Concurrent training remodels the subcutaneous adipose tissue extracellular matrix of people living with HIV: a non-randomized clinical trial.

Evaluate the effect of 12 wks of concurrent training (CT) in the extracellular matrix (ECM) of subcutaneous adipose tissue (SAT) in people living with HIV/AIDS (PLWHA). In the non-randomized clinical trial, 19 participants, 11 healthy (HIV-) and 18 PLWHA under the use of highly active antiretroviral therapy (HAART) for at least 1 year (HIV+). All participants engaged in a moderate-intensity CT program for 12 weeks, 3 times a week. Before and after CT, aerobic and strength performance were assessed, as well as anthropometric and biochemical blood profiles. In addition, SAT biopsies were performed for histologic and morphometric analyses. Statistical analysis was carried out with R Studio, using descriptive and inferential analysis, ANOVA test, and mixed-effect model (P < 0.05). HIV+ showed higher levels of very-low-density lipoproteins and triglycerides and lower levels of high-density lipoproteins at baseline than HIV- (P < 0.05). All groups showed improved aerobic and strength performances (P < 0.05). Both groups showed reduced adipocyte sizes after CT (P < 0.05). Lastly, HIV+ presented smaller adipocytes and higher elastic fiber deposition at baseline and decreased after training only in HIV+, similar to the HIV group. Thus, CT in PLWHA promoted a decrease in the size heterogeneity of adipocytes and elastic fiber deposition, remodeling the ECM, and improving the SAT fibrosis profile. Brazilian Clinical Trials Registry (ensaiosclinicos.gov.br - UTN: U1111-1214-3022). Novelty: Adipose tissue fibrosis is improved by training in people living with HIV. Concurrent training remodels adipose tissue extracellular matrix.

Chemical Interaction and Interface Analysis of Self-Etch Adhesives Containing 10-MDP and Methacrylamide With the Dentin in Noncarious Cervical Lesions.

OBJECTIVES: To characterize the chemical interactions and analyze the interface of adhesive systems containing 10-methacryloyloxydecyl dihydrogen phosphate (10-MDP) and N-methacryloyl glycine (methacrylamide) functional monomers with the dentin in noncarious cervical lesions (NCCLs) compared with artificial defects (ADs). METHODS AND MATERIALS: Twenty human teeth with natural NCCLs on the buccal surface were used. Class V cavities, similar to NCCLs, were created on the lingual surface to serve as controls. Teeth were randomly allocated to two groups according to the functional monomer in the adhesive (N=10): G1, 10-MDP; and G2, methacrylamide. NCCLs and ADs were characterized by their mineral composition (MC) and degree of demineralization (DD) using micro-Raman spectroscopy, adhesive/dentin chemical interactions (CIs) were assessed with infrared photoacoustic spectroscopy, and interface morphology was evaluated with scanning electron and light microscopy. MC, CI, and DD data were submitted to Shapiro-Wilk and Student t-tests ( p<0.05). RESULTS: Compared with ADs, dentin in NCCLs was hypermineralized ( p<0.05). In G1, CI, and DD in the first 2 µm, and adhesive projections in NCCLs and ADs interfaces were similar. Additionally, a thin layer of dentin collagen was observed in ADs, while it was hardly present in NCCLs. In G2, although CI could not be identified, changes in the mineral components were observed. The DD in the ADs and NCCLs were statistically similar, while SEM showed a lack of adhesion at NCCLs interface. DD and collagen exposure in the ADs and NCCLs were more pronounced than in G1. CONCLUSIONS: Results suggest that the G1 adhesive could be applied directly on the superficial sclerotic layer in NCCLs. In contrast, previous cavity preparation should be conducted to improve the micromechanical interaction of G2 with the dentin.

The toxin BjussuLAAO-II induces oxidative stress and DNA damage, upregulates the inflammatory cytokine genes TNF and IL6, and downregulates the apoptotic-related genes BAX, BCL2 and RELA in human Caco-2 cells.

