2nd Neuroscience Ireland Conference 329 The effects of blood–brain barrier disruption on glial cell function in multiple sclerosis Stephen McQuaid*† 1 , Paula Cunnea‡, Jill McMahon§ and Una Fitzgerald§ *Molecular Pathology Laboratory, Institute of Pathology, Royal Group of Hospitals Trust, Grosvenor Road, Belfast BT12 6BL, U.K., †Tissue Core Technology Unit, Queen’s University, 97 Lisburn Road, Belfast BT9 7BL, U.K., ‡Centre for Cancer Research and Cell Biology, Queen’s University, 97 Lisburn Road, Belfast BT9 7BL, U.K., and §National Centre for Biomedical Engineering Science, National University of Ireland, University Road, Galway, Ireland Abstract Dysfunction of the BBB (blood–brain barrier) is a major hallmark of MS (multiple sclerosis). Studies in our laboratories over the last decade have shown that increased BBB permeability is associated with decreased expression of TJ (tight junction) proteins in brain capillary endothelial cells. Results have revealed that TJ abnormalities were most common in active lesions (42% of vessels affected), but were also present in inactive lesions (23%) and in MS normal-appearing white matter (13%). Importantly, TJ abnormality was also positively associated with leakage of the serum protein fibrinogen which has recently been shown to be an activator of microglia. TJ abnormality and the resultant vascular permeability in both lesional and non-lesional white matter may impair tissue homoeostasis, which may have effects on disease progression, repair mechanisms and drug delivery. Introduction MS (multiple sclerosis) is an inflammatory demyelinating disease of the central nervous system associated with the development of focal lesions throughout the neural axis. BBB (blood–brain barrier) leakage has been recognized at the histological level in MS tissue since the early 1960s [1] and cli- nically since the advent of CT (computed tomography) and MRI (magnetic resonance imaging) in the 1980s [2]. Increas- ing evidence shows that BBB impairment and leakage in MS extends well beyond the CELs (contrast-enhancing lesions) which are associated with acute inflammation and demyelin- ation. Resolution of CELs is often considered a surrogate for BBB restoration either naturally or as a result of drug therapy. Lesions are invariably centred on blood vessels, and these become inflamed and leaky from an early stage [3]. Moreover, both MRI and confocal microscopic studies point to BBB disturbances in NAWM (normal-appearing white matter) in MS tissue that precede the formation of demyelinating lesions and also in inflammatory silent inactive lesions [4–8]. Key to the study of BBB impairment is a good under- standing of the structure associated with the endothelial cells comprising the barrier. TJs (tight junctions) are found as par- allel bands, tight apposition and towards the apical side of the lateral membrane contacts between adjacent endothelial cells. The main function of TJs is to restrict the paracellular passage of molecules through the BBB and to maintain endothelial po- larity [9]. Compromised BBB TJs are recognized as hallmarks of neuroinflammatory and other CNS (central nervous sys- Key words: blood–brain barrier, fibrinogen, glia, imaging, multiple sclerosis (MS), tight junction. Abbreviations used: BBB, blood–brain barrier; CEL, contrast-enhancing lesion; CNS, central nervous system; MRI, magnetic resonance imaging; MS, multiple sclerosis; NAWM, normal- appearing white matter; PPMS, primary progressive MS; SPMS, secondary progressive MS; TJ, tight junction; ZO-1, zonula occludens 1. 1 To whom correspondence should be addressed (email s.mcquaid@qub.ac.uk). tem) diseases. Dysfunctional TJs and subsequently impaired endothelial function allow more cells and immunologically active molecules access to the privileged CNS. Microscopical data In our laboratories, we have completed extensive human tissue-based studies using significant numbers of snap-frozen MS and control tissue samples [9–12]. These studies have revealed aspects of the pathological status of the BBB in active and inactive white matter lesions, in NAWM and in grey matter in the two most prevalent clinical presentations of MS: PPMS (primary progressive MS) and SPMS (secondary progressive MS). To achieve this, we have developed robust protocols for the immunocytochemical demonstration and assessment of proteins [ZO-1 (zonula occludens 1), occludin, junctional adhesion molecule-A] that are expressed in the TJs of the vascular endothelium. We also used dual-labelling to systematically assess the relationship between the expression of TJ proteins and leaked plasma protein (fibrinogen) in the different categories of MS tissue. Studies of histologically well-characterized white and grey matter samples in acute, PPMS and SPMS revealed endothelial abnormalities associated predominantly with active lesions (affecting 40% of vessels) and, to a lesser extent, inactive lesions (affecting 23% of vessels in SPMS and 37% of vessels in PPMS) and MS NAWM (13%). TJ abnormalities were more frequent in the normal-appearing grey matter in SPMS (23%) than in PPMS (10%). Vessels of all sizes were affected equally, supporting a causal role for diffusible inflammatory mediators. In active lesions, dual-labelling showed that 41% of vessels with severe TJ alterations showed pronounced fibrinogen leakage. In both NAWM and inactive lesions, the trend towards vessels with Biochem. Soc. Trans. (2009) 37, 329–331; doi:10.1042/BST0370329 C  The Authors Journal compilation C  2009 Biochemical Society Biochemical Society Transactions www.biochemsoctrans.org