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Keywords:

  • choline;
  • cisplatin-induced gastric abnormalities;
  • functional dyspepsia;
  • herbal medicine;
  • muscarinic receptors;
  • prokinetic drugs;
  • stomach;
  • yarrow

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References

Background  Functional dyspepsia (FD) is a highly prevalent gastrointestinal disorder characterized by alterations in gastric motility. Yarrow (Achillea millefolium L., Fam Asteraceae) preparations are traditional remedies used to treat dyspeptic complaints. Herein, we investigated the effect of a standardized dry water extract obtained from A. millefolium flowering tops (AME) on gastric motility.

Methods  The effect of AME on motility was evaluated on the resting tone of the isolated gastric antrum and on gastric emptying in vivo (phenol red meal method) both in control mice and in the model of cancer chemotherapy (cisplatin)-induced gastric abnormalities.

Key Results  The AME contracted mouse and human gastric strips and this action was unaffected by hexamethonium and tetrodotoxin, but strongly reduced by atropine. Among various chemical ingredients in yarrow, choline, but not the flavonoids rutin and apigenin, mimicked the action of AME. Furthermore, AME deprived of choline did not exert a contractile effect. In vivo, AME stimulated gastric emptying both in control and in cisplatin-treated mice, being more active in pathological states.

Conclusions & Inferences  It is concluded that (i) AME exerts a direct spasmogenic effect on gastric antrum; (ii) choline is the chemical ingredient responsible of such effect; (iii) the prokinetic effect of AME observed in vivo could provide the pharmacological basis underlying its traditional use in the treatment of dyspepsia.


Abbreviations:
AME

water extract of Achillea millefolium L. flowering tops

AME-wpc

water extract of Achillea millefolium L. flowering tops deprived of choline and related polar compounds

DMSO

dimethyl sulphoxide

FD

functional dyspepsia

TTX

tetrodotoxin

Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References

Dyspeptic complaints are highly prevalent. Indeed, epidemiological studies suggest that approximately 15% of the general population in Western countries suffers from functional dyspepsia (FD)1 (also labeled non-ulcer or idiopathic dyspepsia), which is also the most common cause of dyspepsia. Several symptom-based criteria for FD [i.e. Rome (I, II, and III) criteria] have recently been developed to facilitate and standardize the diagnosis of FD. The etiology and pathophysiology of FD remains unclear; however, delayed gastric emptying has been proposed to be important in many clinical situations.2–5 Although the clinical course of FD is benign without disease-associated mortality, the impact can be substantial for the affected patients with regard to the decrease of quality of life, and for society with regard to the economical implications.6 Current available agents for the treatment of dyspeptic disturbances include the use of prokinetic drugs, which are a heterogenous class of drugs that act by stimulating smooth muscle contractions and enhancing gastric emptying.7,8 As the available pharmacological therapy for patients with FD is overall largely unsatisfactory,9,10 alternative remedies for symptomatic relief are widely used by dyspeptic patients.11

Herbal products may be an attractive alternative based on the perception of its ‘natural’ approach and low risk of side effects and they are widely used by FD patients.12,13 One of such herbs is yarrow (Achillea millefolium L.), a widespread plant from the Asteraceae family confined to the Northern Hemisphere. In addition to its use in the production of liquors, yarrow is traditionally used for the treatment of inflammatory and gastrointestinal disorders and hepato-biliary complaints.14,15 To rationalize the folklore use of yarrow in the treatment of these ailments, preclinical studies have demonstrated an hepatoprotective, antispasmodic, and gastroprotective effect of this herb.16–19 The British Herbal Pharmacopoeia approved the internal use of yarrow for feverish conditions, common cold, and digestive complaints.20 Similarly, the German commission E – a special committee of the Federal Department of Health, which is a consulting body appointed by the German equivalent of the USA Food and Drug Administration – supports the use of yarrow for a number of diseases, including the treatment of dyspeptic ailments.18 This medicinal use has been attributed to presence in the plant of compounds which could reflexly, via increasing the tone of the vagal system, stimulate the digestive fluids in stomach, pancreas, and liver and thereby alleviate dyspeptic complaints.21

As yarrow preparations are traditional remedies used in the treatment of dyspeptic complaints, herein, we evaluated the action of a standardized dry water extract obtained from the flowering tops of A. millefolium L. (AME) on gastric antrum in vitro (both in mice and humans) and on gastric emptying in vivo [both in control mice or in the model of cancer chemotherapy (cisplatin)-induced gastric abnormalities].

