Startseite Continuous reference intervals for leukocyte telomere length in children: the method matters
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Continuous reference intervals for leukocyte telomere length in children: the method matters

  • Analia Lesmana , Pei Tian , Vasiliki Karlaftis , Stephen Hearps , Paul Monagle , Vera Ignjatovic EMAIL logo , Ngaire Elwood und The Harmonising Age Pathology Parameters in Kids Study Team
Veröffentlicht/Copyright: 15. März 2021
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Clinical Chemistry and Laboratory Medicine (CCLM)
Aus der Zeitschrift Clinical Chemistry and Laboratory Medicine (CCLM) Band 59 Heft 7

Abstract

Objectives

Children with very short telomeres commonly develop bone marrow failure and other severe diseases. Identifying the individuals with short telomeres can improve outcome of bone marrow transplantation, with accurate diagnosis requiring the use of age-matched reference intervals (RIs). This study aimed to establish RIs for telomere length (TL) in children using three commonly used methods for TL measurement.

Methods

Healthy children aged 30 days to 18 years were recruited for assessment using age as a continuous variable. Venous blood samples were collected and leukocyte TL was measured using terminal restriction fragment (TRF) analysis, quantitative PCR (QPCR) and flow cytometry with fluorescence in situ hybridization (Flow-FISH). Fractional polynomial model and quantile regression were performed to generate continuous RIs. Factors that might contribute to variation in TL, such as gender, were also examined.

Results

A total of 212 samples were analyzed. Continuous RIs are presented as functions of age. TRF analysis and QPCR showed significant negative correlation between TL and age (r=−0.28 and r=−0.38, p<0.001). In contrast, Flow-FISH showed no change in TL with age (r=−0.08, p=0.23). Gender did not have significant influence on TL in children.

Conclusions

This study provides three options to assess TL in children by establishing method-specific continuous RIs. Choosing which method to use will depend on several factors such as amount and type of sample available and required sensitivity to age-related change.

Keywords: hematopoiesis; methods; pediatric; reference intervals; telomere length
Corresponding author: Prof. Vera Ignjatovic, Haematology Research, Murdoch Children's Research Institute, 50 Flemington Road, Parkville 3052, VIC, Australia; and Department of Paediatrics, The University of Melbourne, 50 Flemington Road, Parkville 3052, VIC, Australia, Phone: +61 (3) 9936 6520, E-mail:

Funding source: Royal Children’s Hospital Foundation

Award Identifier / Grant number: 2017-923

Acknowledgments

The authors thank staff of the Pathology Collection Department at The Royal Children’s Hospital for obtaining the consent of participants and the collection of samples. The authors thank staff of the Anaesthetic and Surgical Departments at the Royal Children’s Hospital.

  1. Research funding: This work was supported by grants from Royal Children’s Hospital Foundation [2017-923]. The funding organization played no role in the study design; in the collection, analysis and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.

  2. Author contributions: A.L. processed samples, performed research and wrote the manuscript. P.T. established methods, analyzed data and contributed to writing the manuscript. V.K. wrote the study protocol, obtained ethics approval for the study, reviewed the manuscript and is the coordinator of the study. S.H. performed statistical analysis and provided support on statistical analysis and reviewed the manuscript. V.I. contributed to the design of the study and was a major contributor in writing the manuscript. P.M. conceived the study and contributed to the design of the study and was a major contributor in writing the manuscript. N.E. conceived the study and contributed to the design of the study and was a major contributor in writing the manuscript. All authors read and approved the final manuscript. All authors have accepted responsibility for the entire content of this manuscript and approved its submission.

  3. Competing interests: Authors state no conflict of interest.

  4. Informed consent: Informed consent was obtained from parents or guardians of children who met study inclusion criteria.

  5. Ethical approval: The research related to human use has complied with all the relevant national regulations, institutional policies and in accordance with the tenets of the Helsinki Declaration. The study protocol has been approved by The Royal Children’s Hospital, Melbourne, Ethics in Human Research Committee (34183 A).

