Marcellus H.L.P. Souza

Role of the NO/cGMP/K(ATP) pathway in the protective effects of sildenafil against ethanol-induced gastric damage in rats.

Sildenafil is a selective inhibitor of cGMP‐specific phosphodiesterase. Sildenafil, acting via NO‐dependent mechanisms, prevents indomethacin‐induced gastropathy. Activation of ATP‐sensitive potassium channels (KATP) is involved in gastric defence. Our objective was to evaluate the role of the NO/cGMP/KATP pathway in the protective effects of sildenafil against ethanol‐induced gastric damage.

Anal canal anatomy showed by three-dimensional anorectal ultrasonography

BackgroundDemonstrate precisely the anatomic configuration of the anal canal and the length and thickness of the anal sphincters using three-dimensional (3-D) anorectal ultra-sonography in both genders.MethodsTwelve normal volunteer males and 14 females, with a mean age of 52.4 and 50.3 years, respectively, were prospectively enrolled in this study. All individuals from both groups were submitted to anorectal ultra-sonography. The anal canal was analyzed, measuring the length and thickness of the external anal sphincter (EAE), internal anal sphincter (IAS), puborectalis muscle (PR) and the gap (distance from the anterior EAS to the anorectal junction) in the midline longitudinal (ML) and transverse (MT) planes, and the results were compared between quadrants and genders.ResultsThe distribution of sphincter muscles is asymmetric in both genders. The anterior upper anal canal is an extension of the rectal wall with all layers clearly identified. The anterior IAS is formed in the distal upper anal canal and is significantly shorter in female than in male in all quadrants. The anterior IAS length is shorter than the posterior and lateral in both genders. The anterior EAS length is significantly shorter (2.2 cm) and the gap is longer (1.2 cm) in female than in male (3.4 cm) (0.7 cm) (p < 0.05), respectively. The posterior and lateral EAS-PR is significant longer in males (3.6 cm) (3.9 cm) than in females (3.2 cm) (3.5 cm) (p < 0.05), respectively. The lateral EAS-PR is significant longer than the posterior part in both genders. The anterior IAS is significantly thicker in males (0.19 cm) than in females (0.12 cm) (p = 0.04).Conclusion3-D anal endosonography enabled measurement of the different anatomical structures of the anal canal and demonstrated its asymmetrical configuration. The shorter anterior EAS and IAS associated with a longer gap could justify the higher incidence of pelvic floor dysfunction in females, especially fecal incontinence and anorectocele with rectal intussusception.

Gastric damage and granulocyte infiltration induced by indomethacin in tumour necrosis factor receptor 1 (TNF-R1) or inducible nitric oxide synthase (iNOS) deficient mice

Background: Tumour necrosis factor α (TNF-α) is involved in non-steroidal anti-inflammatory drug induced gastropathy. Nitric oxide (NO) is a mediator of gastrointestinal mucosal defence but, paradoxically, it also contributes to mucosal damage. Aims: We optimised the C57BL/6 mouse model of indomethacin induced gastropathy to evaluate the role of TNF-α and inducible nitric oxide synthase (iNOS) generated NO in gastric damage and granulocyte infiltration using tumour necrosis factor receptor 1 (TNF-R1−/−) or iNOS (iNOS−/−) deficient mice. Methods: Different doses of indomethacin (2.5, 5, 10, 20 mg/kg) were administered and animals were assessed 6, 12, or 24 hours later. Gastric damage was measured by the sum of all erosions in the gastric mucosa, and gastric granulocyte infiltration was determined by myeloperoxidase (MPO) activity. Other groups of wild-type mice received thalidomide, dexamethasone, fucoidin, l-NAME, or 1400W, and then indomethacin was administered. Additionally, indomethacin was administered to TNF-R1−/− or iNOS−/−. Gastric damage and MPO activity were evaluated 12 hours later. Results: Indomethacin induced dose and time dependent gastric damage and increase in MPO activity in wild-type mice, with the greatest effect at a dose of 10 mg/kg and after 12 hours. Treatment with thalidomide, dexamethasone, or fucoidin reduced gastric damage and MPO activity induced by indomethacin. After indomethacin administration, TNF-R1−/− had less gastric damage and MPO activity than controls. Genetic (knockout mice) or pharmacological (1400W and l-NAME) inhibition of iNOS activity reduced indomethacin induced gastric damage, despite no reduction in MPO activity. Conclusion: TNF-α, acting via TNF-R1, is involved in indomethacin induced gastric damage and granulocyte infiltration. Furthermore, iNOS generated NO is involved in gastric damage induced by indomethacin.

