Kaempferol Blocks the Skin Fibroblastic Interleukin 1β Expression and Cytotoxicity Induced by 12-O-tetradecanoylphorbol-13-acetate by Suppressing c-jun N-terminal Kinase

Kaempferol, a bioflavonoid present in fruits and vegetables, has a variety of antioxidant and anti-inflammatory capacities, but the functional role of kaempferol in oxidative skin dermal damage has yet to be well studied. In this study, we examine the role of kaempferol during the inflammation and cell death caused by 12-O-tetradecanoylphorbol-13-acetate (TPA) in normal human dermal fibroblasts (NHDF). TPA (5 μM) significantly induced cytotoxicity of NHDF, where a robust increase in the interleukin (IL)-1β mRNA among the various pro-inflammatory cytokines. The skin fibroblastic cytotoxicity and IL-1β expression induced by TPA were significantly ameliorated by a treatment with 100 nM of kaempferol. Kaempferol blocked the production of the intracellular reactive oxygen species (ROS) responsible for the phosphorylation of c-jun N-terminal kinase (JNK) induced by TPA. Interestingly, we found that kaempferol inhibited the phosphorylation of nuclear factor-kappa B (NF-κB) and the inhibitor NF-κB (IκBα), which are necessary for the expression of cleaved caspase-3 and the IL-1β secretion in TPA-treated NHDF. These results suggest that kaempferol is a functional agent that blocks the signaling cascade of the skin fibroblastic inflammatory response and cytotoxicity triggered by TPA.

Efficacy of Chininum Sulphuricum 30C against Malaria: An in vitro and in vivo Study

<b><i>Background:</i></b> New effective, economical and safe antimalarial drugs are urgently needed due to the development of multi-drug-resistant strains of the parasite. Homeopathy uses ultra-diluted doses of various substances to stimulate autoregulatory and self-healing processes to cure various ailments. The aim of the study was to evaluate the in vitro and in vivo antimalarial efficacy of a homeopathic drug, Chininum sulphuricum 30C. <b><i>Methods:</i></b> In vitro antiplasmodial activity was screened against the <i>P. falciparum</i> chloroquine-sensitive (3D7) strain, and cell viability was assessed against normal human dermal fibroblasts and HepG2 cells. Suppressive, preventive and curative studies were carried out against <i>P. berghei</i>-infected mice in vivo. <b><i>Results:</i></b> Chininum sulphuricum (30C) revealed good antiplasmodial activity in vitro, with 92.79 ± 6.93% inhibition against the 3D7 strain. The cell viability was 83.6 ± 0.6% against normal human dermal fibroblasts and 95.22 ± 5.1% against HepG2 cells. It also exhibited suppressive efficacy with 95.56% chemosuppression on day 7 with no mortality throughout the follow-up period of 28 days. It also showed preventive activity against the disease. Drug treatment was also safe to the liver and kidney function of the host as evidenced by biochemical studies. <b><i>Conclusion:</i></b> Chininum sulphuricum 30C exhibited considerable antimalarial activity along with safety to the liver and kidney function of the host.

Attachment and Growth of Fibroblast Cells on Poly (2-Methoxyethyl Acrylate) Analog Polymers as Coating Materials

The regulation of adhesion and the subsequent behavior of fibroblast cells on the surface of biomaterials is important for successful tissue regeneration and wound healing by implanted biomaterials. We have synthesized poly(ω-methoxyalkyl acrylate)s (PMCxAs; x indicates the number of methylene carbons between the ester and ethyl oxygen), with a carbon chain length of x = 2–6, to investigate the regulation of fibroblast cell behavior including adhesion, proliferation, migration, differentiation and collagen production. We found that PMC2A suppressed the cell spreading, protein adsorption, formation of focal adhesion, and differentiation of normal human dermal fibroblasts, while PMC4A surfaces enhanced them compared to other PMCxAs. Our findings suggest that fibroblast activities attached to the PMCxA substrates can be modified by changing the number of methylene carbons in the side chains of the polymers. These results indicate that PMCxAs could be useful coating materials for use in skin regeneration and wound dressing applications.

Lumenato protects normal human dermal fibroblasts from neutrophil-induced collagen-3 damage in co-cultures

Collagen is the major structural protein in the extracellular matrix of skin produced by fibroblasts. UV exposure results in infiltration of neutrophils within the epidermis and dermis, inducing collagen damage and contributing to the process of photo-aging. Collagen-3 is an integral structural component with collagen-1, and is an important regulator of collagen-1 fibrillogenesis. Addition of neutrophils activated with TNFα to normal human dermal fibroblast cultures, but not their supernatant, caused significant collagen-3 damage. To study whether Lumenato can protect from collagen-3 damage, it was added to co-cultures of Normal human dermal fibroblasts and neutrophils activated with TNFα. Lumenato prevented collagen-3 damage induced by activated neutrophils in a dose-dependent manner in the co-cultures. Lumenato also induced a low rate of collagen-3 synthesis in a dose-dependent manner detected by pro-collagen-3 secretion, but did not affect fibroblast cell number. Although Lumenato inhibited MMP-8, MMP-9, and elastase secreted from neutrophils, its main effect was in inhibiting both NADPH oxidase-producing superoxides and MPO activity-producing halides in a dose-dependent manner that correlated with protection from collagen-3 damage. In conclusion, the results suggest that Lumenato induces low levels of collagen-3 that may contribute for skin health and is very effective in defending the co-cultures from collagen-3 damage by inhibiting free radicals secreted from neutrophils, thus, indicating Lumenato's possible potential for skin protection.

