Altered expression of claudin family proteins in Alzheimer's disease and vascular dementia brains

Abstract

Claudins (Cls) are a multigene family of transmembrane proteins with different tissue distribution, which have an essential role in the formation and sealing capacity of tight junctions (TJs). At the level of the blood-brain barrier (BBB), TJs are the main molecular structures which separate the neuronal milieu from the circulatory space, by a restriction of the paracellular flow of water, ions and larger molecules into the brain. Different studies suggested recently significant BBB alterations in both vascular and degenerative dementia types. In a previous study we found in Alzheimer's disease (AD) and vascular dementia (VaD) brains an altered expression of occludin, a molecular partner of Cls in the TJs structure. Therefore in this study, using an immunohistochemical approach, we investigated the expression of Cl family proteins (Cl-2, Cl-5 and Cl-11) in frontal cortex of aged control, AD and VaD brains. To estimate the number of Cl-expressing cells, we applied a random systematic sampling and the unbiased optical fractionator method. We found selected neurons, astrocytes, oligodendrocytes and endothelial cells expressing Cl-2, Cl-5 and Cl-11 at detectable levels in all cases studied. We report a significant increase in ratio of neurons expressing Cl-2, Cl-5 and Cl-11 in both AD and VaD as compared to aged controls. The ratio of astrocytes expressing Cl-2 and Cl-11 was significantly higher in AD and VaD as compared to aged controls. The ratio of oligodendrocytes expressing Cl-11 was significantly higher in AD and the ratio of oligodendrocytes expressing Cl-2 was significantly higher in VaD as compared to aged controls. Within the cerebral cortex, Cls were selectively expressed by pyramidal neurons, which are the ones responsible for cognitive processes and affected by AD pathology. Our findings suggest a new function of Cl family proteins which might be linked to response to cellular stress.

Fig 1

Expression of Cl-2, Cl-5 and Cl-11 in control (CON), AD and VaD brains, in the frontal cortex, Brodmann area 46/9. Cl-expressing cells are brownish, as a result of the immunohistochemical staining method with anti-Cl sera detected with avidin–biotin-peroxidase complex kit and DAB substrate. Expression of Cl-2, Cl-5 and Cl-11 was increased in neurons and neuronal fibres in AD and VaD brains, as compared to control brains. Expression of Cls was noticed mainly in the pyramidal neurons in CON, AD and VaD. Arrows are indicating Cl-expressing neurons. Bars: 30 μm.

Fig 2

Ratios of Cl-2, Cl-5 and Cl-11-expressing cells in control, AD and VaD brains, in the frontal cortex, Brodmann area 46/9. The neuronal, astrocyte or oligodendrocyte ratio was defined as (number of Cl-expressing cells / total number of cells) × 100. To compare the cell ratio between controls, AD and VaD (two by two) factorial ANOVA has been used, followed by the Fisher’s LSD post hoc test. Error bars mark the S.D.

Fig 3

Expression of Cl-2, Cl-5 and Cl-11 in oligodendrocytes in control (CON), AD and VaD brains, in the frontal cortex, Brodmann area 46/9. Cl-expressing cells are brownish, as a result of the immunohistochemical staining method with anti-Cl sera detected with avidin–biotin-peroxidase complex kit and DAB substrate. Oligodendrocytes are indicated by arrows. Bars: 5 μm.

Fig 4

Expression of Cl-2, Cl-5 and Cl-11 in astrocytes in control (CON), AD and VaD brains, in the frontal cortex, Brodmann area 46/9. Cl-expressing cells are brownish, as a result of the immunohistochemical staining method with anti-Cl sera detected with avidin–biotin-peroxidase complex kit and DAB substrate. Astrocytes are indicated by arrows. Bars: 5 μm.

Fig 5

Expression of Cl-2, Cl-5 and Cl-11 in brain microvessels in control (CON), AD and VaD brains, in the frontal cortex, Brodmann area 46/9. Cl-expressing cells are brownish, as a result of the immunohistochemical staining method with anti-Cl sera detected with avidin–biotin-peroxidase complex kit and DAB substrate. Endothelial cells expressing Cl-2, Cl-5 or Cl-11 are indicated by arrows. Bars: 5 μm.