Dual role of eugenol on chronic gastric ulcer in rats: Low-dose healing efficacy and the worsening gastric lesion in high doses.

This study evaluated the gastric healing activity of eugenol, the main bioactive compound from clove (Syzygium aromaticun) essential oil. Five groups of female Wistar rats were submitted to acetic acid-induced ulcer model and treated with Vehicle (1 mL/kg, p.o.), eugenol (1, 10 or 100 mg/kg, p.o) or omeprazole (20 mg/kg, p.o), twice a day, by seven or fourteen days. Macroscopic, microscopic and biochemical analyses were performed in the ulcerated site. Eugenol (1 mg/kg, p.o) administered by 7 or 14 days accelerated the gastric healing process by 33% and 52%, respectively. The healing actions of eugenol were accompanied by the rescue on the histological architecture and the normalization of the superoxide dismutase and catalase activity. Moreover, eugenol (1 mg/kg, p.o) reduced the gastric mucosal myeloperoxidase activity and increased the mucin secretion. In contrast, eugenol at a dose of 100 mg/kg administered by 7 days enhanced 49% the ulcerated area, but at 10 mg/kg did not change the ulcer area after 7 or 14 days of treatment. Thus, despite the undesirable results due to the worsening of the gastric lesion with the use of eugenol in high doses, the antiulcer potential of this compound is evident and manageable in an adequate dose.

Role of the antioxidant properties in the gastroprotective and gastric healing activity promoted by Brazilian green propolis and the healing efficacy of Artepillin C.

Green propolis is a resinous substance used in folk medicine given its anti-inflammatory, antibacterial, and anti-ulcer effects. Our research group has already confirmed the gastroprotective activity of hydroalcoholic extract from green propolis (HEGP), as well as of its main isolated compounds. In continuity, this study evaluated the antioxidant mode of action involved in the preventive effect induced by HEGP, and its therapeutic gastric healing potential on installed ulcers. In addition, the healing effect of its main compound Artepillin C was also investigated. Acute and chronic ulcers were induced in rats by given ethanol or acetic acid, respectively. In acute model, the rats were orally pre-treated with vehicle (water plus 1% Tween, 1 mL/kg), HEGP (30-300 mg/kg), or carbenoxolone (200 mg/kg) 1 h prior the ulcer induction. In the chronic ulcer protocol, the rats received vehicle (water plus 1% Tween, 1 mL/kg), HEGP (300 mg/kg), or omeprazole (20 mg/kg) twice a day by 7 days, whereas groups of mice received vehicle (water plus 1% Tween, 1 mL/kg), Artepillin C (18 mg/kg), or ranitidine (20 mg/kg) twice a day by 4 days. Ulcerated tissue was collected for histological, histochemical, immunostaining, oxidative, and inflammatory analyses. The in vitro scavenger activity of HEGP was also verified using the DPPH assay. The oral pre-treatment with HEGP (100 and 300 mg/kg) prevented the gastric epithelial damage promoted by ethanol. Besides, HEGP (100 and 300 mg/kg) reduced SOD activity about 11% and 26%, respectively, and increased the activity of GST around 20% and CAT in 80%. HEGP (300 mg/kg) also reduced the production of reactive oxygen species, as well as lipoperoxidation levels in the ethanol-ulcerated tissue. In the acetic acid-induced chronic ulcer, the daily treatment with HEGP (300 mg/kg) accelerates the healing process by 71%. In this model, HEGP normalized SOD and CAT activity and increased GST activity by 109% when compared to non-ulcerated rats. In both models, the extract administration increased the mucin PAS staining and reduced the myeloperoxidase activity at the ulcer site. Moreover, the treatment with HEGP enhanced the PCNA immunostaining, but did not alter the concentration of collagen in the acetic acid-ulcerated tissue. The extract had a direct DPPH radical-scavenging ability (LogIC50: 0.56). Besides, as expected, HPLC analysis showed Artepillin C as a major compound and its administration at 18 mg/kg also accelerated the gastric healing ulcer process in mice. Our findings confirm that HEGP displays both gastroprotective and gastric healing properties, contributing to the validation of its popular use as preventive and therapeutic approaches. These actions occur through the increase in mucin production and the reestablishment of the oxidative balance due to a reduction in gastric inflammation.