IGF-1 Upregulates Biglycan and Decorin by Increasing Translation and Reducing ADAMTS5 Expression

Proteoglycan (PG) is a glycosaminoglycan (GAG)-conjugated protein essential for maintaining tissue strength and elasticity. The most abundant skin PGs, biglycan and decorin, have been reported to decrease as skin ages. Insulin-like growth factor-1 (IGF-1) is important in various physiological functions such as cell survival, growth, and apoptosis. It is well known that the serum level of IGF-1 decreases with age. Therefore, we investigated whether and how IGF-1 affects biglycan and decorin. When primary cultured normal human dermal fibroblasts (NHDFs) were treated with IGF-1, protein levels of biglycan and decorin increased, despite no difference in mRNA expression. This increase was not inhibited by transcription blockade using actinomycin D, suggesting that it is mediated by IGF-1-induced enhanced translation. Additionally, both mRNA and protein expression of ADAMTS5, a PG-degrading enzyme, were decreased in IGF-1-treated NHDFs. Knockdown of ADAMTS5 via RNA interference increased protein expression of biglycan and decorin. Moreover, mRNA and protein expression of ADAMTS5 increased in aged human skin tissues compared to young tissue. Overall, IGF-1 increases biglycan and decorin, which is achieved by improving protein translation to increase synthesis and preventing ADAMTS5-mediated degradation. This suggests a new role of IGF-1 as a regulator for biglycan and decorin in skin aging process.

Effect of Ultraviolet Radiation on the Expression of Drug Transporters in In Vitro Skin Models

The majority of skin cancers are caused by over exposure to ultraviolet (UV) radiation. The effects of UV radiation on the expression of drug transporters expressed in human skin has never been studied. In this the effects of UVA and UVB irradiation on the expression of ATP-binding cassette (ABC) transporters and Solute carrier (SLC) transporters was evaluated in normal human epidermal keratinocytes (NHEK) and normal human dermal fibroblasts (NHDF) in primary culture. First experiments were intended to validate the inflammatory reaction in response to stimulation by lipopolysaccharide (LPS) in NHEK, NHDF and 3D-reconstructed human epidermis (3D-RHE) model. LPS treatment has shown to increase the expression of IL-8 and TNF-alpha in all three in vitro models. Expression of the most expressed ABC and SLC transporters was then measured in NHEK and NHDF after UVA (30 J/m&sup2;) and UVB (40 mJ/m&sup2;) irradiation. The most striking result was a significant 29-fold increase of the expression of SLCO4A1 in normal human dermal fibroblasts. In summary, this study shows for the first time a significant regulation of the expression of SLCO4A1 in human dermal fibroblasts induced by UVA irradiation. This finding is of particular interest as most of skin cancers are caused by over exposure to ultraviolet radiation and need to be considered in pharmacokinetic evaluation of topical drugs.

PLGA Membranes Functionalized with Gelatin through Biomimetic Mussel-Inspired Strategy

Electrospun membranes have been widely used as scaffolds for soft tissue engineering due to their extracellular matrix-like structure. A mussel-inspired coating approach based on 3,4-dihydroxy-DL-phenylalanine (DOPA) polymerization was proposed to graft gelatin (G) onto poly(lactic-co-glycolic) acid (PLGA) electrospun membranes. PolyDOPA coating allowed grafting of gelatin to PLGA fibers without affecting their bulk characteristics, such as molecular weight and thermal properties. PLGA electrospun membranes were dipped in a DOPA solution (2 mg/mL, Tris/HCl 10 mM, pH 8.5) for 7 h and then incubated in G solution (2 mg/mL, Tris/HCl 10 mM, pH 8.5) for 16 h. PLGA fibers had an average diameter of 1.37 ± 0.23 µm. Quartz crystal microbalance with dissipation technique (QCM-D) analysis was performed to monitor DOPA polymerization over time: after 7 h the amount of deposited polyDOPA was 71 ng/cm2. After polyDOPA surface functionalization, which was, also revealed by Raman spectroscopy, PLGA membranes maintained their fibrous morphology, however the fiber size and junction number increased. Successful functionalization with G was demonstrated by FTIR-ATR spectra, which showed the presence of G adsorption bands at 1653 cm−1 (Amide I) and 1544 cm−1 (Amide II) after G grafting, and by the Kaiser Test, which revealed a higher amount of amino groups for G functionalized membranes. Finally, the biocompatibility of the developed substrates and their ability to induce cell growth was assessed using Neonatal Normal Human Dermal Fibroblasts.