A Study Of Pathomechanism Of Cerebral Small Vessel In Leukoaraiosis (White Matter Lesions)

Background and Purpose: The term leukoaraiosis (from the Greek leuko, white, and araiosis, rarefaction) was introduced in 1986 by Hachinski, Potter and Merskey to designate bilateral and symmetrical areas in the periventricular and centrum semiovale white matter that appeared hypodense on CT scans and hyperintense on T2-weighted MRI. Leukoaraiosis (LA) is used interchangeably with the term white matter lesions (WMLs) on MRI and seen to some degree in more than half of the routine scans in older persons. Leukoaraiosis can reflect a broad public health problem, which is caused by a cognitive impairment ranging from mild slowness of thinking to full-blown subcortical dementia. As many as 80% of patients with leukoaraiosis have some degree of gait disorder. This association is independent of age, gender, previous stroke and hypertension. Furthermore, deterioration of gait is associated with progression of leukoaraiosis. Other features include bladder instability and mood disturbance. The presence of leukoaraiosis has been identified as a marker of less favorable prognosis also in the acute stroke settings. In particular, white matter changes have been associated with an increased risk of hemorrhagic transformation of the brain infarct in patients subjected to thrombolysis. One-quarter of subjects aged 65 years or over are affected by some degree of white matter changes. The well-known vascular risk factors, however, do not fully explain the incidence of LA. The neuropathological appearances corresponding to leukoaraiosis are neuronal loss, ischemic demyelination, and gliosis. Ischemic demyelination and small-vessel disease have been assumed as underlying pathological processes in the evolution of this entity. The pathogenesis of cerebral small vessel disease (SVD) is poorly understood, but endothelial activation and dysfunction may play a causal role. The exact etiological factors and the pathomechanism of LA remain to be identified.Cerebral arteries not only deliver blood to the brain, but their walls also act as drainage pathways for interstitial fluid (ISF) and solutes from the brain. More specifically, ISF and solutes drain from the brain along the basement membranes between the smooth muscle cells in the tunica media. Amyloid-β(Aβ) is deposited in the basement membranes of arteries in cerebral amyloid angiopathy (CAA) in elderly humans. This strongly suggests that ISF and solutes drain from the human brain along artery walls and that changes occurring in the walls of arteries with age impair such drainage. Several factors may be responsible for the impaired drainage of ISF and solutes from the white matter in LA in elderly brains. Mathematical models suggest that the motive force for the drainage of ISF along artery walls is the contrary wave that follows each pulse wave and that the motive force would decrease when the amplitude of the pulse wave is reduced in stiffened arteriosclerotic arteries. Both arteriosclerosis and arteriolosclerosis in the elderly brain are characterised by loss of smooth muscle cells and deposition of collagen in artery walls. Such a change in architecture would interfere with the drainage pathways that depend upon intact smooth muscle cell basement membranes in the walls of the arteries.Brain microvascular endothelial cells, the inner liner of blood-brain barrier, are involved in many physiological and pathophysiological processes in the brain such as transport of nutrients from blood to brain, export of critical toxins from brain, transmigration of circulating leukocytes and formation of new blood vessels. Endothelial dysfunction, which can lead to breakdown of the blood-brain barrier, impaired cerebral autoregulation and prothrombotic changes, is believed to be important in mediating disease. Intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) are two important adhesion molecules that are upregulated during endothelial activation. Both belong to the immunoglobulin superfamily and have adhesion molecules on leukocytes as ligands, namely the integrins CD11/CD18 for ICAM-1 and very late antigen-4 (VLA-4) for VCAM-1. Cytokines formed immediately after ischemia stimulates the expression of adhesion molecules on endothelial cells and leukocytes, leading to leukocyte adherence and extravasation into brain parenchyma. White matter lesions can be experimentally induced in rat brains under chronic cerebral hypoperfusion by permanent occlusion of both common carotid arteries.WM lesions have a strong relationship with oxidative stress, apoptosis and inflammatory damage. ICAM levels are related to progression of WMH on MRI. A prospective study design indicates the likelihood that this association is causal and supports a role of endothelial cell activation in the pathogenesis of LA. Studies of circulating endothelial markers are beginning to hint at a specific pattern of endothelial function that is associated with leukoaraiosis.Lipoic acid, a neuroprotective metabolic antioxidant has been used to improve glycemic control, treat polyneuropathies associated with diabetes mellitus, and mitigate toxicities associated