Heloísa Sobreiro Selistre de Araújo

Lysine 49 phospholipase A2 proteins.

The structures of several K49 PLA2 proteins have been determined and these differ as a group in several regions from the closely related D49 PLA2 enzymes. One outstanding difference is the presence of a high number of positively charged residues in the C-terminal region which combined with the overall high number of conserved lysine residues gives the molecule an interfacial adsorption surface which is highly positively charged compared to the opposite surface of the molecule. Although some nucleotide sequences have been reported, progress in obtaining active recombinant proteins has been slow. The K49 proteins exert several toxic activities, including myotoxicity, anticoagulation and edema formation. The most studied of these activities is myotoxicity. The myotoxicity induced by the K49 PLA2 proteins is histologically similar to that caused by the D49 PLA2 myotoxins, with some muscle fiber types possibly more sensitive than others. Whereas it is clear that the K49 PLA2 myotoxins lyse the plasma membrane of the affected muscle cell in vivo, the exact mechanism of this lysis is not known. Also, it is not known whether the toxin is internalized before, during or after the initial lysis or ever. The K49 PLA2 toxins lyse liposomes and cells in culture and in the latter, the PLA2 myotoxins exert at least two distinct mechanisms of action, neither of which is well-characterized. While the K49 PLA2 proteins are enzymatically inactive on artificial substrates, the toxins cause fatty acid production in cell cultures. Whether the fatty acid release is due to the enzymatic activity of the K49 PLA2 or stimulation of tissue lipases, is unknown. While there may be a role for fatty acid production in one mechanism of myotoxicity, a second mechanism appears to be independent of enzymatic activity. Although we are beginning to understand more about the structure of these toxins, we still know little about the precise mechanism by which they interact with the skeletal muscle cell in vivo.

Biomarker responses as indication of contaminant effects in Oreochromis niloticus.

The current study investigated oxidative stress parameters (enzymes activities, metallothionein content and lipid peroxidation) in freshwater fish, Oreochromis niloticus, tilapia exposure to Monjolinho River (in 4 months of year: January, April, July and November). One critical site in Monjolinho River (site B) was assessed in comparison to a reference site (site A). Water pH and oxygen concentration was lower than that recommended by CONAMA (Brazilian National Environmental Committee), resolution 357/2005 for protection of aquatic communities, and ammonium and the metals Cu, Zn, Mn and Fe (on all months) concentrations were higher than the maximum concentration recommended. Glutathione peroxidase (GPx) and superoxide dismutase (SOD) activities were significantly decreased in liver and muscle in tilapia from Monjolinho River, throughout the year, in relation to reference except in gills that SOD activity increased. Glutathione S-transferase (GST) activity was significantly increased in liver of the tilapia from Monjolinho River in all sites, in relation to reference except in gills that GST activity increased in July and decreased in November, suggesting that GST activity could be induced to neutralize the pollutants toxicity. On the other hand, GST activity was significantly decreased in white muscle indicating a toxic effect of pollutants, resulting in a decreased ability of tilapia to perform defense reactions associated to GSTs. The decrease of catalase (CAT) activity in gills of the O. niloticus together with the increase of SOD activity, could explain the increased lipid peroxidation (LPO) level in this organ. Metallothionein levels in liver and gills were significantly high in all sites. Results indicate that the exposure to metals caused severe damage to tissues; despite the consensually assumed antioxidant induction as a sign of exposure to contaminants the effects seem in part to be mediated by suppression of antioxidant system with SOD, CAT and GPx as potential candidates for tissues toxicity biomarkers of pollutants.

Low-Level Laser Therapy Induces Differential Expression of Osteogenic Genes During Bone Repair in Rats

OBJECTIVES The aim of this study was to measure the temporal pattern of the expression of osteogenic genes after low-level laser therapy during the process of bone healing. We used quantitative real-time polymerase chain reaction (qPCR) along with histology to assess gene expression following laser irradiation on created bone defects in tibias of rats. MATERIAL AND METHODS The animals were randomly distributed into two groups: control or laser-irradiated group. Noncritical size bone defects were surgically created at the upper third of the tibia. Laser irradiation started 24 h post-surgery and was performed for 3, 6, and 12 sessions, with an interval of 48 h. A 830 nm laser, 50 J/cm(2), 30 mW, was used. On days 7, 13, and 25 post-injury, rats were sacrificed individually by carbon dioxide asphyxia. The tibias were removed for analysis. RESULTS The histological results revealed intense new bone formation surrounded by highly vascularized connective tissue presenting slight osteogenic activity, with primary bone deposition in the group exposed to laser in the intermediary (13 days) and late stages of repair (25 days). The quantitative real-time PCR showed that laser irradiation produced an upregulation of BMP-4 at day 13 post-surgery and an upregulation of BMP4, ALP, and Runx 2 at day 25 after surgery. CONCLUSION Our results indicate that laser therapy improves bone repair in rats as depicted by differential histopathological and osteogenic genes expression, mainly at the late stages of recovery.

