Incorporated into the Pericellular Matrix and Promotes Proliferation in Human Skin Fibroblasts in vitro

Background: Over the past decade injectable hyaluronan (HA) formulations have been widely used to decrease the visibility of skin aging. However, little basic research has been performed to address their effect on dermal skin fibroblasts. Objective: The aim of the present study was to investigate whether human skin fibroblasts are affected by exogenous non-cross-linked HA with respect to proliferation, migration and extracellular matrix composition. Methods: The effect of a non-cross-linked HA on proliferation, migration and gene expression of human dermal fibroblasts was determined. Furthermore, affinity histochemistry of pericellular HA was performed. Results: Proliferation was significantly stimulated by HA whereas migration was not affected. Importantly, exogenous HA was incorporated into fine HA filaments of the pericellular fibroblast matrix. Conclusion: This is the first evidence that a HA formulation used in humans for cosmetic reasons stimulates fibroblast proliferation and is utilized to assemble a supramolecular pericellular structure.

Proliferation of fibroblasts cultured on a hemi-cellulose dressing

In recent times, hemi-cellulose dressing (HD) has been used clinically with satisfactory rates of success [Melandri D, De Angelis A, Orioli R, et-al. Use of a new hemicellulose dressing (Veloderm) for the treatment of split-thickness skin graft donor sites A within-patient controlled study. Burns 2006 Dec;32:964-72.]; however, the effect of cellulose dressings on the wound-healing process is unclear due to paucity of experimental data. This study aimed to determine the adhesion and proliferation of human skin fibroblasts, which were cultured in vitro using the explant technique, on HD. Cells were seeded onto HD discs and evaluated for cell adhesion and cell proliferation after 7, 14 and 21 days. Fibroblasts displayed 70% adhesion to HD after 24h. The HD discs seeded with a density of 5x10(4) cells per well showed a proliferation rate of 12% on day 7, 30% on day 14 and 75% on day 21. The results demonstrated that HD can sustain fibroblast proliferation--a highly desirable characteristic for an ideal skin substitute.

Ferrous Ferric Chloride Stimulates the Proliferation of Human Skin Keratinocytes, Melanocytes, and Fibroblasts in Culture

Ferrous ferric chloride (FFC®) is a distinct form of aqueous iron made up of a complex of ferrous chloride and ferric chloride that participates in both oxidation and reduction reactions. Although FFC is known to stimulate cellular function, its precise mechanism of action is largely unknown. My previous study showed that FFC stimulated the proliferation of cultured epidermal keratinocytes and melanocytes derived from newborn mice, suggesting that FFC stimulates their function of mice. However, it is not known whether FFC stimulates the proliferation of keratinocytes and melanocytes derived from human skin. Moreover, it is unknown whether FFC stimulates the proliferation of human skin fibroblasts. Pairogen® (Akatsuka Co., Tsu, Japan) is a drink comprising FFC, vinegars, and vitamins, Pairogen Gold® (Akatsuka) contains herbal medicines in addition to FFC, vinegars, and vitamins, while Noncalorie Pairogen® (Akatsuka) contains low molecular weight collagen and hyaluronic acid in addition to FFC, vinegars and vitamins. To clarify whether FFC in addition to the natural factors included in these drinks stimulates the proliferation of human skin cells, FFC and these different drinks were added to culture media and tested for their proliferation-stimulating activity on skin cells. FFC, Pairogen, and Pairogen Gold equally doubled the growth rate of cultured human keratinocytes, melanocytes, and fibroblasts. Non-calorie Pairogen tripled the growth rate of human fibroblasts, but Non-calorie Pairogen only doubled the growth rate of human keratinocytes and melanocytes. FFC plus low molecular weight collagen tripled the growth rate of human fibroblasts, whereas FFC plus low molecular weight hyaluronic acid only doubled the growth rate of human fibroblasts. Low molecular weight collagen, with or without hyaluronic acid failed to stimulate the proliferation of fibroblasts. These results suggest that FFC stimulates the proliferation of human skin keratinocytes and melanocytes. Moreover, FFC markedly stimulates the proliferation of human skin fibroblasts synergistically with low molecular weight collagen.