Hydrogen sulfide prevents ethanol-induced gastric damage in mice: role of ATP-sensitive potassium channels and capsaicin-sensitive primary afferent neurons.

The aim of this study was to evaluate the protective effect of hydrogen sulfide (H2S) on ethanol-induced gastric lesions in mice and the influence of ATP-sensitive potassium (KATP) channels, capsaicin-sensitive sensory afferent neurons, and transient receptor potential vanilloid (TRPV) 1 receptors on such an effect. Saline and l-cysteine alone or with propargylglycine, sodium hydrogen sulfide (NaHS), or Lawessons reagent were administrated for testing purposes. For other experiments, mice were pretreated with glibenclamide, neurotoxic doses of capsaicin, or capsazepine. Afterward, mice received l-cysteine, NaHS, or Lawessons reagent. After 30 min, 50% ethanol was administrated by gavage. After 1 h, mice were sacrificed, and gastric damage was evaluated by macroscopic and microscopic analyses. l-Cysteine, NaHS, and Lawessons reagent treatment prevented ethanol-induced macroscopic and microscopic gastric damage in a dose-dependent manner. Administration of propargylglycine, an inhibitor of endogenous H2S synthesis, reversed gastric protection induced by l-cysteine. Glibenclamide reversed l-cysteine, NaHS, or Lawessons reagent gastroprotective effects against ethanol-induced macroscopic damage in a dose-dependent manner. Chemical ablation of sensory afferent neurons by capsaicin reversed gastroprotective effects of l-cysteine or H2S donors (NaHS or Lawessons reagent) in ethanol-induced macroscopic gastric damage. Likewise, in the presence of the TRPV1 antagonist capsazepine, the gastroprotective effects of l-cysteine, NaHS, or Lawessons reagent were also abolished. Our results suggest that H2S prevents ethanol-induced gastric damage. Although there are many mechanisms through which this effect can occur, our data support the hypothesis that the activation of KATP channels and afferent neurons/TRPV1 receptors is of primary importance.

Role of resident mast cells and macrophages in the neutrophil migration induced by LTB4, fMLP and C5a des arg.

In the present study, we have investigated the participation of resident peritoneal cells (macrophages and mast cells) in the neutrophil migration induced in rats by the intraperitoneal administration of LTB4, fMLP or C5a des arg. The intraperitoneal injection of LTB4 (10 nmol), fMLP (10 nmol) and C5a des arg (zymosan-activated plasma, 1 ml) caused an intense neutrophil migration compared to the saline control (1,000, 1,500 and 2,000%, respectively). An 83% depletion in the number of resident cells following peritoneal lavage reduced the LTB4-stimulated neutrophil migration by 73.6% without affecting that caused by fMLP and C5a des arg. Increasing the peritoneal macrophage population (236%) by pretreating the cavities with thioglycollate enhanced the neutrophil migration induced by LTB4 (129%), but did not alter that induced by fMLP and C5a des arg. Similarly, reducing the population of peritoneal mast cells containing toluidine-blue-staining granules by subchronically pretreating the cavities with compound 48/80 diminished the LTB4-induced NM by 69% but had no effect on the responses to fMLP and C5a des arg. Pretreating the animals with dexamethasone strongly inhibited (70%) the neutrophil migration induced by the intraperitoneal injection of LTB4, fMLP and C5a des arg. Indomethacin, BW A4C and NDGA had no such effect. The incubating medium from peritoneal macrophages and mast cells stimulated with LTB4 induced neutrophil migration when injected into the peritoneal cavity of rats. This migration was strongly reduced (70%) by treating the cells with dexamethasone. In contrast, stimulating the cells with fMLP or C5a des arg did not result in the release of any promigratory activity into the incubating fluid. Our results suggest that LTB4 induces neutrophil migration via a mechanism dependent on resident mast cells and macrophages while that induced by C5a des arg and fMLP seems to be independent of such cellular involvement. The neutrophil migration induced by LTB4 is apparently mediated by factor(s) whose release is blocked by dexamethasone. fMLP and C5a appear to cause in vivo migration by the formation of a concentration gradient and by a glucocorticoid-sensitive mechanism different from that stimulated by LTB4.

