Ultraviolet B irradiation of human skin induces an angiogenic switch that is mediated by upregulation of vascular endothelial growth factor and by downregulation of thrombospondin-1.

BACKGROUND: We have previously demonstrated that skin-specific overexpression of the endogenous angiogenesis inhibitor thrombospondin (TSP)-1 prevented chronic ultraviolet (UV) B-induced angiogenesis, inflammatory cell infiltration and cutaneous photodamage in mice. OBJECTIVES: To elucidate the mechanisms by which acute UVB-induced angiogenesis induces dermal damage, and to study the molecular regulation of acute UVB-induced angiogenesis in human skin. METHODS: We subjected five healthy volunteers to acute UVB irradiation (2 minimal erythema doses) and performed histological analysis at 48 h after UVB irradiation. RESULTS: Histology revealed epidermal hyperplasia, infiltration of elastase-producing neutrophils and elastin fibre damage. Immunohistochemistry for CD31 demonstrated pronounced angiogenesis with a significant increase in both vascular density and vessel size, associated with increased endothelial cell proliferation. Whereas constitutive expression of TSP-1 but only weak expression of vascular endothelial growth factor (VEGF) were detected in normal human epidermis, pronounced downregulation of TSP-1 and upregulation of VEGF were observed in epidermal keratinocytes after acute UVB irradiation. These findings were confirmed by quantitative reverse transcription-polymerase chain reaction analysis after UVB irradiation of cultured HaCaT keratinocytes in vitro. CONCLUSIONS: Together, these data indicate that a disruption of the balance between VEGF and TSP-1 expression leads to a UVB-induced angiogenic switch, facilitating the infiltration of elastase-producing leucocytes and cutaneous photodamage.

Molecular bases of odor discrimination: Reconstitution of olfactory receptors that recognize overlapping sets of odorants.

The vertebrate olfactory system discriminates a wide variety of odorants by relaying coded information from olfactory sensory neurons in the olfactory epithelium to olfactory cortical areas of the brain. Recent studies have shown that the first step in odor discrimination is mediated by approximately 1000 distinct olfactory receptors, which comprise the largest family of G-protein-coupled receptors. In the present study, we used Ca(2+) imaging and single-cell reverse transcription-PCR techniques to identify mouse olfactory neurons responding to an odorant and subsequently to clone a receptor gene from the responsive cell. The functionally cloned receptors were expressed in heterologous systems, demonstrating that structurally related olfactory receptors recognized overlapping sets of odorants with distinct affinities and specificities. Our results provide direct evidence for the existence of a receptor code in which the identities of different odorants are specified by distinct combinations of odorant receptors that possess unique molecular receptive ranges. We further demonstrate that the receptor code for an odorant changes with odorant concentration. Finally, we show that odorant receptors in human embryonic kidney 293 cells couple to stimulatory G-proteins such as Galphaolf, resulting in odorant-dependent increases in cAMP. Odor discrimination is thus determined by differences in the receptive ranges of the odorant receptors that together encode specific odorant molecules.

Role of lipophilicity and hydrogen peroxide formation in the cytotoxicity of flavonols.

We compared the lipophilicity and toxicity of the four flavonols, galangin, kaempferol, quercetin and myricetin, which respectively have no, one, two and three hydroxyl groups on the B-ring. The lipophilicity was in the order of myricetin < quercetin < kaempferol < galangin. The cytotoxicity determined by a colony-formation assay with Chinese hamster lung fibroblast V79 cells was in the order of quercetin < kaempferol < galangin < myricetin. Apart from myricetin, the order of lipophilicity was the same as that of cytotoxicity, implying that the cytotoxicity was attributable to the lipophilicity. The cytotoxicity of myricetin was attributable to the hydrogen peroxide formed by autoxidation.

Two nuclear proteins, Cin5 and Ydr259c, confer resistance to cisplatin in Saccharomyces cerevisiae.

In an attempt to identify genes that can confer resistance to cisplatin, we introduced a yeast genomic library into Saccharomyces cerevisiae and selected for transformants that grew in the presence of a normally toxic concentration of cisplatin. Plasmids were rescued from the transformants and were analyzed for the presence of individual open reading frames that conferred resistance to cisplatin. We isolated two genes, CIN5 and YDR259c, that increased resistance to cisplatin when overexpressed in Saccharomyces cerevisiae. These genes encoded two proteins, Cin5 and Ydr259c, that were homologous to yAP-1, a basic leucine zipper transcriptional factor that is known to mediate cellular resistance to various toxic agents. The two proteins exhibited stronger homology to each other (33.2% identity, 49.2% similarity) than to all other gene products in S. cerevisiae. Overexpression of each of these proteins also conferred resistance to two DNA-alkylating agents, methylmethanesulfonate and mitomycin C. An experiment with fusion proteins with green fluorescent protein revealed that Cin5 and Ydr259c were localized constitutively in the nuclei of yeast cells. Our results suggest that Cin5 and Ydr259c might be involved in pleiotropic drug-resistance and might protect yeast against the toxicity of cisplatin and other alkylating agents via a single mechanism. These two nuclear proteins might act as transcriptional factors, regulating the expression of certain genes that confer resistance to DNA-alkylating agents.

Steric effects on interaction of tea catechins with lipid bilayers.

Interaction of tea catechins with lipid bilayers has been investigated with liposome systems. Tea catechins are classified into cis-type and trans-type from the configuration of the two hydrogens at the 2 and 3 positions on the C-ring. The amount of trans-type catechins incorporated into liposomes was less than that of the respective cis-type catechins. Furthermore, the order of the partition coefficients of catechins in an n-octanol/PBS system is the same as that of the amount incorporated into liposomes. These results indicate that in addition to the number of hydroxyl groups on the B-ring and the presence of the galloyl moiety, the stereochemical structure of the C-ring also governs the hydrophobicity and the affinity for lipid bilayers. Trans-type catechins with the galloyl moiety were located on the surface of the lipid bilayer, as well as cis-type catechins with the galloyl moiety, and perturbed the membrane structure. These different stereochemical structures should influence the affinity for lipid bilayers, the alteration of membrane structures, and the difference in the order of the biological activities.

Affinity of polyphenols for lipid bilayers.

Interaction of tea catechins with lipid bilayers has been investigated with liposome systems. Epicatechin gallate had the highest affinity for lipid bilayers, followed by epigallocatechin gallate, epicatechin, and epigallocatechin. Epicatechin gallate and epigallocatechin gallate in the surface of lipid bilayer perturbed the membrane structure.

Rhesus diploid rabies vaccine (adsorbed): a new rabies vaccine using FRhL-2 cells.

The FRhL-2 cell line, a diploid line derived from the lung of a fetal rhesus monkey, was used to prepare a potent rabies vaccine by adapting the Kissling strain of rabies virus to FRhL-2 cells, growing the virus in quantity, inactivating the virus with beta-propiolactone, and concentrating the virus by adsorption to aluminum phosphate. High levels of antibody to rabies virus, induced by the vaccine in both guinea pigs and humans at 14 days after immunization, were determined to be IgG. Data from postexposure protocols with guinea pigs and simulated postexposure protocols in humans showed protection and antibody response even when rabies immune globulin was administered at the time of vaccination.