Materials and Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References

Animals

Experiments were carried out on male Swiss mice (20–25 g), purchased from Harlan Italy (San Pietro al Natisone, Italy). The mice were maintained under controlled conditions of temperature (24 ± 2 °C) and humidity (60%), and had free access to water and food until used. All experiments complied with the Italian D.L. no. 116 of 27 January 1992 and associated guidelines in the European Communities Council Directive of 24 November 1986 (86/609/ECC).

Isolated mouse stomach

Isolated stomach preparations were obtained as previously described.22,23 Briefly, mice were killed by asphyxiation with CO2 and the stomach was rapidly dissected from the abdomen and placed in Krebs solution (mmol L−1: NaCl 119, KCl 4.75, KH2PO4 1.2, NaHCO3 25, CaCl2 2.5, MgSO4 1.5, and glucose 11). Two full-thickness strips (about 1 cm) were cut in the direction of the longitudinal muscle layer from each gastric antrum. The strips were suspended in an organ bath containing 20 mL Krebs equilibrated with 95% O2, 5% CO2 at 37 °C, and connected to an isometric transducer (load 0.5 g). The transducer was coupled to a PowerLab system (Ugo Basile, Comerio, Italy). After a minimum 60-min equilibration period, the tissues were subjected to a maximum contraction with acetylcholine 10−3 mol L−1. Stomachs not showing contractions were rejected (rejection percentage: <5%). After removal of acetylcholine through wash out, the stomach responses were observed in the presence of increasing cumulative concentrations of AME (1–30 000 μg mL−1). In some experiments, after at least three stable concentration-effect control contractions, the effect of AME was evaluated in the presence of tetrodotoxin (TTX, 3 × 10−7 mol L−1), hexamethonium (10−4 mol L−1) or atropine (10−6 mol L−1) (contact time for each antagonist: 30 min). The concentrations of TTX, hexamethonium, and atropine were selected on the basis of previous works.24–26 In some experiments, the effect of AME deprived of choline and related polar compounds (AME-wpc) was also evaluated.

Finally, we evaluated the effect of a number of yarrow ingredients, namely choline (3 × 10−4 to 10−2 mol L−1, corresponding to 117.3–39100 μg mL−1), rutin (10−7 to 10−2 mol L−1), and apigenin (10−7 to 3 × 10−4 mol L−1), on the stomach preparation (contact time: 10 min for each concentration). In the set of experiments in which a relaxing effect was observed, isoproterenol (10−5 mol L−1) was added at the end of each experiment. The 10−5 mol L−1 isoproterenol concentration produced maximal relaxation of the stomach tissue. In some experiments, the effect of choline was evaluated in the presence of TTX (3 × 10−7 mol L−1), hexamethonium (10−4 mol L−1) or atropine (10−6 mol L−1).

Isolated human stomach

Strips of stomach antrum (that include the entire muscularis propria) were taken from six patients who underwent total or subtotal gastrectomy for gastric carcinoma.

Segments of 1–2 cm of the stomach antrum smooth muscle free of disease were set up in the bath under a resting tension of 1 g. The experimental protocol, including drug administration and contact time, was identical to that described for mouse stomach. The research followed guidelines of the Declaration of Helsinki and Tokyo for humans; informed consent was received from all patients, as well as the approval of the ethics committee at our institution.

Gastric emptying

Gastric emptying was evaluated as previously described.27 Briefly, after an overnight fast, the animals received by gavage 1 mL 100 g−1 of a solution of 50 mg phenol red in 100 mL 1.5% carboxymethylcellulose, which was constantly stirred and held at 37 °C. After 20 min, mice were euthanized and the stomach was quickly ligated at the lower esophageal sphincter and pyloric region and removed. The stomach was opened, and its contents were poured in a test tube and washed with 4 mL distilled water. At the end of the experiment, 2 mL NaOH (1 mol L−1) was added to each tube to develop the maximum intensity of color. The solutions were assayed using a spectrometer at 560 nm. Percent gastric emptying was calculated according to the following formula: 100 × (1 − amount of phenol red recovered after 20 min per amount of phenol red recovered after 0 min). AME (1–30 mg kg−1) was given intraperitoneally 30 min before the administration of the marker (phenol red).

In another set of experiments, gastric emptying was abnormally delayed by administration of cisplatin, a model of cancer chemotherapy-induced gastric abnormalities.28 We used a 10 mg kg−1 i.p. cisplatin dose on the basis of our preliminary experiments in which we observed a submaximal inhibition of gastric emptying. Mice were injected with cisplatin and gastric emptying was measured after 48 h.28 AME (1–30 mg kg−1) was given intraperitoneally 30 min before the administration of the marker (phenol red).