Peripheral neuropathy and neurological disorders in an unselected Brazilian population-based cohort of IBD patients.

Background: Several neurological disorders have been described in inflammatory bowel disease (IBD) patients, but their exact frequency is unknown. Methods: We prospectively studied the prevalence of neurological disorders (especially peripheral neuropathy) in a group of 82 patients with Crohns disease (CD, n = 31) or ulcerative colitis (UC, n = 51) from 2 Brazilian tertiary care university clinics and followed them through a period of at least 1 year. All patients were interviewed and had complete neurological evaluations. Results: Large‐fiber sensory or sensorimotor polyneuropathy (PN) was observed in 16.1% of the CD and 19.6% of the UC patients. PN was usually mild, predominantly symmetric, and distal with axonal involvement. One patient had demyelinating PN at the diagnosis of CD. Mild carpal tunnel syndrome was common in female UC patients. Sensory symptoms without electromyography abnormalities, suggestive of small‐fiber neuropathy or subclinical myelopathy, affected 29% and 11.8%, respectively. After excluding other known etiological or contributory factors for PN, 13.4% of the IBD patients had otherwise unexplained large‐fiber or small‐fiber PN (7.3% with large‐fiber SM PN). Nondebilitating headache was the most common neurological complaint. Three patients had ischemic strokes, 5 were epileptic, and 1 transient chorea. Conclusions: Neurological disorders, especially PN, are common in our Brazilian cohort of IBD patients. They are diverse, multifactorial, and more common in women. Despite the mild phenotype in most cases, attention should be given by the general practitioner and gastroenterologist since they are frequently undiagnosed. Further studies are necessary to confirm these findings in populations with different genetic and nutritional backgrounds.

Antiinflammatory and antinociceptive effects in mice of a sulfated polysaccharide fraction extracted from the marine red algae Gracilaria caudata

Many algal species contain relatively high concentrations of polysaccharide substances, a number of which have been shown to have anti-inflammatory and/or immunomodulatory activity. In this study, we evaluated the anti-inflammatory and antinociceptive effects in mice of a sulfated polysaccharide fraction (PLS) extracted from the algae Gracilaria caudata. The antiinflammatory activity of PLS was evaluated using several inflammatory agents (carrageenan, dextran, bradykinin, and histamine) to induce paw edema and peritonitis in Swiss mice. Samples of the paw tissue and peritoneal fluid were removed to determine myeloperoxidase (MPO) activity or TNF-α and IL-1β levels, respectively. Mechanical hypernociception was induced by subcutaneous injection of carrageenan into the plantar surface of the paw. Pretreatment of mice by intraperitoneal administration of PLS (2.5, 5, and 10 mg/kg) significantly and dose-dependently reduced carrageenan-induced paw edema (p < 0.05) compared to vehicle-treated mice. Similarly, PLS 10 mg/kg effectively inhibited edema induced by dextran and histamine; however, edema induced by bradykinin was unaffected by PLS. PLS 10 mg/kg inhibited total and differential peritoneal leukocyte counts following carrageenan-induced peritonitis. Furthermore, PLS reduced carrageenan-increased MPO activity in paws and reduced cytokine levels in the peritoneal cavity. Finally PLS pretreatment also reduced hypernociception 3–4 h after carrageenan. We conclude that PLS reduces the inflammatory response and hypernociception in mice by reducing neutrophil migration and cytokines concentration.

Sildenafil prevents indomethacin-induced gastropathy in rats: role of leukocyte adherence and gastric blood flow

