Inhibition of HMGB1 With Neutralizing Antibody Reduces Retinal Cell Viability and Disturbs Structural Integrity of the Retina

Abstract

Purpose: : High-mobility group box 1 protein (HMGB1) is thought to be required for proper transcriptional regulation in the nucleus as well as cellular motility in the cell surface. The purpose of this study is to determine whether recombinant HMGB1 or inhibition of HMGB1 with neutralizing antibody affects the retinal neuron in vitro and in vivo.

Methods: : An immortalized retinal precursor R28 cell was used to assess the impacts of recombinant HMGB1 and antibody blockade of HMGB1. The expression of RAGE, one of the receptors for HMGB1, was analyzed by immunoblotting. Cells were treated by recombinant HMGB1 or HMGB1-neutralizing/ control IgY, then cell viability was measured by MTT assay. Brown Norway rats received either a subretinal injection of HMGB1-neutralizing IgY or control IgY. Retinas were evaluated using morphology. The distributions of IgY in the treated eyes were confirmed by immunohistochemistry.

Results: : Although RAGE was detected in R28 cells, recombinant HMGB1 did not affect cell viability with relatively low concentrations in vitro. Whereas, antibody blockade of HMGB1 significantly reduced the viability. HMGB1-neutralizing IgY also disrupted the retinal structural integrity and promoted loss of photoreceptors in vivo, compared to the control.

Conclusions: : Retinal neuronal cell was affected by antibody blockade of HMGB1, but not recombinant HMGB1. Thus, endogenous HMGB1 might be sufficient for its roles in the retina. These findings suggest that HMGB1 is an essential protein for retinal cell survival and the structural integrity of the retina.

Keywords: retina • cell survival • retinal detachment