Trifloxystrobin induces tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis in HaCaT,human keratinocyte cells

As the outermost layer of the body, the skin plays an important role in exposure to pesticides, which could have negative impacts on human health. Trifloxystrobin is a widely used fungicide of the strobilurin class, however, there is little information regarding the skin contact-associated toxic mechanism. Therefore, the present study was performed in order to identify the skin toxicity mechanism of trifloxystrobin using HaCaT (keratinocyte of human skin) cells. Following 24 or 48?h treatment, cell viability, and subsequent Annexin V-FITC/propidium iodide assay, TUNEL assay and Western blotting were performed to investigate the cell death mechanism of trifloxystrobin. Exposure to trifloxystrobin resulted in diminished viability of HaCaT cells in both a time- and concentration-dependent manner. The cell death was derived through apoptotic pathways in the HaCaT cells. Furthermore, we explored the effect of trifloxystrobin on TRAIL-mediated extrinsic apoptosis using siRNA transfection. Knockdown of death receptor 5 suppressed trifloxystrobin-provoked apoptosis. These results indicate that trifloxystrobin induces TRAIL-mediated apoptosis and has an inhibitory effect in keratinocytes that can interfere with the barrier function and integrity of the skin. This could be proposed as a mechanism of skin toxicity by trifloxystrobin and considered in the management of pesticide exposure.     

Effect of a Gelatin Hydrogel Incorporating Epiregulin on Human Keratinocyte Growth

Abstract

Ionic hydrogels are biocompatible interesting candidates for tissue-engineering applications, such as the creation of artificial skin, as they can also be used, along with growth factors and cells grown in vitro, for developing bioengineered tissues to be implanted. Among the growth factors that can be used to induce keratinocytes growth in vitro, epiregulin, a broad-specificity epidermal growth factor (EGF) family member, has been shown to be more effective than EGF and transforming growth factor-alpha (TGF-α) in promoting re-epithelization in vitro. To produce a drug-delivery hydrogel for epiregulin, bovine gelatin was cross-linked with poly(glutamic acid) (PLG) in the presence of epiregulin (5–50 ng/ml). Spontaneously immortalized human keratinocytes (HaCaT) were seeded on unloaded and epiregulin-loaded hydrogels and cell adhesion was evaluated after 6 h. Moreover, cell proliferation and stratification, cytokeratins (K5, K10), differentiation markers (filaggrin and transglutaminase-1 (TG-1)) and matrix metalloproteinases (MMP-2, MMP-9 and MMP-28) expression were evaluated after 7 days. The presence of epiregulin induced an increase in cell proliferation, stratification and K5 expression along with MMP-9 and MMP-28 expression, while all differentiation markers expression (K10, filaggrin, TG-1) was decreased. These data indicated that a simple hydrogel loaded with epiregulin could be an effective tool for skin tissue engineering.     

A new tool to test active ingredient using lactic acid in vitro,a help to understand cellular mechanism involved in stinging test: An example using a bacterial polysaccharide (Fucogel®)

The stinging test is an in vivo protocol that evaluates sensitive skin using lactic acid (LA). A soothing sensation of cosmetics or ingredients can be also appreciated through a decrease in stinging score. To predict the soothing sensation of a product before in vivo testing, we developed a model based on an LA test and substance P (SP) release using a co‐culture of human keratinocytes and NGF‐differentiated PC12 cells. A bacterial fucose‐rich polysaccharide present in Fucogel^® was evaluated as the soothing molecule in the in vivo stinging test and our in vitro model. Excluding toxic concentrations, the release of SP was significant from 0.2% of lactic acid for the PC12 cells and from 0.1% of lactic acid for the keratinocytes. When the pH was adjusted to approximately 7.4, LA did not provoke SP release. At these concentrations of LA, 0.1% of polysaccharide showed a significant decrease in SP release from the two cellular types and in co‐cultures without modifying the pH of the medium. In vivo, a stinging test using the polysaccharide showed a 30% decrease in prickling intensity vs the placebo in 19 women between the ages of 21 and 69. Our in vitro model is ethically interesting and is adapted for cosmetic ingredients screening because it does not use animal experimentation and limits human volunteers. Moreover, Fucogel^® reduced prickling sensation as revealed by the in vivo stinging test and inhibits the neurogenic inflammation as showed by our new in vitro stinging test based on SP release.     

Keratinocyte growth factor is an endogenous stimulant of rabbit gastric epithelial cell proliferation and migration in primary culture

Mesenchymal-epithelial interactions are important in the gastric mucosal repair. However, specific factors responsible for such interactions have not been established. In the present study, keratinocyte growth factor (KGF) significantly stimulated proliferation of gastric epithelial cells dose dependently and synergistically with hepatocyte growth factor (HGF), epidermal growth factor (EGF) and insulin. Restitution of gastric epithelial monolayers was also assessed, using a round wound restitution model. Keratinocyte growth factor facilitated the restitution of gastric epithelial cells significantly but did not have any effects on gastric fibroblasts. Keratinocyte growth factor receptor mRNA was expressed by gastric epithelial cells, indicating that these effects were elicited by the specific receptor mediated pathway. Northern blot analysis revealed the expression of KGF mRNA in gastric fibroblasts but not in gastric epithelial cells, indicating the production of KGF. These results suggest that KGF might be involved in gastric mucosal repair, through mesenchymal-epithelial interaction.