Supplementation with L-Glutamine and L-Alanyl-L-Glutamine Changes Biochemical Parameters and Jejunum Morphophysiology in Type 1 Diabetic Wistar Rats.

We evaluated the effects of the supplementation with L-glutamine and glutamine dipeptide (GDP) on biochemical and morphophysiological parameters in streptozotocin-diabetic rats. For this purpose, thirty animals were distributed into six groups treated orally (gavage) during thirty days: non diabetic rats (Control) + saline, diabetic + saline; Control + L-glutamine (248 mg/kg), Diabetic + L-glutamine (248 mg/kg), Control + GDP (400 mg/kg), Diabetic + GDP (400 mg/kg). Diabetes was induced by an intravenous injection of streptozotocin (60 mg/kg) and confirmed by fasting glucose ≥ 200 mg/dL. Physiological parameters, i.e., body mass, food intake, blood glucose, water intake, urine and faeces were evaluated during supplementation. After the period of supplementation, the animals were euthanized. The blood was collected for biochemical assays (fructosamine, transaminases, lipid profile, total protein, urea, ammonia). Moreover, the jejunum was excised and stored for morphophysiological assays (intestinal enzyme activity, intestinal wall morphology, crypt proliferative index, number of serotoninergic cells from the mucosa, and vipergic neurons from the submucosal tunica). The physiological parameters, protein metabolism and intestinal enzyme activity did not change with the supplementation with L-glutamine or GDP. In diabetic animals, transaminases and fructosamine improved with L-glutamine and GDP supplementations, while the lipid profile improved with L-glutamine. Furthermore, both forms of supplementation promoted changes in jejunal tunicas and wall morphometry of control and diabetic groups, but only L-glutamine promoted maintenance of serotoninergic cells and vipergic neurons populations. On the other hand, control animals showed changes that may indicate negative effects of L-glutamine. Thus, the supplementation with L-glutamine was more efficient for maintaining intestinal morphophysiology and the supplementation with GDP was more efficient to the organism as a whole. Thus, we can conclude that local differences in absorption and metabolism could explain the differences between the supplementation with L-glutamine or GDP.

Injuries in pacu embryos (Piaractus mesopotamicus) after freezing and thawing.

Although the sperm cryopreservation of freshwater and marine teleosts has been feasible for years, the cryopreservation of some fish embryos still remains elusive. Thus, the objective of this experiment was to analyze the embryo morphology after freezing and thawing 40 embryos of Piaractus mesopotamicus immersed into methanol and ethylene glycol, both at 7, 10 and 13% plus 0.1 M sucrose for 10 min. Soon after thawing, three embryos were treated with historesin, stained with hematoxylin-eosin and analyzed under an optical microscope. From every treatment, one palette containing embryos was thawed and incubated, but none of the eggs hatched. Samples containing two embryos were immersed into 10% methanol or 10% ethylene glycol both in association with sucrose, and embryos immersed into only water or sucrose solution were frozen, processed and analyzed using scanning electron microscopy (SEM). In both cases, the control group was immersed into only water. Although the embryos had the chorion, vitello, yolk syncytial layer and blastoderm, all of them were found altered under the optical microscope and by SEM. The chorion was irregular and injured; there was no individuality in the yolk granules; the yolk syncytial layer had an irregular shape, thickness and size; the blastoderm showed injuries in the nucleus shape and sometimes was absent; the blastoderm was located in atypical areas and absent in some embryos. In conclusion, no treatment was effective in preserving the embryos, and none of the embryos avoided injury from intracellular ice formation. These morphological injuries during the freezing process made the P. mesopotamicus embryos unfeasible for hatching.

Crude extract of Fusarium oxysporum induces apoptosis and structural alterations in the skin of healthy rats.

We evaluate the biological and physicochemical effects of a Fusarium oxysporum crude extract (CE) on the skin of healthy rats. The CE is topically applied and subsequently the skin is collected after 3, 6, 12, and 24 h. The samples are analyzed by Fourier transform infrared photoacoustic spectroscopy (FTIR-PAS) and histomorphometric analysis. Terminal dUTP nick end labeling (TUNEL assay) is performed to detect both the cells in apoptosis and proliferation. There is a thickening of the epidermis after 6, 12, and 24 h and dermis after 12 and 24 h of CE application. A reduction of the dermis thickness is observed at 3 and 6 h. The treated skin shows higher labeling intensity by TUNEL at 3 h, while a higher intensity by proliferating cell nuclear antigen occurs at 3 and 12 h. FTIR-PAS data support the histology observations showing an increase in the absorption peaks in the dermis after the application of the CE. F. oxysporum CE permeated through the epidermis and the dermis, reaching the subcutaneous tissue, inducing cell apoptosis, and causing physicochemical changes in the organic molecules located in the dermis. This is the first known study associating histopathological and physical chemistry changes on healthy skin after the application of F. oxysporum CE.