Colorectal carcinoma is one of the most common cancers in adults. As chemotherapy, the first-choice treatment for colorectal carcinoma, is often infeasible due to acquired tumor resistance and several adverse effects, it is important to discover and explore new molecules with better therapeutic action. Snake venom toxins have shown promising results with high cytotoxicity against tumor cells, but their mechanisms of action remain unclear. Here we examined how BjussuLAAO-II, an L-amino acid oxidase isolated from Bothrops jararacussu snake venom, exerts cytotoxicity towards colorectal adenocarcinoma human cells (Caco-2) and human umbilical vein endothelial cell line (HUVEC). A 24-h treatment with BjussuLAAO-II at 0.25 - 5.00 µg/mL diminished cell viability by decreasing (i) mitochondrial activity, assessed by reduction of 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide and resazurin; (ii) the activity of acid phosphatases; and (iii) lysosomal function, assessed by neutral red uptake. BjussuLAAO-II also increased intracellular levels of reactive oxygen species and DNA damage, as assessed by fluorescence and the comet assay, respectively. BjussuLAAO-II altered the expression of cell proliferation-related genes, as determined by RT-qPCR: it elevated the expression of the inflammatory cytokine genes TNF and IL6, and lowered the expression of the apoptotic-related genes BAX, BCL2, and RELA. Therefore, BjussuLAAO-II induces Caco-2 cells death by acting on multiple intracellular targets, providing important data for further studies to assess whether these effects are seen in both tumor and normal cells, with the aim of selecting this drug for possible therapeutic purposes in the future.

Chemopreventive effect and lack of genotoxicity and mutagenicity of the exopolysaccharide botryosphaeran on human lymphocytes.

Carbohydrate biopolymers of fungal-origin are an important natural resource in the search for new bioagents with therapeutic and nutraceutical potential. In this study the mutagenic, genotoxic, antigenotoxic and antioxidant properties of the fungal exopolysaccharide botryosphaeran, a (1→3)(1→6)-ß-D-glucan, from Botryosphaeria rhodina MAMB-05, was evaluated. The mutagenicity was assessed at five concentrations in Salmonella typhimurium by the Ames test. Normal and tumor (Jurkat cells) human T lymphocyte cultures were used to evaluate the genotoxicity and antigenotoxicity (Comet assay) of botryosphaeran alone and in combination with the mutagen methyl methanesulfonate (MMS). The ability of botryosphaeran to reduce the production of reactive oxygen and nitrogen species (RONS) generated by hydrogen peroxide was assessed using the CM-H2DCFDA probe in lymphocyte cultures under different treatment times. None of the evaluated botryosphaeran concentrations were mutagenic in bacteria, nor induced genotoxicity in normal and tumor lymphocytes. Botryosphaeran protected lymphocyte DNA against damage caused by MMS under simultaneous treatment and post-treatment conditions. However, botryosphaeran was not able to reduce the RONS generated by H2O2. Besides the absence of genotoxicity, botryosphaeran exerted a protective effect on human lymphocytes against genotoxic damage caused by MMS. These results are important in the validation of botryosphaeran as a therapeutic agent targeting health promotion.

Creatine supplementation in trained rats causes changes in myenteric neurons and intestinal wall morphometry

Creatine is widely used by athletes as an ergogenic resource. The aim of this study was to evaluate the influence of creatine supplementation on the duodenum of rats submitted to physical training. The number and myenteric neuronal cell bodies as well mucosal and muscular tunic morphometry were evaluated. Control animals received a standard chow for 8 weeks, and the treated ones received the standard chow for 4 weeks and were later fed with the same chow but added with 2% creatine. Animals were divided in groups: sedentary, sedentary supplemented with creatine, trained and trained supplemented with creatine. The training consisted in treadmill running for 8 weeks. Duodenal samples were either processed for whole mount preparations or for paraffin embedding and hematoxylin-eosin staining for histological and morpho metric studies of the mucosa, the muscular tunic and myenteric neurons. It was observed that neither creatine nor physical training alone promoted alterations in muscular tunic thickness, villus height or crypts depth, however, a reduction in these parameters was observed when both were associated. The number of myenteric neurons was unchanged, but the neuronal cell body area was reduced in trained animals but not when training and creatine was associated, suggesting a neuroprotector role of this substance.

Densitometry, radiography, and histological assessment of collagen as methods to evaluate femoral bones in an experimental model of osteoporosis.