  6. Data sharing statement: Available data can be obtained by contacting the corresponding author.

References

1. Moyzis, RK, Buckingham, JM, Cram, LS, Dani, M, Deaven, LL, Jones, MD, et al.. A highly conserved repetitive DNA sequence, (TTAGGG) n, present at the telomeres of human chromosomes. Proc Natl Acad Sci U S A 1988;85:6622–6. https://doi.org/10.1073/pnas.85.18.6622.Suche in Google Scholar

2. Olovnikov, AM. A theory of marginotomy: the incomplete copying of template margin in enzymic synthesis of polynucleotides and biological significance of the phenomenon. J Theor Biol 1973;41:181–90. https://doi.org/10.1016/0022-5193(73)90198-7.Suche in Google Scholar

3. Ohki, R, Tsurimoto, T, Ishikawa, F. In vitro reconstitution of the end replication problem. Mol Cell Biol 2001;21:5753–66. https://doi.org/10.1128/mcb.21.17.5753-5766.2001.Suche in Google Scholar

4. Lindsey, J, McGill, NI, Lindsey, LA, Green, DK, Cooke, HJ. In vivo loss of telomeric repeats with age in humans. Mutat Res 1991;256:45–8. https://doi.org/10.1016/0921-8734(91)90032-7.Suche in Google Scholar

5. Takubo, K, Nakamura, K-I, Izumiyama, N, Furugori, E, Sawabe, M, Arai, T, et al.. Telomere shortening with aging in human liver. J Gerontol A Biol Sci Med Sci 2000;55:B533–B6. https://doi.org/10.1093/gerona/55.11.b533.Suche in Google Scholar

6. Takubo, K, Nakamura, K-I, Izumiyama, N, Sawabe, M, Arai, T, Esaki, Y, et al.. Telomere shortening with aging in human esophageal mucosa. Age 1999;22:95–9. https://doi.org/10.1007/s11357-999-0011-6.Suche in Google Scholar

7. Frenck, RW, Blackburn, EH, Shannon, KM. The rate of telomere sequence loss in human leukocytes varies with age. Proc Natl Acad Sci U S A 1998;95:5607–10. https://doi.org/10.1073/pnas.95.10.5607.Suche in Google Scholar

8. Canela, A, Vera, E, Klatt, P, Blasco, MA. High-throughput telomere length quantification by FISH and its application to human population studies. Proc Natl Acad Sci U S A 2007;104:5300–5. https://doi.org/10.1073/pnas.0609367104.Suche in Google Scholar

9. di Fagagna, FA, Reaper, PM, Clay-Farrace, L, Fiegler, H, Carr, P, Von Zglinicki, T, et al.. A DNA damage checkpoint response in telomere-initiated senescence. Nature 2003;426:194–8. https://doi.org/10.1038/nature02118.Suche in Google Scholar

10. Longhese, MP. DNA damage response at functional and dysfunctional telomeres. Genes Dev 2008;22:125–40. https://doi.org/10.1101/gad.1626908.Suche in Google Scholar

11. Collado, M, Blasco, MA, Serrano, M. Cellular senescence in cancer and aging. Cell 2007;130:223–33. https://doi.org/10.1016/j.cell.2007.07.003.Suche in Google Scholar

12. Childs, BG, Durik, M, Baker, DJ, Van Deursen, JM. Cellular senescence in aging and age-related disease: from mechanisms to therapy. Nat Med 2015;21:1424–35. https://doi.org/10.1038/nm.4000.Suche in Google Scholar

13. Panossian, L, Porter, V, Valenzuela, H, Zhu, X, Reback, E, Masterman, D, et al.. Telomere shortening in T cells correlates with Alzheimer’s disease status. Neurobiol Aging 2003;24:77–84. https://doi.org/10.1016/s0197-4580(02)00043-x.Suche in Google Scholar