Administering ascorbic acid to rats undergoing ageing processes: effects on myosin-V immunoreactive myenteric neurons.

During the ageing process the enteric nervous system undergoes morphofunctional changes, such as enteric neurodegeneration. Neuronal death can be attributed to increase radicals free, and ascorbic acid (AA), known antioxidant, could minimize damage cause by oxidative stress. The objective of this study is to analyse the behaviour of morphoquantative myenteric neurons in the duodenum of adult Wistar rats with aged 90 (C90), 345 (E345) and 428 (E428) days, as well as animals of the same age who received ascorbic acid supplementation for 120 days (EA345 and EA428). Whole-mount preparations of muscle layer from the duodenum of the animals were immunostained by the method myosin V. 80 microscopic fields were quantified (14.8 mm2/animal) and measured 100 neuronal cell bodies per animal. During the aging process, there was a reduction in neuronal density in all animals groups, indicating that the effects of age were not attenuated with AA supplementation. The increase in the neuronal area of the cell bodies in 428-day-old animals proved the influence of age on this parameter. There was no observed a neuroprotective effect of AA (1 mL/g body weight) on the neuronal population myenteric myosin V immunoreactive.

Antihypernociceptive activity of anethole in experimental inflammatory pain.

Anethole has been reported to have antioxidant, antibacterial, antifungal, antiinflammatory, and anesthetic properties. In the present study, we evaluated the effects of anethole in two pain models of inflammatory origin: acute inflammation induced by carrageenan and persistent inflammation induced by Complete Freund's adjuvant. We evaluated the effects of anethole (125, 250, and 500 mg/kg) on the development of paw oedema and mechanical hypernociception. The liver was collected for histological analysis. Paw skin was collected to determine the levels of the cytokines tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), interleukin-17 (IL-17), and myeloperoxidase activity. Blood was collected to assess alanine transaminase (ALT) and aspartate transaminase (AST). The chemical composition of star anise oil was determined by gas chromatography/mass spectrometry (GC/MS), showing a presence of anethole of 98.1%. Oral pretreatment with anethole in mice inhibited paw oedema, mechanical pernociception, myelopewroxidase activity, TNF-α, IL-1ß and IL-17 levels in acute and persistent inflammation models. Additionally, anethole treatment did not alter prostaglandin E2-induced mechanical hypernociception. Possible side effects were also examined. Seven-day anethole treatment did not alter plasma AST and ALT levels, and the histological profile of liver tissue was normal. The present study provides evidence of the antiinflammatory and analgesic activities of anethole in acute and persistent inflammation models.

Characterization of the myenteric neuronal population and subpopulation of the duodenum of adult wistar rat fed with hypoproteic chow.

The effects of severe protein malnutrition (4%) on myenteric neurons of Wistar rat duodenum, in relation to a standard 22%-protein diet for rodents, were assessed in this study. Segments of the duodenum from 10 rats from each nutritional group were submitted to the elaboration of whole mounts - 5 stained with Giemsa to determine the total population of myenteric neurons and the others stained by a histochemical method to detect nervous cells through the NADPH-diaphorase enzyme activity for studying the subpopulation of nitrergic neurons. The area of 100 neurons per animal, totalizing 2,000 neurons, were randomly measured by using the Image Pro-Plus(®)software. Malnourished rats presented 34.38% lower body weight and 10.60% duodenum length reduction when compared to the control group. Quantitative analysis demonstrated no significant differences between control and malnourished group by using Giemsa; however, as the organ reduction was not followed by an increase inversely proportional to the density of neurons, the condition imposed suggests the loss of neurons from the total population. Nevertheless, through NADPH-d histochemistry, there was a neuronal density increase for the malnourished group. There was no significant difference between the groups for both techniques with respect to the morphometric analysis of the body cell.