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Fig. 4 | Journal of Neuroinflammation

Fig. 4

From: Lipocalin 2 induces neuroinflammation and blood-brain barrier dysfunction through liver-brain axis in murine model of nonalcoholic steatohepatitis

Fig. 4

Lcn2 is correlated with increased HMGB1 secretion from brain cells. a Immunoreactivity of HMGB1-DAMP as shown by immunohistochemistry in brain slices from mice fed with CHOW diet serves as a control and MCD diet as NASH. Images were taken in × 20 in cerebral cortex area of brain. Immunoreactivity was indicated with black arrows. b Morphometric analysis of HMGB1 immunoreactivity in CHOW and MCD fed mouse groups. Morphometric analysis was performed by taking mean % ROI values from three separate fields (designed on y-axis). c Co-localization and morphometric analysis of HMGB1 reactivity with endothelial cells, microglia, astrocytes, and neurons marked as HMGB1/CD31, HMGB1/CD11b, and HMGB1/GFAP. HMGB1/NeuN in brain slices from MCD mouse group. Co-localization was displayed yellow dots in C, highlighted by white circles. Images were taken at × 40. Significance was tested between the groups by unpaired t test, *p < 0.05, **p < 0.01, ***p < 0.001. e Immunoblot analysis of HMGB1 and Lcn2 with protein extracts from cerebral cortex from CHOW and MCD fed mouse group. f Morphometry analysis of HMGB1 and Lcn2 immunoblot with protein extracts from cerebral cortex, normalized against β-actin, and plotted as bar graph. g HMGB1 ELISA was performed with supernatants from brain endothelial cells (24p3R+), and 24p3RsiRNA exposed brain endothelial cells (24p3R−) treated with VEHICLE (0.05% DMSO), mouse recombinant Lcn2 (100 ng/ml), 20 μl of serum from CHOW, and MCD fed mouse. HMGB1 concentration (ng/ml) was plotted as bar graph. Significance was tested by unpaired t test and pair t test between the groups of 24p3R+ and 24p3R− ,*p < 0.05, **p < 0.01, ***p < 0.001, ns, nonsignificant; p ≥ 0.05

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