SUMMARY: All methods to detect experimental loss of bone present technique limitations. The sensitivities of image and histological analyses to detect the effects of teriparatide in rats with bone loss after ovariectomy were evaluated. All methods were qualitatively valid. INTRODUCTION: The standardization of methods to assess bone loss after ovariectomy is crucial to establish the degree of experimental osteoporosis. In general, methods per image or histological techniques are used. To validate these two ways to determine the degree of bone loss in ovariectomized rats, we evaluated the sensitivities of bone densitometry, conventional radiography, and histological analysis of the area occupied by collagen, detecting the effects of teriparatide treatment in the femur of ovariectomized rats with bone loss. METHODS: Wistar rats were divided into three groups: a control group, in which the animals were only subjected to laparotomy; an ovariectomized group, in which bilateral removal of the ovaries was performed; and an ovariectomized + teriparatide group, in which bilateral removal of the ovaries was performed, and the animals were treated with 3 µg/100 g/day of teriparatide. Three months following the ovariectomy, bone densitometry, radiographic densitometry, and histological analysis of the area occupied by collagen fibers were carried out in the femur diaphysis. RESULTS: The bone densitometry revealed 11.2% reduction in femur density; in the conventional radiography, the loss of bone mass was 14.5%, and with the histological analysis, a 40.9% reduction in the area occupied by collagen was detected in the femur diaphysis. CONCLUSIONS: In conclusion, histological analysis could not be quantitatively compared with the methods of bone densitometry and conventional radiography; however, all of these methods were qualitatively valid for assessing the loss of bone stemming from ovariectomy and the therapeutic effect of teriparatide in the prevention of osteoporosis.

Toxic thermoresistant metabolites of Fusarium oxysporum are capable of inducing histopathological alterations in Wistar rats

The genus Fusarium is known to produce mycotoxins that cause fusariosis in plants, animals and humans. Mycotoxins are among the virulence factors of this genus. Metabolic extracts of Fusarium oxysporum, isolated from a patient with onychomycosis and sterilized by filtration or autoclave, were inoculated intradermally into Wistar rats at concentrations of 0.25, 0.5 and 1 µg/µL, and the effects on their tegument were observed at 24 and 72 hours. After histological procedures and staining by hematoxylineosin, the sections were studied for their inflammatory-reaction intensity and for evidence of injury and tissue distortion. Inflammatory reactions in the dermis and the subcutaneous tissue were observed at all concentrations of the inoculated extract tested. There was a significant influx of neutrophils, mastocytes and lymphocytes, as well as a large quantity of macrophages. Apoptotic bodies and hyperemic blood vessels were observed. This reaction was directly related to the extract concentration, and was most intense in animals that received the 1 mg/µL dose. The maximum peak was observed at 24 hours. The autoclaved metabolic extract produced the same effects as the untreated one, indicating the presence of heat-resistant metabolites. In conclusion, the metabolic extracts obtained from sterilized culture filtrates of F. oxysporum are capable of inducing an inflammatory response within 24 hours in the dermis and subcutaneous tissue of rats.(AU)

Dynamics of reepithelialisation and penetration rate of a bee propolis formulation during cutaneous wounds healing.

The aim of this work is to investigate the dynamics of reepithelialisation and the penetration rate of a propolis ointment formulation during cutaneous wounds healing. The experiments were performed as a function of the treatment time in a well controlled group of rats. We observed that the propolis ointment influenced the healing process stimulating keratinocytes cell proliferation as compared to the control group. It was shown that the propagation of the bee propolis was dependent on the wound healing stages. In addition, the photoacoustic spectroscopy showed that the applied substances reached the deep wound region, highlighting once again the particular characteristic of this technique to evaluate the penetration rate of substances through the skin.

Ex vivo evaluation of the percutaneous penetration of proanthocyanidin extracts from Guazuma ulmifolia using photoacoustic spectroscopy.

In this work photoacoustic spectroscopy has been applied to determine ex vivo the percutaneous penetration of proanthocyanidins present in extracts obtained from Guazuma ulmifolia, in rats. Lotion formulations containing 0.0663 mg of procyanidin B2 day(-1)animal(-1) were topically applied during 7, 10 and 13 days in each group of the animals. After the end of treatment the animals were killed, the skin dissected to remove the basal content, and the measurements were carried out as a function of the period of time of treatment. The results showed that despite the very low concentration of the active principle (procyanidin B2) in the lotion, the photoacoustic method was able to show the presence of optical absorption bands from this substance in the dermis region, evidencing once again that this method may be useful for studies of topically applied formulations of interest in the pharmacokinetic area.

Ileal VIP submucous neurons: confocal study of the area enlargement induced by myenteric denervation in weanling rats.

