Wy-14,643 stimulates hepatic protein kinase C activity.

The mechanism by which hypolipidemic drugs and industrial plasticizers cause hepatic tumors in rodents remains unknown. Protein kinase C is elevated during hepatic cell turnover, and sustained cellular replication has been shown to correlate with an increase in hepatic tumors. Therefore, the effect of [4-chloro-6-(2,3-xylidino)-2-pyrimidinylthio]acetic acid (Wy-14,643) on protein kinase C activity was examined. Female Sprague-Dawley rats were given 100 mg/kg Wy-14,643 in olive oil (i.g.), while control rats received equal volumes of oil vehicle. After 24 h, the activity of protein kinase C was estimated in isolated hepatic fractions by measuring the binding of 3H-phorbol-12,13-dibutyrate, a specific ligand for protein kinase C. Administration of Wy-14,643 significantly increased protein kinase C activity nearly 2-fold in microsomal fractions. Thus, it is possible that Wy-14,643 increases cell proliferation and causes tumors by mechanisms involving protein kinase C.     

The effect of nifedipine,nimodipine and nisoldipine on agonistand trauma-stimulated vascular prostacyclin synthesis in vitro

Summary The present study shows that nifedipine, nimodipine and nisoldipine inhibit in vitro agonist-induced prostacyclin (PGI₂) release from rat aortic rings. The agonists used were: U46619 (a thromboxane A₂ analogue); A23187 (a calcium ionophore) and adrenaline.In contrast, these calcium channel blockers did not inhibit in vitro trauma-or arachidonic acid (AA)-induced PGI₂ release. Therefore, in vitro PGI₂ release, models may be calcium channel dependent (adrenaline, U46619, A23187) or independent (trauma, AA), a property which is relevant to calcium channel blocker research. It is suggested that calcium channel dependent PGI₂ release is relevant to modulating the relaxation phase of muscle contraction, while calcium channel independent PGI₂ release is relevant to limiting the extension of thrombi following local vascular trauma.     

Activation of Erk mitogen-activated protein kinase proteins by vascular serotonin receptors

We previously demonstrated that 5-hydroxytryptamine 2A (5-HT 2A ) receptor-mediated rat arterial contraction was dependent on activation of tyrosine kinases, including mitogen-activated protein kinase (MAPK) kinase. In the current study, we examined arterial smooth muscle for the presence of serotonin (5-hydroxytryptamine, 5-HT) 5-HT 1B, 5-HT 1D, 5-HT 1F, 5-HT 2A, 5-HT 2B, and 5-HT 7 receptor mRNA and hypothesized that, if present, activation of these receptors would stimulate the extracellular signal-regulated kinase (Erk) MAPK pathway and an Erk MAPK-dependent contraction. RT-PCR analyses of rat aortic smooth muscle cells, cultured and fresh, indicated the presence of 5-HT 1B, 5-HT 1D, 5-HT 1F, 5-HT 2A, 5-HT 2B, and 5-HT 7 receptor mRNA. The 5-HT 1B agonists RU24969 and CGS12066B, 5-HT 1B/1D/1F receptor agonist sumatriptan, and 5-HT 2B receptor agonist BW723C86 (10(-9) - 10(-4) M ) did not contract the aorta, nor did the 5-HT 7 receptor antagonist LY215840 leftward shift 5-HT-induced contraction. The 5-HT 1E/1F receptor agonist BRL54443 induced contraction, but this was abolished by the 5-HT 2A/2C receptor antagonist ketanserin (10 nM ); contraction was not observed with a different 5-HT 1F receptor agonist, LY344864. 5-HT and alpha-methyl-5-HT produced a concentration-dependent increase in Erk MAPK activity in cultured aortic smooth muscle cells and in aorta contracted with these agonists. All other agonists were inactive; a high concentration of BRL54443 (10 microM ) stimulated Erk MAPK activation (150% basal). Thus, while mRNA and possibly protein for multiple 5-HT receptors are present in aortic smooth muscle, only the 5-HT 2A receptor plays a significant role in directly modulating contractility and activating the Erk MAPK pathway.     

