Received 19 May 2016; accepted 9 July 2016; published 12 July 2016
Electroplating waste water is produced in the electroplating industries such as the machinery manufactured, light and electronic industries. Due to its harmful components such as chromium, cadmium and cyanide and other toxic chemicals, it is considered to pollute the environments and impair human health   .
Some researchers have reported the harmful effects of electroplating waste water in shrimps and fishes. For example, electroplating waste water has harmful influence on the embryos and juveniles of shrimps and fishes  -  . Many fishes died in a river near the city of
Phytohaemagglutinin (PHA) response has been used to evaluate mammalian cellular immunity   . Immune organs such as thymus and spleen are also indicative of immune function  -  . Thymus is crucial for primary T cell development  , and a larger spleen implies a stronger immune system  . In addition, total white blood cells (WBC), which are important for immune response against pathogens, are also useful to assess the overall health  . Due to the scarce research about the effect of electroplating waste water on rodents, we tested the hypothesis that electroplating waste water would suppress immune function in mice in the present study.
2. Materials and Methods
2.1. Animals and Experimental Design
All animal procedures were licensed under the Institutional Animal Care and Use Committee of Qufu Normal University. Adult male Kunming mice (age: 5 - 6 months) used in this study were obtained from Experiment Animal Center in Shandong Lukang Pharmaceutical Co., Ltd. The animals were housed individually in plastic cages (30 cm × 15 cm × 20 cm) with sawdust as bedding under a natural light. The experiment was carried out from May 10th to June 15th in 2014. Standard rat pellets chow (Shandong Lukang Pharmaceutical Co., Ltd., Jining,
2.2. Organs and Body Composition
Organs were examined as described previously  . In brief, after mice were killed, the visceral organs, including heart, thymus, lungs, liver, spleen, kidneys, adrenal glands, testes, epididymis, seminal vesicals and the digestive organs with contents (i.e., stomach, small intestine, caecum and colon) were dissected and weighed (±1 mg). The stomach, small intestine, caecum and colon were rinsed with saline to eliminate all the gut contents, before being weighed.
2.3. Cellular Immunity Assays
PHA response was assessed as described previously  . Specifically, Kunming mice in the control and experimental groups on day 24 were caught, then we measured their footpad thickness of the left hind foot with a micrometer (Digimatic Indicator ID-C Mitutoyo Absolute cod. 547-301, Japan) to ±
2.4. White Blood Cells Assays
White blood cells were counted as described previously  . Briefly, at the end of the experiment, 20 µL whole blood was diluted immediately in 0.38 mL solution containing 1.5% glacial acetic acid, 1% crystal violet (Sigma) and the leukocytes were counted in an improved Neubauer chamber using microscope. The total number of WBC was determined by counting all leucocytes in the four corner large-squares of the Neubauer chamber, and multiplying the raw data by 5 × 107 to obtain the final values (109 cells/L).
2.5. Statistical Analysis
Data were analyzed using SPSS 13.0 software (SPSS Inc.,
3.1. Body Mass
Body mass did not differ between the control and experiment groups from the beginning to the end of the experiment (day 0: t = −0.592, df = 21, P = 0.560; day 27: t = 0.829, df = 21, P = 0.417) (Figure 1).
Electroplating waste water increased the masses of stomach with its content, small intestine, small intestine with its contents and colon with its contents (Table 1). However, masses of immune organs (i.e., thymus and spleen) and other organs were not influenced by electroplating waste water (Table 1).
3.3. Cellular Immunity
Electroplating waste water suppressed 6 h (t = 2.781, df = 21, P = 0.011) and 12 h (t = 2.185, df = 21, P = 0.040) of PHA responses (Figure 2). PHA responses of 24 h (t = 1.547, df = 21, P = 0.137), 48 h (t = 1.787, df = 21, P = 0.088), 72 h (t = 1.303, df = 21, P = 0.207) did not differ between the experimental and control groups.
3.4. White Blood Cells
WBC in the experiment group was not different from the control group (t = 0.140, df = 21, P = 0.890) (Figure 3).
As expected, cellular immunity in Kunming mice was impaired by electroplating waste water, however other
Figure 1. Effect of electroplating waste water on body mass in Kunming mice.
Figure 2. Effect of electroplating waste water on PHA response in Kunming mice. An asterisk (*) indicates statistical differences at P < 0.05.
Figure 3. Effect of electroplating waste water on total white blood cells in Kunming mice.
immunological parameters including white blood cells, thymus and spleen mass did not differ between the control and experimental mice.
4.1. Body Mass, Organs and Waste Water
Electroplating waste water had no effect on body mass and most organ masses. Stomach with its content, small intestine, small intestine with its contents and colon with its contents increased upon electroplating waste water, implying that the digestive capacity of mice might improve.
4.2. Immunity and Waste Water
Cellular immunity in mice was impaired by electroplating waste water, indicating their capacity of fighting against pathogens decreased. Other immunological parameters including white blood cells, thymus and spleen mass were all not response to electroplating waste water. These results demonstrated that different components of immune system had distinct sensitivities to electroplating waste water. Due to the complex components of electroplating waste water in our study, it was difficult to clarify which component played the major role in affecting immune function in mice. Effect of single heavy metal on immunity has been conducted in some researches. For example, dietary chromium can increase the levels of immunoglobulin M in juvenile rockfish  . Lower dietary nickel concentrations suppressed while higher nickel concentrations increased immune responses of Fabricius larvae  . In addition, negative correlation was observed between elevated lead levels and the percentage of natural killer cells, blood hepatitis B surface antibody levels in children from an electronic waste recycling area   . Although copper is recognized as an essential trace element, uncertainties remain regarding copper reference values for humans. For instance, very low copper intake may impair immune function in human  . However, low levels of copper contamination may enhance immunity of estuarine bivalves  . Zinc is an essential trace element which is important for maintenance and development of immune cells of both the innate and adaptive immune system  . Synergistic or antagonistic actions among different metals in electroplating waste water on immunity in mice need further researches in the future.
Table 1. Effect of electroplating waste water on wet organ mass in Kunming mice.
Values are means ± SE. Values for a specific parameter that share different superscripts are significantly different at P < 0.05, determined by General Linear Model multivariate analysis followed by Bonferroni post hoc tests with body mass as the covariate ns, not significant.
In summary, electroplating waste water harmed cellular immunity in mice, whereas other immunological parameters (i.e., white blood cells, thymus and spleen mass) were not response to electroplating waste water. Due to the complex constituents of electroplating waste water, further research is required to study synergistic or antagonistic actions among different metals of electroplating waste water on immune function in the future. Taken together, electroplating waste water had distinct impacts on different immunological parameters.
The present study was supported by grants from the National Natural Science Foundation of China (No. 31370427), Natural Science Foundation of Shandong province (ZR2013CM019) and Undergraduate Scientific Research and Training Program of Qufu Normal University (2014A054) and the Crosswise Project (hxkj- 2014001).