Elevated levels of 3-nitrotyrosine in brain from subjects with amnestic mild cognitive impairment: implications for the role of nitration in the progression of Alzheimer's disease

Abstract

A number of studies reported that oxidative and nitrosative damage may be important in the pathogenesis of Alzheimer's disease (AD). However, whether oxidative damage precedes, contributes directly, or is secondary to AD pathogenesis is not known. Amnestic mild cognitive impairment (MCI) is a clinical condition that is a transition between normal aging and dementia and AD, characterized by a memory deficit without loss of general cognitive and functional abilities. Analysis of nitrosative stress in MCI could be important to determine whether nitrosative damage directly contributes to AD. In the present study, we measured the level of total protein nitration to determine if excess protein nitration occurs in brain samples from subjects with MCI compared to that in healthy controls. We demonstrated using slot blot that protein nitration is higher in the inferior parietal lobule (IPL) and hippocampus in MCI compared to those regions from control subjects. Immunohistochemistry analysis of hippocampus confirmed this result. These findings suggest that nitrosative damage occurs early in the course of MCI, and that protein nitration may be important for conversion of MCI to AD.

Figure 1

Tyrosine nitration as indexed by 3-NT immunoreactivity. ‘A’ represents the histogram obtained from Control and MCI hippocampus and IPL. ‘B’ is the slot blot results for IPL, while ‘C’ is the slot blot for hippocampus. (D) Slot blots showing standards that consist of BSA treated with defined concentrations of peroxynitrite along with control and MCI IPL *p<0.01. (E) Standard graph prepared with nitration of BSA with known concentration of peroxynitrite. The figure also shows the mean level of 3-NT in MCI hippocampus and IPL and associated concentration of peroxynitrite needed to produce these levels. Data are presented as the mean ± SEM. N=6.

Figure 2

Immunohistochemical staining. ‘A’ represents the negative control using rabbit IgG. ‘B’ and ‘C’ are representative micrographs of immunohistochemistry obtained with a polyclonal antibody for 3-nitrotyrosine in control hippocampus and MCI hippocampus, respectively (×20 magnification). Intense nitrotyrosine staining is present in MCI hippocampus, whereas staining is far less prominent in control hippocampus. Nitrotyrosine is localized predominantly in neurons (arrows).