| During the past two decades, there have been increasing evidence that a bidirect ional flow of information exists between the immune and central nervous systems, representing an important homeostatic mechanism in the body. Cytokines seem to play a crucial role in this communication. Interferon-alpha (IFNα) not only pos s esses immunoregulatory activity, but also has important effects on central nervo us system. Investigations in vivo and in vitro have elucidated the facts that IF N( and IFNα receptors have been detected in the brain. IFNα affects sleeping a nd physiologic behavior, has analgesic effect assemble to endogenous opioid peptid e molecules, and affects the hypothalamic-pituitary axes. There are many reports about neuroregulatory effect of IFNα, but among which less is about the struct ural bases. In 1981, Blalock found that human leukocyte IFN had opioid-like analgesia. Our p revious work also showed that IFNα had opioid-like analgesic effect, and had tw o distinct domains that mediated its immunoactivity and analgesic action respecti vely. And then how about the molecular structural basis for the analgesic effect of IFNα? Which acid residue is composed of the analgesic domain? In this study, the analgesic effect of IFNα (IFNα2b) and its mechanism were in ve stigated using stereotaxic technique, pain threshold measurement. Techniques of molecular biology and methods of protein engineering including site-directed mut agenesis, gene recombination and expression, purification of protein and so on, were performed to study the structure and analgesic function relationship, and t o explore the molecular structure of analgesic effect of IFNα. Our previous work showed that the analgesia of IFNα was related to α opioid re ce ptor. In this study, we tested the changes of the content of cAMP, PGE2 and the temperature of rats, to elucidate the possible receptor pathway for the opioid-l ike action of IFNα. The main results of our research are as follows:Acording to the tertiary structural map of IFNα, we changed all of Phe and Pro r esidues around the 122nd Tyr residue in the tertiary structure of IFNα. The mut a nts of IFNα were obtained by using site-directed mutagenesis, and were used to determine the antiviral activity and the analgesic activity respectively. It was suggested that the analgesic domain is composed of the 122nd Tyr, the 36th Phe, the 38th Phe and the 39th Pro resid ues of IFNα. The CHO cells transfected with α opioid receptor were cultured. After incubati n g with IFNα and its mutants, the cells were used to measure the content of cAMP . The wild-type IFNα, the mutants having analgesic activity (129Ser-IFNα) and m or phine could decrease the content of cAMP, and naloxone could block these effects . The mutant that has no analgesic activity (38Leu-IFNα), failed to decrease th e content of cAMP. It is suggested that the analgesic domain could interact with α opioid receptor. SD rats were stereotaxically implanted with steel guide cannulae to permit late ral cerebral ventricular infusion, and the rectal temperature was determined. Wi ld-type IFNα and 129Ser-IFNα could increase the temperature, and naloxone coul d block the effects. It is suggested that the fever induced by IFNα might be rela ted to its analgesic domain binding to opioid receptors. After incubating with IFNα and its mutants, the hypothalamus slices were used t o determine the release of PGE2. The wild-type IFNα and 129Ser-IFNα could incr ea se the release of PGE2, and naloxone could bloc...
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