Liposomal binding of lead, mercury, cadmium, gadolinium and copper and their effect on the toxicity of heavy metals in a cell culture model

Toxic heavy metals are mainly released into the environment by human industrial activities. Both acute heavy metal poisoning and chronic heavy metal accumulation continue to increase and jeopardise human health. Therapeutically used chelators, such as D-penicillamine, are a less effective therapy with many side effects. In principle, toxins can also be removed from the human organism by extracorporeal procedures such as dialysis and apheresis. Mathematical modelling at the Institute of Clinical Chemistry at the University of Freiburg has shown that the use of heavy metal-binding liposomes with their subsequent removal by lipid apheresis could lead to a significant reduction in the heavy metal load in plasma and tissues such as the liver, brain and kidneys. In this work, the binding of the heavy metals Pb${}^{2+}$, Hg${}^{2+}$, Cd${}^{2+}$, Gd${}^{3+}$ and Cu${}^{2+}$ to differently functionalized liposomes was investigated by fluorimetric heavy metal concentration measurements with rhodamine 5N and calcein. The liposomal binding of essential ions was quantified by concentration measurements in human serum. To evaluate heavy metal binding in HepG2 cell culture, the viability was analysed by MTT tests and the impairment of cellular metabolism and cellular oxidative stress by fluorescent resorufin. The liposomes analysed had different potential binding groups with free electron pairs such as amino, thiol, hydroxy and glutaryl groups or covalently bound chelators and were compared with a neutral liposome as a control. The control liposomes bound 0-5.5 mol% of the ions used. The liposomes with chelator molecules such as DTPA (heavy metal binding of 86.9 - 100 mol%) and NTA (heavy metal binding of 83 - 100 mol%) showed the highest heavy metal affinity. Even after modification of the chelator liposomes to extend their half-life in vivo, they retained their high heavy metal affinity and reached maximum heavy metal binding at 21 and 176μM Hg${}^{2+}$. These concentrations are well above the concentrations expected in poisoning. The functionalized liposomes showed a very low affinity to other essential ions and led to a slight reduction in their concentration of 0-5% in serum samples. In cell culture, evidence of positive effects of liposomes could be observed by reducing heavy metal toxicity and decreasing metabolic disorders and oxidative stress. This work proves that functionalized liposomes can bind heavy metals with high affinity. In vitro, some evidence for the reduction of heavy metal toxicity by liposomal treatment was found. The therapeutic use of functionalized liposomes to reduce the heavy metal load in vivo should therefore be further investigated as a promising new therapeutic approach.