Inflammatory responses after endothelin B (ETB) receptor activation in human monocytes: new evidence for beneficial anti-inflammatory potency of ETB-receptor antagonism.

Endothelin (ET) stimulates potent ETA/ETB receptors important in the pathogenesis of pulmonary arterial hypertension (PAH) and fibrosis. Though therapy with ET-receptor antagonists is well established uncertainty exists whether selective ETA or dual ETA/ETB-receptor antagonism is superior in PAH. The objective of this study was to further elucidate the pro-inflammatory effects of ET-1 on ETB receptors in cultured human monocytes (10(5)/20 h) compared with non-specific stimulation with LPS in vitro and to define the antagonizing effects of bosentan, a dual ETA/ETB-receptor antagonist, on inflammatory mediator production. We further hypothesized that ETB-receptor antagonism reduces the requirement of PGE2 to control inflammatory mediator production. Activation of the monocyte ETB subtype by ET (1 ng/ml) concentration-dependently stimulated TNF-alpha (744%) >PGE2 (570%) > IL-1 beta (112%) and had no effect on 5-lipoxygenase metabolism. Compared with ET a different profile of IL-1 beta >TNF-alpha >PGE2 was induced by LPS. ETB-receptor antagonism attenuated ET- and LPS-responses in monocytes, in particular of TNF-alpha and PGE2 to a similar extend (40%) that were only demonstrable following LPS at therapeutic plasma concentrations of bosentan and had no effect on IL-1 beta. Inhibition of ETB receptors in LPS-stimulated monocytes by bosentan was responded with suppression of PGE2 and increased production of leukotrienes indicating strong effects in the cyclooxygenase pathway that is known to control cellular ET transcription. These data suggest an important signaling pathway between ET-induced cytokine production following ETB-receptor activation with no further control of ET transcription by PGE2 required following ETB receptor antagonism. Therefore, in states of inflammation increased ETB-receptor expression and activation mediated by elevated ET concentrations may be an underestimated mechanism, which warrants the application of combined ETA/ETB-receptor antagonists.

New evidence of H1-receptor independent COX-2 inhibition by fexofenadine HCl in vitro.

BACKGROUND/AIMS: Fexofenadine HCl (FEX) has previously been shown to have anti-inflammatory properties in relieving nasal congestion in allergic rhinitis. The objective of this study was to further elucidate the mechanism of action behind the anti-inflammatory properties of FEX in addition to its H(1)-receptor antagonism. METHODS: The effects of two antihistamines, FEX and loratadine (LOR), were investigated on cyclooxygenase (COX)-1 and -2 enzymes in vitro. FEX (10(-9)-10(-3) mol/l) and LOR (10(-9)-10(-4) mol/l) were incubated with arachidonic acid in a COX screening assay with either ovine COX-1 or COX-2 or human COX-2. COX-2 enzyme inhibitory activity for the antihistamines was compared with the known selective COX-2 inhibitor DuP-679. RESULTS: High concentrations of FEX (10(-3) mol/l) significantly inhibited arachidonic acid-mediated ovine COX-1 activity, but low concentrations had no effect. Low concentrations of FEX (10(-8) mol/l) inhibited ovine COX-2 activity, and this inhibition decreased with increasing concentrations. The inhibition of COX-2 activity by FEX was similar to that seen with the selective COX-2 inhibitor, DuP-679. Conversely, LOR inhibited COX-1 activity at low concentrations (10(-8) mol/l), but had little inhibitory effect on COX-1 at high concentrations. LOR (10(-5) mol/l) markedly stimulated COX-2 activity. CONCLUSION: FEX showed selective arachidonic acid-mediated COX-2 inhibitory enzyme activity, which differed markedly from the COX inhibitory enzyme activity of LOR. This selective COX-2 inhibitor activity by FEX may contribute to its anti-inflammatory properties in relieving nasal congestion in allergic rhinitis.