14. Forero, DA, González-Giraldo, Y, López-Quintero, C, Castro-Vega, LJ, Barreto, GE, Perry, G. Meta-analysis of telomere length in Alzheimer’s disease. J Gerontol A Biol Sci Med Sci 2016;71:1069–73. https://doi.org/10.1093/gerona/glw053.Suche in Google Scholar

15. Oh, H, Wang, SC, Prahash, A, Sano, M, Moravec, CS, Taffet, GE, et al.. Telomere attrition and Chk2 activation in human heart failure. Proc Natl Acad Sci U S A 2003;100:5378–83. https://doi.org/10.1073/pnas.0836098100.Suche in Google Scholar

16. Van Der Harst, P, van der Steege, G, de Boer, RA, Voors, AA, Hall, AS, Mulder, MJ, et al.. Telomere length of circulating leukocytes is decreased in patients with chronic heart failure. J Am Coll Cardiol 2007;49:1459–64. https://doi.org/10.1016/j.jacc.2007.01.027.Suche in Google Scholar

17. Armanios, M, Blackburn, EH. The telomere syndromes. Nat Rev Genet 2012;13:693–704. https://doi.org/10.1038/nrg3246.Suche in Google Scholar

18. Parry, EM, Alder, JK, Qi, X, Chen, JJ-L, Armanios, M. Syndrome complex of bone marrow failure and pulmonary fibrosis predicts germline defects in telomerase. Blood 2011;117:5607–11. https://doi.org/10.1182/blood-2010-11-322149.Suche in Google Scholar

19. Vulliamy, TJ, Knight, SW, Mason, PJ, Dokal, I. Very short telomeres in the peripheral blood of patients with X-linked and autosomal dyskeratosis congenita. Blood Cells Mol Dis 2001;27:353–7. https://doi.org/10.1006/bcmd.2001.0389.Suche in Google Scholar

20. Stanley, SE, Armanios, M. The short and long telomere syndromes: paired paradigms for molecular medicine. Curr Opin Genet Dev 2015;33:1–9. https://doi.org/10.1016/j.gde.2015.06.004.Suche in Google Scholar

21. Armanios, M. Telomeres and age-related disease: how telomere biology informs clinical paradigms. J Clin Invest 2013;123:996–1002. https://doi.org/10.1172/jci66370.Suche in Google Scholar

22. Alder, JK, Hanumanthu, VS, Strong, MA, DeZern, AE, Stanley, SE, Takemoto, CM, et al.. Diagnostic utility of telomere length testing in a hospital-based setting. Proc Natl Acad Sci U S A 2018;115:E2358–E65. https://doi.org/10.1073/pnas.1720427115.Suche in Google Scholar

23. Ballew, BJ, Joseph, V, De, S, Sarek, G, Vannier, J-B, Stracker, T, et al.. A recessive founder mutation in regulator of telomere elongation helicase 1, RTEL1, underlies severe immunodeficiency and features of Hoyeraal Hreidarsson syndrome. PLoS Genet 2013;9:e1003695. https://doi.org/10.1371/journal.pgen.1003695.Suche in Google Scholar

24. Jonassaint, NL, Guo, N, Califano, JA, Montgomery, EA, Armanios, M. The gastrointestinal manifestations of telomere‐mediated disease. Aging Cell 2013;12:319–23. https://doi.org/10.1111/acel.12041.Suche in Google Scholar

25. Aubert, G, Hills, M, Lansdorp, PM. Telomere length measurement—Caveats and a critical assessment of the available technologies and tools. Mutat Res 2012;730:59–67. https://doi.org/10.1016/j.mrfmmm.2011.04.003.Suche in Google Scholar

26. Lai, TP, Wright, WE, Shay, JW. Comparison of telomere length measurement methods. Philos Trans R Soc Lond B Biol Sci 2018;373:20160451. https://doi.org/10.1098/rstb.2016.0451.Suche in Google Scholar

27. Montpetit, AJ, Alhareeri, AA, Montpetit, M, Starkweather, AR, Elmore, LW, Filler, K, et al.. Telomere length: a review of methods for measurement. Nurs Res 2014;63:289–99. https://doi.org/10.1097/nnr.0000000000000037.Suche in Google Scholar

