In order to analyse telomere length in subsets of human peripheral blood lymphocytes and monocytes, we modified a recently developed multicolor flow- fluorescent in situ hybridization (FISH) methodology that combines flow-FISH and antibody staining for cell surface antigens. We analysed telomere length of peripheral blood mononuclear cells in a group of 22 patients with systemic lupus erythematosus (SLE) and 20 age-matched healthy donors. We found that neither CD4+, CD8+, CD19+ cells nor CD14+ monocytes have significantly shorter telomeres compared with their healthy counterparts. On the basis of these findings, we then used monocyte telomere length as internal reference in order to control for intra-individual variability in telomere length. By using this approach, we could demonstrate significant telomere shortening in all three lymphocyte subsets (in all cases P < 0.05) compared with monocytes. However, these differences did not vary significantly between SLE patients and controls. In summary, telomere lengths in subpopulations of hematopoietic cells can be monitored in patients with SLE using multicolor flow-FISH. While confirming data by other groups on telomere length in lymphocyte subpopulations, our data argue against an increased proliferation rate of peripheral blood monocytes reflected by accelerated telomere shortening in patients with SLE. Lupus (2007) 16, 955—962.
Blackburn EHStructure and function of telomeres. Nature1991; 350: 569—573.
2.
Hayflick L.Aging under glass. Mutat Res1991; 256: 69—80.
3.
Levy MZ, Allsopp RC, Futcher AB, Greider CW, Harley CBTelomere end-replication problem and cell aging. J Mol Biol1992; 225: 951—960.
4.
Allsopp RC, Vaziri H., Patterson C. et al. Telomere length predicts replicative capacity of human fibroblasts . Proc Natl Acad Sci USA1992; 89: 10114—10118.
5.
Brummendorf TH, Balabanov S.Telomere length dynamics in normal hematopoiesis and in disease states characterized by increased stem cell turnover. Leukemia2006; 20: 1706—1716.
6.
Mills JASystemic lupus erythematosus. N Engl J Med1994; 330: 1871—1879.
7.
Crispin JC, cocer-Varela J.Interleukin-2 and systemic lupus erythematosus — fifteen years later . Lupus1998; 7: 214—222.
8.
Lipsky PESystemic lupus erythematosus: an autoimmune disease of B cell hyperactivity . Nat Immunol2001; 2: 764—766.
9.
Rus V., Via CSTelomeres, telomerase, and lupus: the long and short of it. Clin Immunol2001; 99: 195—197.
10.
Blanco P., Palucka AK, Gill M., Pascual V., Banchereau J.Induction of dendritic cell differentiation by IFN-alpha in systemic lupus erythematosus. Science2001; 294: 1540—1543.
Schonland SO, Lopez C., Widmann T. et al. Premature telomeric loss in rheumatoid arthritis is genetically determined and involves both myeloid and lymphoid cell lineages. Proc Natl Acad Sci USA2003; 100: 13471—13476.
13.
Steer SE, Williams FM, Kato B. et al. Reduced telomere length in rheumatoid arthritis is independent of disease activity and duration. Ann Rheum Dis2007; 66: 476—480.
14.
Wu K., Higashi N., Hansen ER et al. Telomerase activity is increased and telomere length shortened in T cells from blood of patients with atopic dermatitis and psoriasis. J Immunol2000; 165: 4742—4747.
15.
Fritsch RD, Shen X., Illei GG et al. Abnormal differentiation of memory T cells in systemic lupus erythematosus. Arthritis Rheum2006; 54: 2184—2197.
16.
Honda M., Mengesha E., Albano S. et al. Telomere shortening and decreased replicative potential, contrasted by continued proliferation of telomerase-positive CD8 + CD28(lo) T cells in patients with systemic lupus erythematosus. Clin Immunol2001; 99: 211—221.
17.
Klapper W., Moosig F., Sotnikova A. et al. Telomerase activity in B and T lymphocytes of patients with systemic lupus erythematosus. Ann Rheum Dis2004; 63: 1681—1683.
18.
Kurosaka D. , Yasuda J., Yoshida K. et al. Telomerase activity and telomere length of peripheral blood mononuclear cells in SLE patients. Lupus2003; 12: 591—599.
19.
Kurosaka D. , Yasuda J., Yoshida K. et al. Abnormal telomerase activity and telomere length in T and B cells from patients with systemic lupus erythematosus. J Rheumatol2006; 33: 1102—1107.
20.
