POZylation: a new approach to enhance nanoparticle diffusion through mucosal barriers

The increasing use of nanoparticles in the pharmaceutical industry is generating concomitant interest in developing nanomaterials that can rapidly penetrate into, and permeate through, biological membranes to facilitate drug delivery and improve the bioavailability of active pharmaceutical ingredients. Here, we demonstrate that the permeation of thiolated silica nanoparticles through porcine gastric mucosa can be significantly enhanced by their functionalization with either 5 kDa poly(2-ethyl-2-oxazoline) or poly(ethylene glycol). Nanoparticle diffusion was assessed using two independent techniques; Nanoparticle Tracking Analysis, and fluorescence microscopy. Our results show that poly(2-ethyl-2-oxazoline) and poly(ethylene glycol) have comparable abilities to enhance diffusion of silica nanoparticles in mucin dispersions and through the gastric mucosa. These findings provide a new strategy in the design of nanomedicines, by surface modification or nanoparticle core construction, for enhanced transmucosal drug delivery.

Structural changes in the epithelium of the small intestine and immune cell infiltration of enteric ganglia following acute mucosal damage and local inflammation

An acute enteritis is commonly followed by intestinal neuromuscular dysfunction, including prolonged hyperexcitability of enteric neurons. Such motility disorders are associated with maintained increases in immune cells adjacent to enteric ganglia and in the mucosa. However, whether the commonly used animal model, trinitrobenzene sulphonate (TNBS)-induced enteritis, causes histological and immune cell changes similar to human enteric neuropathies is not clear. We have made a detailed study of the mucosal damage and repair and immune cell invasion following intralumenal administration of TNBS. Intestines from untreated, sham-operated and TNBS-treated animals were examined at 3 h to 56 days. At 3 h, the mucosal surface was completely ablated, by 6 h an epithelial covering was substantially restored and by 1 day there was full re-epithelialisation. The lumenal epithelium developed from a squamous cell covering to a fully differentiated columnar epithelium with mature villi at about 7 days. Prominent phagocytic activity of enterocytes occurred at 1-7 days. A surge of eosinophils and T lymphocytes associated with the enteric nerve ganglia occurred at 3 h to 3 days. However, elevated immune cell numbers occurred in the lamina propria of the mucosa until 56 days, when eosinophils were still three times normal. We conclude that the disruption of the mucosal surface that causes TNBS-induced ileitis is brief, a little more than 6 h, and causes a transient immune cell surge adjacent to enteric ganglia. This is much briefer than the enteric neuropathy that ensues. Ongoing mucosal inflammatory reaction may contribute to the persistence of enteric neuropathy.

Ileal mucosal mast cell, eosinophil, and goblet cell populations during Hymenolepis diminuta infection of the rat

The population dynamics in the enteric connective tissues of eosinophils, mucosal mast cells (MMC), and in the mucosal epithelium of goblet cells were examined morphometrically in fixed ileal tissue of outbred Sprague Dawley rats during the first 32 days of infection with the tapeworm Hymenolepis diminuta. MMC and eosinophils were present in the lamina propria and submucosa; however, only eosinophils were also present in the muscularis externa. Eosinophilic infiltrate was first observed in the lamina propria at 15 days postinfection (dpi) and the numbers of eosinophils remained elevated through 32 dpi. Initial mucosal mastocytosis was detected on 6 dpi and MC numbers continued to rise over the study period without reaching a plateau. Goblet cell hyperplasia occurred only at 32 dpi. In contrast to some intestinal nematode infections where these same 3 cell types are associated with the host's expulsion responses, H. diminuta is not lost by a rapid host response in the outbred Sprague Dawley rat strain used in these experiments. We suggest that either the induction of hyperplasia of these host effector cells in ileum tissue during H. diminuta infection is not capable of triggering parasite rejection mechanisms, or the function of the induced hyperplasia is necessary for some as yet unassociated physiological or tissue architecture change in the host's intestine.

Hymenolepis diminuta: Praziquantel removal of adult tapeworms is followed by apoptotic down-regulation of mucosal mastocytosis

