Lead Exposure Induces Telomere Instability in Human Cells

<div><p>Lead (Pb) is an important environmental contaminant due to its widespread use over many centuries. While it affects primarily every organ system of the body, the most pernicious effects of Pb are on the central nervous system leading to cognitive and behavioral modification. Despite decades of research, the mechanisms responsible for Pb toxicity remain poorly understood. Recent work has suggested that Pb exposure may have consequences on chromosomal integrity as it was shown that Pb exposure leads to the generation of γH2Ax foci, a well-established biomarker for DNA double stranded break (DSB formation). As the chromosomal localization of γH2Ax foci plays an important role in determining the molecular mechanism responsible for their formation, we examined the localization of Pb-induced foci with respect to telomeres. Indeed, short or dysfunctional telomeres (uncapped or damaged telomeres) may be recognized as DSB by the DNA repair machinery, leading to “telomere-Induced Foci” (TIFs). In the current study, we show that while Pb exposure did not increase intra-chromosomal foci, it significantly induced TIFs, leading in some cases, to chromosomal abnormalities including telomere loss. The evidence suggests that these chromosomal abnormalities are likely due to perturbation of telomere replication, in particular on the lagging DNA strand. We propose a mechanism by which Pb exposure leads to the loss of telomere maintenance. As numerous studies have demonstrated a role for telomere maintenance in brain development and tissue homeostasis, our results suggest a possible mechanism for lead-induced neurotoxicity.</p></div

Lead induces Telomere-Induced Foci (TIFs).

<p>Results obtained with B3 cells following 24 h exposure with the indicated Pb(NO₃)₂ concentrations. <b>A.</b> Representative images obtained following IF-FISH. TIFs (yellow) represent co-localisation of γH2Ax (red) and telomere (green) signals. The upper panel shows a representative image of a single metaphase. The lower panel shows representative images of each type of staining pattern obtained from several metaphases <b>B.</b> Localisation of γH2Ax and telomere staining using IF-FISH. *<i>p</i><0.05, ** <i>p</i><0.01. The different combinations of staining are shown in the legend on the right.</p

Lead induces mainly lagging strand instability.

<p><b>A.</b> Principle of telomere hybridization by CO-FISH. <b>B.</b> Mean telomere instability observed with B3 cells after 24 h exposure with indicated Pb(NO₃)₂ concentrations. *<i>p</i><0.05. Representative images obtained with B3 cells are shown in the inset.</p

Lead induces γH2Ax foci in B3 cells.

<p>Results obtained with B3 cells following 24 h exposure with the indicated Pb(NO₃)₂ concentrations. <b>A.</b> Number of γH2Ax foci per cell. Inserts show representative γH2Ax signals observed for 0 and 1 mM Pb(NO₃)₂. <b>B.</b> Distribution of the number of γH2Ax foci per nucleus as a function of Pb concentration. Violet, 0–9 foci/nucleus; Yellow, 10–19 foci/nucleus; Green, 20–29 foci/nucleus; Blue, >30 foci/nucleus.</p

Lead induces telomere instability.

<p><b>A–C.</b> Mean telomere loss observed in B3 cells after 24 h exposure with the indicated Pb(NO₃)₂ concentrations and normalized to the correspondent mitotic index. <b>A.</b> Mitotic index. <b>B.</b> Loss of one telomere on one chromatid. <b>C.</b> Loss of two telomeres on two chromatids. <b>D.</b> Representative images obtained with B3 cells following the indicated fluorescent <i>in situ</i> hybridization procedure. Subtelomere markers are: p-arm (FITC), q-arm (Texas Red).</p

Retrospective cohort study and biobanking of patients treated for hemangioma in childhood – telomeres as biomarker of aging and radiation exposure

<p><b>Purpose:</b> Cohorts allowing joint epidemiological and biological analyses are essential for radiation risk assessment. The French Hemangioma Cohort (FHC), studied within the European project EpiRadBio, is one of the rare cohorts suitable for studying the effect of low dose radiation exposure (<100 mGy at organs), with a long-term follow-up. This highly homogeneous cohort consists of healthy individuals belonging to a normal population, except for the presence of skin hemangioma (age at exposure: between 6 months and 3 years of age). Published epidemiological studies have demonstrated that the risk of developing cancer is three times higher in the exposed individuals than in the general population. Here, we present the biobanking of samples (nucleated blood cells, cytogenetic slides of T and B lymphocytes) from the FHC and a primary feasibility study of biomarker analysis focusing on mean telomere length (MTL). Telomeres act as an internal clock, regulating the lifetime of the cell by their shortening during cell division. MTL is thus a biomarker of age. Many in vitro studies have linked MTL and radiosensitivity. The FHC will make it possible to discriminate between the effects of aging and radiation on this biomarker.</p> <p><b>Conclusion:</b> The establishment of a biobank of essentially healthy individuals (369 in total), exposed 40–70 years before, during their early childhood, is a logistical challenge. Even among those who previously participated to a self-questionnaire based study, the response rate was only 30%. The first biomarker to be studied was the MTL to discriminate age effects from those of radiation exposure. MTL showed significant variation within age groups (4–11 kb) in both the exposed and non-exposed groups. MTL within the limited age window (i.e. 40–73 year) examined, showed age-dependent changes of 46 bp/year, consistent with the age-dependent decline of 41 bp/year previously reported. We observed no significant changes in MTL according to the average active bone marrow dose. However, we were able to demonstrate that exposure to radiation causes the loss of cells with, on average, shorter telomeres, by applying a model in which both the heterogeneity of the individual dose received at the bone marrow and the heterogeneity of the intercellular distribution of MTL were taken into account.</p