Telomere Length In Granulocyte, Mononuclear, CD34 and CD3 Cell Fractions In Primary Myelofibrosis and Age Matched Controls

Telomeres are repetitive, noncoding DNA sequences that cap the ends of chromosomes to provide genome protection during cell division. Telomere shortening is a normal component of cellular aging and shortened telomeres are also associated with cancer. Leukocyte telomere lengths in patients with myeloproliferative neoplasms (MPN), especially in those with myelofibrosis (MF) and polycythemia vera (PV), have been shown to be shorter than normal controls and inversely correlate to JAK2V617F allele burden (Experimental hematology. 2013;41:627). There is inadequate information on the contribution of fractioned peripheral blood samples to the relative measurement of telomere length in MF, especially in the context of age-matched controls.

Peripheral blood collected from each individual was subjected to density gradient centrifugation to enrich for granulocyte and mononuclear cell fractions. Antibody labeled magnetic bead separation was applied to a portion of the mononuclear cells to enrich for CD34+ and CD3+ fractions. DNA was isolated and telomere length was determined using quantitative PCR (qPCR) with primer sets specific to telomere repeat regions and a single copy gene. Relative telomere length of each sample was determined by calculating the Ct ratio of the telomere region and single copy gene. Quantitative PCR was used to measure JAK2V617F allele burden.

The study population included 20 patients with MF and 16 normal controls. Telomere length characterization in age-stratified normal controls indicated a clear effect of aging on telomere length, with significant differences for all cell fractions, especially when comparing age <50 years to age >65 years, and most pronounced for CD34-positive cells (Figure 1a; p=0.003). In MF, patients age 50-65 years displayed significantly shorter telomere lengths, in all cell fractions, compared to age-matched controls (Figure 1b; p=0.0001). The difference was most pronounced in CD34-positive cells (Figure 1b). However, older MF patients (age >65 years) did not differ in their telomere lengths of the aforementioned cell fractions, when compared to their age-matched controls; a trend for significance was seen for CD34-positive cells (Figure 1c; p=0.07). In MF, JAK2V617F allele burden was inversely correlated to telomere lengths of both granulocytes and mononuclear cells, in the younger (age 50-65 years; p=0.03) but not the older (age >65 years; p=0.76).

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We demonstrate the importance of age, age-matched controls, and specific peripheral blood cell fractions in the assessment of telomere length in MF. We identify CD34-positive cell fractions as the most reliable in assessing telomere length in MF but its value might be limited in patients older than age 65 years. It should be noted, however, that our results do not undermine the value of intra-patient comparison of telomere lengths, as might be necessary to monitor during treatment affecting telomerase activity.

No relevant conflicts of interest to declare.