Differential effects of fexofenadine on arachidonic acid metabolism in cultured human monocytes.

The relief of nasal congestion with the antihistamine fexofenadine in seasonal allergic rhinitis is thought to be due to its additional anti-inflammatory properties. The objective of this study was to evaluate the in vitro effects of fexofenadine on stimulated arachidonic acid metabolism. Human monocytes, isolated from blood and donated by 5 healthy volunteers, were either incubated for 20 h with 10 microg/ml lipopolysaccharide, with and without fexofenadine (10(-8)-10(-3) mol/l, n = 8-19), or were incubated for 20 h, with and without fexofenadine, and then stimulated with 0.5 mg/ml zymosan for 2 h. Leukotriene B4 (LTB4), LTC4, LTD4 and LTE4, prostaglandin E2 (PGE2) and F2alpha (PGF2alpha) production was determined by enzyme immunoassay. Zymosan-stimulated production of LTC4, LTD4 and LTE4 was significantly inhibited by clinically relevant concentrations of fexofenadine HCl: 10(-7) mol/l (22% inhibition vs. control, p = 0.008) and 10(-6) mol/l (24% inhibition vs. control, p = 0.020). Higher concentrations of fexofenadine (10(-4) and 10(-3) mol/l) inhibited LTB(4) generation. Lipopolysaccharide-stimulated production of PGE2 was significantly inhibited by fexofenadine HCl 10(-6) mol/l (26% inhibition, p = 0.035) and 10(-5) mol/l (40% inhibition, p = 0.001). Higher concentrations of fexofenadine HCl (10(-4) and 10(-3) mol/l) significantly inhibited PGF2alpha production by 50% (p = 0.026) and 63% (p = 0.001), respectively. Fexofenadine, at both clinically relevant and higher concentrations, inhibits LTC4, LTD4, LTE4 and PGE2 in cultured human monocytes. These additional anti-inflammatory properties may underlie the relief of nasal congestion observed in clinical studies.

Inhibitory activity of 1,8-cineol (eucalyptol) on cytokine production in cultured human lymphocytes and monocytes.

BACKGROUND: The therapeutic value of secretolytic agents in COPD and asthma is still disputed. For this reason, in a preclinical study we aimed to test the potential anti-inflammatory efficacy of 1,8-cineol (eucalyptol) in inhibiting polyclonal stimulated cytokine production by human unselected lymphocytes and LPS-stimulated monocytes. METHODS: Cytokine production was determined following 20 h of incubation cells with 1,8-cineol simultaneously with the stimuli in culture supernatants by enzyme immunoassay. RESULTS: Therapeutic concentrations of 1,8-cineol (1.5 microg/ml=10(-5)M) inhibited significantly (n=13-19, p=0.0001) cytokine production in lymphocytes of TNF-alpha > IL-1beta> IL-4> IL-5 by 92, 84, 70, and 65%, respectively. Cytokine production in monocytes of TNF-alpha > IL-1beta> IL-6> IL-8 was also significantly (n=7-16, p<0.001) inhibited by 99, 84, 76, and 65%, respectively. In the presence of 1,8-cineol (0.15 microg/ml=10(-6)M) production of TNF-alpha>IL-1beta by monocytes and of IL-1beta> TNF-alpha by lymph-ocytes was significantly inhibited by 77, 61 and by 36, 16%, respectively. 1,8-cineol (10(-6)M) had a larger impact on TNF-alpha and IL-1beta-production in monocytes compared to lymphocytes (p<0.03) and similar effects (p>0.59) at therapeutically relevant concentrations of 1,8-Cineol (10(-5)M). CONCLUSION: These results characterize 1,8-cineol as strong inhibitor of TNF-alpha and IL-1beta and suggest smaller effects on chemotactic cytokines. This is increasing evidence for the role of 1,8-cineol to control airway mucus hypersecretion by cytokine inhibition, suggesting long-term treatment to reduce exacerbations in asthma, sinusitis and COPD.