Cigarette smoking during pregnancy: Chromosome translocations and phenotypic susceptibility in mothers and newborns

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Abstract

The effects of maternal cigarette smoking during pregnancy on structural chromosome aberrations were evaluated in peripheral lymphocytes from 239 mothers and their 241 newborns to determine whether smoking during pregnancy, genetic susceptibility, and race are associated with chromosome aberrations including translocations. Demographic information and cigarette smoking data were obtained via questionnaire. There were 119 Caucasian Americans, 118 African Americans, and 2 Asian Americans. The average maternal age was 24.9 ± 5.8 (mean ± S.D.) years. Thirty-nine percent of the Caucasian Americans and 45.4% of the African Americans self-reported that they were active smokers during the index pregnancy. The average number of cigarettes smoked per day was 2.65 ± 5.75 and 1.37 ± 3.17 for Caucasian and African American mothers, respectively. Peripheral blood lymphocytes from the mother and from the fetal side of the placenta were evaluated for chromosome aberrations by whole chromosome painting. Aliquots from the same blood samples were also used to assess genetic susceptibility with an in vitro bleomycin challenge assay. Spontaneous translocation frequencies in both maternal and newborn lymphocytes were not associated with cigarette smoking, socioeconomic status, or education. The absence of a smoking effect may be attributable to the low level of cigarette usage in these subjects. The average bleomycin-induced damage in the maternal and newborn populations was 0.37 ± 0.27 and 0.15 ± 0.14 breaks per cell, respectively, a difference that was highly significant (p < 0.0001). In newborns there was a positive association between bleomycin sensitivity and the frequencies of aberrations as measured by chromosome painting: p  0.0007 for dicentrics and fragments, and p  0.002 for translocations. Caucasian American newborns demonstrated a significant association between dicentrics and fragments as measured by painting, and bleomycin sensitivity (p  0.0002), but no such association was observed for African American newborns. The results of this study indicate that while differences were observed between African Americans and Caucasian Americans, race does not appear to be a major contributor to chromosome damage in newborns or their mothers. However, peripheral lymphocytes in pregnant women are more susceptible to genetic damage than peripheral lymphocytes in newborns.

Introduction

The first report of cancer caused by tobacco was made in 1761 when Hill reported a correlation between the use of snuff and oral cancer [1]. The same report stated that cancer could be prevented by avoiding the use of tobacco products. Two hundred thirty-nine years later, tobacco smoking, environmental tobacco smoke and smokeless tobacco were for the first time included in the Ninth Report on Carcinogens (RoC) as known human carcinogens [2]. The RoC listings are based on observed causal relationships between tobacco exposure and cancer or other illnesses. The International Agency for Research on Cancer (IARC) has linked active tobacco smoking with cancer causation at more than a dozen organ sites [3]. Approximately 4000 chemicals have been identified in tobacco smoke, many of which are classified as known or suspected human carcinogens [3].

Babies born to women who smoke during pregnancy are at increased risk for poor lung development, sudden infant death syndrome, and low birth weight. Whereas some studies report an association between tobacco exposure and childhood cancer, others do not [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14], [15]. Epidemiologic studies provide evidence that environmental tobacco exposure is harmful to children, but few reports address the health effects of maternal tobacco exposures upon children during gestation. Analyses are challenging because the gestational exposures in epidemiologic studies typically cannot be separated from pre- and post-gestational maternal and paternal exposures.

Several molecular epidemiologic studies have sought to correlate maternal smoking with biomarkers associated with tobacco exposure as measured in fetal cord blood or amniocytes [16]. Maternal smoking before and during pregnancy has been associated with increased chromosomal instability in amniocytes [17]. Cotinine, DNA adducts, gene mutations, chromosome aberrations, and micronuclei have all been examined [18], [19], [20], [21], [22], [23] to evaluate the effects of transplacental exposures in fetal cells and to determine whether tobacco exposure during embryological and fetal development can increase the risks of disease later in life.

Exposures during fetal development may influence cancer risks. There is a potentially increased incidence of cancer among children whose mothers were exposed during pregnancy. Compounds in passive and active tobacco smoke and their metabolites have been known for many years to be capable of crossing the placenta [24], [25]. Diethylstilbestrol is a powerful transplacental carcinogen, increasing the risk of vaginal clear cell carcinoma and testicular cancer in the children of exposed mothers [26], [27], [28]. Gestational exposure to alcohol is associated with childhood leukemia [11]. A recent pair of review articles [29], [30] summarize the literature on environmental exposures in children, including newborns. The results suggest that children are more susceptible than adults to the effects of exposure.

Cancer is influenced by interactions among frequency and timing of exposures and genetic susceptibility [31], [32]. Rare genetic variants with high penetrance contribute to approximately 5% of all cancers, whereas the remaining cases are largely attributed to gene–gene and gene–environment interactions. Genetic polymorphisms that increase the risk for cancer among smokers have been identified [33], [34], [35], [36]. Inherent genetic susceptibility to the effects of genotoxic exposures can be assessed by evaluating the sensitivity of peripheral blood lymphocytes to bleomycin in vitro [37], [38], [39]. Cells from susceptible individuals are compromised in their ability to repair chromosomal damage compared to non-susceptible people. Cloos et al. [40] compared gene expression patterns between mutagen-susceptible and mutagen-resistant cells. They describe expression differences in a spectrum of genes involved in response to bleomycin with potential for involvement in cancer.

Structural chromosome aberrations have long been known to be widely present in tumor cells, e.g. [41], and chromosome aberrations in peripheral lymphocytes of healthy individuals are significantly associated with increased cancer risks [42], [43]. Chromosome translocations are of particular interest in assessing chronic exposure because most translocations are stable through cell division and have been shown to accumulate under conditions of protracted or highly fractionated exposure [44], [45]. Translocation frequencies have been shown to be elevated in smokers compared to non-smokers, e.g. [46], [47], although other studies have not observed such an association, e.g. [48]. Recently, an international effort to collect information on translocation frequencies in normal, healthy subjects showed a statistically significant increase in translocations among cigarette smokers compared to non-smokers [49].

Here we investigated potential interactions between maternal cigarette smoking during pregnancy, genetic susceptibility, and race, and evaluated the influence of smoking and susceptibility on translocations and other types of chromosome aberrations in mothers and their newborns. Chromosome aberrations were evaluated by simultaneously painting chromosomes 1, 2, and 4 in red and chromosomes 3, 5 and 6 in green. Genetic susceptibility was assessed using the in vitro bleomycin assay, and tobacco smoke exposure was assessed by several approaches including the use of a self-reported questionnaire, and the measure of serum cotinine levels in maternal blood samples.

Section snippets

Research subjects

Two hundred thirty-nine healthy pregnant female patients between the ages of 15 and 43 were recruited for the University of Pittsburgh Prenatal Exposures and Preeclampsia Prevention (PEPP) study [18]. Following informed consent, patients who volunteered for the study completed detailed questionnaires that were administered by trained interviewers. The questionnaires provided information about tobacco exposure, alcohol use, diet and nutrition, physical activity, race, and socio-demographics.

Demographics

The cohort from the larger PEPP study consisted of 239 mothers and their 241 newborns (including two sets of twins). A summary of the demographic data, both discrete and continuous, collected from the questionnaire is presented in Table 1. Specifically, the variables considered in this study included newborn gender, maternal race, smoking status, number of cigarettes smoked per day, socioeconomic status, education, and age. The study participants self-identified race and consisted of 49.4%

Discussion

This report demonstrates for the first time an association between genetic susceptibility, as measured by a bleomycin challenge assay, and the frequency of spontaneous chromosome aberrations including translocations measured by chromosome painting, in a cohort of human newborns. We also show that peripheral blood lymphocytes from newborns, as a group, are inherently more resistant to bleomycin-induced damage than peripheral blood lymphocytes from their mothers.

A positive association was

Conflict of interest

The authors declare that there are no conflicts of interest.

Acknowledgments

This work was performed in part under the auspices of the U.S. Department of Energy by the University of California, Lawrence Livermore National Laboratory under contract W-7405-ENG-48. Funding from the California Tobacco Related Disease Research Program Grant #8RT-0070 to JDT and NIH grant R01 HD33016 to WLB is gratefully acknowledged. We thank J. Montgomery for expert technical assistance with slide preparation and J. Williams, B. Cutter, F. Zamora, J. Coffey, S. Sun, J. Lamberton, B. Lear,

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