Investigating the effects of the heavy metals mercury, nickel and manganese, alone and in combination, on human erythrocytes and components of the coagulation system

Abstract

Heavy metals are major pollutants across the globe. Heavy metals are used in various industries and the waste matter is discarded in a manner that allows them to enter the soil, rivers and other parts of the environment. The widespread usage of heavy metals within different sectors such as agricultural, domestic and industrial areas raises an alarming concern over their impact on the environment and human health because of the wide spread contamination. Some metals are known to pose a danger to human health, while others such as copper and chromium are considered essential metals to humans and animals. However, chronic exposure to heavy metals may result in various ailments such as cardiovascular diseases, neurological- and behavioural disorders, and those affecting the immune system and kidneys. Blood cells play a crucial role in cardiovascular health where erythrocytes are responsible for the transportation of oxygen in the body and platelets and fibrin networks play a crucial role in the coagulation system also known as haemostasis. Abnormalities in haemostasis can result in haemorrhage or thrombosis. Heavy metals have been reported to cause alterations in erythrocytes influencing their function, morphology and distribution across the body, thus contributing to the pathophysiology of cardiovascular diseases. The aim of this study was to investigate the effects that the metals mercury, nickel and manganese alone and in combination, have on erythrocyte morphology and other components of the coagulation system by using the haemolysis assay, scanning electron microscopy and confocal microscopy. The metals in the study where chosen based on the likelihood of being exposed to them in South Africa. In this study, human blood was exposed to the heavy metals mercury, nickel and manganese ex vivo at concentration ranges of 1x, 10x, 100x, 1000x and 10000x the World Health Organization safety level standards for each respective metal. The World Health Organization safety level standards for these metals are: mercury = 6µg/L; nickel = 20µg/L; manganese = 400 µg/L. Blood samples were obtained from healthy male donors after written informed consent was obtained. Exposure of mercury caused increased haemolysis compared to nickel and manganese alone and in combination. At the highest concentration of 10000x all metals including the double and triple combination caused increased haemolysis. Results indicate that the erythrocytes membrane integrity was compromised resulting in haemoglobin leaking. Results provided by ultrastructural analysis indicated that the heavy metals (mercury, nickel and manganese) significantly impact on the shape of erythrocytes and structure of platelets and fibrin networks altering the coagulation system. Scanning electron microscopy results showed that with an increase in heavy metal exposure the erythrocytes lose their typical biconcave morphology and become echinocytic with a bulging appearance visible with increased membrane roughness. Platelet membrane spreading and presence of pseudopodia increases, and fibrin networks appeared unorganized with increased membrane roughness observed in all the single metal groups (Hg, Ni and Mn) and combinations (Hg+Ni, Hg+Mn, Mn+Ni and Hg+Ni+Mn). Confocal microscopy results showed that the membrane phospholipid, phosphatidylserine, was translocated from the internal membrane leaflet to the outer surface of the membranes of erythrocytes exposed to all three metals alone (Hg, Ni and Mn) and in combination (Hg+Ni, Hg+Mn, Mn+Ni and Hg+Ni+Mn), this could indicate eryptosis of erythrocytes. The morphological changes of erythrocytes, platelets and fibrin networks may cause weakened capacity for erythrocytes to carry oxygen effectively, impair the coagulation system resulting in thrombosis and additionally prevent the restoration of homeostasis of the body. The outcome of this study provides a better understanding of the effect of these metals on the coagulation system and the negative impact on human health.