Elevated expression of galectin-3 in microglia exacerbated neuron apoptosis via promoting TNF-α release through the TLR4/NF-κB signaling pathway

High blood levels of galectin-3 (Gal-3) predict poor outcomes after intracerebral hemorrhage (ICH). Our previous study also showed that Gal-3 could aggravate ICH-induced brain injury through increasing neuroinflammatory activation and nerve cell death. In this study, we focus on the role of Gal-3 in nerve cell death after ICH. An ICH mice model and an in vitro co-stimulation model were established to study Gal-3's effect on neuron cell death via toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB) pathway. Western blot and immunofluorescence (IF) staining were applied for neuron apoptosis evaluation. Enzyme-linked immunosorbent assay (ELISA) was used to measure the production of neuroinflammation factors. Gal-3 expression in microglia was increased and positively correlated with the severity of neurological impairment after ICH. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and NeuN (or MAP2) double staining assay results revealed that the increasing of neuron cell apoptosis after Gal-3 treatment both in in vivo and in vitro co-stimulation experiments could be reversed by treatment with Gal-3 inhibitor MCP, TLR4 inhibitor TAK-242, NF-κB inhibitor PDTC, or TNF-α inhibitor C87 effectively. ELISA results revealed the same trends of TNF-α release changes from microglia after Gal-3 or inhibitor treatment both in vivo and in vitro. WB results confirmed the Gal-3's role on apoptosis by the expression level of proteins such as FADD, Apaf-1, Bax, Cytochrome C, Caspase-8, and cleaved-Caspase-3 in neuron cells. The upregulation of Gal-3 in microglia after ICH could aggravate neuron cell apoptosis through increasing TNF-α release via TLR4/NF-κB pathway.