Autolysosomes accumulate during in vitro CD8+ T-lymphocyte aging and may participate in induced death sensitization of senescent cells.
As autophagic inclusions accumulate in senescent fibroblasts, we wondered whether an increase in cellular fragility during in vitro lymphocyte aging may be related to an autolysosome accumulation. We established that, during long-term cultures, repeatedly stimulated T-lymphocytes acquired characteristics of replicative senescence and became progressively intolerant to activation. Cell death following stimulations: (i) corresponded to apoptosis, associated with necrosis at the end of the culture; (ii) was not, for its main part, mediated through CD95/CD178 or TNFRII/TNF alpha interactions; and (iii) occurred in spite of bcl-2 increased expression. After 14 weeks of culture, the percentage of lymphocytes containing at least one autophagic inclusion (p<0.0001) and the lipofuscin autofluorescence in lymphocytes (p<0.0001) were significantly increased. The expression of several genes regulating autophagy did not significantly vary with the age of the culture. Forty-eight hours after each stimulation, the percentage of induced cell death rose while, in the remaining living cells, the percentage of lymphocytes with autophagic vacuoles (p<0.05), with beta-galactosidase activity and the lipofuscin autofluorescence (p<0.001) significantly decreased, suggesting the preferential death of cells with autophagy. Our data support the view that the accumulation of autolysosomes in senescent lymphocytes might aggravate cellular fragility, leading to apoptosis and necrosis mainly induced by lymphocyte activation.
Gerland
Luc-Marie
LM
Laboratoire de Cytologie Analytique, Faculté de Médecine, INSERM U 590, 69373 Lyon Cedex 08, France.
Genestier
Laurent
L
Peyrol
Simone
S
Michallet
Marie-Cécile
MC
Hayette
Sandrine
S
Urbanowicz
Iwona
I
Ffrench
Patrick
P
Magaud
Jean-Pierre
JP
Ffrench
Martine
M
D017209:Apoptosis / Q000502:physiology; D001343:Autophagy / Q000502:physiology; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D018414:CD8-Positive T-Lymphocytes / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D016923:Cell Death / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000187:drug effects / Q000502:physiology*; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D008062:Lipofuscin / Q000096:biosynthesis; D008247:Lysosomes / Q000502:physiology*; D008854:Microscopy, Electron / Q000379:methods; D009336:Necrosis; D010835:Phytohemagglutinins / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000032:analysis; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology; D014617:Vacuoles / Q000502:physiology; D019014:fas Receptor / Q000502:physiology
D008062:Lipofuscin; D010835:Phytohemagglutinins; D019253:Proto-Oncogene Proteins c-bcl-2; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor