HMGB1 is a Potential Mediator of Astrocytic TLR4 Signaling Activation following Acute and Chronic Focal Cerebral Ischemia

Figure 4

(a) Representative area of GFAP/TLR4 and iNOS/TLR4 staining following HMGB1 cortical injection. (b) Increase in TLR4 and iNOS expression in astrocytes following cortical injection of HMGB1 (n = 3). (A) Upregulation of TLR4 expression in perilesional astrocytes 48 hours following injection of recombinant HMGB1 into normal rat cortex. Immunofluorescence staining of representative 10 μm coronal brain sections; GFAP (green) expression and TLR4 (red) expression in perilesional astrocytes (merged) (upper panel). TLR4 expression in perilesional astrocytes following injection of PBS into normal rat cortex. GFAP (green) expression and TLR4 (red) expression in perilesional astrocytes (merged) in the contralateral hemisphere (lower panel). (B) Upregulation of iNOS expression in perilesional astrocytes 48 hours following injection of recombinant HMGB1 into normal rat cortex. Immunofluorescence staining of representative 10 μm coronal brain sections; GFAP (green) expression and iNOS (red) expression, in perilesional astrocytes (merged) (upper panel). iNOS expression in perilesional astrocytes following injection of phosphate buffered saline into normal rat cortex. GFAP expression (green) and TLR4 expression (red) in perilesional astrocytes (merged) in the contralateral hemisphere (lower panel). Brain sections are representative sections from 3 animals each injected with HMGB1 and 3 animals injected with PBS. Scale bar = 50 μm. (c) Percentage of TLR4-postitive astrocytes in the peri-injection site following injection of rHMGB1 into normal rat cortex. There is a trend towards an almost 2-fold increase in the percentage of TLR4-positive astrocytes in the peri-injection site following injection of rHGMB1 compared to injection of PBS; 69.7 ± 10.5% compared to 35.9 ± 31.4%; (n = 3 each per group for rHMGB1 injection and PBS injection).

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