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Conditional immortalization of human B cells by CD40 ligation.Tumor-derived microvesicles in sera of patients with head and neck cancer and their role in tumor progression.
Head Neck 31, 371-380 (2009)
It is generally assumed that human differentiated cells have a limited life-span and proliferation capacity in vivo, and that genetic modifications are a prerequisite for their immortalization in vitro. Here we readdress this issue, studying the long-term proliferation potential of human B cells. It was shown earlier that human B cells from peripheral blood of healthy donors can be efficiently induced to proliferate for up to ten weeks in vitro by stimulating their receptor CD40 in the presence of interleukin-4. When we applied the same stimuli under conditions of modified cell number and culture size, we were surprised to find that our treatment induced B cells to proliferate throughout an observation period of presently up to 1650 days, representing more than 370 population doublings, which suggested that these B cells were immortalized in vitro. Long-term CD40-stimulated B cell cultures could be established from most healthy adult human donors. These B cells had a constant phenotype, were free from Epstein-Barr virus, and remained dependent on CD40 ligation. They had constitutive telomerase activity and stabilized telomere length. Moreover, they were susceptible to activation by Toll-like receptor 9 ligands, and could be used to expand antigen-specific cytotoxic T cells in vitro. Our results indicate that human somatic cells can evade senescence and be conditionally immortalized by external stimulation only, without a requirement for genetic manipulation or oncoviral infection. Conditionally immortalized human B cells are a new tool for immunotherapy and studies of B cell oncogenesis, activation, and function. BACKGROUND: Tumor-derived membranous vesicles (MV) isolated from sera of the patients with squamous cell carcinomas of the head and neck (HNSCC) induce apoptosis of activated CD8(+) T cells. We tested if MV molecular profile and activity correlate with disease progression. METHODS: CD8(+) Jurkat cells were incubated with MAGE 3/6(+), FasL(+), MHC class I(+) MV isolated from sera of 60 patients with HNSCC and 25 normal controls by exclusion chromatography and ultracentrifugation. Z-VAD-FITC binding to Jurkat was measured and correlated with clinical data. RESULTS: MV from patients' sera, but not from sera of normal controls, induced Jurkat cell apoptosis. Forty-five percent T cells+MV from patients with N(1)-N(3) disease were apoptotic versus 18% T cells+MV from patients with N(0) disease (p < .008). MV from patients with active disease (AD) expressed higher FasL levels than MV from patients with no evident disease (NED) or normal controls (p CONCLUSION: MAGE 3/6(+), FasL(+), and MHCI(+) MV in sera of patients induced T-cell apoptosis, which correlated with disease activity and the presence of lymph node metastases.
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Publikationstyp
Artikel: Journalartikel
Dokumenttyp
Wissenschaftlicher Artikel
ISSN (print) / ISBN
1043-3074
e-ISSN
1097-0347
Zeitschrift
Head and Neck
Quellenangaben
Band: 31,
Heft: 3,
Seiten: 371-380
Verlag
Wiley
Nichtpatentliteratur
Publikationen
Begutachtungsstatus
Peer reviewed
Institut(e)
Research Unit Gene Vector (AGV)