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UV irradiation is a major insult to the skin. We have shown previously that exogenous vitamin C (ascorbate) accumulates in HaCaT keratinocytes, thus conferring the ability to prevent radical formation and cell death elicited by UV-B. Here, we have investigated the potential mechanisms accounting for the cytoprotective effects exerted by this antioxidant. Using a cDNA microarray hybridization, we identified several genes whose expression was up-regulated by ascorbate. We focused on the fra-1 gene, a member of the Fos family of transcription factors that down-regulates activator protein-1 (AP-1) target genes. Both in HaCaT and in normal human epidermal keratinocytes, we found Fra-1 mRNA induction as early as 2h after ascorbate loading. Electrophoretic mobility-shift assay and antibody supershift analysis revealed that ascorbate modulates AP-1 DNA-binding activity and that Fra-1 is in AP-1 complexes in treated cells. Furthermore, transient-transfection studies, using an AP-1 reporter construct, showed that ascorbate was able to inhibit both basal and UV-B-induced AP-1-dependent transcription. Ascorbate also modulates UV-B-induced AP-1 activity by preventing the phosphorylation and activation of the upstream c-Jun N-terminal kinase (JNK), thus inhibiting phosphorylation of the endogenous c-Jun protein. These data suggest that ascorbate mediates cellular responses aimed at counteracting UV-mediated cell damage and cell death by interfering at multiple levels with the activity of the JNK/AP-1 pathway and modulating the expression of AP-1-regulated genes.

In the last decade, the neurovascular effects exerted by endocannabinoids (eCBs) have attracted growing interest, because they hold the promise to open new avenues of therapeutic intervention against major causes of death in Western society. Several actions of eCBs are mediated by type-1 (CB₁) or type-2 (CB₂) cannabinoid receptors, yet there is no clear evidence of the presence of these proteins in platelets. To demonstrate that CB₁ and CB₂ are expressed in human platelets, we analyzed their protein level by Western blotting and ELISA, visualized their cellular localization by confocal microscopy, and ascertained their functionality by binding assays. We found that CB₁, and to a lesser extent CB₂, are expressed in highly purified human platelets. Both receptor subtypes were predominantly localized inside the cell, thus explaining why they might remain undetected in preparations of plasma membranes. The identification of authentic CB₁ and CB₂ in human platelets supports the potential exploitation of selective agonists or antagonists of these receptors as novel therapeutics to combat neurovascular disorders. It seems remarkable that some of these substances have been already used in humans to treat disease states.

X-ray photoelectron spectroscopy (XPS) was used to define the chemical composition of the outermost surface layer and the surface modification of a plasma-coated phospho-silicate glass (identified as BVA) when immersed in K-phosphate buffer or in phosphate buffered human albumin solution. Its behavior was compared with that of a soda-lime-based glass (identified as BVH) treated in the same way. The surface % composition of plasma-sprayed glass was consistent with bulk composition. After incubation with buffer, a Ca–P-rich layer developed only on the surface of BVA glass. Human serum albumin was bound reversibly to both glasses maintaining its native state. However, the protein completely covered the BVA glass surface within 24 h, with the formation of a mixed albumin–Ca–P layer, while 4 days incubation was necessary for complete coverage of BVH glass surface. Murine fibroblasts seeded on plasma-coated BVA glass showed a proliferation pattern similar to that of control cells grown on Petri dish, while cells seeded on BVH had more restricted growth. A limited response was induced in polymorphonuclear granulocytes by both bulk glasses powder. In conclusion, the glass identified as BVA has the suitable characteristics of its surface layers to be considered biologically active from both a chemical and a cellular point of view.

Glasses incorporating increasing amounts of bovine serum albumin were prepared by sol-gel techniques from a tetra methoxy silane precursor. The surface of the glass samples was studied by X-ray photoelectron spectroscopy, revealing that the protein is present also in the superficial layer of the silica network. Moreover, the protein is distributed in a dose-dependent way, since the N/Si atomic ratio increases linearly with the albumin concentration in the reaction mixture. Angle-dependent measurements show that the protein distribution occurs homogeneously and is the same at different sampling depths. Protein incorporation in the bulk SiO2 network, with a uniform protein distribution between bulk and surface, is confirmed by infrared spectroscopy measurements, performed both in reflectance and transmittance mode. The reaction with a specific antibody and the adhesivity assay of osteoblastic cells show that embedded albumin present on the glass surface is able to interact with other proteins.

X-ray photoelectron spectroscopy (XPS) was used to define the chemical composition of the outermost surface layer and the surface modification of a plasma-coated phospho-silicate glass (identified as BVA) when immersed in potassium phosphate buffer or in phosphate-buffered human albumin solution. Its behaviour was compared with that of a soda-lime-based glass (identified as BVH) treated in the same way. The surface percentage composition of plasma-sprayed glass was consistent with the bulk composition. After incubation with buffer, a Ca-P-rich layer developed only on the surface of BVA glass. Human serum albumin was bound reversibly to both glasses; however, the protein completely covered the BVA glass surface within 24 h, with the formation of a mixed albumin-Ca-P layer, whereas 4 days of incubation were necessary for complete coverage of the BVH glass surface. The thickness of the organic/inorganic overlayer has also been estimated.