Competitive PCR quantification of pro- and anti-inflammatory cytokine mRNA in the central nervous system during autoimmune encephalomyelitis.
Experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease of the central nervous system that can be induced by immunization with myelin basic protein (MBP)/complete Freund's adjuvant and serves as a model for multiple sclerosis. Recent studies have suggested that cytokines play a crucial role in the clinical course of EAE. To clarify the roles of cytokines in EAE, we examined levels of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta1 (TGF-beta1) and interleukin-10 (IL-10) mRNA in isolates from infiltrating inflammatory cells in EAE lesions induced in Lewis rats. The non-radioactive and sensitive competitive PCR method was employed to quantify the relative amounts of cytokine mRNA. Levels of both IFN-gamma and TNF-alpha mRNA were increased at the early stage of EAE and rapidly decreased at the peak stage. On the other hand, TGF-beta1 mRNA was demonstrated throughout the course of EAE as well as under normal conditions and its amount paralleled the severity of EAE. IL-10 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR) under normal conditions, but was below the level of detection of competitive PCR. IL-10 mRNA expression peaked at the early stage of EAE and declined gradually thereafter. Taken together, these results suggest that IFN-gamma and TNF-alpha might play a crucial role in the development of EAE. Furthermore, it appears that the peak expression of IL-10 mRNA at the early stage and the following marked TGF-beta1 expression at the peak stage might represent an important endogenous mechanism to limit the extent of inflammation and to prevent relapse in the course of acute monophasic EAE.
Tanuma
N
N
Department of Neuropathology, Tokyo Metropolitan Institute for Neuroscience, Fuchu City, Japan.
Kojima
T
T
Shin
T
T
Aikawa
Y
Y
Kohji
T
T
Ishihara
Y
Y
Matsumoto
Y
Y
D000818:Animals; D002469:Cell Separation; D002490:Central Nervous System / Q000378:metabolism* / Q000473:pathology; D016207:Cytokines / Q000235:genetics* / Q000378:metabolism*; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000378:metabolism* / Q000473:pathology; D005260:Female; D005434:Flow Cytometry; D007150:Immunohistochemistry; D007371:Interferon-gamma / Q000235:genetics / Q000378:metabolism; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D011917:Rats, Inbred Lew; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism
D016207:Cytokines; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma