Cytogenetics data in adult men involved in the recycling of electronic wastes

In this data article, 146 villagers (exposed group) were randomly selected from the workers who involved in the e-wastes recycling directly as a daily job in Tianjin. Control group, including 121 villagers, came from another town without e-waste disposal sites. Chromosomal aberrations (CA) and cytokinesis blocking micronucleus (CBMN) were performed to detect the cytogenetic effect for each subject. DNA damage was detected using comet assay; the DNA percentage in the comet tail (TDNA%), tail moment (TM), and Olive tail moment (OTM) were recorded to describe DNA damage to lymphocytes and spermatozoa. Routine semen analysis, spermatozoa motility and morphology were analyzed. The RT2Profiler PCR array was used to measure levels of expression of 84 genes related to quality of DNA. It showed significant relationships between CA, CBMN, DNA damage and exposure time in exposure subjects. The alteration of sperm motility rate, abnormality rate and total sperm counts had association with exposure time and age.


Specifications
Semen and blood were sampled from the two groups in our lab.

Experimental features
Lymphocytes were cultured in RPMI 1640 medium for CA and CBMN assay. Spermatozoa or lymphocytes were suspended in PBS for comet assay. Data source location Institute of Radiation Medicine, Chinese Academy of Medical Sciences & Peking Union Medical College Data accessibility All the data are in this data article.
Value of the data.
The data were helpful to understand the positive associations between both CA and CBMN and the duration of working with e-wastes.
When stratified for age, for each of the age sub-groups, a statistically significant difference was observed between the group exposed to e-waste and the reference group.
Semen quality was worse in the workers who recycled e-wastes than that of reference subjects.

Data
A largest electronic waste disposal centers in northern China had been found recently years. Components of e-wastes such as electronic circuit boards or microchips were illegally burned or heated for reclaimable materials (Fig. 1).
The exposure and reference group were both divided by age into three sub-groups (20-29, 30-39 and 440 years old). For each age sub-group, significant differences were found between exposure and reference groups ( Fig. 2A-F). No significant difference was observed among age-groups in either the exposure or reference group ( Fig. 2A-F). The exposure group was stratified into three sub-groups according to their exposure time (r 3, 3-6 and 4 6-year groups). The statistical significant relationships between DNA damage (TDNA%, TM) and duration of exposure for DNA damage were found in both lymphocytes and spermatozoa ( Fig. 3A and B). ** ** ** Fig. 2. DNA damage detected by comet assay in lymphocytes and spermatozoa for different age sub-groups. The group of workers recycling e-wastes and reference group were both divided into three sub-groups by age (20-29, 30-39 and 440 years old). A-C: TDNA%, TM and OTM in lymphocytes between the group of workers recycling e-wastes and reference group for the sub-groups divided by age. D-F: TDNA%, TM and OTM in spermatozoa between the group of workers recycling e-wastes and reference group for the sub-groups divided by age. **: P o 0.01. Two way ANOVA was also used to test the interactions between age and DNA damage in lymphocytes and spermatozoa, respectively. (lymphocytes: F ¼ 2.13, P ¼ 0.15; spermatozoa: For each of the sub-groups divided by age, there was significantly higher of CA and CBMN in the e-waste workers compared to the reference group ( Fig. 4A and B). No significant difference was found among sub-groups in either the exposure or reference group ( Fig. 4A and B).
Statistically significant was found between CA, CBMN and exposure time (Fig. 5A). A classical micronucleus in a binucleated lymphocyte, a dicentric chromosome and an acentric fragment are shown in Fig. 5B and C.
Sperm motility rate, abnormality rate and total sperm counts were analyzed in the three subgroups divided by age for exposure and reference groups. For the same age sub-groups, significant difference was found between exposure and reference group (Fig. 6A, B and C). The sperm parameters above also showed significant difference among different sub-groups in exposure or reference group respectively (Fig. 6A1, B1 and C1).
Relationship between semen alteration and exposure time of e-wastes was analyzed in exposure group. For the three sub-groups divided by exposure time ( r 3, 3-6 and 46years groups), semen parameters were analyzed for every two sub-groups by Wilcoxon rank-sum test. Sperm motility rate, semen volume, sperm concentration and total sperm count decreased significantly with exposure time, however, sperm abnormality rate increased significantly with e-wastes exposure time (Fig. 7). RNA of peripheral blood cells was isolated by use of the RNeasy Mini kit (Qiagen, Hilden, Germany) as instructed by the manufacturer. Integrity of RNA was assessed by means of the Bioanalyzer 2100 (Agilent Technologies, Palo Alto, CA). 84 key genes (Table 1)  A-B: CA and CBMN in lymphocytes between the group of workers recycling e-wastes and reference group for the sub-groups divided by age. There is no difference of CA and CBMN among the age sub-groups. **: P o 0.01. Two way ANOVA was also used to test the interactions between age and CA, CBMN in lymphocytes, respectively. (CA:

Instruments and reagents
Agarose gels with normal and low melting points were purchased from the Biowest Company (Miami, FL, USA). Tris-HCl, DMSO, NaHCO3, formaldehyde (A.P.), trypan blue and TritonX-100 were purchased from Sigma (St. Louis, MO, USA). The electrophoresis apparatus was purchased from BIO-RAD (Hercules, CA, USA), and the Nikon90i fluorescence microscope was purchased from NIKON (Tokyo, Japan). The comet slides were purchased from Trevigen. Inc. (Gaithersburg, MD, USA).

Routine semen analysis
The procedure of routine semen analysis was performed according to the standard methods in the WHO manual [2]. Briefly, the semen samples were examined immediately after liquefaction or within one hour of ejaculation. All the semen samples were ensured to be homogeneous by mixing thoroughly. A fixed volume of 10 μl semen was delivered onto a clean glass slide and covered with a coverslip. Scanning the slide and estimating the number of spermatozoa per 400× magnification field gives an approximate sperm concentration in 106/ml. This estimate is used to decide the dilution (1:5, 1:10, 1:20, 1:50) for determining the sperm concentration by hemocytometry. The spermatozoa concentration was determined using the hemocytometer method on two separate preparations of the semen sample. The diluted semen sample was dropped onto the hemocytometer and covered with a coverslip, and was placed in a humid chamber for about five minutes to prevent drying out. The cells sedimented during this time and were then counted. The count only included complete spermatozoa (heads with tails). Any sperm lying on the line dividing two adjacent squares was counted only if it was on the upper or the left side of the square being assessed.  Fig. 6. Sperm motility rate, abnormality rate and total sperm counts for different age sub-groups. A-C: Sperm motility rate, abnormality rate and total sperm counts between the group of workers recycling e-wastes and reference group for the subgroups divided by age. There is significant difference of sperm motility rate, abnormality rate and total sperm counts between the group of workers recycling e-wastes and reference group in the age sub-groups, respectively. A1-C1: Line chart showed the change of sperm motility rate, abnormality rate and total sperm counts along with age. There is significant difference of sperm motility rate, abnormality rate and total sperm counts among the age sub-groups in exposure or reference group respectively. **: P o 0.01, *: P o 0.05. Two way ANOVA was also used to test the interactions between age and sperm motility rate, abnormality rate and total sperm counts, respectively. (sperm motility rate: F ¼ 3.24, P ¼ 0.07; abnormality rate: F ¼ 3.13, P ¼ 0.08; total counts: F ¼ 2.89, P ¼ 0.12).

Alkaline Comet assay
Spermatozoa or lymphocytes were suspended in PBS at a concentration of 1 × 105/ml. The comet assay, also called single cell gel electrophoresis, was performed as previously reported [3]. Briefly, comet slides were coated with 100 μl of 0.75% (w/v) normal-melting-point agarose. Once the first agarose layer was coagulated, a mixture of 75 µl of low-melting-point agarose and 25 µl of spermatozoa suspension was applied as the second layer. The comet slides were immersed in cold lysis buffer (2.5 M NaCl, 0.5 M EDTA, 10 mM Tris HCl pH 10.0 containing 1% Triton X-100, 40 mM dithiothreitol and 100 μg/ml proteinase K) for 2 h at room temperature to remove any DNA-associated proteins.
After lysis, double-distilled water was used to rinse away excess salt. All the comet slides were then placed in buffer for 20 min in a horizontal electrophoresis tank that was pre-filled with cold alkaline buffer (1 mM Na2EDTA and 0.3 M NaOH, pH 13.0) to loosen the tight double-helical structure of DNA for electrophoresis. Electrophoresis was then performed at 25 V and 10 mA for 20 min in electrophoresis buffer at room temperature. The slides were then rinsed twice with distilled water and stained with ethidium bromide (2 µg/ml). All of the above procedures were carried out in the dark to avoid additional DNA damage. The comets were viewed using a Nikon 90i fluorescence microscope,  and images of 100 comets were collected for each subject using a digital imaging system. Cells that overlapped were not counted. All the comet images were analyzed using Comet Assay Software Project (CASP, Wroclaw University, Poland) [4] and the DNA percentage in the comet tail (TDNA%), the tail moment (TM) and the Olive tail moment (OTM) were recorded to describe the DNA damage to the spermatozoa or lymphocytes.