प्रायोगिक Autoimmune Encephalomyelitis (EAE) प्रतिरोधी माउस खींच में दत्तक स्थानांतरण और प्रतिजनी चैलेंज के द्वारा पर काबू पाने unresponsiveness
Summary
कुछ माउस उपभेदों के माइलिन बुनियादी प्रोटीन साथ प्रयोगात्मक के autoimmune मस्तिष्क – सुषुम्नाशोथ (EAE) के शामिल होने का विरोध कर रहे हैं. यहाँ वर्णित एक सरल प्रतिरक्षण प्रोटोकॉल है कि unresponsiveness के पराजयों और कई ठेठ EAE प्रतिरोधी माउस दाग के झोले के मारे बीमारी लाती है.
Abstract
Experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease of the central nervous system (CNS) and has been used as an animal model for study of the human demyelinating disease, multiple sclerosis (MS). EAE is characterized by pathologic infiltration of mononuclear cells into the CNS and by clinical manifestation of paralytic disease. Similar to MS, EAE is also under genetic control in that certain mouse strains are susceptible to disease induction while others are resistant. Typically, C57BL/6 (H-2b) mice immunized with myelin basic protein (MBP) fail to develop paralytic signs. This unresponsiveness is certainly not due to defects in antigen processing or antigen presentation of MBP, as an experimental protocol described here had been used to induce severe EAE in C57BL/6 mice as well as other reputed resistant mouse strains. In addition, encephalitogenic T cell clones from C57BL/6 and Balb/c mice reactive to MBP had been successfully isolated and propagated.
The experimental protocol involves using a cellular adoptive transfer system in which MBP-primed (200 μg/mouse) C57BL/6 donor lymph node cells are isolated and cultured for five days with the antigen to expand the pool of MBP-specific T cells. At the end of the culture period, 50 million viable cells are transferred into naive syngeneic recipients through the tail vein. Recipient mice so treated normally do not develop EAE, thus reaffirming their resistant status, and they can remain normal indefinitely. Ten days post cell transfer, recipient mice are challenged with complete Freund adjuvant (CFA)-emulsified MBP in four sites in the flanks. Severe EAE starts to develop in these mice ten to fourteen days after challenge. Results showed that the induction of disease was antigenic specific as challenge with irrelevant antigens did not induce clinical signs of disease. Significantly, a titration of the antigen dose used to challenge the recipient mice showed that it could be as low as 5 μg/mouse. In addition, a kinetic study of the timing of antigenic challenge showed that challenge to induce disease was effective as early as 5 days post antigenic challenge and as long as over 445 days post antigenic challenge. These data strongly point toward the involvement of a “long-lived” T cell population in maintaining unresponsiveness. The involvement of regulatory T cells (Tregs) in this system is not defined.
Protocol
Discussion
चूहों में EAE के अध्ययन अक्सर neuroantigens, माइलिन मूल प्रोटीन (MBP), proteolipid प्रोटीन (पीएलपी) या माइलिन oligodendrocyte प्रोटीन (MOG) का उपयोग. पहले के अध्ययनों से ज्यादातर MBP इस्तेमाल किया. पीएलपी SJL चूहों से मजबूत और लगातार प्रतिक्?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
स्वास्थ्य के राष्ट्रीय संस्थान से अनुदान (: NS37253 R01 और NS055167 N01) और राष्ट्रीय मल्टीपल स्केलेरोसिस सोसायटी (+३२८८ A6 आरजी) द्वारा भाग में समर्थित है.
Materials
| Name of the reagent | Company | Catalogue number | Comments |
| Guinea pig spinal cords | Rockland Immuno-chemicals, Inc. Gilbertsville, PA 19525 www.rockland-inc.com |
GP-T065 | frozen |
| Freund’s Adjuvant, complete H37Ra | Difco Laboratories Detroit, MI |
231131 | |
| Multifit interchange-able syringe | B-D www.bd.com/us |
512133 | |
| RPMI 1640 | Mediatech Inc. Manassas, VA www.cellgro.com |
10-040-CM | |
| OVA | Sigma-Aldrich www.sigmaaldrich.com |
A5503 | |
| Mice | Jackson Laboratory Bar Harbor, Maine |
B6 mice: 0000664 |
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