The rat tapeworm, Hymenolepis diminuta, induces mastocytosis, hypertrophy of enteric smooth muscle, alteration of enteric myoelectric activity, and slowed enteric transit of the rat host's intestine. This report examines the resolution of both tapeworm-induced mastocytosis and tissue changes during the period following removal of the tapeworm with Praziquantel (PZQ). The dynamics of the mucosal mast cell (MMC) population following removal of the tapeworms was assessed by histochemical identification of MMC and morphometric techniques. As a possible mechanism of MMC population regulation, MMC apoptosis was examined over the same experimental period using the in situ nick end labeling of fragmented DNA (TUNEL). Shifts in MMC numbers were correlated with functional and morphological changes of the intestine following removal of the adult-stage tapeworm. Ileal tissues from rats infected 32 days with H. diminuta (the beginning of plateau phase of tapeworm-induced chronic mastocytosis) were harvested 1, 2, 3, and 4 weeks after the PZQ treatment. Control ilea were obtained either from rats which were never infected and never treated with PZQ or from rats infected with H, diminuta for 32 days but not treated with PZQ. In order to detect MMC and apoptosis, tissue sections of ileum were doubled stained sequentially with Astra blue for MMC granules followed by a modification of the TUNEL technique. No alteration in MMC numbers were observed in PZQ-treated animals until 3 weeks after the removal of the tapeworms. The decline of MMC occurred in the mucosa and submucosa. MMC numbers first approached uninfected control levels at 4 weeks posttreatment. Coincident with the decline in mucosal MMC numbers, the rate of MMC entering apoptosis also declined. Simultaneously, ileal smooth muscle layers, hypertrophied by infection, and mucosal structures began the process of involution and atrophy. Apoptosis of MMC in the submucosa and muscularis mucosa was not detected. In conclusion, H. diminuta elicited mastocytosis and increased thickness of both mucosa and muscularis externa do not begin a decline toward control Values until 3 weeks after the parasites are gone and normal intestinal motility is restored. These data are consistent with the lack of MMC mediation of altered motility, and the decline in the rate of MMC apoptosis at 3 weeks post-PZQ suggests that apoptosis may play an important role in the involution of tapeworm-induced mastocytosis. (C) 1999 Academic Press.

High laryngeal mask airway pressures resulting from nitrous oxide do not increase pharyngeal mucosal injury in dogs

Purpose: During general anesthesia, nitrous oxide (N2O) diffuses rapidly into the air-filled laryngeal mask airway (LMA) cuff, increasing intracuff pressure. There is no clear correlation between LMA intracuff pressure and pressure on the pharynx. We have studied the effects of high LMA intracuff pressures secondary to N2O on the pharyngeal mucosa of dogs.Methods: Sixteen mongrel dogs were randomly allocated to two groups: G1 (intracuff volume, 30 mL; n = 8) breathed a mixture of O-2 (1 L.min(-1)) and air (1 L.min(-1)) and G2 (intracuff volume, 30 mL; n=8) a mixture of O-2 (1 L.min(-1)) and N2O (1 L.min(-1)). Anesthesia was induced and maintained with pentobarbitone. LMA cuff pressure was measured at zero (control), 30, 60, 90 and 120 min after #4 LMA insertion. The dogs were sacrificed, and biopsy specimens from seven predetermined areas of the pharynx in contact with the LMA cuff were collected for light (LM) and scanning electron microscopic (SEM) examination by a blinded observer.Results: LMA intracuff pressure decreased with time in G1 (P < 0.001) and increased in G2 (P < 0.001). There was a significant difference between the groups (P < 0.001). In both groups, the LM study showed a normal epithelium covering the pharyngeal mucosa and mild congestion in the subepithelial layer There were no differences between the groups (P > 0.10) or among the areas sampled (P > 0.05). In both groups, the SEM study showed a normal pharyngeal mucosa with mild superficial desquamation. Few specimens in G1 and G2 showed more intense epithelial desquamation.Conclusion: High LMA intracuff pressures produced by N2O do not increase pharyngeal mucosal injury in dogs.

Gastric mucosal perfusion in dogs: Effects of halogenated anesthetics and of hemorrhage

The gastrointestinal tract is one of the first organs affected by hypoperfusion during hemorrhagic shock. The hemodynamics and oxygen transport variables during hemorrhagic shock and resuscitation can be affected by the anesthetics used. In a model of pressure-guided hemorrhagic shock in dogs, we studied the effects of three halogenated anesthetics - halothane, sevoflurane, and isoflurane - at equipotent concentrations on gastric oxygenation. Thirty dogs were anesthetized with 1.0 minimum alveolar anesthetic concentration (MAC) of either halothane, sevoflurane, or isoflurane. A gastric tonometer was placed in the stomach to determine mucosal gastric CO2 (PgCO(2)) and for the calculation of gastric-arterial PCO2 gradient (PCO2 gap). The dogs were splenectomized and hemorrhaged to hold mean arterial pressure at 40-50 mm Hg over 45 min and then resuscitated with the shed blood volume. Hemodynamics, systemic oxygenation, and PCO2 gap were measured at baseline, after 45 min of hemorrhage, and at 15 and 60 min after blood resuscitation. Hemorrhage induced reductions of mean arterial pressure and cardiac index, while systemic oxygen extraction increased (p < .05), without significant differences among groups (p > .05). Halothane group showed significant lower PCO2 gap values than the other groups (p < .05). After 60 min of shed blood replacement, all groups restored hemodynamics, systemic oxygenation, and PCO2 gap to the prehemorrhage levels (p > .05), without significant differences among groups (p > .05). We conclude that halothane is superior to preserve the gastric mucosal perfusion in comparison to isoflurane and sevoflurane, in dogs submitted to pressure-guided hemorrhagic shock at equipotent doses of halogenated anesthetics.

