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Juan C. Cendan, MD; Lyle L. Moldawer, PhD; Wiley W. Souba, MD, ScD; Edward M. Copeland, III, MD; D. Scott Lind, MD

Arch Surg. 1994;129(12):1296-1300.

Abstract
Background
L-Arginine is the sole precursor of nitric oxide (NO). Bacterial lipopolysaccharide (endotoxin) (LPS) stimulates carrier-mediated L-arginine transport in porcine pulmonary artery endothelial cells (PAECs) through an autocrine pathway that involves interleukin-1 (IL-1) and tumor necrosis factor (TNF-).

Objectives
To determine if Escherichia coli LPS stimulates NO synthesis in PAECs and, if so, if LPS stimulation of NO production is also mediated by autocrine secretion of IL-1 and TNF-.

Design
Monolayers of PAECs were incubated with various concentrations of LPS, recombinant human TNF-, or IL-1, and total nitrate-nitrite accumulation was measured at different time points with the Greiss reagent following cadmium reduction. Release of TNF- and IL-1 release by LPS-stimulated PAECs were measured using the WEHI (for TNF-) and A375.S2 (for IL-) bioassays. The PAECs were then incubated with saline solution or LPS in the presence or absence of either a polyclonal antibody to human TNF or IL-1 receptor antagonist, and nitratenitrite accumulation was measured at 48 hours.

Results
Production of NO by PAECs was increased 230% by LPS (1 µg/mL), 350% by TNF- (1000 U/mL), and 240% by IL-1 (1000 U/mL) (P<.05 vs control). The LPS-stimulated NO production was inhibited by IL-1 receptor antagonist (100 µg/mL) or antibody to TNF (10 µg/mL) to control levels (P<.05 vs LPS; difference vs saline solution was not significant). The LPS-stimulated TNF- secretion by PAECs and TNF- activity were maximal at 6 hours (400±42 pg/mL). The IL-1 activity was not detectable in LPS-stimulated PAECs by the A375.S2 bioassay.

Conclusions
Endotoxin, TNF-, and IL-1 stimulated NO synthesis in PAECs. Endotoxin-stimulated NO synthesis through an autocrine pathway involving the cytokines TNF- and IL-1.

(Arch Surg. 1994;129:1296-1300)

Author Affiliations
From the Department of Surgery, University of Florida, Gainesville (Drs Cendan, Moldawer, Copeland, and Lind), and the Division of Surgical Oncology, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston (Dr Souba).

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