In situ polymerization of tropoelastin in the absence of chemical cross-linking

Tropoelastin, the polypeptide monomer precursor of elastin, is covalently cross-linked to give stable elastic structures. We show here that elastic biomaterials can be generated from tropoelastin in the absence of the classically accepted cross-linking pathway. Under alkaline conditions tropoelastin proceeds through a sol–gel transition leading to the formation of an irreversible hydrogel. This does not occur at neutral pH. The resulting biomaterial is stable, elastic and flexible. Scanning electron microscopy revealed that the hydrogel forms through the coalescence of ∼1 μm quantized protein spheres. These spheres resemble the tropoelastin-rich globules that accumulate on cultured cell surfaces during elastin formation. In vitro cell culture studies demonstrate that the hydrogel can support human skin fibroblast proliferation. In vivo studies demonstrate that following injection, the tropoelastin solution undergoes rapid localized gelation to form a persistent mass. These subcutaneous rodent injection data establish the material's potential as a novel cell-compatible elastic scaffold that can be formed in situ.

Effect of γδ T cell supernatant on human skin fibroblast proliferation and collagen production – Possible role of transforming growth factor‐β and basic fibroblast growth factor

Perivascular mononuclear cell infiltration, vascular injury and diffuse interstitial fibrosis has been shown in systemic sclerosis (SSc). Objective To study a role of cellular immune mechanism inducing fibroblast proliferation and collagen synthesis. The interaction of gammadelta T-cell receptor bearing lymphocyte supernatant to human fibroblast proliferation and collagen synthesis was studied. The fibroblasts added with gammadelta T cell supernatant showed increased proliferation compared with those without it and that added with alphabeta supernatant. The endothelial cells added with gammadelta T cell supernatants, however, showed significantly decreased proliferation compared with those without it and that added with alphabeta T cell supernatant. The fibroblasts added with endothelial cell supernatant showed increased proliferation compared with those without it. A doubling of collagen synthesis was observed when human skin fibroblasts were maintained in all of the supernatant. The collagen synthesis was inhibited by antitransforming growth factor-beta antibody and antibasic fibroblast growth factor antibody. gammadelta T-cell-receptor bearing lymphocyte has proliferative and collagen synthesis activity to human skin fibroblasts, but suppressive to endothelial cells.

Parthenolide Inhibits Proliferation of Fibroblast-Like Synoviocytes In Vitro

Parthenolide is a bioactive constituent of an aromatic herb Feverfew (Tanacetum parthenium). It has been found that both parthenolide and extract of feverfew have anti-inflammatory and antinociceptive properties. Moreover, they demonstrate antiproliferative activities on different human tumour cells. The massive hyperplasia of synovial fibroblasts is the one of the most striking features of rheumatoid arthritis. It is not known whether this is due to the proliferation of synovial fibroblasts or to defective apoptosis. We investigated the effect of parthenolide on the proliferation of rabbit synoviocytes cell line HIG-82, rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) and human skin fibroblasts (HSF) in vitro. Cell proliferation was assessed by means of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and 5'-bromo-2'-deoxy-uridine methods. Parthenolide inhibited proliferation of HIG-82 and human RA-FLS. The proliferation of HSF was inhibited less effectively. The antiproliferative potential of parthenolide was demonstrated.

Inhibitors of Dipeptidyl Peptidase IV-Like Activity Mediate Antifibrotic Effects in Normal and Keloid-Derived Skin Fibroblasts

Suppression of collagen and matrix synthesis and inhibition of the fibrogenic cytokine transforming growth factor-beta(1) (TGF-beta(1)) is a major therapeutic goal in the treatment of fibrosis and keloids. Inhibitors of dipeptidyl peptidase IV (DP IV)-like activity affect cell growth and cytokine production and are currently under investigation for the treatment of metabolic, autoimmune and inflammatory diseases. We show here that the inhibitors of DP IV-like activity, Lys[Z(NO(2))]-thiazolidide and Lys[Z(NO(2))]-pyrrolidide, suppress proliferation in human skin fibroblasts and keloid-derived skin fibroblasts in vitro. They significantly decrease TGF-beta(1) expression and secretion of procollagen type I C-terminal peptide in supernatants of both cell types. Furthermore, they abrogate the TGF-beta(1)-induced stimulation of collagen synthesis, matrix deposition, and TGF-beta(1) and fibronectin expression. Both inhibitors lead to dephosphorylation of mitogen-activated protein kinases pp38 and pERK1/2, which are activated upon TGF-beta1 stimulation and have been implicated in fibrogenesis. In a mouse model of dermal fibrosis, induced by repetitive intracutaneous injections of TGF-beta(1), the profibrotic effect of TGF-beta(1) detected by dermal thickening, collagen I, and alpha-smooth muscle actin expression, is significantly suppressed in the presence of inhibitors. Inhibition of DP IV-like enzymatic activity may therefore represent a promising therapeutic approach for the treatment of fibrotic skin disorders and keloids.