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Titolo:
RESPONSE OF CD4(-CELLS FROM MYASTHENIC PATIENTS AND HEALTHY-SUBJECTS OF BIOSYNTHETIC AND SYNTHETIC SEQUENCES OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR() T)
Autore:
DIETHELMOKITA B; WELLS GB; KURYATOV A; OKITA D; HOWARD J; LINDSTROM JM; CONTIFINE BM;
Indirizzi:
UNIV MINNESOTA,COLL BIOL SCI,DEPT BIOCHEM ST PAUL MN 55108 UNIV MINNESOTA,COLL BIOL SCI,DEPT BIOCHEM ST PAUL MN 55108 UNIV MINNESOTA,SCH MED,DEPT PHARMACOL MINNEAPOLIS MN 55455 UNIV N CAROLINA,DEPT NEUROL CHAPEL HILL NC 00000 UNIV PENN,MED CTR,DEPT NEUROSCI PHILADELPHIA PA 19104
Titolo Testata:
Journal of autoimmunity
fascicolo: 2, volume: 11, anno: 1998,
pagine: 191 - 203
SICI:
0896-8411(1998)11:2<191:ROCFMP>2.0.ZU;2-3
Fonte:
ISI
Lingua:
ENG
Soggetto:
ALPHA-SUBUNIT; BACULOVIRUS VECTORS; EPITOPE REPERTOIRE; IMMUNE-RESPONSES; GRAVIS; EXPRESSION; PROTEINS; INTERLEUKIN-12; HETEROGENEITY; DETERMINANTS;
Keywords:
MYASTHENIA GRAVIS; NICOTINIC RECEPTOR; T-CELL EPITOPES; SYNTHETIC T-EPITOPES; BACTERIAL EXPRESSED ANTIGENS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
41
Recensione:
Indirizzi per estratti:
Citazione:
B. Diethelmokita et al., "RESPONSE OF CD4(-CELLS FROM MYASTHENIC PATIENTS AND HEALTHY-SUBJECTS OF BIOSYNTHETIC AND SYNTHETIC SEQUENCES OF THE NICOTINIC ACETYLCHOLINE-RECEPTOR() T)", Journal of autoimmunity, 11(2), 1998, pp. 191-203

Abstract

We investigated the suitability of pools of overlapping synthetic peptides spanning the complete alpha(1) subunit sequence of the human muscle acetylcholine receptor (AChR) (alpha(1) pool) or the extracellulardomain (residues 1-218, alpha(1)1-218 pool), and of biosynthetic alpha 1 constructs from E. coli, as stimulants of human CD4(+) cells from myasthenia gravis (MG) patients and healthy subjects. A construct corresponding to residues alpha(1)1-209 was obtained as solubilized inclusion bodies (ib alpha(1)1-209), or purified by SDS gel electrophoresis (pur alpha(1),1-209). A second construct included the extracellular, cytoplasmic and carboxyl-terminal domains plus histidine residues, and was obtained as inclusion bodies (ib alpha(1)NoTrans) or purified by gel permeation and histidine tag affinity chromatography (pur alpha(1)NoTrans). A biosynthetic extracellular domain of the neuronal AChR alpha(7) subunit (ib alpha(7)1-206) isolated from E. coli as inclusion bodies served as control for bacterial contaminants. We used ib alpha(1)1-209, pur alpha(1)1-209 and peptide pools to propagate CD4(+) lines from two MG patients. The lines obtained using pur alpha(1)1-209 and thepeptide pools recognized the peptide pools and alpha(1) constructs tested well, but ib alpha(7)1-206 poorly or not at all. These lines recognized peptides known to form CD4(+) epitopes in these patients. The ib alpha(1)1-209 lines recognized ib alpha(1)1-209 and ib alpha(7)1-206strongly, but recognized poorly pur alpha(1)1-209 and the alpha(1)1-218 pool. We propagated T-cell lines from a healthy subject using pur alpha(1)1-209 and ib alpha(1)1-209. The pur alpha(1)1-209 line recognized pur alpha(1)1-209 and the alpha(1)1-218 pool, but not ib alpha(1)1-209 or ib alpha(7)1-206. The ib alpha(1)1-209 line recognized ib alpha(1)1-209 and ib alpha(7)1-206, but not pur alpha(1)1-209 or the alpha(1)1-218 pool. We tested blood CD4(+) cells from six MG patients and eight healthy subjects with ib alpha(1)1-209, pur alpha(1)1-209, the alpha(1)1-218 pool and-in the healthy subjects-also ib alpha(7)1-206, ib alpha(1)NoTrans and pur alpha(1)NoTrans. In both populations, the alpha(1)1-218 pool elicited low and sporadic responses, while the constructs elicited clear responses that were frequently higher for ib alpha(1)1-209 than pur alpha(1)1-209. The responses to ib alpha(7)1-206 were strong and comparable to those to ib alpha(1)1-209, ib alpha(1)NoTrans, and pur alpha(1)NoTrans. These results indicate that even purified constructs from E. coli contain bacterial contaminants recognized by CD4(+) cells. They should not be used to test unselected blood CD4(+) cells, because they may evoke strong CD4(+) responses to the bacterial antigens. Purified recombinant sequences may be suitable for propagation of CD4(+) cell lines, if the specificity of the Lines can be verified using different antigen prepar; ations. Short synthetic peptide sequences can be safely used for propagation of specific CD4(+) cells. Although they are poor stimulants for unselected blood CD4(+) cells, the low responses they elicit are probably due to these cells. (C) 1998 Academic Press Limited.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 28/09/20 alle ore 15:40:20