Achillea millefolium water extract preparation

Dried flowering tops of A. millefolium (165.2 g) were repeatedly extracted at room temperature with H2O/MeOH 1 : 1 (4 × 1 L) to give a hydromethanolic extract which, evaporated to dryness, provided 12.9 g of a brownish solid. This extract was partitioned between H2O and ethyl acetate (EtOAc) (3 × 500 mL) to give an EtOAc extract (4.1 g of a brown-colored viscous oil) and a water extract (named AME = 8.8 g). Then, the AME (H2O) extract was partitioned against n-BuOH (3 × 500 mL) to give a BuOH extract (5.2 g) and a water extract (W2 = 3.6 g). The latter partitioning resulted in the concentration in the water phase W2 of inorganic salts, choline, and other small polar metabolites (e.g., amino acids). In particular, a comparison (thin layer chromatography, nuclear magnetic resonance) with an authentic sample of choline chloride confirmed the presence of choline in the water phase, whereas it was practically undetectable in the butanol phase (therefore called A. millefolium extract without choline and polar compounds = AME-wpc). Chromatographic purification of choline from the water W2 phase was partially accomplished through preparative thin layer chromatography (eluent BuOH/H2O/AcOH 50 : 40 : 10) and Amberlite IRC50 column. As a result of these purifications, 315.0 mg of choline (3.58% of AME extract; 0.19% of the dried flowering tops) could be obtained.

Data evaluation and statistical analysis

Data are expressed as the mean ± SEM. Contractile responses were expressed as % of the maximal contractile response produced by 10−3 mol L−1 acetylcholine; relaxant responses were expressed as % of the maximal relaxant response produced by 10−5 mol L−1 isoproterenol. To determine statistical significance, Student’s t-test was used for comparing a single treatment mean with a control mean, and a one-way analysis of variance (ANOVA) followed by a Tukey–Kramer multiple comparisons test was used for analysis of multiple treatment means. Two-way ANOVA was used to compare different cumulative concentration effect curves. A P-value less than 0.05 was considered significant. The concentration of AME that produced 50% of contraction (EC50), the dose of AME that increased gastric emptying of 50% (ED50) and maximal contractile/gastric emptying effect (Emax) were used to characterize its potency and efficacy, respectively. EC50, ED50, and Emax values were calculated using the Graph Pad Instat program version 4.01 (GraphPad Software Inc., La Jolla, CA, USA).

Drugs

Flowering tops of A. millefolium were obtained from D. Ulrich Spa, Via Pisacane, 910042 Nichelino, Torino, Italy. Voucher specimen (50 g of flowering tops; reg. number: 09–03) is deposited in the Department of Experimental Pharmacology, Via D. Montesano 49, 80131 Naples, Italy. Achillea millefolium extract (AME, standardized dry water extract prepared from the flowering tops of A. millefolium) and AME without polar compounds including choline (AME-wpc) were obtained as described above. Acetylcholine hydrochloride, tetrodotoxin, atropine sulfate, hexamethonium bromide, rutin, apigenin, isoproterenol hydrochloride, cisplatin, and phenol red were purchased from Sigma (Milan, Italy). AME, AME-wpc, choline, acetylcholine, tetrodotoxin, atropine, hexamethonium, and isoproterenol were solubilized in distilled water (for in vivo experiments, AME was dissolved in water). Rutin and apigenin were dissolved in dimethyl sulphoxide (DMSO) to give a 1 mol L−1 stock solution; subsequent dilutions were made in distilled water. Cisplatin was dissolved in DMSO. Drugs were added in volumes less than 0.01%in vitro and given in the amount of 0.1 mL mouse−1 (water) or 10 μL mouse−1 (DMSO) in vivo. The drug vehicles had no effect on the responses under study, both in vitro and in vivo.