Mechanisms underlying Eugenia mattosii D. Legrand leaves extract, fractions and compounds induce relaxation of the aorta from normotensive and hypertensive rats.

The present study aimed to verify the effect of methanolic extract, fractions, and phenolic compounds of Eugenia mattosii D. Legrand leaves on the aorta relaxation. Isometric tensions were measured on the aorta of normotensive (NTR) and spontaneously hypertensive rats (SHR). The results showed that both methanolic extracts of leaves and stems, as well as, fractions obtained from leaves were able to induce a concentration-dependent relaxation in both endothelium-intact and -denuded aortas. The methanolic extract of leaves (ME-leaves) was the most effective since the maximal relaxation (≈ 83%) obtained was at the concentration of 300 µg/mL. As the endothelium-dependent relaxation was more significant, we investigated the mechanisms by which ME-leaves induced this effect. After the pretreatment with LNAME, ME-leaves-induced relaxation was decreased in the aorta of NTR and SHR. However, the pretreatment with methylene blue only reduced the relaxation in the aorta of NTR. Furthermore, pretreatment with ME-leaves decreased phenylephrine-induced contraction in preparation Ca2+-free only in aortic rings from NTR. This study also reveals that both compounds, cryptostrobin isolated from chloroform fraction and catechin from the ethyl acetate fraction induced a marked relaxation in endotheliumintact aortic rings of NTR. In conclusion, ME-leaves induces relaxation in the rat aorta involves the modulation of NO/cGMP dependent signaling pathway, this mechanism may at least, in part, explain the endothelium-dependent relaxation. Furthermore, cryptostrobin and catechin also induced relaxation, which may contribute synergistically to the vasorelaxation effect of the ME-leaves.

Cryptostrobin and catechin isolated from Eugenia mattosii D. Legrand leaves induce endothelium-dependent and independent relaxation in spontaneously hypertensive rat aorta.

BACKGROUND: Considering the therapeutic potential of phenolic compounds, the purpose of the present study was to investigate the mechanisms involved in the relaxation induced by cryptostrobin and catechin, isolated from Eugenia mattosii D. Legrand leaves, in the aorta of spontaneously hypertensive rats (SHR). METHODS: The thoracic aorta was isolated from SHR and kept in the organ bath system by recording contractile or relaxant responses. RESULTS: The addition of cumulative concentrations of cryptostrobin and catechin induced endothelium-dependent and-independent relaxation in aorta rings from SHR, as well as both compounds were effective in reducing phenylephrine-induced contraction. Pretreatment of aortic rings with Nω-nitro-l-arginine methylester (L-NAME, an inhibitor of nitric oxide synthase) or 1H-[1,2,4] oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, an inhibitor of soluble guanylate cyclase), resulted in a significant change of relaxant effect induced by catechin, and a slight influence on cryptostrobin-induced relaxation. Muscarinic receptor and potassium channels are involved in catechin-induced relaxation as assessed using atropine (a muscarinic receptor antagonist), tetraethylammonium (a non-selective K+ channel blocker) and glibenclamide (an ATP-sensitive K+ channel blocker). Conversely, cryptostrobin, but not catechin, blunted the contraction induced by the addition of phenylephrine in a calcium-free solution. Besides that, cryptostrobin attenuated the contraction of rat aorta rings induced by internal Ca2+ release and external Ca2+ influx. CONCLUSIONS: These findings indicated that cryptostrobin and catechin alter vascular smooth muscle reactivity, and this effect may be involved, at least in part, by enhancing the endothelium NO/cGMP pathway and potassium channels activation. In addition, cryptostrobin reduced the phenylephrine, KCl and CaCl2-induced contractions in a calcium-free solution.

1,3,5,6-Tetrahydroxyxanthone, a natural xanthone, induces diuresis and saluresis in normotensive and hypertensive rats.

