Effect of 5-Aminosalicylic Acid and Analogous Substances on Superoxide Generation and Intracellular Free Calcium in Human Neutrophilic Granulocytes

Abstract

Activated polymorphonuclear leukocytes (PMNs), which are found in the inflammatory lesions of chronic inflammatory bowel disease, produce tissue-destructive oxygen-derived free radicals. The influence of 5-aminosalicylic acid (5-ASA), its acetylated metabolite (Ac-5-ASA), sulfasalazine (SAZ), and olsalazine (OLZ) (5-ASA dimer linked by an azo group) in pharmacologically relevant concentrations (0.1-10 mM) were tested on PMN superoxide production with either the receptor-specific agent formyl-methionyl-leucyl-phenylalanine (fMLP) or the protein kinase C activator phorbol myristate acetate (PMA). Inhibition of receptor-specific superoxide production occurred at 0.07, 0.32, and 0.63 mM (IC50 values) for 5-ASA, SAZ, and OLZ, respectively. No inhibitory effects of SAZ and OLZ were observed when PMA was applied as stimulus for PMN superoxide production. The results indicate that the signal to which PMNs respond by generating superoxide is primarily due to calcium release from intracellular stores. They further suggest that SAZ and OLZ may affect the oxygen-derived free radical production in human PMNs by unspecific cytotoxicity or by interference with the nicotinamide adenine dinucleotide phosphate, reduced (NADPH) oxidase system, whereas 5-ASA itself is a free radical scavenger.

Flavonoids and 5-Aminosalicylic Acid Inhibit the Formation of Neutrophil Extracellular Traps

“…The concentrations of antioxidants were chosen either based on the literature [22, 23, 36–41] or on our own preliminary studies. To exclude toxic or apoptosis-inducing effects, freshly isolated human neutrophils were incubated with the antioxidants over 5 h and double-stained with annexin V-FITC and propidium iodide.…”

“…This hypothesis is underlined by the demonstration of extracellular DNA in large bowel tissues from patients suffering from a bacterial gastrointestinal infection and Crohn's disease [65] and by activation of NET release in response to long pentraxin 3 (PTX3) [66, 67]. 5-ASA is an effective scavenger of neutrophil-derived ROS such as superoxide anions, hydroxyl radicals, singlet oxygen, amino chlorides, and especially hypochlorite [25, 36, 68, 69]. An effect of 5-ASA on MPO is discussed controversly.…”

Superoxide production and expression of NAD(P)H oxidases by transformed and primary human colonic epithelial cells

“…16 In brief, blood was sampled in EDTA containing vials and subjected to sequential Dextran sedimentation (Lymphoprep; Nycomed, Oslo, Norway) and gradient centrifugation. Contaminating erythrocytes were lysed using hypotonic saline, and normal osmolality was restored by the addition of hypertonic saline followed by centrifugation at 2000 g for 10 minutes at 4°C.…”

Prevention of post‐operative recurrence of Crohn’s disease requires adequate mucosal concentration of mesalazine

“…Moreover, mesalazine inhibits the synthesis of many cytokines such as IL‐1, TNF‐alpha IFN‐gamma, which are primarily involved in the pathogenesis of Crohn’s disease 26 –28 . All these anti‐inflammatory effects require an in vitro threshold concentration of the drug and are dose‐dependent 29 –32 . In vivo data are lacking.…”