Assessing Lead Effects on Fisher-344 Rats Using ICP-MS and Histology

As a way of laying the foundation for research in analyzing global gene expression patterns in Fisher 344 rat liver, the effects of lead toxicity on Fisher 344 rats were assessed using histopathology and tissue residues were measured for metal ions using ICP-MS. Accumulation of lead in blood, liver, kidney and bone marrow increased significantly in lead exposed groups. With the exception of kidney, the 90-days treatment groups showed markedly high levels of lead in blood, liver and marrow than the 30days exposed groups. Potential interactions of calcium, iron and zinc and lead examined showed positive and negative correlations. Hepatic histopathology produced no evidence of necrosis nor changes in architecture of hepatocytes in the 0 ppm and 50 ppm for the 30-day duration of exposure in the case of the liver. In contrast, necrosis and alterations in the structure and disposition of the liver and kidney tissues were observed for the 500 ppm treatment group. INSTITUTION WHERE WORK DONE Department of Chemistry, Western Michigan University, Kalamazoo, Michigan. This work was supported by grants to Dr. Jay C. Means (EPA grant #XP97528301-0 to the Environmental Institute at Western Michigan University) INTRODUCTION TO STUDY The most ancient and relevant environmental poison to be used by man is lead. According to Jernigan et al [1], hundreds of millions of people have been affected by the toxicity of lead during the last 4500 years either as mining slaves, or as consumers of adulterated wine and food or from breathing urban air. Written archeological evidence exists that lead was used widely in the ancient world. In recent times, lead has been used in gasoline which makes it widely distributed in the environment. The environmental significance of lead as a chemical species exhibiting various form of toxicity in humans has been well documented. [2-5]. As a result of its past uses history, lead is widely distributed in water, soil and air. This is particularly of great importance considering lead use in gasoline and paint was curtailed more than two decades ago. As a result, the potential exposure to significant lead levels, especially infants in the population, is high [6]. Pb exposure targets organ systems such as the skeletal, hematopoeitic, renal, endocrine and nervous systems [7], thereby partitioning between soft and hard tissues in the body with approximately 95 % and 70 % being found in the bones and teeth of adults and children, respectively. Bone then serves as a pool to replenish excreted lead from blood. Some adverse conditions associated with lead poisoning include DNA damage, neurological impairment, abnormal heart function, osteoporosis among others [8-9]. In addition, a weakened immune defense system, sterility in male and females, abnormal fetal development, and glycosuria are also associated with chronic lead poisoning. Lead perturbs the functions of enzymes and proteins of varied classes. Studies have shown that, lead exerts its influence physiologically and biochemically as a mimetic agent substituting for essential elements participating in metabolism such as calcium, iron and zinc. For instance, it directly interferes with zinc and iron in the biosynthesis of heme, in the function sulfhydryl group rich protein enzymes and in protein synthesis in general either directly or indirectly [8-9]. Lead binds to different kinds of transport proteins including metallothionein, transferin, calmodulin and calcium-ATPase. By associating with these proteins, it is transported to specific tissues where it causes its damage. Lead transport and assimilation are optimum when there is the dietary deficiency of essential metals such as iron or calcium or zinc. This is because lead is able to displace these metals in transport proteins and during specific proteinmediated physiologic processes [10]. Assessing Lead Effects on Fisher-344 Rats Using ICP-MS and Histology 2 of 11 Liver and kidney damage have been linked to lead toxicity although the exact toxicity mechanism is not entirely understood. Other than the use of histopathology to assess the effects of lead poison in hepatocytes, the use of other methods has been inconclusive. The objective of this study is to assess the interaction of lead with calcium, iron, copper, zinc, cobalt and nickel in blood, liver, kidney and bone marrow using rat model. Histopathology of the liver and kidney will also be examined. Inductively-coupled plasma spectrometry (ICP-MS) is a useful analytical tool for quantifying multi-elements from such matrices as geological, environmental and biological samples at sub parts per billion. That is, it has a quantitation limit of 1-100 parts per trillion and a linear dynamic range of about eight orders of magnitude. The range of analytes that can be employed in ICP-MS has recently been extended to both metals and nonmetals including radionuclides, rare earths, and some halogens like Br and I. ICP-MS works by generating ions in the plasma which are directed into the ion focusing region using turbomoelcular pumps backed by rotary pumps. Then electrostatic ion and extraction lenses sort the negative and positive ions so that positive ions are directed towards the quadrupole. Ions then enter the separation hardware, the quadrupole mass spectrometer where the electric field forces them into wavelike motion. Ions with stable trajectories are filtered according to their mass to charge ration (m/z) in the quadrupole. Finally, individual ions are detected by ion counting electron multiplier [11]. The Fisher 344 rat inbred strain was developed in 1920 to address the lack of reproducible animal model for cancer research. In 1970, the National Cancer Institute selected Fisher 344 rat as a replacement for the Osborne-Mendel rat model in cancer bioassay program because tumor latency due to chemical exposure is relatively short whilst maintaining good survivability. Fairly recent literature has indication that F344 rats are prone to exhibit inflammatory effects and mononuclear cell leukemia due to exposure of a range of chemicals and pharmaceuticals. Nevertheless, this animal model has been employed in as many cancer, toxicological, aging, neurological, organ transplant, heart disease etc studies in the literature [12-15]. This experiment is part of a larger project investigating the overall effects of Pb2+ on gene expression in the rat. Results from this investigation will enable us to determine what specific tissue levels of Pb in rat lead to alterations in gene regulation.