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References

Isolated mouse stomach

The AME (1–30000 μg mL−1) contracted the isolated mouse stomach antrum in a concentration-dependent manner, with an effective threshold concentration of 100 μg mL−1 (Fig. 1). The contractions induced by AME had an average latency of 30–60 s. The time of contact of AME to reach a maximum contractile effect depended from the concentration of AME used and varied from 1–3 min for the higher concentrations tested (i.e., starting from 1000 μg mL−1) and 8–10 min for the lower concentration tested (1–300 μg mL−1). When the tissue bath was drained and Krebs was added, there was rapid relaxation of the muscle to baseline resting tension (Fig. 1 insert). A typical trace showing the effect of AME (1000 μg mL−1) is reported in the insert of Fig. 1. The EC50 and Emax values were 617.5 ± 12.1 μg mL−1 and 89.2 ± 4.5%, respectively.

image

Figure 1.  Effect of a standardized dry water extract prepared from the flowering tops of Achillea millefolium (AME, 1–30 000 μg mL−1) and AME deprived of choline and related polar compounds (AME-wpc) on the resting tone of the isolated mouse stomach antrum. The responses are expressed as % of the maximal contraction induced by acetylcholine 10−3 mol L−1. Each point represents the mean of 7–8 experiments; vertical lines show SEM. ***P < 0.001 vs AME (significance between the curves). Insert: Representative experimental trace showing the contractile response to 1000 μg mL−1 AME.

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The contractile effect of AME was unaffected by tetrodotoxin (3 × 10−7 mol L−1) and hexamethonium (10−4 mol L−1) (Fig. 2). However, treatment of the tissue with the muscarinic receptor antagonist atropine (10−6 mol L−1) abolished the contractile response of AME on mouse stomach antrum (Fig. 2).

image

Figure 2.  Contractile effect of a standardized dry water extract prepared from the flowering tops of Achillea millefolium (AME, 1–30 000 μg mL−1) alone (Vehicle) or in the presence of tetrodotoxin (TTX, 3 × 10−7 mol L−1), hexamethonium (10−4 mol L−1), or atropine (10−6 mol L−1) in the isolated mouse stomach antrum. The ordinate shows the % of the maximal contractile response induced by acetylcholine 10−3 mol L−1. Each point represents the mean of 8–9 experiments; vertical lines show SEM ***P < 0.001 vs AME alone (significance between the curves).

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Among the chemical compounds tested, choline (3 × 10−4 to 10−2 mol L−1) contracted, in a concentration-dependent manner, the isolated tissue (threshold concentration: 5.62 × 10−4 mol L−1, latency 30–60 s, maximum contractile effect within 1–3 min) (Fig. 3). The EC50 and Emax values were 1.84 × 10−3 ± 1.12 mol L−1 and 98.4 ± 5.7%, respectively. The contractile effect of choline on the stomach antrum was significantly reduced by atropine (10−6 mol L−1), but not by tetrodotoxin (3 × 10−7 mol L−1) or hexamethonium (10−4 mol L−1) (Fig. 3). Fig. 4A shows a comparison between the contractile effect of pure choline and the choline contained in AME, in which it appears that the choline contained in AME was more active than the pure choline. In addition, AME-wpc (1–30 000 μg mL−1) did not cause contractions of the stomach antrum (Fig. 1). Finally, rutin (10−7 to 10−2 mol L−1) and apigenin (10−7 to 3 × 10−4 mol L−1), two yarrow ingredients, relaxed the isolated stomach strip in a concentration-dependent manner (Fig. 5A).

image

Figure 3.  Effect of choline (3 × 10−4 to 0−2 mol L−1) alone (Vehicle) or in the presence of tetrodotoxin (TTX, 3 × 10−7 mol L−1), hexamethonium (10−4 mol L−1), or atropine (10−6 mol L−1) in the isolated mouse stomach antrum. The ordinate shows the % of the maximal contractile response induced by acetylcholine 10−3 mol L−1. Each point represents the mean of 8–9 experiments; vertical lines show SEM. ***P < 0.001 vs choline alone (significance between the curves).

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image

Figure 4.  Comparative contractile effect of choline contained in Achillea millefolium extract (choline contained in AME) and pure choline on the isolated mouse (A) and human (B) gastric antrum. The ordinate shows the % of the maximal contractile response induced by acetylcholine 10−3 mol L−1. Each point represents the mean of 7–8 experiments; vertical lines show SEM. **P < 0.01 and ***P < 0.001 vs choline contained in AME (significance between the curves).

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image

Figure 5.  Relaxant effect of rutin (10−7 to 10−2 mol L−1) and apigenin (10−7 to 3 × 10−4 mol L−1) on the isolated mouse (A) and human (B) antrum. The ordinate shows the percentage of relaxation induced by isoproterenol 10−5 mol L−1. Each point represents the mean of 7–8 experiments; vertical lines show SEM.

