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Titolo:
GENERATION OF MATURE DENDRITIC CELLS FROM HUMAN BLOOD - AN IMPROVED METHOD WITH SPECIAL REGARD TO CLINICAL APPLICABILITY
Autore:
ROMANI N; REIDER D; HEUER M; EBNER S; KAMPGEN E; EIBL B; NIEDERWIESER D; SCHULER G;
Indirizzi:
INNSBRUCK UNIV,DEPT DERMATOL,ANICHSTR 35 A-6020 INNSBRUCK AUSTRIA INNSBRUCK UNIV,DEPT INTERNAL MED A-6020 INNSBRUCK AUSTRIA UNIV WURZBURG,DEPT DERMATOL D-8700 WURZBURG GERMANY
Titolo Testata:
Journal of immunological methods
fascicolo: 2, volume: 196, anno: 1996,
pagine: 137 - 151
SICI:
0022-1759(1996)196:2<137:GOMDCF>2.0.ZU;2-Y
Fonte:
ISI
Lingua:
ENG
Soggetto:
EPIDERMAL LANGERHANS CELLS; COLONY-STIMULATING FACTOR; TUMOR-NECROSIS-FACTOR; HUMAN BONE-MARROW; T-LYMPHOCYTES; IN-VITRO; RESPONSES; ANTIGENS; INVITRO; PROGENITORS;
Keywords:
DENDRITIC CELL; MATURATION, DENDRITIC CELL; BLOOD; CONDITIONED MEDIUM; IMMUNOTHERAPY; (HUMAN);
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
31
Recensione:
Indirizzi per estratti:
Citazione:
N. Romani et al., "GENERATION OF MATURE DENDRITIC CELLS FROM HUMAN BLOOD - AN IMPROVED METHOD WITH SPECIAL REGARD TO CLINICAL APPLICABILITY", Journal of immunological methods, 196(2), 1996, pp. 137-151

Abstract

Two methods to generate human dendritic cells from hematopoietic precursor cells in peripheral blood have recently been published. One approach utilizes the rare CD34(+) precursors and GM-CSF plus TNF-rr. The other method makes use of the more abundant CD34(-) precursor population and GM-CSF plus IL-4. Here we report a method that is based on the latter approach. However, the GM-CSF and IL-4 treated cells are not stable mature dendritic cells, e.g., the characteristic morphology and nonadherence of dendritic cells is lost if the cytokines are removed. We describe the need for a monocyte-conditioned medium to generate fully mature and stable dendritic cells. This is achieved by adding a 3 day 'maturation culture' to the initial 6-7 day culture in the presence of GM-CSF and IL-4. Macrophage-conditioned medium contains the critical maturation factors. Mature dendritic cells are defined by their pronounced display of motile cytoplasmic processes ('veils'), their high capacity to induce proliferative responses in resting T cells, particularly in naive umbilical cord T cells, their down-regulated antigen processing ability, and their characteristic phenotype: expression of CD83, high levels of MHC molecules and CD86, lack of CD115 and perinuclear dot-like CD68 staining. These features are stable for at least 3 days upon withdrawal of cytokines and conditioned media. IL-4 can be replaced by IL-13. When CD34(+) progenitors are depleted from blood, thereis only a minor reduction in the yield of dendritic cells by this method. We have adapted the method to consider several variables that arepertinent to clinical use, including a change from fetal calf serum to human plasma and to media approved for clinical use like X-VIVO or AIM-V. 1% plasma and RPMI 1640 are currently optimal. Additional reagents used for cell culture (Ig, cytokines) and cell separation (immunomagnetic beads) are approved for or already used in clinical applications. For 40 ml blood, the yield is 0.8-3.3 x 10(6) mature dendritic cells as defined by the expression of the new dendritic cell-restricted marker CD83, CD83(+) cells constitute between 30 and 80% of all cells recovered at the end of the culture period. Yields can be enhanced up tosix-fold if the blood donors are pretreated with G-CSF. Stable, mature dendritic cells generated by this method should be a powerful tool for active immunotherapy.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 27/11/20 alle ore 22:18:53