The aim of the present study was to evaluate the diuretic effect of 1,3,5,6-tetrahydroxyxanthone (THX), isolated from preparations of Garcinia achachairu Rusby (Clusiaceae) branches, in rats. Wistar normotensive (NTR) and spontaneously hypertensive rats (SHR) received a single oral treatment with THX, hydrochlorothiazide (HCTZ) or just vehicle (VEH). The effects of THX in combination with diuretics of clinical use, as well as with l-NAME, atropine, and indomethacin were also explored. Cumulative urine volume and urinary parameters were measured at the end of the 8-h or 24-h experiment. THX was able to stimulate 8-h and 24-h diuresis in both NTR and SHR, as well as urinary Na+ and K+ excretion, at a dose of 0.1 mg/kg; while 8-h urinary Cl- levels were only significantly increased in the group of animals treated with THX at the dose of 0.3 mg/kg. In addition, Ca2+ content was reduced in the 24-h urine of THX-treated NTR and SHR, like that obtained in the HCTZ (10 mg/kg) group. The combination with HCTZ or furosemide, but not with amiloride, significantly enhanced THX-induced diuresis. The diuretic effect with HCTZ plus THX treatment was accompanied by an increase of the urinary Na+, K+, and Cl- excretion. On the other hand, when given THX in combination with amiloride, there was a significant increase in Na+ and a decrease in K+ excretion, an effect characteristic of this class of diuretics. Moreover, the diuretic effect of THX was heightened after pretreatment with l-NAME, and its ability to induce diuresis was prevented neither in the presence of indomethacin nor in the presence of atropine. However, the pretreatment with atropine completely avoided the saluretic effect stimulated by THX, suggesting, at least in part, the role of muscarinic receptors in the renal effects of THX disclosed in this study.

Bauhinia forficata link, a Brazilian medicinal plant traditionally used to treat cardiovascular disorders, exerts endothelium-dependent and independent vasorelaxation in thoracic aorta of normotensive and hypertensive rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Bauhinia forficata Link, commonly known as "cow's paw", is a native plant from South America. Its leaves are widely used in Brazilian folk medicine to treat diabetes and cardiovascular disorders. Although this species' biological potential has been extensively proven as an antidiabetic, anti-inflammatory and antioxidant agent, there is a lack of studies to evidence its action on the cardiovascular system. AIM OF THE STUDY: This study was designed to investigate the vascular effects of B. forficata leaves preparations and its majority compound kaempferitrin, as well as its aglycone form kaempferol, in rat aortic rings of normotensive (NTR) and hypertensive (SHR) rats. MATERIALS AND METHODS: Aorta rings from NTR and SHR precontracted with phenylephrine were exposed to cumulative concentrations of B. forficata extract, fractions (1-50 µg/mL) and compounds (0.001-0.3 µg/mL). The mechanisms involved in the vasorelaxant effect of ethyl-acetate plus butanol fraction (EAButF) were also evaluated. RESULTS: Although kaempferitrin is the most abundant compound found in both methanolic extract and EAButF, 24 minor phenolic compounds were identified in B. forficata leaves, including kaempferol. EAButF was the only with endothelium-dependent and independent vasorelaxant properties in both NTR and SHR. The incubation with L-NAME or ODQ completely blocked EAButF-induced vasorelaxation. On the other hand, the incubation with propranolol, atropine, indomethacin, glibenclamide or barium chloride did not change the vasorelaxant activity of EAButF (50 µg/mL). Nevertheless, the incubation with tetraethylammonium and 4-aminopyridine significantly influenced the EAButF activity. It was also shown that Ca2+ influx or efflux is not related to EAButF vasorelaxation potential. Kaempferitrin and kaempferol were also able to relax the rat aortic rings in 34.70% and 40.54%, respectively. CONCLUSIONS: This study shows, for the first time, the vasorelaxant effect of EAButF from B. forficata leaves, an effect that may be attributed to the modulation of vascular tone through nitric oxide/soluble guanylate cyclase pathway, and potassium channels. The bioactive kaempferitrin and kaempferol seem to be important for the effects observed with the fraction. Finally, preparations obtained from the leaves of B. forficata may be interesting candidates for new or complementary strategies regarding cardiovascular diseases.

Hydroalcoholic extract of Tagetes erecta L. flowers, rich in the carotenoid lutein, attenuates inflammatory cytokine secretion and improves the oxidative stress in an animal model of ulcerative colitis.