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Isolated human stomach

The AME (1–30 000 μg mL−1) contracted the isolated human stomach antrum in a concentration-dependent manner (Fig. 6A). The EC50 and Emax values were 581.4 ± 12.4 μg mL−1 and 97.7 ± 5.3%, respectively. Atropine (10−6 mol L−1), but not tetrodotoxin (3 × 10−7 mol L−1) or hexamethonium (10−4 mol L−1), significantly reduced the contractile response of AME on human stomach antrum (Fig. 6A).

image

Figure 6.  Effect of a standardized dry water extract prepared from the flowering tops of Achillea millefolium (AME, 1–30 000 μg mL−1) (A) or choline (10−5 to 10−2 mol L−1) (B) alone or in the presence of tetrodotoxin (TTX, 3 × 10−7 mol L−1), hexamethonium (10−4 mol L−1) or atropine (10−6 mol L−1) in the isolated human antrum. The ordinate shows the percentage of the maximal contractile response induced by acetylcholine 10−3 mol L−1. Each point represents the mean of 8–9 experiments; vertical lines show SEM. ***P < 0.001 vs AME or choline alone (significance between the curves).

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Choline (10−5 to 10−2 mol L−1), in a concentration-dependent manner, contracted the human tissues antrum (Fig. 6B). The EC50 and Emax values were 1.43 × 10−3 ± 1.42 mol L−1 and 99.3 ± 15.2%, respectively. The effect of choline was abolished by atropine (10−6 mol L−1), but not by tetrodotoxin (3 × 10−7 mol L−1) or hexamethonium (10−4 mol L−1) (Fig. 6B). Similar to the mouse experiments, choline contained in AME was more active than the pure choline (Fig. 4B). Rutin (10−7 to 10−2 mol L−1) and apigenin (10−7 to 3 × 10−4 mol L−1) did not display any significant contractile effect on human tissues, but rather relaxed the tissue (Fig. 5B).

Gastric emptying

Twenty minutes after the administration of phenol red, the percentage of gastric emptying was about 50% (Fig. 7A). AME (1–30 mg kg−1, per os) produced a dose-dependent increase in gastric emptying (Fig. 7A). A significant prokinetic effect was achieved starting from the 10 mg kg−1 dose. By contrast, AME-wpc did not induce any significant effect on gastric emptying (% of gastric emptying: control 47.6 ± 4.2, AME-wpc 10 mg kg−1 46.3 ± 3.9, AME-wpc 30 mg kg−1 46.3 ± 3.8, n = 10–12 animals).

image

Figure 7.  Effect of a standardized dry water extract prepared from the flowering tops of Achillea millefolium (AME, 1–30 mg kg−1, per os) on gastric emptying in control mice (A) and in mice with delayed gastric motility induced by the anticancer drug cisplatin (10 mg kg−1 i.p.) (B). Results (mean ± SEM of 10–12 animals for each experimental group) are expressed as a percentage of gastric emptying. The insert shows the comparative effect of AME (1–30 mg kg−1) on gastric emptying – expressed as % of increase of corresponding control values – both in control and in cisplatin-treated mice. **P < 0.01, ***P < 0.001, and #P < 0.001 vs control; °°P < 0.01 and °°°P < 0.001 vs cisplatin alone.

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Cisplatin (10 mg kg−1) caused a significant reduction in gastric emptying 48 h after its i.p. administration (Fig. 7B). AME (1–30 mg kg−1, per os) produced an increase in gastric emptying in cisplatin-treated animals. The effect was statistically significant starting from the 1 mg kg−1 dose (Fig. 7B). AME was significantly more potent (ED50 values: healthy mice 6.2 ± 0.5 mg kg−1, dyspeptic mice 1.6 ± 0.3 mg kg−1, P < 0.001) and effective (Emax values: healthy mice 45.1 ± 0.8%, dyspeptic mice 56.6 ± 1.1%P < 0.001) in animals treated with the anticancer drug (see also Fig. 7, insert). At the doses used, AME and AME-wpc did not cause evident side effects in mice.

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References

Preparations from A. millefolium are traditionally used for the treatment of dyspeptic complaints.29 Disturbances of gastric motility are often observed in dyspeptic patients and drugs which enhance gastric emptying may be an effective treatment.7,8 Here, we report that a standardized dry water extract obtained from the flowering tops of A. millefolium contracts mouse and human stomach strips as well as stimulate gastric emptying in vivo, both in control and in cisplatin-treated mice.