Relationship among oxidative DNA damage, gastric mucosal density and the relevance of cagA, vacA and iceA genotypes of Helicobacter pylori

The aim of this study was to evaluate the relationship among oxidative DNA damage, density of Helicobacter pylori and the relevance of cagA, vacA and iceA genotypes of H. pylori. Gastric epithelial cells were isolated from 24 uninfected patients, 42 H. pylori infected patients with gastritis, and 61 patients with gastric cancer. Oxidative DNA damage was analyzed by the Comet assay, the density of H. pylori was measured by real-time polymerase chain reaction (PCR), and allelic variants of cagA, vacA and iceA were identified using the PCR. Infected patients by Helicobacter pylori cagA(+), vacAs1 m1 and iceA1 genotype showed higher levels of oxidative DNA damage than infected patients with H. pylori cagA(-), vacAs2 m2 and iceA2 genotypes and uninfected patients. Density of H. pylori did not influence oxidative DNA damage. Our results indicate that H. pylori genotype is more relevant than density for oxidative DNA damage.

Mucosal Bridge of the Vocal Fold: Difficulties in the Diagnosis and Treatment

Mucosal bridges are rare laryngeal lesions probably of genetic origin. They may cause dysphonia of varying degrees, especially when associated with other laryngeal lesions such as vocal sulci and cysts. Reports on mucosal bridges are rare, and the better treatment is inconclusive.Aim. To report the authors' experience in 14 cases of mucosal bridge showing details on endoscopic examinations and treatment.Study Design. Retrospective study.Methods. We reviewed the medical records of 14 patients with a diagnosis of mucosal bridge confirmed by video-laryngostroboscopy and direct laryngoscopy who attended the Outpatient Clinic of Voice Disorders of the Discipline of Otorhinolaryngology, Botucatu Medical School, São Paulo State University, São Paulo. Data collected included information on gender, age, symptoms, time of onset, history of intubation, smoking status, alcohol intake, associated laryngeal lesions, treatment, and GRBAS (grade of hoarseness, roughness, breathiness, asthenia, and stress) scale ratings.Results. of 14 patients, 10 were females and four were males. There was a prevalence of adults (n = 12), with only two of the patients being younger than 13 years (10 and 13 years). Mucosal bridges showed no correlations with smoking, alcohol intake, or gastroesophageal and sinonasal symptoms. Voice abuse was reported in 50% of the cases that consisted of patients who had high-voice demand occupations. In seven cases, mucosal bridges were associated with other laryngeal lesions, particularly vocal cysts and sulci. All patients who underwent surgery and phonotherapy showed improved vocal quality.Conclusions. We documented 14 patients with dysphonia caused by mucosal bridge. Promising results were obtained with surgery.

Chlorhexidine induces DNA damage in rat peripheral leukocytes and oral mucosal cells

Objective: Chlorhexidine digluconate is widely used in dental practice for decreasing plaque control, controlling gingivitis and disinfecting root canals. However, the undesirable effects of chlorhexidine digluconate regarding its genotoxicity are conflicting in the literature. Thus, the aim of this study was to investigate the genotoxicity of chlorhexidine digluconate in rat peripheral blood and oral mucosal cells by the single cell gel (comet) assay and micronucleus assay.Methods: Thirty male Wistar rats were distributed into three groups: negative control; experimental group orally treated with 0.5 ml of 0.12% chlorhexidine digluconate, twice daily, during 8 days; and positive control, which received 4-nitroquinoline 1-oxide at 0.5 g/l by drinking water.Results: A statistically significant increase of DNA damage was observed in leukocytes and oral mucosal cells of the chlorhexidine digluconate treated group, as assessed by the comet assay. However, no increase of micronucleated cells was detected in reticulocytes from peripheral blood cells.Conclusions: Taken together, the data indicate that chlorhexidine digluconate is able to induce primary DNA damage in leukocytes and in oral mucosal cells, but no chromosome breakage or loss in erythrocytes.

Mechanisms of the Gastric Antiulcerogenic Activity of Anacardium humile St. Hil on Ethanol-Induced Acute Gastric Mucosal Injury in Rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Effect of age on the gastrointestinal-associated mucosal immune response of humans

Age-related changes in gastrointestinal-associated mucosal immune response have not been well studied. Thus, we investigated the effect of age on this response and compared these responses to those of peripheral immune cells. Saliva, blood, and intestinal biopsies were collected from young and old healthy subjects to determine immunoglobulin (Ig) levels and to isolate peripheral blood mononuclear cells, intraepithelial lymphocytes (IELs), and lamina propria lymphocytes (LPLs). Although subject age did not influence the level of total IgA found in saliva, IgA levels in serum increased (p < .05) with age. Older subjects' peripheral blood mononuclear cell proliferation and IL-2 production were significantly lower than those of young subjects. LPLs from older subjects produced significantly less IL-2 in response to all stimuli than did that from the young. IEL's ability to proliferate and produce IL-2 was not affected by subject age. Thus, LPL but not IEL demonstrated an age-related decline in immune function similar to that seen in peripheral lymphocytes.

A comparison of mucosal surface area and villous histology in small intestines of the brazilian free-tailed bat (Tadarida brasiliensis) and the mouse (Mus musculus)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)