28. Dietz, A, Orchard, P, Baker, K, Giller, R, Savage, S, Alter, B, et al.. Disease-specific hematopoietic cell transplantation: nonmyeloablative conditioning regimen for dyskeratosis congenita. Bone Marrow Transplant 2011;46:98–104. https://doi.org/10.1038/bmt.2010.65.Suche in Google Scholar

29. Mangaonkar, AA, Patnaik, MM. Short telomere syndromes in clinical practice: bridging bench and bedside. Mayo Clin Proc 2018;93:904–16. https://doi.org/10.1016/j.mayocp.2018.03.020.Suche in Google Scholar

30. Yamaguchi, H, Calado, RT, Ly, H, Kajigaya, S, Baerlocher, GM, Chanock, SJ, et al.. Mutations in TERT, the gene for telomerase reverse transcriptase, in aplastic anemia. N Engl J Med 2005;352:1413–24. https://doi.org/10.1056/nejmoa042980.Suche in Google Scholar

31. Du, HY, Pumbo, E, Ivanovich, J, An, P, Maziarz, RT, Reiss, UM, et al.. TERC and TERT gene mutations in patients with bone marrow failure and the significance of telomere length measurements. Blood 2009;113:309–16. https://doi.org/10.1182/blood-2008-07-166421.Suche in Google Scholar

32. Gutierrez-Rodrigues, F, Santana-Lemos, BA, Scheucher, PS, Alves-Paiva, RM, Calado, RT. Direct comparison of flow-FISH and qPCR as diagnostic tests for telomere length measurement in humans. PLoS One 2014;9:e113747. https://doi.org/10.1371/journal.pone.0113747.Suche in Google Scholar

33. Aubert, G, Baerlocher, GM, Vulto, I, Poon, SS, Lansdorp, PM. Collapse of telomere homeostasis in hematopoietic cells caused by heterozygous mutations in telomerase genes. PLoS Genet 2012;8:e1002696. https://doi.org/10.1371/journal.pgen.1002696.Suche in Google Scholar

34. Zeichner, SL, Palumbo, P, Feng, Y, Xiao, X, Gee, D, Sleasman, J, et al.. Rapid telomere shortening in children. Blood 1999;93:2824–30. https://doi.org/10.1182/blood.v93.9.2824.Suche in Google Scholar

35. Slagboom, PE, Droog, S, Boomsma, DI. Genetic determination of telomere size in humans: a twin study of three age groups. Am J Hum Genet 1994;55:876–82.Suche in Google Scholar

36. Okuda, K, Bardeguez, A, Gardner, JP, Rodriguez, P, Ganesh, V, Kimura, M, et al.. Telomere length in the newborn. Pediatr Res 2002;52:377–81. https://doi.org/10.1203/00006450-200209000-00012.Suche in Google Scholar

37. Nguyen, MT, Lycett, K, Vryer, R, Burgner, DP, Ranganathan, S, Grobler, AC, et al.. Telomere length: population epidemiology and concordance in Australian children aged 11–12 years and their parents. BMJ Open 2019;9:118–26. https://doi.org/10.1136/bmjopen-2017-020263.Suche in Google Scholar

38. Hoq, M, Karlaftis, V, Mathews, S, Burgess, J, Donath, SM, Carlin, J, et al.. A prospective, cross-sectional study to establish age-specific reference intervals for neonates and children in the setting of clinical biochemistry, immunology and haematology: the HAPPI Kids study protocol. BMJ Open 2019;9:e025897. https://doi.org/10.1136/bmjopen-2018-025897.Suche in Google Scholar

39. O’Callaghan, NJ, Fenech, M. A quantitative PCR method for measuring absolute telomere length. Biol Proc Online 2011;13:3. https://doi.org/10.1186/1480-9222-13-3.Suche in Google Scholar