We have shown that AME evoked contractions of strips isolated from mouse stomach antrum. The contractile response was not affected by tetrodotoxin, indicating that the herbal extract exerts a direct effect on intestinal smooth muscle and that activation of sodium channels is not essential for AME to induce contraction. However, the effect of AME was completely abolished by atropine, which blocks the action of acetylcholine on muscarinic receptors located on smooth muscles. Taken together, our results suggest that AME could evoke contractions through a cholinergic mechanism involving a direct activation of muscarinic receptors and/or a stimulatory release of acetylcholine from non-neuronal sources. Indeed, although acetylcholine is regarded as a classical neurotransmitter, a non-neuronal cholinergic system also exists.30 Evidence for acetylcholine synthesis is not only provided by positive anti-ChAT immunoreactivity, but ChAT enzyme activity and/or acetylcholine content have also been determined in the majority of the cells, including in the digestive tract.30

We tried to investigate which was the chemical component of AME responsible for the gastric contractile effect. The chemical composition of yarrow has been analyzed in detail and extracts of this plant have been demonstrated to contain a number of pharmacological active ingredients, including alkaloids, such as choline, and flavonoids such as rutin and apigenin. We first focused our attention toward choline, the natural precursor of acetylcholine. Choline, by itself, is known to activate both nicotinic and muscarinic receptors and thus, it may contract gastrointestinal tissue either through direct activation of muscarinic receptors on smooth muscle or through the release of acetylcholine following neuronal nicotinic receptor activation.31 We found that choline contracted, in a concentration-dependent manner, the stomach strip. Similar to AME, the effect of choline was completely blocked by atropine, but insensitive to tetrodotoxin or hexamethonium, suggesting that choline evokes contractions through a direct activation of muscarinic receptors located on smooth muscle. The hypothesis that choline was one of the active ingredients responsible of AME action was further supported by the observation that AME-wpc, which is an extract without choline related polar compounds, displayed no pharmacological activity. Overall, such results indicate that choline, or a related polar compound, is the chemical ingredient responsible of AME pharmacological activity in the gastric strips. Other compounds present in yarrow, such as the flavonoids rutin and apigenin, unlikely contribute to the spasmogenic effect of the extract, as these compounds exerted a relaxing effect on mouse gastric strips. The relaxant effect of flavonoids on the mouse isolated stomach has been previously documented.32–34

We also confirmed in human tissues the main findings reported for the mouse stomach. We found that AME as well as choline contracted, in a tetrodotoxin- and hexamethonium-insensitive and in an atropine-sensitive manner the human gastric antrum. The spasmogenic effect of AME on the human stomach let us hypothesize that yarrow might exert antidyspeptic effects in humans through a stimulatory action on gastric motility. In addition, we found both in mice and humans that the potency of choline contained in the extract was superior to the potency of choline as pure compound (see Fig. 4). Therefore, despite the presence in the extract of muscle relaxant flavonoids, synergistic/additive effects may occur among choline and other not-yet-identified gastric spasmogenic ingredients. For example, the contribution of choline esters, in some cases found to co-occur with choline35,36 cannot be excluded.

As drugs that contract gastric antrum may affect gastric emptying and in the light of the observation that alterations in gastric emptying are observed in dyspeptic patients,7,8 we evaluated the effect of AME on gastric emptying in healthy and dyspeptic mice in vivo. We found that AME (but not AME-wpc) stimulated gastric emptying in healthy mice, which is relevant in the light of the observation that delayed gastric emptying is found in up to 30% of patients with FD.6 Notably, prokinetic drugs, such as 5-HT4 receptor agonists or dopamine D2 agonists, which are widely used in the treatment of FD8 are known to stimulate gastric emptying in mice.37,38 When AME was evaluated in a model of gastric abnormalities induced by a cancer chemotherapic drug, we found that it was more active and effective in pathologic than in physiologic state.

In conclusion, we have shown that a standardized dry water extract obtained from the flowering tops of A. millefolium is able to contract mice and humans gastric antrum. The alkaloid choline, through its ability to activate muscarinic receptors, could contribute, at least in part, to the pharmacological action of the extract. The contractile/prokinetic action of AME observed both in mice and human tissues as well as AME ability to normalize the gastric emptying altered by the antitumoral agent cisplatin could provide a pharmacological basis for the traditional use of AME in the treatment of dyspeptic ailments.

Author Contribution

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References

FB and AAI conceived and designed the experiments; IF, RC, OT, BR, GA performed the experiments; FB, IF and RC analyzed the data; FB, AAI wrote the manuscript; GA, ECB, FC contributed reagents/materials/analysis tools.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. Author Contribution
  8. Competing Interests
  9. References