Given the role of oxidative stress in ulcerative colitis (UC) etiology, and the amount of lutein (a carotenoid with antioxidant properties) in the dry hydroalcoholic extract of Tagetes erecta flowers (DHETE), this study investigated the intestinal anti-inflammatory properties of DHETE in an animal model of UC. The amount of lutein in the extract was determined by 1H-nuclear magnetic resonance spectroscopy, and total phenols, radical scavenger capability, cytotoxicity, and effects on reactive oxygen species and nitric oxide production were evaluated in vitro. Experimental UC was established by adding 5% dextran sulfate sodium (DSS) to drinking water, with the effects of DHETE (30-300 mg/kg, once a day for 7 days) on the morphological (colon length and weight), clinical (disease activity index and body weight loss), microscopic (histological score and mucin levels), and biochemical parameters analyzed. The lutein concentration found in DHETE was 8.2%, and DHETE scavenged 2,2-diphenyl-1-picrylhydrazyl radicals at 1000 µg/mL The exposure of intestinal epithelial cells to DHETE did not change its viability but reduced reactive oxygen species and nitric oxide production after lipopolysaccharide stimulation. In vivo, DHETE (300 mg/kg) attenuated weight loss, disease activity index, colon shortening, and histopathological changes promoted by DSS intake. Moreover, DHETE increased mucin colonic staining. The treatment with DHETE decreased myeloperoxidase activity as well as tumor necrosis factor and interleukin-6 levels. The extract also increased reduced glutathione levels and catalase activity and normalized superoxide dismutase and glutathione-S-transferase activities. In conclusion, DHETE reduced colitis severity by attenuating inflammatory cytokine secretion and improved the endogenous antioxidant defense in DSS-induced UC in mice.

Preclinical evaluation of the diuretic and saluretic effects of (-)-epicatechin and the result of its combination with standard diuretics.

Several studies have suggested that (-)-epicatechin-containing foods and plant extracts benefit conditions that affect the cardiovascular system, such as hypertension and endothelial dysfunction. However, no study was conducted so far to evaluate the potential of this flavonoid on diuretic activity assay. For that, female Wistar normotensive (NTR) and spontaneously hypertensive rats (SHR) received a single oral treatment with (-)-epicatechin (EPI), hydrochlorothiazide (HCTZ) or just vehicle (VEH). The effects of EPI in combination with diuretics for clinical use, as well as with L-NAME, atropine and indomethacin were also explored. Cumulative urine volume, plasma and urinary parameters were evaluated at the end of 8 h experiment. When given to NTR and SHR, at doses of 0.3, 1 and 3 mg/kg, EPI was able to stimulate both diuresis and saluresis (Na+, K+ and Cl-), without interfering with plasma electrolyte content or urinary pH and uric acid values, when compared with VEH-treated only rats. The combination with HCTZ, but not with furosemide or amiloride, successfully strengthened EPI-induced diuresis. This effect was not accompanied by a potentiation of the saluretic effects. On the other hand, when given EPI in combination with amiloride, a significant increase in Cl- excretion and maintenance of the potassium-sparing effects characteristic of this class of diuretics were detected. In addition, the diuretic effect of EPI was enhanced after pretreatment with L-NAME and its action was significantly precluded in the presence of indomethacin, a cyclooxygenase inhibitor. In conclusion, this study shows the diuretic and saluretic properties of EPI in rats, adding another biological activity whose effect may contribute to the different positive actions already described.

Artepillin C, drupanin, aromadendrin-4'-O-methyl-ether and kaempferide from Brazilian green propolis promote gastroprotective action by diversified mode of action.

