Figure 9. Increased IL-1β, HMGB1 and HMGB1/IL-1β complexes in the hippocampus of human alcoholics.
Postmortem human hippocampal brain tissue was obtained from the New South Wales Tissue Bank. Protein was isolated from frozen tissue and assessed for IL-1β and HMGB1 protein level by ELISA. (A) Human alcoholics showed a 42% increase in IL-1β above healthy controls: 404.5 ± 51.5 vs 573.4 ± 99.2pg/mg, Control vs Ethanol, *p<0.05, paired two-tailed t-test, N=10 per group. (B) Human alcoholics showed a 24% increase in HMGB1 (157.8 ±24.7 vs 195.5± 60.1 ng/mg, Control vs Ethanol, *p<0.05, paired two-tailed t-test, N=10 per group. Note: Data published previously from this same cohort of patients in a different form in Coleman et al J Neuroinflammation 2017. (C) Co-Immunoprecipitation for HMGB1 and IL-1β was performed on postmortem human hippocampus from healthy controls and alcoholics. Eluate was probed for either HMGB1 or IL-1β. HMGB1+ bands were observed at the expected 25–29kD MW. An IL-1β+ band was also observed at ~17kD consistent with cleaved IL-1β, indicating the presence of HMGB1-IL-1β complexes. Quantification of optical densitometry of each band found an increased ratio of IL-1β to HMGB1 in eluate consistent with increased HMGB1-IL-1β complex formation in the hippocampus of human alcoholics. Alcoholics showed a 275% increase in IL-1β association with HMGB1 relative to control. *p<0.05, paired t-test, N=4 per group (D) Theoretical model of enhanced HMGB1/IL-1β signaling