Effect of curcumin on platelet aggregation and vascular prostacyclin synthesis

In vitro and ex vivo effects of 1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione (diferuloylmethane, curcumin) and acetylsalicylic acid (ASA) on the synthesis of prostacyclin (PGI2) and on platelet aggregation has been studied in rat. Both drugs inhibited adenosine diphosphate (ADP)-, epinephrine (adrenaline)- and collagen-induced platelet aggregation in monkey plasma. Pretreatment with ASA (25-100 mg/kg), but not curcumin (100-300 mg/kg), inhibited PGI2 synthesis in rat aorta. In the in vitro system, too, curcumin caused a slight increase in the synthesis of PGI2, while ASA inhibited it. Curcumin may, therefore, be preferable in patients prone to vascular thrombosis and requiring antiarthritic therapy.     

Platelet-activating factor stimulates phosphoinositide turnover in neurohybrid NCB-20 cells: involvement of pertussis toxin-sensitive guanine nucleotide-binding proteins and inhibition by protein kinase C.

Platelet-activating factor (PAF) is an unusually potent phospholipid known to be produced by neuronal cells and to modulate cerebral blood flow and metabolism. In previous studies with NCB-20 cells, we reported that PAF induced a significant mobilization of intracellular free Ca2+ ([Ca2+]i), which was inhibited by PAF antagonists. The increase was the result of release from intracellular stores and influx from extracellular sources. The present study was designed to characterize further PAF receptor-mediated cellular signal-transduction mechanisms in myo-[3H]inositol-labeled cells. PAF induced a concentration-dependent increase in phosphatidylinositol (Pl) metabolism, with EC50 values of 1.96 +/- 0.62 nM and 1.12 +/- 0.50 nM for inositol trisphosphate (IP3) and inositol monophosphate (IP1) formation, respectively (four experiments). The maximal production of IP3 and IP1 induced by 50 nM PAF was 254 +/- 34% and 178 +/- 25% over the basal, respectively (four experiments). PAF-induced Pl metabolism was concentration-dependently inhibited by the PAF antagonist BN50739, with an IC50 value of 6.48 +/- 0.52 nM (four experiments). The protein kinase C (PKC) activator phorbol 12,13-dibutyrate concentration-dependently inhibited PAF-induced Pl metabolism and [Ca2+]i mobilization in NCB-20 cells, of NCB-20 cells with pertussis toxin (PTX) resulted in a concentration-dependent inhibition of PAF-induced IP3 production and intracellular Ca2+ release, with a maximal reduction of 66.9 +/- 3.5% and 63 +/- 6.1%, respectively, at 300 ng/ml PTX. PTX in the presence of [32P]NAD specifically [32P]ADP-ribosylated a 38-kDa protein in membranes prepared from NCB-20 cells. Pretreatment of the cells with PTX resulted in a concentration-dependent inhibition of subsequent 32P-labeling of the toxin substrate in the membranes and correlated with the uncoupling of PAF-induced IP3 formation. PAF (0.01-10 nM) elicited a concentration-related stimulation in guanosine 5'-O-(3-[35S]) triphosphate ([35S]GTP gamma S) binding to G alpha i(1,2) proteins, which was inhibited by the PAF antagonist BN50739. PAF at 10 nM also increased [35S]GTP gamma S binding to G alpha s and G alpha o. PAF-evoked activation of G alpha i(1,2) and G alpha o was reduced by preincubation with PTX. Our results reveal that neuronal cells possess PAF receptors linked through guanine nucleotide-binding proteins to phospholipase C and receptor-operated Ca2+ channels that are regulated by PKC. Both PTX-sensitive and -insensitive guanine nucleotide-binding proteins appear to couple the PAF receptor to activation of phospholipase C and the increase in [Ca2+]i. These results contribute to the further understanding of the mechanisms behind PAF actions on neuronal cells.     

A concise synthesis of a promising protein kinase C inhibitor: D-erythro-sphingosine

In this study, we explored a convenient and concise route for synthesis of D-erythro-sphingosine 1 from commercially available and cheap L-serine. The key steps are simple preparation of amino ketone 5 from Weinreb amide 3 and high diastereoselective reduction of amino ketone 5 to give the natural erythro-(anti-) isomer.     

Muscarinic receptor-mediated prostacyclin and cGMP synthesis in cultured vascular cells.