ETHOPHARMACOLOGICAL RELEVANCE: The propolis is extensively used in folk medicine in natura or to prepare pharmaceutical formulations since ancient time to improve health or prevent diseases, among them gastrointestinal disorders. Aiming to contribute in the scientific validation about the popular use of Brazilian Green propolis (BGP) against gastritis and gastric ulcer, this work evaluated the antiulcer potential of isolated compounds from BGP, three prenylated p-coumaric acid derivatives and two flavonoids, respectively named: 3,5 diprenyl-4-hydroxycinnamic acid (artepillin C) (1), 3-prenyl-4-dihydroxycinnamoiloxy cinnamic acid (baccharin) (2), 3-prenyl-4-hydroxycinnamic acid (drupanin) (3), aromadendrin-4'-O-methyl-ether (4) and kaempferide (5). MATERIAL AND METHODS: The compounds were characterized by nuclear magnetic resonance and mass spectrometry. Their gastroprotective effects were evaluated against ethanol/HCl- and indomethacin-induced ulcer in mice. Further, histological, histochemical, oxidative and inflammatory parameters were analyzed at ulcerated tissue. Acid antisecretory activities also were also assessed. RESULTS: Compound 2 did not reduce the ethanol/HCl- induced ulcer at 30 mg/kg (p.o), whereas the minimum oral gastroprotective doses of 1, 3, 4 and 5 were 0.3, 0.3, 3 and 3 mg/kg, respectively. Besides, these compounds prevented ethanol/HCl-induced ulcer by intraperitoneal route, as well as indomethacin-induced ulcer by oral route. The gastroprotection was accompanied by normalization of superoxide dismutase, catalase and glutathione-S-transferase activities and reduction in myeloperoxidase activity. Moreover, the compounds 4 and 5 increased the gastric mucin content and 1 reduced TNF amount. Furthermore, 1, 3, 4 and 5 decreased volume, pH, total acidity and pepsin activity of the gastric juice from rats. CONCLUSIONS: Together, our findings showed a diversified mode of action elicited by 1, 3, 4 and 5 on the gastroprotection and contribute to explain the anti-ulcer activity reported for BGP.

Gallic acid, a phenolic compound isolated from Mimosa bimucronata (DC.) Kuntze leaves, induces diuresis and saluresis in rats.

Although present in the leaves of Mimosa bimucronata (DC.) and many other medicinal plants commonly used to augment urinary volume excretion, the effects of gallic acid as a diuretic agent remain to be studied. Wistar rats were orally treated with vehicle, hydrochlorothiazide, or gallic acid. The effects of gallic acid in the presence of hydrochlorothiazide, furosemide, amiloride, L-NAME, atropine, and indomethacin were also investigated. Diuretic index, pH, conductivity, and electrolyte excretion were evaluated at the end of the experiment (after 8 or 24 h). Gallic acid induced diuretic and saluretic (Na+ and Cl-) effects, without interfering with K+ excretion, when orally given to female and male rats at a dose of 3 mg/kg. These effects were associated with increased creatinine and conductivity values while pH was unaffected by any of the treatments. Plasma Na+, K+, and Cl- levels were not affected by any of the acute treatments. The combination with hydrochlorothiazide or furosemide was unable to intensify the effects of gallic acid when compared with the response obtained with each drug alone. On the other hand, the treatment with amiloride plus gallic acid amplified both diuresis and saluresis, besides to a marked potassium-sparing effect. Its diuretic action was significantly prevented in the presence of indomethacin, a cyclooxygenase inhibitor, but not with the pretreatments with L-NAME or atropine. Although several biological activities have already been described for gallic acid, this is the first study demonstrating its potential as a diuretic agent.

Brazilian green propolis hydroalcoholic extract reduces colon damages caused by dextran sulfate sodium-induced colitis in mice.

This study investigated the effects of Brazilian green propolis hydroalcoholic extract (BPE) in 3% w/v dextran sodium sulfate (DSS)-induced colitis in mice. The effects of BPE (3, 30 and 300 mg/kg, p.o, by 7 days) on the morphological (colon length and colon weight), clinical (disease activity index and weight loss), microscopic (histological score and mucin levels) and biochemical parameters were determined. The effects of BPE (300 mg/kg, p.o) in the gastrointestinal transit of mice were also evaluated. As expected, the DSS ingestion damaged the colonic tissue, lowered the body weight, decreased the mucin levels, increased MPO activity, reduced SOD activity and GSH amount. In contrast, the treatment with BPE (300 mg/kg) significantly reduced macroscopic colonic injury and the mucosal damage in colon on histopathological examination and reversed the decrease in mucin levels induced by DSS. It also significantly normalized the SOD activity and the levels of GSH, but did not elicit any effect on MPO activity in the colon. In addition, BPE did not change the gastric emptying or the intestinal transit rate of mice. Together, these results suggested that BPE reduced the signs of DSS-induced colitis in mice through maintenance of intestinal mucin barrier and favoring intestinal antioxidant defenses.

