Supplementary Material for: Glial Network Responses to Polymicrobial Invasion of Dentin

This study investigated the distribution patterns of glial networks disclosed by reactivity for glial fibrillary acidic protein (GFAP) and S100B in healthy and carious human teeth. The objective was to determine the assembly and collapse of glial networks in response to encroaching infection. 15 healthy and 37 carious posterior teeth from adults were studied. Immediately after extraction, teeth were cleaned and vertically split and the half with pulp fixed and prepared for resin or frozen sections. Sections were stained with toluidine blue and for immunofluorescence, with observation by confocal laser microscopy and analysis by ImageJ software. Carious teeth were subdivided into three groups according to degree of carious involvement: microbial penetration through enamel (stage A), extension into dentin (stage B) and advanced penetration into dentin but without invasion of underlying pulp tissue (stage C). In stage A lesions there was marked increase in glial networks in dental pulp tissue that extended beyond the zone of microbial invasion. This response was maintained in stage B lesions. In advanced stage C lesions these networks were degraded in the zone of invasion in association with failure to contain infection. Cells expressing the glial markers GFAP and S100B showed a response to initial microbial invasion of dentin by increase in number and altered anatomical arrangement. The late stage of dentinal caries was marked by collapse of these networks in the region adjacent to advancing bacteria. This behaviour is important for understanding and explaining the defensive response of the neurosensory peripheral dental pulp apparatus to infection