The purpose of the present study was to determine the subtype of muscarinic receptor involved in the action of cholinergic stimuli on synthesis of prostacyclin, measured as immunoreactive 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), and cGMP in bovine aortic endothelial and rabbit vascular smooth muscle cells. Acetylcholine and arecaidine propargyl ester, a selective M2 agonist, produced a dose-dependent increase in 6-keto-PGF1 alpha output and cGMP formation in confluent endothelial cells but not in confluent vascular smooth muscle cells. McN-A-343, a selective M1 agonist, failed to alter basal 6-keto-PGF1 alpha or cGMP synthesis. Acetylcholine- and arecaidine propargyl ester-induced 6-keto-PGF1 alpha synthesis and cGMP formation in endothelial cells were attenuated by atropine, AF-DX 116 (M2 antagonist), and hexahydrosiladifenidol (M3 antagonist) but not by pirenzepine (M1 antagonist). The cyclooxygenase inhibitor indomethacin abolished 6-keto-PGF1 alpha synthesis but not the increase in cGMP formation elicited by the cholinergic stimuli. Our data suggest that the effect of cholinergic stimuli to enhance prostacyclin and cGMP synthesis is mediated via activation of M2 and M3 receptors located on endothelial cells and that the increase in cGMP production is independent of prostaglandins.     

MDL 27,032 relaxes vascular smooth muscle and inhibits protein kinase C.

The smooth muscle relaxant effect of MDL 27,032, 4-propyl-5-(4-pyridinyl)-2(3H)-oxazolone, was studied in vitro using strips of femoral arteries and saphenous veins from dogs and trachea from guinea pigs. MDL 27,032 (10(-6)-10(-3) M) produced a concentration-dependent relaxation of arterial and venous strips contracted by carbachol. MDL 27,032 also antagonized contractions of arterial and venous strips produced by phorbol 12,13-dibutyrate (PDB), a protein kinase C activator, both in normal-Ca2+ and zero-Ca2+ medium. The compound inhibited protein kinase C in soluble extracts prepared from saphenous veins of dogs, with an IC50 value of 36.6 microM. MDL 27,032 was more effective against the contractions produced by phenylephrine and serotonin than by KCl in arteries, but no such selectivity was noted in veins. MDL 27,032 (10(-3) M) also inhibited accumulation of inositol phosphates in femoral artery but not in saphenous vein, and this effect may have contributed to the arterial-relaxant effect. Because the vascular smooth muscle relaxant effect of MDL 27,032 was endothelium independent, did not involve blockade of alpha-adrenoceptors or inhibition of cyclic nucleotide phosphodiesterases, stimulation of beta-adrenergic receptors, stimulation of adenosine A2-receptors, or activation of K+ channels, these data suggest that the relaxant effects of MDL 27,032 primarily involve inhibition of protein kinase C.     

The role of protein kinase C activation and the vascular complications of diabetes

Diabetes mellitus is a chronic disease caused by inherited and/or acquired deficiency in production of insulin by the pancreas, and by resistance to insulin's effects. Such a deficiency results in increased concentrations of glucose and other metabolites in the blood, which in turn damages many of the body's systems, in particular the eyes, kidneys, nerves, heart and blood vessels. There are two major types of diabetes mellitus: Type 1 diabetes (insulin-dependent diabetes, IDDM or juvenile onset diabetes) and Type 2 diabetes (non-insulin-dependent diabetes, NIDDM or adult-onset). Chronic hyperglycemia is a major initiator of diabetic micro- and cardiovascular complications, such as retinopathy, neuropathy and nephropathy. Several hyperglycemia-induced mechanisms may induce vascular dysfunctions, which include increased polyol pathway flux, altered cellular redox state, increased formation of diacylglycerol (DAG) and the subsequent activation of protein kinase C (PKC) isoforms and accelerated non-enzymatic formation of advanced glycated end products. It is likely that each of these mechanisms may contribute to the known pathophysiologic features of diabetic complications. Others and we have shown that activation of the DAG-PKC pathway is associated with many vascular abnormalities in the retinal, renal, neural and cardiovascular tissues in diabetes mellitus. DAG-PKC pathway affects cardiovascular function in many ways, such as the regulation of endothelial permeability, vasoconstriction, extracellular matrix (ECM) synthesis/turnover, cell growth, angiogenesis, cytokine activation and leucocyte adhesion, to name a few. Increased DAG levels and PKC activity, especially alpha, beta1/2 and delta isoforms in retina, aorta, heart, renal glomeruli and circulating macrophages have been reported in diabetes. Increased PKC activation have been associated with changes in blood flow, basement membrane thickening, extracellular matrix expansion, increases in vascular permeability, abnormal angiogenesis, excessive apoptosis and changes in enzymatic activity alterations such as Na(+)-K(+)-ATPase, cPLA(2), PI3Kinase and MAP kinase. Inhibition of PKC, especially the beta1/2 isoform has been reported to prevent or normalize many vascular abnormalities in the tissues described above. Clinical studies have shown that ruboxistaurin, a PKCbeta isoform selective inhibitor, normalize endothelial dysfunction, renal glomerular filtration rate and prevented loss of visual acuity in diabetic patients. Thus, PKC activation involving several isoforms is likely to be responsible for some of the pathologies in diabetic retinopathy, nephropathy and cardiovascular disease. PKC isoform selective inhibitors are likely new therapeutics, which can delay the onset or stop the progression of diabetic vascular disease with very little side effects.     

