Introduction
Recent studies indicate that cerebrovascular pathologies, ranging from structural alterations, atherosclerotic lesions and impaired hemodynamic responses, are another if not a primary feature of Alzheimer's disease (AD). Oxidative stress mediated by amyloid beta (Aß) and chronic inflammation associated with increased levels of transforming growth factor-beta 1 (TGF-ß1), have been implicated in these dysfunctions. Using transgenic mice of different ages (4, 12 and/or 18/21 months) that overexpress Aß or TGF-ß1, we studied the age-related changes in cerebrovascular responsiveness, assessed their relationships with alterations in specific proteins, tested in vitro their reversibility at an advanced stage of the pathology, and compared them to changes seen in cortical microvessels, including from neuropathologically confirmed cases of AD.
Methods
Responsiveness of the middle cerebral artery to increasing concentrations of serotonin (5-HT), endothelin-1 (ET-1), or calcitonin gene-related peptide (CGRP), and to NOS inhibition (10μM L-NNA) was assessed in vitro in transgenic APP (Swedish and Indiana mutated forms of the amyloid precursor protein) and TGF-ß1 mice. Underlying mechanisms were explored by i) measuring, by Western blot, changes in protein levels of markers of inflammation, oxidative stress, angiogenesis, or blood vessel wall components, ii) localizing, by immunohistochemistry, changes in some of these or other markers in cortical microvessels of transgenic mice and AD brains, and iii) pharmacologically attempting to reverse cerebrovascular deficits in vitro by MCA superfusion (30–60 min) with superoxide dismutase (SOD) or catalase (120 or 1000U/ml). Wild-type littermate mice and non-demented elderly brains served as controls.
Results
Middle cerebral arteries from both APP and TGF-ß1 mice showed age-impaired ability to dilate to CGRP and constrict upon NOS inhibition. The contractile responses to 5-HT and ET-1 were preserved except in aged TGF-ß1 mice. SOD normalized the response to NOS inhibition in APP mice but SOD and catalase had no beneficial effect in TGF-ß1 mice. Protein changes in pial and intracortical vessels of APP mice were limited to upregulation in manganese SOD (MnSOD) that typically distributed in cuffs along and around penetrating and small intracortical microvessels at all ages, being more salient in aged mice. In contrast, TGF-ß1 mice exhibited no change in MnSOD but increases in vascular content of VEGF, total collagen and collagen type IV together with a decrease in eNOS protein and alkaline phosphatase activity. In AD brains, there was no perivascular upregulation of MnSOD, but increased microvascular collagen content and decreased alkaline phosphatase activity that occurred independently from vascular Aß plaques.
Conclusion
Increased vasocontractile tone, as seen in APP and TGF-ß1 mice, can result from different underlying mechanisms. Oxidative stress, mediated primarily by superoxide anions, was the main culprit in APP mice, while structural changes in the vessel wall, exemplified by decreased eNOS and alkaline phosphatase activity and increased VEGF and collagen content, characterized TGF-ß1 mice and, also, AD cortical microvessels. We suggest that pathological vascular remodeling – and not oxidative stress – is a key determinant in AD cerebrovascular dysfunctions, and that it may be an attractive therapeutic target for AD. CIHR (MOP-64194, EH) and Alzheimer Society of Canada.