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FP5

T-CIA Report Summary

Project ID: QLK6-CT-2002-02283
Funded under: FP5-LIFE QUALITY
Country: United Kingdom

Proteomic analysis of T-cell clone ageing and senescence

T-cell senescence occurs during ageing of the immune system and is an important contributor to dysregulation and dysfunction of cellular processes observed in ageing. These include alterations in cellular growth, replication, metabolism, cell-cell communication, signalling, cytokine secretion, DNA repair and apoptosis. However, little is known about the mechanisms and molecular alterations underlying these functional changes.

In the current study, we have applied surface-enhanced laser desorption/ionisation (SELDI)-ToF-MS ProteinChip Array technology for protein profiling of early and senescent T-cells in search of protein or peptide bio-markers of immunosenescence. We used the Ciphergen ProteinChip® PCS4000 SELDI system, a combination of affinity chromatography and mass spectrometry, to study protein profile changes in T-cell lysates that occur during in vitro ageing and immunosenescence with an aim to find protein or peptide biomarkers of immunosenescence.

H50 chips (reversed-phase chromatography) and Q10 chips (anion-exchange chromatography) were used to target hydrophobic and negatively charged proteins respectively in T-cell lysates. Biomarker analysis using the CiphergenExpressTM software identified differential expression of a variety of peaks associated with in vitro T-cell ageing. A consistent pattern of differential protein expression was observed between both early and late passage T-cell clones grown in vitro, and from T-cell clones derived from young and old donors. The corresponding proteins were subsequently identified by a combination of SELDI-TOF-MS, peptide mass fingerprinting MALDI-TOF-MS and Nanospray-IonTrap-MS/MS.

Through a combination of SELDI/MALDI-TOF-MS and nano-ESI-MS/MS we have identified several candidate proteins that are differentially expressed in ageing and senescent T-lymphocytes. These factors belong to a variety of functional processes including lipid metabolism, cytoskeleton remodelling, histone organisation, calcium signalling and protein degradation and represent potential bio-markers of T-cell ageing and immunosenescence. Of interest, several actin polymerising/ depolymerising factors were differentially expressed in ageing T-cells, indicating that alterations in cytoskeleton remodelling may be an important step in progression toward cellular senescence.

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Dawn MAZZATTI
Tel.: +44-1234-222372
Fax: +44-1234-248010
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