40. CLSI. Defining, establishing, and verifying reference intervals in the clinical laboratory; approved guideline-third edition. CLSI document EP28-A3c. Wayne, PA: Clinical and Laboratory Standards Institute; 2008:72 p.Suche in Google Scholar

41. Altman, DG, Bland, JM. Measurement in medicine: the analysis of method comparison studies. J R Stat Soc 1983;32:307–17. https://doi.org/10.2307/2987937.Suche in Google Scholar

42. Aviv, A, Hunt, SC, Lin, J, Cao, X, Kimura, M, Blackburn, E. Impartial comparative analysis of measurement of leukocyte telomere length/DNA content by Southern blots and qPCR. Nucleic Acids Res 2011;39:e134. https://doi.org/10.1093/nar/gkr634.Suche in Google Scholar

43. Cawthon, RM. Telomere measurement by quantitative PCR. Nucleic Acids Res 2002;30:e47. https://doi.org/10.1093/nar/30.10.e47.Suche in Google Scholar

44. von Zglinicki, T. Role of oxidative stress in telomere length regulation and replicative senescence. Ann N Y Acad Sci 2000;908:99–110. https://doi.org/10.1111/j.1749-6632.2000.tb06639.x.Suche in Google Scholar

45. Allsopp, RC, Vaziri, H, Patterson, C, Goldstein, S, Younglai, EV, Futcher, AB, et al.. Telomere length predicts replicative capacity of human fibroblasts. Proc Natl Acad Sci U S A 1992;89:10114–8. https://doi.org/10.1073/pnas.89.21.10114.Suche in Google Scholar

46. Kimura, M, Cherkas, LF, Kato, BS, Demissie, S, Hjelmborg, JB, Brimacombe, M, et al.. Offspring’s leukocyte telomere length, paternal age, and telomere elongation in sperm. PLoS Genet 2008;4:e37. https://doi.org/10.1371/journal.pgen.0040037.Suche in Google Scholar

47. Unryn, BM, Cook, LS, Riabowol, KT. Paternal age is positively linked to telomere length of children. Aging Cell 2005;4:97–101. https://doi.org/10.1111/j.1474-9728.2005.00144.x.Suche in Google Scholar

48. De Meyer, T, Rietzschel, ER, De Buyzere, ML, De Bacquer, D, Van Criekinge, W, De Backer, GG, et al.. Paternal age at birth is an important determinant of offspring telomere length. Hum Mol Genet 2007;16:3097–102. https://doi.org/10.1093/hmg/ddm271.Suche in Google Scholar

49. Whiteman, VE, Goswami, A, Salihu, HM. Telomere length and fetal programming: a review of recent scientific advances. Am J Reprod Immunol 2017;77:e12661. https://doi.org/10.1111/aji.12661.Suche in Google Scholar

Supplementary Material

The online version of this article offers supplementary material (https://doi.org/10.1515/cclm-2021-0059).

Received: 2021-01-13
Accepted: 2021-02-25
Published Online: 2021-03-15
Published in Print: 2021-06-25

© 2021 Walter de Gruyter GmbH, Berlin/Boston

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  5. Updates on liquid biopsy: current trends and future perspectives for clinical application in solid tumors
  6. The underestimated issue of non-reproducible cardiac troponin I and T results: case series and systematic review of the literature
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  17. High-resolution pediatric reference intervals for 15 biochemical analytes described using fractional polynomials
  18. Continuous reference intervals for leukocyte telomere length in children: the method matters
  19. Hematology and Coagulation
  20. Using machine learning to identify clotted specimens in coagulation testing
  21. Cardiovascular Diseases
  22. Long term pronostic value of suPAR in chronic heart failure: reclassification of patients with low MAGGIC score
  23. Infectious Diseases
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  27. Corrigendum
  28. Applying the concept of uncertainty to the sFlt-1/PlGF cut-offs for diagnosis and prognosis of preeclampsia
  29. Letters to the Editors
  30. Additional approaches for identifying non-reproducible cardiac troponin results
  31. Paediatric reference intervals for ionised calcium – a data mining approach
  32. A case of interference in testosterone, DHEA-S and progesterone measurements by second generation immunoassays
  33. Lack of cross-reactivity between anti-A IgG isoagglutinins and anti-SARS-CoV-2 IgG antibodies
  34. Artefactual bands on urine protein immunofixation gels
  35. A case of methaemoglobinaemia interference on the WDF channel on Sysmex XN-Series analysers
  36. Soluble fms-like tyrosine kinase-1: a potential early predictor of respiratory failure in COVID-19 patients
  37. Serendipitous detection of α1-antitrypsin deficiency: a single institution’s experience over a 32 month period
  38. The activated partial thromboplastin time may not reveal even severe fibrinogen deficiency
  39. Influence of C-reactive protein on thrombin generation assay
  40. Inappropriate extrapolations abound in fecal microbiota research
https://doi.org/10.1515/cclm-2021-0059
Schlagwörter für diesen Artikel
hematopoiesis; methods; pediatric; reference intervals; telomere length

