Interleukin 1β and tumour necrosis factor α inhibit acid secretion in cultured rabbit parietal cells by multiple pathways

Abstract

Background—The cytokines interleukin 1β (IL-1β) and tumour necrosis factor α (TNF-α) are inhibitors of gastric acid secretion when administered systemically. 
Aims—To investigate the inhibitory effect of IL-1β and TNF-α on cultured, acid secreting parietal cells in order to determine the mechanism of this inhibition. 
Methods—Rabbit parietal cells were prepared by collagenase-EDTA digestion and counter flow elutriation. Acid secretory activity was assessed by aminopyrine accumulation. 
Results—IL-1β and TNF-α inhibited basal and stimulated acid secretion in a dose dependent manner; near maximal effects were seen with both at 10 ng/ml. Inhibition was maximal with 15 minutes pretreatment but seen with up to 18 hours of preincubation. Both cytokines inhibited histamine, carbachol, gastrin, forskolin, and A23187 stimulated acid secretion but had no effect on stimulation by dibutyryl-cAMP. Inhibition of acid secretion was not accompanied by a change in radioligand binding to histamine H₂ or gastrin/CCK_B receptors. Pertussis toxin abolished the inhibitory effects on histamine and forskolin stimulation. The tyrosine kinase inhibitor herbimycin reduced the inhibitory effects of TNF-α against all stimuli but only reduced the effects of IL-1β against histamine and forskolin stimulation. 
Conclusions—IL-1β and TNF-α seem to inhibit parietal cell acid secretion by multiple pathways; the inhibition occurs at postreceptor level and involves pertussis toxin and tyrosine kinase dependent and independent pathways. Mucosal production of cytokines may be important in the regulation of gastric acid secretion. 



Keywords: acid secretion; aminopyrine; cytokines; interleukin 1β; tumour necrosis factor α; parietal cell

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Figure 1 .

Effect of increasing concentrations of TNF-α on aminopyrine (AP) uptake in cultured rabbit parietal cells. Cells were incubated with cytokine for 15 minutes and then aminopyrine accumulation was measured over 30 minutes. Uptake of aminopyrine was measured in the basal state and when stimulated by histamine (10⁻⁴ M) and carbachol (10⁻⁴ M). Results are expressed as percentage (mean (SEM)) of basal control (n=6). *p<0.05; **p<0.01. 


Figure 2 .

Effect of increasing concentrations of IL-1β on aminopyrine (AP) uptake in cultured rabbit parietal cells. Cells were incubated with cytokine for 15 minutes and then aminopyrine accumulation was measured over 30 minutes. Uptake of aminopyrine was measured in the basal state and when stimulated by histamine (10⁻⁴ M) and carbachol (10⁻⁴ M). Results are expressed as percentage (mean (SEM)) of basal control (n=6). *p<0.05; **p<0.01. 


Figure 3 .

Effect of TNF-α (10 ng/ml) on aminopyrine accumulation by parietal cells in the basal state and when stimulated by histamine (His; 10⁻⁴ M), forskolin (Fsk; 10⁻⁵ M), dibutyryl-cAMP (dbcAMP; 10⁻⁴ M), carbachol (Cbh; 10⁻⁴ M), gastrin-17 (Gas; 10⁻⁷ M), or A23187 (10⁻⁶ M). Cells were exposed to TNF-α for 15 minutes before stimulation. Results are expressed as percentage of basal aminopyrine uptake in the absence of cytokine (mean (SEM)), n=5-6. *p<0.05; **p<0.01. 


Figure 4 .

Effect of IL-1β (10 ng/ml) on aminopyrine (AP) accumulation by parietal cell in the basal state or stimulated by histamine (His; 10⁻⁴ M), forskolin (Fsk; 10⁻⁵ M), dibutyryl-cAMP (dbcAMP; 10⁻⁴ M), carbachol (Cbh; 10⁻⁴ M), gastrin-17 (Gas; 10⁻⁷ M), or A23187 (10⁻⁶ M). Cells were exposed to IL-1β for 15 minutes before stimulation. Results are expressed as percentage of basal aminopyrine uptake in the absence of cytokine (mean (SEM)), n=5-6. *p<0.05; **p<0.01. 


Figure 5 .

Effect of TNF-α and IL-1β (both 10 ng/ml) on binding of ³H-histamine to rabbit parietal cells. Cells were incubated in appropriate cytokine for 15 minutes before performing the binding experiments. Data are expressed as percentage of binding in the absence of unlabelled histamine (B/Bo). Results are the mean (SEM) of three separate animal preparations. 


Figure 6 .

Effect of TNF-α and IL-1β (both 10 ng/ml) on binding of ¹²⁵I-CCK to rabbit parietal cells. Cells were incubated in appropriate cytokine for 15 minutes before performing the binding experiments. Data are expressed as percentage of binding in the absence of unlabelled gastrin (B/Bo). Results are the mean (SEM) of three separate animal preparations. 


Figure 7 .

Effect of pertussis toxin (PTX; 200 ng/ml) on TNF-α (10 ng/ml) inhibition of aminopyrine (AP) accumulation in the basal state or stimulated by histamine (His; 10⁻⁴ M), forskolin (Fsk; 10⁻⁵ M), carbachol (Cbh; 10⁻⁴ M), gastrin-17 (Gas; 10⁻⁷ M), or A23187 (10⁻⁶ M). Cells were incubated with PTX for two hours before addition of cytokine. Data are expressed as mean (SEM), n=6. *p<0.05 compared with the effect of TNF-α in the absence of PTX. 


Figure 8 .

Effect of pertussis toxin (PTX; 200 ng/ml) on IL-1β (10 ng/ml) inhibition of aminopyrine (AP) accumulation in the basal state or stimulated by histamine (His; 10⁻⁴ M), forskolin (Fsk; 10⁻⁵ M), carbachol (Cbh; 10⁻⁴ M), gastrin-17 (Gas; 10⁻⁷ M), or A23187 (10⁻⁶ M). Cells were incubated with PTX for two hours before addition of cytokine. Data are expressed as mean (SEM), n=6. *p<0.05 compared with the effect of IL-1β in the absence of PTX. 


Figure 9 .

Effect of the tyrosine kinase inhibitor herbimycin (10⁻⁶ M) on TNF-α (10 ng/ml) induced inhibition of aminopyrine (AP) uptake in rabbit parietal cells in the basal state or stimulated with histamine (His; 10⁻⁴ M), forskolin (Fsk; 10⁻⁵ M), carbachol (Cbh; 10⁻⁴ M), or A23187 (10⁻⁶ M). Herbimycin was added 60 minutes before TNF-α. Data are expressed as mean (SEM), n=4. *p<0.05 compared with the effect of TNF-α in the absence of herbimycin. 


Figure 10 .

Effect of herbimycin (10⁻⁶ M) on IL-1β (10 µg/ml) inhibition of aminopyrine (AP) accumulation in rabbit parietal cells in the basal state or stimulated with histamine (His; 10⁻⁴ M), forskolin (Fsk; 10⁻⁵ M), carbachol (Cbh; 10⁻⁴ M), or A23187 (10⁻⁶ M). Herbimycin was added 60 minutes before IL-1β. Data are expressed as mean (SEM), n=5. *p<0.05 compared with effect of IL-1β in the absence of herbimycin.