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Figure 4 | BMC Physiology

Figure 4

From: Angiotensin II directly regulates intestinal epithelial NHE3 in Caco2BBE cells

Figure 4

Type I AII receptors mediate AII effects on NHE3 through activation of phospholipase C, epoxygenation of arachidonic acid, stimulation of the EGF receptor, and activation of phosphatidyl inositol 3 kinase and Akt. (A) RNA was harvested from a cell monolayer, reverse transcribed, and analyzed for AII type I and II receptors by PCR. (B) To determine the mechanism of action of AII, cell monolayers were treated basolaterally with the type I receptor antagonist losartan (100 μM) or the type II receptor antagonist PD123319 (100 μM) or with inhibitors of phosholipase C (U73122, 30 μM), cytochrome P450 fatty acid epoxygenase (ketoconazole, 30 μM), a metalloproteinase inhibitor (TAPI-1, 100 μM), EGF receptor tyrosine kinase (Tyrphostin AG1478, 30 μM), MEK-1 (PD98059, 30 μM), phosphatidyl inositol 3 kinase (wortmannin, 30 μM or LY294002, 30 μM), Akt (API-1, 30 μM) all for 30 min prior to addition of AII (1 nM). Fluxes were measured 1 hr after basolateral addition of AII. Values are means ± SEM for four separate experiments. * P < 0.05 + P < 0.01 as designed for comparisons by analysis of variance using a Bonferroni correction. (C) Cells were treated as for flux studies in B, however, cells were used for apical surface biotinylation as described in Methods. Image shown is representative of those of four separate experiments. Densitometric values were calculated by setting the intensity for no addition to 100% in each experiment and calculating densitometric changes of other groups (decreases or increases) relative to this value. Values are means ± SE. * P < 0.05 + P < 0.01 as designed for comparisons by analysis of variance using a Bonferroni correction.

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