Artikel in diesem Heft

  1. Frontmatter
  2. Editorial
  3. Machine learning and coagulation testing: the next big thing in hemostasis investigations?
  4. Reviews
  5. Updates on liquid biopsy: current trends and future perspectives for clinical application in solid tumors
  6. The underestimated issue of non-reproducible cardiac troponin I and T results: case series and systematic review of the literature
  7. Opinion Paper
  8. Benefits, limitations and controversies on patient-based real-time quality control (PBRTQC) and the evidence behind the practice
  9. Genetics and Molecular Diagnostics
  10. ctDNA from body fluids is an adequate source for EGFR biomarker testing in advanced lung adenocarcinoma
  11. General Clinical Chemistry and Laboratory Medicine
  12. Incidence, characteristics and outcomes among inpatient, outpatient and emergency department with reported high critical serum potassium values
  13. Clinical usefulness of drug-laboratory test interaction alerts: a multicentre survey
  14. Integrating quality assurance in autoimmunity: the changing face of the automated ANA IIF test
  15. Plasma thiol/disulphide homeostasis changes in patients with restless legs syndrome
  16. Reference Values and Biological Variations
  17. High-resolution pediatric reference intervals for 15 biochemical analytes described using fractional polynomials
  18. Continuous reference intervals for leukocyte telomere length in children: the method matters
  19. Hematology and Coagulation
  20. Using machine learning to identify clotted specimens in coagulation testing
  21. Cardiovascular Diseases
  22. Long term pronostic value of suPAR in chronic heart failure: reclassification of patients with low MAGGIC score
  23. Infectious Diseases
  24. Monocyte distribution width (MDW) parameter as a sepsis indicator in intensive care units
  25. A low level of CD16pos monocytes in SARS-CoV-2 infected patients is a marker of severity
  26. Thrombin generation in patients with COVID-19 with and without thromboprophylaxis
  27. Corrigendum
  28. Applying the concept of uncertainty to the sFlt-1/PlGF cut-offs for diagnosis and prognosis of preeclampsia
  29. Letters to the Editors
  30. Additional approaches for identifying non-reproducible cardiac troponin results
  31. Paediatric reference intervals for ionised calcium – a data mining approach
  32. A case of interference in testosterone, DHEA-S and progesterone measurements by second generation immunoassays
  33. Lack of cross-reactivity between anti-A IgG isoagglutinins and anti-SARS-CoV-2 IgG antibodies
  34. Artefactual bands on urine protein immunofixation gels
  35. A case of methaemoglobinaemia interference on the WDF channel on Sysmex XN-Series analysers
  36. Soluble fms-like tyrosine kinase-1: a potential early predictor of respiratory failure in COVID-19 patients
  37. Serendipitous detection of α1-antitrypsin deficiency: a single institution’s experience over a 32 month period
  38. The activated partial thromboplastin time may not reveal even severe fibrinogen deficiency
  39. Influence of C-reactive protein on thrombin generation assay
  40. Inappropriate extrapolations abound in fecal microbiota research
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