Receptor for advanced glycation endproducts (RAGE) modulates glyoxalase-1 enzyme activity in mouse models.
Gore, Smruti Satishchandra
University of Kansas
Pharmacology & Toxicology
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Alzheimer’s disease (AD) is the most common cause of dementia in the world. AD is characterized pathologically by the presence of Amyloid-β (Aβ) plaques and tau neurofibrillary tangles (NFTs). Advanced glycation endproducts (AGEs), which are derived from alpha-dicarbonyls such as methylglyoxal (MG), form endogenously with physiological aging. These AGEs have been observed to co-localize with both the plaques and NFTs in AD patients’ brains. Receptor for advanced glycation endproducts (RAGE) has been implicated in the pathogenesis of AD. However, the exact mechanism by which RAGE contributes to AD pathology is only partially known. Glyoxalase-1(GLO1) is an important enzyme involved in the detoxification of precursors of AGEs, which serve as the highly reactive primary ligands for RAGE. GLO1 is found to be downregulated in the brains of advanced stage AD patients whereas RAGE is overexpressed in such brains. Aging is associated with increased generation and deposition of AGEs, resulting from non-enzymatic glycation (or oxidation) of proteins and lipids. Higher AGE formation is associated with a multitude of cellular and synaptic disturbances. Thus, to see the direct effect of age on enzyme activity, we first performed a study of wild type (WT) animals from 3 to 30 months and found that beyond 12 months of age, GLO1 activity significantly decreases. This shows that aging can be a major factor contributing to AD pathology through the downregulation of GLO1 activity. For uniformity, we used 12 months old mice for all further studies. In order to investigate whether RAGE directly modulates GLO1 enzyme activity and protein expression, we used previously generated multiple transgenic (Tg) non-AD mouse models by either genetic deletion of RAGE (RAGE knockout) or the introduction of signal deficient dominant-negative mutant RAGE (DNRAGE). They were, in the past, characterized for RAGE and subsequent RAGE-mediated signal transduction in our lab. GLO1 enzyme activity was measured spectrophotometrically while GLO1 protein expression was determined with western blotting. The Tg mice displayed either a) an increase in GLO1 enzyme activity, and/or b) an increase in GLO1 protein expression when compared with age-matched WT controls. Global RAGE knockout (RO) and neuronal RAGE knockout (nRKO) mice showed significantly higher GLO1 enzyme activity compared to WT controls. RO mice showed a significant increase in the protein expression but nRKO mice did not. Similarly, the mice with DNRAGE targeted to cortical neurons (neuronal DNRAGE) and microglia (DNMSR) exhibited an increase in GLO1 enzyme activity compared with WT mice but showed no significant change in protein expression. This differential effect on protein expression can be due to the difference in post-translational modification such as disulfide bridge formation or presence of GLO1 variations. It could also be the effect of normal or higher GLO1 activity in non-neuronal or microglial cells. Given the clear modulation of GLO1 by RAGE variation, we performed the GLO1 enzyme activity and expression assays in Tg mice modeling amyloid plaque development by expressing a mutant form of human APP leading to overproduction of Aβ (mAPP) to see if similar effects are observed in a mouse model showing AD-like pathology. We also crossed these mAPP mice with RO mice to develop RAGE-deficient mAPP mice (mAPP/RO). The mAPP mice showed a significant reduction in both GLO1 enzyme activity and protein expression. mAPP/RO mice and RO mice showed higher GLO1 activity and protein expression compared to mAPP mice. These findings highlight that RAGE-dependent signaling downregulates GLO1 enzyme activity and the deletion of RAGE protects against this decrease. Also, it supports the hypothesis that loss of RAGE-mediated signaling leads to an increase in GLO1 activity, whereas specific APP mutation (and Aβ overproduction) contributes to the decrease in GLO1 activity. Taken together, these data show that RAGE functions as an active modulator of GLO1 enzyme activity, thereby providing new insights into a mechanism by which the RAGE-dependent signaling cascade contributes to the pathogenesis of AD. Thus, RAGE deletion or blockade of RAGE signaling may be a potential target for developing treatments for preventing the progression of AD and related degenerative disorders through modulation of GLO1 function.
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