Vasoactive Intestinal Peptide (VIP) neurons are maturing during suckling and weaning periods and the neuropeptide VIP is thought to be neurotrophic during ontogenesis. We have previously demonstrated that suckling rats with myenteric ablation have significantly higher mitotic index and an increase on villus height and crypt depth 15 days after treatment. In the current study, we measured the area of VIP neurons of submucous plexus in the ileum of weanling rats, in which myenteric neurons were ablated by serosal application of benzalkonium chloride (BAC). The area of VIP immunoreactive cell bodies, reconstructed under confocal microscope, was significantly increased in response to denervation. This result suggests that the myenteric plexus may have an inhibitory role over submucous plexus in the normal intestine. The enhanced production of VIP may be correlated with the increased epithelial proliferation induced by denervation in a critical period of life, from suckling to weaning time.

Goblet cell number in the ileum of rats denervated during suckling and weaning

The enteric nervous system plays a role on the stimulation of secretory cells of intestinal epithelia. We have demonstrated that ablation of ENS stimulates epithelial cell proliferation. As goblet cells are important constituents of the epithelial sheet, it is mandatory to investigate separately this cell type. The myenteric plexus of the ileum of rats in postnatal development was partially removed by the serosal application of benzalkonium chloride (BAC). Three groups of animals were used: those where BAC application was at 13 days and sacrifice was at 15 (13/28-day-old) or 23 days after treatment (13/36-day-old), and those where BAC was applied at 21 days and rats were killed 15 days after treatment (21/36-day-old) . The number of goblet cells in the ileum was estimated in sections stained by periodic acid-Schiff (PAS) histochemistry. In the 13/28 and 21/36 groups, the number of goblet cells was significantly higher after BAC treatment. These results suggest that the myenteric denervation may have an acute effect on the number of goblet cell in suckling and weanling rats, probably through submucous plexus.

Streptozotocin-induced diabetes duration is important to determine changes in the number and basophily of myenteric neurons.

The aim of present study was to evaluate the number and basophily of cell bodies of myenteric neurons in the ileum of rats with diabetes mellitus induced by streptozotocin. Four groups of rats were used: diabetes was induced in two (D) whereas the other two worked as controls (N). Animals were sacrificed six (6N, 6D) or nineteen (19N, 19D) weeks after diabetes induction. A segment of the terminal portion of the ileum of each rat was obtained and stained with Giemsa's solution, for whole-mount preparation studies. Forty fields were analyzed in each animal, and the number and basophily intensity of cell bodies were recorded. After counting, the following mean numbers of neurons/mm2 were obtained: 6N=593.1 +/- 95.75, 6D=639.1 +/- 130.8, 19N=580.1 +/- 175.6 and 19D=402.0 +/- 144.8. The analysis of basophily shown that highest frequency of neurons with weak/intermediary basophily was verified in 6D group (55.3%), whereas the groups 6N, 19N e 19D presented 38%, 36% e 40% respectively. The statistical analysis showed that a long period is necessary to decrease the number of neurons/mm2 in the rat ileum after diabetes induction, and that there was a reduction in basophily intensity in diabetic rats after 6 weeks of treatment, and such cells do not recover after a longer period (19 weeks).

Effect of myenteric denervation on intestinal epithelium proliferation and migration of suckling and weanling rats.

The effects of myenteric denervation on the cell kinetics of the intestinal epithelium of suckling and weanling rats were investigated. The myenteric plexus of an ileal segment was partially ablated by serosal application of benzalkonium chloride (BAC) in three groups of rats: those that underwent surgery at 13 days and were killed 15 (13/28-day-old) or 23 (13/36-day-old) days after treatment, and those that were operated at 21 days (21/36-day-old) and were killed 15 days after treatment. The extent of denervation was assessed in whole-mount preparations. The cell bodies of myenteric neurones were stained by NADH-diaphorase histochemical technique. Cell proliferation was estimated by the mitotic index (MI) and morphometric analysis of villus and crypt lengths using an image analysis system. Thickness of the muscle layers was also assessed by morphometry. Cell migration on the villi was estimated by the position of the leading labelled cell 24 h after tritiated thymidine injection. The number of neurones was reduced by around 80% in rats operated at 13 days, and reduced by 98% in those operated at 21 days. The thickness of the muscle layers was increased in all groups of treated animals. MI was significantly higher 15 days after BAC-treatment in the 13/28 group. Morphological changes in the intestinal mucosa were observed 15 days after BAC-treatment, when there was an increase in villus height (13/28 group) and crypt depth (13/28 and 21/36 groups). Cell migration rate was accelerated in the 21/36 group. No differences where found in the 13/36 group. These results show the strong effect of myenteric ablation on cell proliferation and migration in the ileal epithelium in the first 15 days of treatment in suckling and in weanling rats, and the subsequent recovery of intestinal mucosa homeostasis later on.