Figure 1.

HMGB1 present in necrotic extract induces HIV-1 replication in U1 cells. (a) Western blot of cell supernatants (necrotic extracts) obtained after freeze-thawing cycles of peripheral blood mononuclear cells (PBMC) (30 × 10⁶ cells/mL) from healthy donors: Molecular weight marker (I); supernatants after immune depletion of HMGB1 with nonspecific rabbit polyclonal antibody (II); depletion with anti-HMGB1 antibody −5 μg (III) and 10 μg (IV); necrotic extract loaded 20 μL (V), 10 μL (VI), and 5 μL (VII); 100 ng (VIII) and 75 ng (IX) of recombinant HMGB1; cell debris (X). Numbers to the left depict positions of molecular mass markers (in kDa). (b) Levels of HIV p24 protein in cell culture supernatants after 72 h incubation of U1 cells with necrotic extract (HMGB1 concentration 1 μg/mL): HMGB1-depleted necrotic extract and mock cells. PMA served as a positive control (20 nM). The levels of viral replication were approximately 2-fold higher after stimulation by necrotic extract compared to the mock cells (P = 0.002). Results from three independent experiments in duplicates are presented.