Nitric oxide (NO) is an important mediator of gastric mucosal defense. Sildenafil (SILD), a cyclic GMP‐specific phosphodiesterase inhibitor, promotes an increase in cGMP concentrations in the gastrointestinal tract. cGMP mediates many of the biological actions of NO. We tested the hypothesis that SILD could increase mucosal defense against indomethacin‐induced gastropathy in rats. SILD (1, 4 or 10 mg kg−1, p.o.) pretreatment significantly reduced (P<0.01) the gastric damage and the increase in gastric myeloperoxidase (MPO) activity elicited by indomethacin (20 mg kg−1 p.o.), with the maximal effect at the dose of 10 mg kg−1. L‐NAME (3, 10 or 20 mg kg−1, i.p.) dose dependently reversed the protective effects of SILD, an effect not seen when L‐arginine (L‐ARG) (200 mg kg−1, i.p.) was co‐administered with L‐NAME. Indomethacin‐induced leukocyte adhesion, assessed by intravital microscopy, was decreased (P<0.01) by SILD, and this effect was reversed by L‐NAME cotreatment. Indomethacin elicited a decrease in gastric blood flow and in gastric PGE2 levels. SILD was able to prevent the decrease in gastric blood flow (P<0.01), without diminishing the inhibitory effect of indomethacin on prostaglandin synthesis. These results indicate that SILD, acting via NO‐dependent mechanisms, prevents indomethacin‐induced gastropathy, possibly through a reduction of leukocyte adhesion and maintenance of gastric blood flow.

Inhaled 1,8-Cineole Reduces Inflammatory Parameters in Airways of Ovalbumin-Challenged Guinea Pigs

Eucalyptol, also known as 1,8-cineole, is a monoterpene traditionally used to treat respiratory disorders due to its secretolytic properties. In addition to its myorelaxant effects, it also has anti-inflammatory actions in vitro. In this study, we aimed to evaluate the efficacy of acute treatment with 1,8-cineole on reducing airway inflammatory parameters. Ovalbumin (OVA)-sensitized guinea pigs were submitted to antigenic challenge (OVA) with or without pre-treatment with a single dose of 1,8-cineole administered by inhalation. Airway inflammatory parameters were reduced or absent in 1,8-cineole-treated animals as compared with untreated guinea pigs. Acute treatment with 1,8-cineole impaired the development of airway hyperresponsiveness to carbachol in isolated tracheal rings. Levels of the pro-inflammatory cytokines TNFα and IL-1β was lower in bronchoalveolar lavage fluid (BALF) of 1,8-cineol-treated guinea pigs than in untreated animals. 1,8-Cineole impaired the OVA-induced increase of the myeloperoxidase activity in BALF. 1,8-Cineole also prevented the reduction of the mucociliary clearance induced by the antigen presentation. The present investigation provides evidence that inhaled 1,8-cineole prevents hyperresponsiveness and inhibits inflammation in airways of ovalbumin-challenged guinea pigs.

The involvement of macrophage‐derived tumour necrosis factor and lipoxygenase products on the neutrophil recruitment induced by Clostridium difficile toxin B

Clostridium difficile (Cd) toxins appear to mediate the inflammatory response in pseudomembranous colitis and/or colitis associated with the use of antibiotics. In contrast to Cd Toxin A (TxA), Cd Toxin B (TxB) has been reported not to promote fluid secretion or morphological damage in rabbits and hamsters and also does not induce neutrophil chemotaxis in vitro. However, TxB is about 1000 times more potent than TxA in stimulating the release of tumour necrosis factor‐α (TNF‐α) by cultured monocytes. In the present study, we investigated the ability of TxB to promote neutrophil migration into peritoneal cavities and subcutaneous air‐pouches of rats. We also examined the role of resident peritoneal cells in this process as well as the inflammatory mediators involved. TxB caused a significant and dose‐dependent neutrophil influx with a maximal response at 0·1 μg/cavity after 4 hr. Depleting the peritoneal resident cell population by washing the peritoneal cavity or increasing this population by pretreating the animals with thioglycollate blocked and amplified the TxB‐induced neutrophil migration, respectively. Pretreating the animals with MK886 (a lipoxygenase inhibitor), NDGA (a dual cyclo‐ and lipoxygenase inhibitor) or the glucocorticoid, dexamethasone, but not with indomethacin (a cyclo‐oxygenase inhibitor), or BN52021 (a platelet‐activating factor antagonist), inhibited the neutrophil migration evoked by TxB. Pretreatment with dexamethasone or the administration of anti‐TNF‐α serum into the air‐pouches also significantly reduced the TxB‐induced neutrophil migration. Supernatants from TxB‐stimulated macrophages induced neutrophil migration when injected into the rat peritoneal cavity. This effect was attenuated by the addition of either MK886 or dexamethasone to the macrophage monolayer and by preincubating the supernatants with anti‐TNF‐α serum. TxB also stimulated the release of TNF‐α by macrophages. Overall, these results suggest that TxB induces an intense neutrophil migration which is mediated by macrophage‐derived TNF‐α and lipoxygenase products.