2003 Progress Report: Aluminum in Drinking Water Induces Neuronal Apoptosis Via Endoplasmic Reticulum StressEPA Grant Number: R829782
Title: Aluminum in Drinking Water Induces Neuronal Apoptosis Via Endoplasmic Reticulum Stress
Investigators: Savory, John , Boyd, James C. , Exley, Christopher , Ghribi, Othman , Herman, Mary M.
Institution: University of Virginia
EPA Project Officer: Page, Angela
Project Period: August 1, 2002 through July 31, 2005 (Extended to July 31, 2006)
Project Period Covered by this Report: August 1, 2003 through July 31, 2004
Project Amount: $821,249
RFA: Health Effects of Chemical Contaminants in Drinking Water (2001) RFA Text | Recipients Lists
Research Category: Drinking Water , Health Effects , Water
The overall objective of this research project is to test the hypothesis that aluminum (Al) in drinking water can induce neuronal injury, eventually resulting in the death of some neurons via programmed cell death (apoptosis). It also is hypothesized that aging results in increased susceptibility to Al in drinking water. Additional experiments will be conducted to study how the neurotoxic effect of Al in drinking water is modulated by silicic acid and fluoride.
The drinking water formulation has been optimized so that precipitation of Al complexes does not occur. The basic formulation of the water was given in the October 2003 report. Five additional groups of rabbits, 12 animals per group, now have been studied with the following drinking water treatments:
- Group 1: 12 New Zealand white rabbits (2-3 years of age) were administered drinking water containing 0.5 mM Al + 1.0 mM silicic acid treatment. This group is designated AS1.
- Group 2: 12 New Zealand white rabbits (2-3 years of age) were administered drinking water containing 0.5 mM Al + 0.1 mM silicic acid treatment. This group is designated AS0.1.
- Group 3: 12 New Zealand white rabbits (2-3 years of age) were administered control (low Al concentration; 3.7 μM) drinking water containing 1.0 mM silicic acid treatment. This group is designated S1.
- Group 4: 12 New Zealand white rabbits (2-3 years of age) were administered drinking water containing 0.5 mM Al + 0.25 mM silicic acid treatment. This group is designated AS0.25.
- Group 5: 12 New Zealand white rabbits (2-3 years of age) were administered control (low Al concentration; 3.7 μM ) drinking water containing 0.1 mM silicic acid treatment. This group is designated S0.1.
The five groups of animals have been treated and sacrificed, and many of the tissue, blood, and urine analyses have been completed.
- S0.1 group (Group 5 above): One rabbit died on 10/23/03 from unknown causes.
- The animals slowly gained some weight with no differences between the five groups of animals studied.
- Pathological examination of tissue specimens: No significant changes were observed between the groups of animals studied so far. A comprehensive study has been made of various brain regions as well as the liver, kidney, spleen, and lung of all animals sacrificed so far. These histological studies have been conducted on blinded slides by two pathologists in a semiquantitative manner using a numbering system of 0 - 3. No evidence of the formation of intraneuronal filamentous aggregates, neuritic pathology, or vasculitis has been seen in any of the groups.
- Gadd 153: No evidence of Gadd 153 changes, a measure of endoplasmic reticulum stress, is observed between any of the groups of rabbits tested so far. Additional studies of endoplasmic reticulum stress, particularly activation of caspase-12, are underway.
- Bcl-2 and Bax (antiapoptosis and proapoptosis regulatory proteins) show no distinctive changes between any of the groups of animals studied so far.
- NF-κB, which is an apoptosis regulatory protein but also an indicator of inflammation, shows no apparent changes between the groups of animals studied in the present phase of this research project.
- Terminal deoxynucleotidyl Transferase-mediated deoxyuridine-triphosphate nick end labeling (TUNEL) staining for DNA fragmentation, which is a marker of apoptosis, was performed on the five groups of rabbits studied in the present phase of this research project. No apparent difference between the various drinking water groups and the control groups are observed.
- 8-Hydroxyguanine immunostaining, which is a measure of oxidation of DNA and hence a marker of oxidative stress, shows no evidence of increased oxidative stress in any of the high Al groups compared to the controls.
- Al measurements. Complete analyses of Al content have been performed on the brain, liver, spleen, bone, and plasma of all animals studied so far. Statistical analyses remain to be conducted, but at this time it does not appear that there are differences between the groups of animals studied.
- Clinical chemistry and hematology results on the five groups of animals studied in the present phase of this research project: Statistical analysis remains to be performed on these data, but so far it does not appear that there is evidence of increased Al accumulation in any of the groups of animals studied so far.
At this time, seven groups of animals have been treated: two in Year 1 of the project and five in Year 2 of the project. So far, there is no marked evidence of neuronal injury by any of the drinking water formulations either by careful neuropathological studies, by markers of endoplasmic reticulum and oxidative stress, or by tests for apoptosis. No suggestion of Al accumulation has been observed.
Future Activities: The following groups of animals as included in the original proposal will be studied as they are sacrificed. Also, the timetable for the future studies is outlined.
|Group 3: Al and Fluoride Treatment||Starting Date||Date of Sacrifice|
|0.5 mM Al + 0.05 mM Fluoride Treatment||6/7/04||2/14/05|
|0.05 mM Fluoride Treatment||6/7/04||2/14/05|
|0.5 mM Al + 0.5 mM Fluoride Treatment||7/12/04||3/21/05|
|0.5 mM Fluoride Treatment||7/12/04||3/21/05|