HYPOXIA INDUCES A HUMAN MACROPHAGE CELL-LINE TO RELEASE TUMOR-NECROSIS-FACTOR-ALPHA AND ITS SOLUBLE RECEPTORS IN-VITRO

Tissue hypoxia following hemorrhage and trauma is a possible initiating factor of the generalized inflammatory response seen after shock. The role of hypoxia in the release from a human monocyte cell line (THP-1) of tumor necrosis factor-α (TNFα) and its soluble membrane receptors (TNFαR) in- vitro is investigated in this study. Flat-bottom plates with 500,000 THP-1 cells/ml were placed in air-tight sealed boxes and exposed to hypoxia (O2 = 1%) or controls (O2 = 9%) for up to 24 hr. Supernatants were tested for TNFα, as well as 55- and 75-kDa soluble receptors for TNFα, by ELISA. Cell viability was assessed by vital dye uptake and was found to be maintained throughout hypoxic exposure. Medium pH levels were within normal range. In eight experiments conducted in duplicate, minimal change over 24 hr occurred in control samples. Control mean and SD were: TNFα = 12.0 ± 4.2, 55-kDa R = 34.6 ± 2.03, and 75-kDa R = 38.88 ± 9.68 pg/ml. During hypoxia, TNFα was released as early as the first 30 min of exposure (41.3 ± 2.3 pg/ml) with a small peak at 1 hr (52 ± 5.0 pg/ml) and a later more pronounced peak at 18 hr (526 ± 48 pg/ml). Both 55- and 75-kDa R were released by the hypoxic monocytes; release was progressive and was maximal at 24 hr in this study. Maximal release value of 55-kDa R was 236 ± 15 pg/ml, while for 75-kDa R it was 2450 ± 63 pg/ml. By one-way analysis of variance followed by the Student-Newman-Keuls test for multiple comparisons, P values were <0.01 at all but the 15-min time period, where P > 0.05. We conclude that when exposed to complete or nearly complete absence of oxygen, human monocytes spontaneously release TNFα, followed by TNFαR. © 1993 Academic Press. All rights reserved.