Prevention of vascular nitroglycerin tolerance by inhibition of protein kinase C.

We investigated whether in vivo inhibition of protein kinase C (PKC) can prevent the development of vascular tolerance and restore the sensitivity of isolated vessels to nitroglycerin (NTG). Tolerance was induced in male Wistar rats by a constant i.v. infusion of NTG 1 mg kg-1 h-1, a dose which did not alter blood pressure. After 72 h, the aorta was removed and the sensitivity of aortic rings to NTG tested. Chronic NTG infusion resulted in a 5.5 fold decrease in NTG-sensitivity as compared with controls (vehicle), indicating the development of vascular tolerance. The simultaneous in vivo administration of the specific PKC inhibitor N-benzoyl-staurosporine (30 mg kg-1 day-1) prevented this decrease in NTG sensitivity. These results suggest a role for PKC activation in the development of vascular NTG tolerance.     

Rho kinase and protein kinase C involvement in vascular smooth muscle myofilament calcium sensitization in arteries from diabetic rats

Background and purpose:

Diabetes mellitus (DM) causes multiple dysfunctions including circulatory disorders such as cardiomyopathy, angiopathy, atherosclerosis and arterial hypertension. Rho kinase (ROCK) and protein kinase C (PKC) regulate vascular smooth muscle (VSM) Ca²⁺ sensitivity, thus enhancing VSM contraction, and up-regulation of both enzymes in DM is well known. We postulated that in DM, Ca²⁺ sensitization occurs in diabetic arteries due to increased ROCK and/or PKC activity.

Experimental approach:

Rats were rendered hyperglycaemic by i.p. injection of streptozotocin. Age-matched control tissues were used for comparison. Contractile responses to phenylephrine (Phe) and different Ca²⁺ concentrations were recorded, respectively, from intact and chemically permeabilized vascular rings from aorta, tail and mesenteric arteries.

Key results:

Diabetic tail and mesenteric arteries demonstrated markedly enhanced sensitivity to Phe while these changes were not observed in aorta. The ROCK inhibitor HA1077, but not the PKC inhibitor chelerythrine, caused significant reduction in sensitivity to agonist in diabetic vessels. Similar changes were observed for myofilament Ca²⁺ sensitivity, which was again enhanced in DM in tail and mesenteric arteries, but not in aorta, and could be reduced by both the ROCK and PKC blockers.

Conclusions and implications:

We conclude that in DM enhanced myofilament Ca²⁺ sensitivity is mainly manifested in muscular-type blood vessels and thus likely to contribute to the development of hypertension. Both PKC and, in particular, ROCK are involved in this phenomenon. This highlights their potential usefulness as drug targets in the pharmacological management of DM-associated vascular dysfunction.     