Acid-gastric antisecretory effect of the ethanolic extract from Arctium lappa L. root: role of H+, K+-ATPase, Ca2+ influx and the cholinergic pathway.

BACKGROUND: Arctium lappa L., popularly known as burdock, is a medicinal plant used worldwide. The antiulcer and gastric-acid antisecretory effects of ethanolic extract from roots of Arctium lappa (EET) were already demonstrated. However, the mechanism by which the extract reduces the gastric acid secretion remains unclear. Therefore, this study was designed to evaluate the antisecretory mode of action of EET. MATERIALS AND METHODS: The effects of EET on H+, K+-ATPase activity were verified in vitro, whereas the effects of the extract on cholinergic-, histaminergic- or gastrinergic-acid gastric stimulation were assessed in vivo on stimulated pylorus ligated rats. Moreover, ex vivo contractility studies on gastric muscle strips from rats were also employed. RESULTS: The incubation with EET (1000 µg/ml) partially inhibited H+, K+-ATPase activity, and the intraduodenal administration of EET (10 mg/kg) decreased the volume and acidity of gastric secretion stimulated by bethanechol, histamine, and pentagastrin. EET (100-1000 µg/ml) did not alter the gastric relaxation induced by histamine but decreased acetylcholine-induced contraction in gastric fundus strips. Interestingly, EET also reduced the increase in the gastric muscle tone induced by 40 mM KCl depolarizing solution, as well as the maximum contractile responses evoked by CaCl2 in Ca2+-free depolarizing solution, without impairing the effect of acetylcholine on fundus strips maintained in Ca2+ -free nutritive solution. CONCLUSION: Our results reinforce the gastric antisecretory properties of preparations obtained from Arctium lappa, and indicate that the mechanisms involved in EET antisecretory effects include a moderate reduction of the H+, K+-ATPase activity associated with inhibitory effects on calcium influx and of cholinergic pathways in the stomach muscle.

Vitamin C Improves Gastroparesis in Diabetic Rats: Effects on Gastric Contractile Responses and Oxidative Stress.

BACKGROUND: Diabetic gastroparesis is a common complication of diabetes mellitus, which mainly affects women. Previous studies have demonstrated that oxidative stress is involved in its onset and development. AIMS: This study evaluated the role of vitamin C on diabetes-associated gastric dysmotility. METHODS: Female rats with streptozotocin-induced diabetes were treated with vehicle (water, 1 mL/kg, p.o.), vitamin C (300 mg/kg/day, p.o.), or insulin (6 IU/day, s.c.). Gastric emptying, in vitro gastric contractility, and biochemistry parameters were analyzed at the end of the treatment (i.e. 8 weeks after the diabetes induction). RESULTS: Vitamin C reversed the delayed gastric emptying of diabetic rats to normal levels, and avoided the changes in the contractile responses to acetylcholine (0.1 nM-1 µM), but not to 5-hydroxytryptamine (0.1 nM-1 µM), in the pylorus and fundus from diabetic rats. Moreover, the contraction evoked by KCl (40 mM) in the fundus, but not in the pylorus, was intensely increased in diabetic rats treated with vitamin C. Notably, the vitamin C reestablished the reduced glutathione levels by 77% and decreased the reactive oxygen species content by 60% in the gastric tissue from diabetic rats. Despite the effects on gastric motility, vitamin C treatment did not change the fasting glycaemia or the glycated hemoglobin of diabetic rats. Unsurprisingly, insulin treatment normalized all parameters evaluated. CONCLUSIONS: Vitamin C exhibited a remarkable beneficial effect on gastric emptying dysfunction in diabetic rats, which was mediated by attenuation of oxidative stress and maintenance of the cholinergic contractile responses in fundus and pylorus.