Cadmium and Lead Induced Proinflammatory Cytokine Polarization in Petroleum Products Occupationally Exposed Nigerians

Keywords

Cytokine responses • Cadmium • Lead • Antioxidant trace • Ooccupationally exposed • Cytokine balance

Introduction

Study area

This study was carried out in Nyanya, Abuja Municipal Area Council (AMAC), which is one of the six area councils of the Federal Capital Territory, Abuja, Nigeria. The case-control study randomly selected sixty-three occupationally exposed (test group) and sixty (60) non exposed individuals (control group). The occupationally exposed individuals, consisting of twentyfour automobile mechanics, eighteen generator mechanics and twenty-one petrol station attendants in filling stations who consented to participate in this study. These occupations are among those likely to expose individuals to the heavy metals studied. The sixty controls were apparently healthy individuals of Nyanya, Abuja who were not occupationally exposed to petroleum products. The length of exposure was captured by administering a self-developed semistructured questionnaire.

Inclusion criteria: All the subjects were within the age group of eighteen to fifty years. The subjects were actively serving as petrol station attendants, automobile mechanics or generator mechanics with a minimum of one year exposure.

Exclusion criteria: Individuals who are not presently serving as petrol station attendants, auto mechanics or generator mechanics were excluded from the occupationally exposed group. Also excluded from the occupationally exposed groups were those residing in and around petrol stations and traffic congestion area.

Informed consent and ethical approval: Informed consent was obtained from all subjects for this study and ethical approval obtained from the Health Research Ethics Committee of the Directorate of Health and Human Services, Federal Capital Territory, Abuja.

Collection of sample: For the heavy metals determination, about 3 ml of venous blood samp les were collected aseptically into Potassium ethylene diamine tetra acetic acid (K-EDTA) anticoagulant tubes. The samples were labelled and immediately placed in ice pack at the site of collection and subsequently transferred into a refrigerator at 4ºC. For the full blood count (FBC) and Interleukin assays, 4 ml of venous blood were collected into K-EDTA tubes, mixed and FBC carried out immediately. The blood samples were centrifuged within 30 minutes of collection for 15 minutes at 2000 rounds per minute (RPM) and plasma separated from the cells into plain tubes and refrigerated at 2-8ºC [1-7].

Determination of blood cadmium, lead, zinc, copper levels and cytokines: A calibrated Buck 205 flame atomic absorption spectrophotometer (Perkin-Elmer, HGA-2100) was used to analyze the blood sample for Cd, Pb, Zn and Cu as described by Inyengar et al. (7) at Yitzhak Rabin Laboratory for Advanced Micropropagation and Biotechnology Research Centre, Nnamdi Azikiwe University Awka, Nigeria. This method is based on the principle that atoms of the element when aspirated into Atomic Absorption Spectrophotometer vaporizes and absorbed light of the same wavelength as that emitted by the element when in the excited state. The amount of light absorbed can be correlated in a linear fashion to the concentration of the analyte in the sample. The levels of IL-1β and IL-4 were determined using enzyme-linked immunosorbent assay (ELISA) method as described by Alfred E F, et al. [8].

Statistical analysis: All numerical results were analyzed with one-way ANOVA with post hoc multiple comparisons test while student’s t- test was used to compare independent variables.

Results

(Table 1) shows the demographic characteristics of the subjects. It shows the total number of subjects that participated in the study per occupation. It also shows the mean levels of ages (years) of the subjects in the different occupational groups. It also showed the mean exposure (years) of the subjects in the individual occupations. (Table 2) shows the mean ± (SD) levels of heavy metals in the different occupational groups as compared with the control group. The levels of cadmium were significantly higher in auto mechanics and petrol station attendants as compared with the control group (p<0.002). The mean levels of Lead were significantly higher in auto mechanics and generator mechanics (0.04), while the level of zinc and copper were significantly lower in auto mechanics and generator mechanics as compared with the control group (P<0.002) but within the acceptable levels. (Table 3) shows the mean ± (SD) levels of haematological indices of the subjects in different occupational groups as compared with the control group. The levels of total white blood cells were significantly higher in auto-mechanics and generator mechanics as compared with the control group (p<0.001). The levels of granulocytes was significantly higher in generator mechanics and petrol station attendants compared with the control group (p<0.001), while the levels of red blood cells count and packed cell volume were significantly lower in generator mechanics and petrol station attendants as compared with control group (p<0.001). The mean levels of MCH were significantly higher in generator mechanics when compared with the control group (P<0.003). (Table 4) shows the mean ± (SD) levels of cytokines in different occupational groups compared with the control group. The levels of IL-1α and IL-4 were significantly higher in auto mechanics, generator mechanics and petrol station attendants as compared with the control group (p<0.01; 0.017) respectively.