Comparison of the effects of auranofin, heavy metals and retinoids on protein kinase C in vitro and on a protein kinase C mediated response in macrophages

The effect of auranofin (AF), retinoic acid (RA), and three heavy metals reacting with thiol groups (Hg, Cd, Pb) has been compared on a PKC mediated response of intact macrophages (i.e. plasminogen activator (PA) induction) and on purified PKC activity. AF, cadmium chloride, and lead nitrate directly inhibit PKC and hence prevent the induction of PA activity in macrophages stimulated with PMA. In vitro, and in absence of chelators, mercuric chloride is also a potent inhibitor of PKC. However, at the cellular level, the PKC mediated response (PA induction) was not inhibited by non-cytotoxic concentrations of mercury possibly due to interference of the metal with additional cellular mechanisms such as calcium mobilisation. Direct inhibition of PKC is probably not the mechanism by which retinoids block the activation of macrophages.     

Estrogen stimulates release of secreted amyloid precursor protein from primary rat cortical neurons via protein kinase C pathway

Aim: To investigate the mechanism of the action of estrogen, which stimulates the release of secreted amyloid precursor protein α (sAPPα) and decreases the generation of amyloid-β protein (Aβ), a dominant component in senile plaques in the brains of Alzheimer‘s disease patients. Methods: Experiments were carried out in primary rat cortical neurons, and Western blot was used to detect sAPPα in aculture medium and the total amount of cellular amyloid precursor protein (APP) in neurons. Results: 17β-Estradiol (but not 17α-estradiol) and β-estradiol 6-(0-carboxymethyl) oxime: BSA increased the secretion of sAPPα and this effect was blocked by protein kinase C (PKC) inhibitor caiphostin C, but not by the classical estrogen receptor antagonist ICI 182,780. Meanwhile, 17β-estradiol did not alter the synthesis of cellular APP. Conclusion: The effect of 17β-estradiol on sAPPα secretion is likely mediated through the membrane binding sites, and needs molecular configuration specificity of the ligand. Furthermore, the action of the PKCdependent pathway might be involved in estrogen-induced sAPPα secretion.     

Nitric oxide stimulates a large-conductance Ca-activated K+ channel in human skin fibroblasts through protein kinase G pathway

In order to investigate the large-conductance Ca(2+)-activated K(+) (BK(Ca)) channel and determine the effects of nitric oxide (NO) on the channel in human skin fibroblasts, we performed electrophysiological patch clamp recordings on 5th-passage cells of human genital skin cultures. The whole-cell outward K(+) current was increased with depolarization, and proved to be sensitive to NS1619 (a selective BK(Ca) channel activator) and iberiotoxin (a specific BK(Ca )channel inhibitor). The single-channel currents showed 226 pS of mean conductance in symmetrical K(+). Sodium nitroprusside (SNP; an NO donor) significantly increased the K(+) current amplitude in the whole-cell mode, and open probability of the channel (NPo) in the cell-attached mode, but not in the inside-out mode. S-nitroso-N-acetylpenicillamine (an NO donor) and 8-Br-cGMP (a membrane-permeant cGMP analogue) also increased the BK(Ca )channel activity. The stimulatory effect of SNP on BK(Ca) channels was inhibited by pretreatment with 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (a soluble guanylyl cyclase inhibitor), or KT5823 [a specific protein kinase G (PKG) inhibitor]. Cytoplasmic PKG also increased the channel activity in inside-out patches. In conclusion, the present data indicate that BK(Ca) channels constitute a significant fraction of K(+) current in human skin fibroblasts, and that NO increases NPo of BK(Ca) channels, which are mediated via the cGMP/PKG pathway, without direct effects on the channel.     

蛋白激酶C在肿瘤中的作用及其抑制剂研究进展

蛋白激酶C(protein kinase C,PKC)在肿瘤的发生、发展和转移中均发挥着重要作用,因此,PKC是开发治疗肿瘤疾病药物的潜在分子靶点。目前,几种PKC抑制剂已经进入临床研究阶段,但是同时也为靶向PKC抗肿瘤药物的研究开发带来了新的挑战。该文拟对近年来蛋白激酶C在肿瘤中的作用及其抑制剂研究进展作一综述。     

New dimensions in G protein signalling: G beta 5 and the RGS proteins

The beta gamma complex of G-proteins regulates effectors independently of the G alpha subunits, such that upon activation G proteins give may signal downstream along one or both pathways. The G beta 5 isoform exhibits much less homology with other G beta isoforms (approximately 50%) and is preferentially expressed in brain. The G beta 5 isoform exhibits novel properties in its activation of effector pathways such as MAPK, phospholipase C-beta, and adenylyl cyclase type II when compared to G beta 1. Recently specific native complexes between G beta 5 and the regulator of G protein signaling (RGS) protein-7 (RGS7) and between G beta 5L (a splice variant with a 42 amino acid N-terminal extension) and RGS9 have been isolated from different retinal fractions. Such findings are not accounted for by current models as only the G alpha subunits and not G beta had been previously implicated in RGS protein function. These recent novel observations further reinforce the view of G beta 5 as a unique and highly specialized G protein subunit.     

