New Concepts of Fluorescent Probes for Specific Detection of DNA Sequences: Bis-Modified Oligonucleotides in Excimer and Exciplex Detection

The detection of single base mismatches in DNA is important for diagnostics, treatment of genetic diseases, and identification of single nucleotide polymorphisms. Highly sensitive, specific assays are needed to investigate genetic samples from patients. The use of a simple fluorescent nucleoside analogue in detection of DNA sequence and point mutations by hybridisation in solution is described in this study. The 5′-bispyrene and 3′-naphthalene oligonucleotide probes form an exciplex on hybridisation to target in water and the 5′-bispyrene oligonucleotide alone is an adequate probe to determine concentration of target present. It was also indicated that this system has a potential to identify mismatches and insertions. The aim of this work was to investigate experimental structures and conditions that permit strong exciplex emission for nucleic acid detectors, and show how such exciplexes can register the presence of mismatches as required in SNP analysis. This study revealed that the hybridisation of 5′-bispyrenyl fluorophore to a DNA target results in formation of a fluorescent probe with high signal intensity change and specificity for detecting a complementary target in a homogeneous system. Detection of SNP mutations using this split-probe system is a highly specific, simple, and accessible method to meet the rigorous requirements of pharmacogenomic studies. Thus, it is possible for the system to act as SNP detectors and it shows promise for future applications in genetic testing

Regulation of kinin receptors in airway epithelial cells by inflammatory cytokines and dexamethasone

The two kinin receptors, B1 and B2, are upregulated in inflammation and may play a role in diseases such as asthma. In pulmonary A549 cells, TNF-α or interleukin-1β dramatically increased bradykinin B1 and B2 receptor mRNA expression and this response was prevented by dexamethasone. In primary human bronchial epithelial cells, bradykinin B1 receptor mRNA expression showed a similar trend, whereas bradykinin B2 receptor showed almost constitutive expression. Radioligand-binding studies revealed significant increases in bradykinin B2 receptor protein expression following both interleukin-1β and TNF-α treatment of A549 cells; however, no evidence was found for bradykinin B1 receptor. Functionally, the bradykinin B2 receptor ligand, bradykinin, but not the B1 ligand, des-Arg10-kallidin, produced a marked increase in prostaglandin E2 release when administered following interleukin-1β treatment. Arachidonic acid release in response to bradykinin was markedly enhanced by prior incubation with interleukin-1β and this was prevented by the prior addition of dexamethasone. © 2002 Elsevier Science B.V. All rights reserved.link_to_subscribed_fulltex