Preparation of recombinant rat interleukin-5 by baculovirus expression system and analysis of its biological activities.
Rat interleukin-5 (IL-5) cDNA was subcloned from peritoneal cells collected 4 h after intraperitoneal injection of Ascaris suum antigen solution into the immunized rats. Cysteine proteinase-deleted (CPd) rat IL-5 recombinant virus was constructed by inserting rat IL-5 cDNA into CPd virus having a deletion in the cysteine proteinase gene of the silkworm Bombyx mori nuclear polyhedrosis virus. On infection with the CPd rat IL-5 recombinant virus, the silkworm B. mori larvae produced rat IL-5 as a dimeric form in hemolymph. Recombinant rat IL-5 was purified more than 95.5% by anion-exchange chromatography and hydrophobic chromatography. The purified recombinant rat IL-5 promoted the proliferation of T88-M cells in a concentration-dependent manner, and its effect was inhibited by an anti-murine IL-5 neutralizing polyclonal antibody. When bone marrow cells from normal rats were incubated with recombinant rat IL-5 in medium containing methylcellulose, the colony formation by eosinophilic cells was induced. Furthermore, when rat peritoneal eosinophils were incubated with recombinant rat IL-5, the spontaneous decrease in the eosinophil viability was inhibited in time- and concentration-dependent manners. In addition, the recombinant rat IL-5-induced eosinophil survival was inhibited by an anti-murine IL-5 neutralizing polyclonal antibody. These findings suggest that rat IL-5 acts as B-cell growth factor II (BCGF-II), eosinophil differentiation factor (EDF), and eosinophil survival-enhancing factor.
Ishihara
K
K
Department of Pathophysiological Biochemistry, Graduate School of Pharmaceutical Sciences, Tohoku University, Aoba Aramaki, Aoba-ku, Sendai, Miyagi 980-8578, Japan.
Satoh
I
I
Nittoh
T
T
Kanaya
T
T
Okazaki
H
H
Suzuki
T
T
Koyama
T
T
Sakamoto
T
T
Ide
T
T
Ohuchi
K
K
D000818:Animals; D001402:B-Lymphocytes / Q000187:drug effects; D016367:Baculoviridae / Q000235:genetics; D015153:Blotting, Western; D012831:Bombyx / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003001:Cloning, Molecular; D003114:Colony-Forming Units Assay; D004591:Electrophoresis, Polyacrylamide Gel; D004804:Eosinophils / Q000187:drug effects; D006458:Hemolymph / Q000378:metabolism; D015848:Interleukin-5 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism
D015848:Interleukin-5; D011994:Recombinant Proteins