Angiotensin II stimulates phosphorylation of 4E-binding protein 1 and p70 S6 kinase in cultured vascular smooth muscle cells

INTRODUCTIONAngiotensin II (Ang II) potently stimulates pro-tein synthesis, the major hallmark of hypertrophy, invascular smooth muscle cells (VSMC) by acting throughthe G protein-coupled Ang II type 1 receptor (AT1receptor)[1]. However, the molecular basis for this hy-pertrophic effect remains largely unknown.Recent studies have established the initiation phaseof mRNA translation as a pivotal site of regulation forglobal rates of protein synthesis[2]. At the heart of thetranslat…     

Activation of mGluRII induces LTD via activation of protein kinase A and protein kinase C in the dentate gyrus of the hippocampus in vitro

The involvement of metabotropic glutamate receptor group II (mGluRII) in the induction of long-term depression (LTD) was investigated in the medial perforant path of the rat dentate gyrus, a region with a very high density of mGluRII. Perfusion of either of two potent mGluRII agonists, (2S,1R,2R,3R)-2-(2S, 1'R, 2'R, 3'R)-2 (2' 3'-dicarboxycyclopropyl)glycine (DCG-IV) or (+)-2- aminobicyclo[3.1.0]hexane-2-6-dicarboxylic acid (LY354740) induced a reversible inhibition of the field EPSP followed, upon washout of the agonist, by LTD. The reversible inhibition was associated with a change in paired pulse depression, indicating an underlying presynaptic reduction in the probability of transmitter release, whereas the LTD was not associated with a change in paired pulse depression, indicating either a presynaptic reduction in the number of active release sites, or a postsynaptic change. Further evidence that the DCG-IV-induced LTD was generated by activation of mGluRII was the finding that the mGluRII antagonist (RS)-alpha-methylserine-O-phosphate monophenylphosphoryl ester (MSOPPE) prevented the induction of the LTD induced by DCG-IV. The DCG-IV-induced LTD showed mutual occlusion with LFS-induced LTD. The generation of the agonist-induced LTD required, in part, activation of N-methyl-D-aspartate receptors (NMDAR), as LTD induction was partially blocked in the presence of the NMDAR antagonist D-2-amino-5-phosphonopentanoate (AP5). Evidence for involvement of protein kinase C (PKC) and protein kinase (PKA) in the induction of LTD by activation of mGluRII was obtained by showing an inhibition of the DCG-IV-induced LTD by the PKC inhibitors Ro-31-8220 and bisindolylmaleimide I, and also by the PKA inhibitor H-89. The study demonstrates that activation of mGluRII induces LTD via activation the PKA and PKC pathways in the medial perforant path of the dentate gyrus.     

Role of sulfhydryl groups in the stimulatory effect of captopril on vascular prostacyclin synthesis

The effect of captopril on vascular prostacyclin production was studied, evaluating which of its components--sulfhydryl (SH) group or proline--is responsible for this effect. Rat aortas were incubated with captopril (10-100 microM), 2-mercaptoethanol or proline (10 microM), and captopril plus the SH-binding reagents N-ethylmaleimide or ethacrynic acid (50 microM). Prostacyclin was measured by radioimmunoassay of 6-keto-prostaglandin F1 alpha. Captopril stimulated prostacyclin production. This effect was associated with an enhanced conversion of arachidonate to prostacyclin and was not related to bradykinin. Since 2-mercaptoethanol increased vascular prostacyclin per se and proline did not, the stimulatory effect of captopril appears to be dependent upon the SH group; in addition, both SH blockers, N-ethylmaleimide and ethacrynic acid, antagonized this effect. This study shows that captopril stimulates vascular prostacyclin synthesis directly and that the SH group plays a key role in this action. This stimulation of prostacyclin synthesis may contribute to the antihypertensive action of captopril.