with heavy metal poisoning. Biological assays and genomic analysis revealed that lipoic acid can modulate endothelial cell behavior and gene expression. Lipoic acid can inhibit expression of ICAM-1 and VCAM-1 at the mRNA level in cultured human aortic endothelial cells Administration of lipoic acid to patients with the metabolic syndrome can improve endothelial function and reduce proinflammatory markers.In the present study, (1) we induced WMLs in a rat chronic cerebral hypoperfusion model by permanent occlusion of both common carotid arteries. We then tested the hypothesis that the ICAM-1 and VCAM-1 upregulation after chronic cerebral hypoperfusion correlates with endothelial dysfunction, which leads to inflammatory damage and subsequent WM lesions. (2) we test the hypothesis that disruption of the structure of artery walls by the deposition of collagen is significantly greater in patients with LA than in normal white matter in age-matched controls. We performed immunohistochemistry on the components of extracellular matrix of the walls of arterioles in LA patients and in controls. In addition to immunohistochemical studies, we present a quantitative analysis of the pathology of arterioles, including the external diameter of arterioles, diameter of arteriolar lumina, thickness of arteriolar walls and sclerotic indices of arterioles in the white matter and cerebral cortex.Methods Part I: One hundred and fourty-four adult male Wister rats weighing 190 to 220g were divided at random into the following 3 groups: (1)the Lipoic acid group (n=60) were provided daily 1.1ml s.c. injections of LA at 100 mg/kg after ligation of both common carotid arteries (LBCCA); (2) the saline ( vehicle ) group: rats of this group (n=60) underwent LBCCA but were provided daily 1.1 ml s.c. injections of saline( vehicle ); and (3) the control sham-operated vehicle- and lipoic acid-treated groups: these rats (n=6 for each subgroup) underwent the same aforementioned protocol except for LBCCA. RT-PCR and double immunofluorescence for ICAM-1, VCAM-1, endothelial cells (staining positive for von Willebrand factor, vWF), reactive astrocyte (staining positive for glial fibrillary acidic protein, GFAP) and activated microglia/macrophages (CD11b/c staining) were analyzed at baseline and at 1, 3, 7, 14 and 28 days after hypoperfusion. The severity of the WM lesions in the corpus callosum, internal capsule, and external capsule of both hemispheres was graded by Luxol fast blue staining. Part II: Twenty brains were obtained from elderly LA patients (12 females and 8 males, mean age 66.4±2.9 years, range 62 to 78 years); ten brains from elderly individuals (5 females and 5 males, mean age 64.3±7.1 years, range 60 to 73 years) without known cerebrovascular disease served as controls. (1)We performed a quantitative analysis of the pathology of arterioles, including the external diameter of arterioles, diameter of arteriolar lumina, thickness of arteriolar walls and sclerotic indices of arterioles in the white matter and cerebral cortex. (2)We used immunocytochemistry to quantify changes to the extracellular matrix in arterioles of cerebral white matter (WM) in 20 autopsy cases with leukoaraiosis and in 10 controls.Results: (1) RT-PCR and double immunofluorescence analysis of white matter from rats that had received lipoic acid (100 mg/kg/day) exhibited markedly reduced expression of ICAM-1 and VCAM-1 over endothelial cells compared with that of rats receiving saline( vehicle) (P<0.01). In the rats treated with lipoic acid, the WM lesions after chronic cerebral hypoperfusion were significantly less severe, and the number of reactive astrocyte (GFAP staining) and activated microglia/macrophages (CD11b/c staining) were also significantly lower as compared with the saline(vehicle)-treated rats (P<0.01). (2) The ratio of the area immunoreactive for collagen types I , III, V, and VI to the cross-sectional area of arterioles was significantly higher in the LA patients compared to controls (P<0.001). Collagen IV and laminin were increased in the basement membrane. The walls of WM arterioles were significantly thicker (P<0.01), and lumina were significantly smaller (P<0.01). Arterioles had a significantly higher more sclerotic index (1-(internal/external diameter)) in LA than in adjacent cortex or control white matter (P<0.01).Conclusions: (1) These findings indicate that endothelial dysfunction plays a critical role in overexpression of ICAM-1 and VCAM-1, glial cell activation and WM lesions after chronic cerebral hypoperfusion and suggest the potential value of lipoic acid as a therapeutic tool in cerebrovascular WM lesions. (2) Our results also provide support for endothelial activation being involved in early pathogenesis of WM lesions and suggest that therapies that stabilize the endothelium may have a role in preventing WM lesions progression. (3) Our results suggest that fibrosis and thickening of the walls of arterioles in cerebral white matter in LA are due to the accumulation of extracellular matrix components. Although these changes may result in decreased perfusion, they could also impede perivascular drainage of interstitial fluid from white matter in LA.