Discussion

Depending on the degree and length of exposure, cadmium and lead exposure can have a variety of biological consequences, including toxic effects on the haematological, cardiovascular, neurological, and reproductive systems [9-14]. According to the findings of this study, cadmium levels are significantly higher in auto mechanics and gas station attendants, whereas lead levels are significantly higher in car mechanics and generator mechanics. This could be related to the group's heavy lubricant usage, which exposed them to high levels of cadmium and lead. Lead and cadmium exposure may also come through practices like hand cleaning with petroleum products and sucking. Fenga C, et al. [15] and Babalola OO, et al. [16] previously found elevated levels of lead among petroleum product occupationally exposed individuals, which were linked to the continued use of leaded gasoline for automobiles in Nigeria. However, prolonged exposure to these petroleum products as seen in (table 1). could also be a contributing factor to the accumulation of cadmium and lead to these group of individual. It had also been observed that prolonged exposure to petroleum compounds via ingestion, gastrointestinal tract, and inhalation from burning fossil fuel might raise heavy metal levels in exposed patients such as cadmium and lead [17]. A study by Musa AM, et al., [18], observed a lower cadmium level in the same group of subjects in Zaria, Nigeria but these group of individuals where acutely exposed to petroleum products, while the level of zinc and copper were significantly lower in auto mechanics and generator mechanics. This observation could be linked to cadmium and lead induced competitive inhibition of metalloenzyme on the site of absorption of zinc and copper which resulted to poor absorption of zinc and copper from the gastrointestinal tract. A study carried out by Flora SJS, et al., [19], showed that shows that high levels of calcium can competitively inhibit the absorption of zinc and lead from the gastrointestinal tract. Studies in both animals and humans have shown that a deficiency in essential metals such as zinc [20], calcium [21] or iron [22] can lead to greater absorption and toxicity of Cd and Pb. It had earlier been reported that zinc has similar chemical and physical properties to Cd and Pb, it competes for the binding sites of metal absorptive and enzymatic proteins [21]. Therefore it could be postulated that the supplementation with zinc could have provide protective effects against Cd and Pb accumulation but these is not within the scope of this present study but studies have shown that appropriate concentrations of essential metal supplementation are helpful in preventing complications associated with Cd and Pb accumulations [23,24].

Furthermore, the observed increased in the granulocytes count and decrease in red blood count and packed cell volume in the generator mechanics and petrol station attendant’s couples with the significantly increased levels of IL-1α and IL-4 across the three different occupational exposed groups (Tables 2-4), could be linked to Cd-Pb induced cytokines dysfunction caused by the accumulation of Cd and Pb in the peripheral blood. It had been reported that Cd and Pb exposure cause the loss of essential metals which are needed to prevent oxidative stress as seen in cases of iron-deficiency anaemia and osteoporosis [25,26]. However, it obvious from these study that the accumulation of Cd and Pb acts as an immunotoxic agent by selectively up-regulating the proinflammatory cytokines while down-regulating the anti-inflammatory cytokines. Studies have shown that the accumulation of heavy metals such as cd and Pb promotes the expression of the pro-inflammatory factors IL-1b, IL-6, and TNF-α in immunological cell by increasing ROS production, while inhibiting that of the anti-inflammatory cytokine IL-10 [27-30]. Cao X, et al. [31] showed that Cd accumulation can activates the mitochondria-mediated internal apoptosis pathway in BEAS2B cells, thus reducing their viability, causing reactive oxygen species (ROS) accumulation, inducing apoptosis, suppressing Bcell lymphoma-2 expression, and enhancing B-cell lymphoma-2- associated X and cleaved caspase-3 protein expression. Pathak N and Khandelwal S [29] showed that Cd-treated splenocytes and thymocytes produce large amounts of ROS, which not only serve as a key mediator of Cd-induced apoptosis/ cellular damage. Furthermore, Cd exposure inhibited the activity of chicken peritoneal macrophages and promoted the expression of IL-1b, IL-6, and TNF-a in both inactivated macrophages and cells in response to LPS stimuli Yucesoy B, et al. [27]. Thus, it is possible that Cd and Pd accumulation can cause cytokine dysfunction and then activate abnormal cytokine signaling pathways to produce the abnormal cytokine responses observed in this study. A better understanding of the pathways of Cd and Pb induced cytokines dysfunction, as well as interventions using inhibitors such as Zn and Cu, should be developed to effectively reduce Cd and Pb induced cytokines dysfunction seen in individuals exposed to petroleum products on the job in Nigeria.

Conclusion

Regardless of profession, prolonged exposure to petroleum products tends to shift the cytokine balance toward a pro-inflammatory pattern. Thus, when handling such materials, personal protective equipment should be worn to avoid the accumulation of cadmium and lead, which might act as mediators.

Competing Interests

The authors declare that they have no competing interests.

Authors Contributions

This work was carried out in collaboration between all authors. Author EFA and MMD designed the study and performed the statistical analysis. Authors EFA, MMD, DAK, UNE, EJI, EAO and URE conducted and managed the Laboratory analysis. All authors read and approved the final manuscript.

Acknowledgements

We acknowledge the cooperation of the members of staff of Iykenson Medicals and Research Centre, Awka, Nigeria.

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