Wu CH, Hsieh SC, Li KJ, Lu MC, Yu CLPremature telomere shortening in polymorphonuclear neutrophils from patients with systemic lupus erythematosus is related to the lupus disease activity . Lupus2007; 16: 265—272.
21.
Rufer N., Dragowska W., Thornbury G., Roosnek E., Lansdorp PMTelomere length dynamics in human lymphocyte subpopulations measured by flow cytometry. Nat Biotechnol1998; 16: 743—747.
22.
Baerlocher GM, Mak J., Tien T., Lansdorp PMTelomere length measurement by fluorescence in situ hybridization and flow cytometry: tips and pitfalls. Cytometry2002; 47: 89—99.
23.
Baerlocher GM, Lansdorp PMTelomere length measurements using fluorescence in situ hybridization and flow cytometry. Methods Cell Biol2004; 75: 719—750.
24.
Bartolovic K., Balabanov S., Berner B. et al. Clonal heterogeneity in growth kinetics of CD34+. Stem Cells2005; 23: 946—957.
25.
Batliwalla FM, Damle RN, Metz C., Chiorazzi N., Gregersen PKSimultaneous flow cytometric analysis of cell surface markers and telomere length: analysis of human tonsilar B cells. J Immunol Methods2001; 247: 103—109.
26.
Beier F., Balabanov S., Buckley T. et al. Accelerated telomere shortening in glycosylphosphatidylinositol (GPI)-negative compared with GPI-positive granulocytes from patients with paroxysmal nocturnal hemoglobinuria (PNH) detected by proaerolysin flow-FISH . Blood2005; 106: 531—533.
27.
Hartmann U. , Brummendorf TH, Balabanov S. et al. Telomere length and hTERT expression in patients with acute myeloid leukemia correlates with chromosomal abnormalities . Haematologica2005; 90: 307—316.
28.
Rufer N., Brummendorf TH, Kolvraa S. et al. Telomere fluorescence measurements in granulocytes and T lymphocyte subsets point to a high turnover of hematopoietic stem cells and memory T cells in early childhood. J Exp Med1999; 190: 157—167.
29.
Harley CB, Vaziri H., Counter CM, Allsopp RCThe telomere hypothesis of cellular aging. Exp Gerontol1992; 27: 375—382.
30.
Blasco MA, Lee HW, Hande MP et al. Telomere shortening and tumor formation by mouse cells lacking telomerase RNA. Cell1997; 91: 25—34.
31.
Knight SW, Heiss NS, Vulliamy TJ et al. X-linked dyskeratosis congenita is predominantly caused by missense mutations in the DKC1 gene. Am J Hum Genet1999; 65: 50—58.
32.
Vulliamy T. , Marrone A., Goldman F. et al. The RNA component of telomerase is mutated in autosomal dominant dyskeratosis congenita. Nature2001; 413: 432—435.
33.
Rufer N., Brummendorf TH, Chapuis B. et al. Accelerated telomere shortening in hematological lineages is limited to the first year following stem cell transplantation. Blood2001; 97: 575—577.
34.
Andrew T., Aviv A., Falchi M. et al. Mapping genetic loci that determine leukocyte telomere length in a large sample of unselected female sibling pairs. Am J Hum Genet2006; 78: 480—486.
35.
Slagboom PE , Droog S., Boomsma DIGenetic determination of telomere size in humans: a twin study of three age groups. Am J Hum Genet1994; 55: 876—882.
36.
Lansdorp PMStress, social rank and leukocyte telomere length . Aging Cell2006; 5: 583—584.
37.
Cherkas LF, Aviv A., Valdes AM et al. The effects of social status on biological aging as measured by white-blood-cell telomere length. Aging Cell2006; 5: 361—365.
38.
Valdes AM, Andrew T., Gardner JP et al. Obesity, cigarette smoking, and telomere length in women. Lancet2005; 366: 662—664.
39.
Brummendorf TH, Holyoake TL, Rufer N. et al. Prognostic implications of differences in telomere length between normal and malignant cells from patients with chronic myeloid leukemia measured by flow cytometry. Blood2000; 95: 1883—1890.
40.
Drummond M. , Lennard A., Brummendorf T., Holyoake T.Telomere shortening correlates with prognostic score at diagnosis and proceeds rapidly during progression of chronic myeloid leukemia. Leuk Lymphoma2004; 45: 1775—1781.
41.
Fritsch RD, Shen X., Sims GP et al. Stepwise differentiation of CD4 memory T cells defined by expression of CCR7 and CD27. J Immunol2005; 175: 6489—6497.