Low-level laser therapy (LLLT) (660 nm) alters gene expression during muscle healing in rats

INTRODUCTION The effects of LLLT were studied during muscle regeneration through gene expression. METHODS It was evaluated 10 and 50J/cm(2) doses during 7, 14 and 21days post cryoinjury, through histopathological analysis and mRNA MyoD, Myogenin, Vegf and Cox-2 expression. RESULTS Irradiated groups presented less inflammatory process than control group after 14 and 21days. Cox-2 levels were downregulated in all irradiated groups after 7, 14 and 21days. On day 7, both treated groups had a downregulation of Vegf levels, and an upregulation after 14 and 21days, mainly with 50J/cm(2). The MyoD levels were upregulated with high dose in all periods and with low dose after 21days. Myogenin expression was downregulated in both treated groups after 7days, and was upregulated with 10J/cm(2) after 21days. CONCLUSION These responses suggest that LLLT can improve the skeletal muscle regeneration through the gene expression stimulation.

Effects of low-level laser therapy on the expression of osteogenic genes related in the initial stages of bone defects in rats

Abstract. We evaluate the effects of low-level laser therapy (LLLT) on the histological modifications and temporal osteogenic genes expression during the initial phase of bone healing in a model of bone defect in rats. Sixty-four Wistar rats were divided into control and treated groups. Noncritical size bone defects were surgically created at the upper third of the tibia. Laser irradiation (Ga-Al-As laser 830 nm, 30 mW, 0.028  cm2, 1.071  W/cm2, 1 min and 34 s, 2.8 Joules, 100  J/cm2) was performed for 1, 2, 3, and 5 sessions. Histopathology revealed that treated animals presented higher inflammatory cells recruitment, especially 12 and 36 h postsurgery. Also, a better tissue organization at the site of the injury, with the presence of granulation tissue and new bone formation was observed on days three and five postsurgery in the treated animals. The quantitative real time polymerase chain reaction showed that LLLT produced a significantly increase in mRNA expression of Runx-2, 12 h and three days post-surgery, a significant upregulation of alkaline phosphatase mRNA expression after 36 h and three days post-surgery and a significant increase of osteocalcin mRNA expression after three and five days. We concluded that LLLT modulated the inflammatory process and accelerated bone repair, and this advanced repair pattern in the laser-treated groups may be related to the higher mRNA expression of genes presented by these animals.

The effects of exercise modalities on adiposity in obese rats

OBJECTIVE: The aim of the present study was to evaluate the effect of both swimming and resistance training on tumor necrosis factor-alpha and interleukin-10 expression, adipocyte area and lipid profiles in rats fed a high-fat diet. METHODS: The study was conducted over an eight-week period on Wistar adult rats, who were divided into six groups as follows (n = 10 per group): sedentary chow diet, sedentary high-fat diet, swimming plus chow diet, swimming plus high-fat diet, resistance training plus chow diet, and resistance training plus high-fat diet. Rats in the resistance training groups climbed a vertical ladder with weights on their tails once every three days. The swimming groups swam for 60 minutes/day, five days/week. RESULTS: The high-fat diet groups had higher body weights, a greater amount of adipose tissue, and higher tumor necrosis factor-alpha expression in the visceral adipose tissue. Furthermore, the high-fat diet promoted a negative change in the lipid profile. In the resistance training high-fat group, the tumor necrosis factor-alpha expression was lower than that in the swimming high-fat and sedentary high-fat groups. Moreover, smaller visceral and retroperitoneal adipocyte areas were found in the resistance training high-fat group than in the sedentary high-fat group. In the swimming high-fat group, the tumor necrosis factor-alpha expression was lower and the epididymal and retroperitoneal adipocyte areas were smaller compared with the sedentary high-fat group. CONCLUSION: The results showed that both exercise modalities improved the lipid profile, adiposity and obesity-associated inflammation in rats, suggesting their use as an alternative to control the deleterious effects of a high-fat diet in humans.

Analysis of a cDNA sequence encoding a novel member of the snake venom metalloproteinase, disintegrin-like, cysteine-rich (MDC) protein family from Agkistrodon contortrix laticinctus.

In this paper, we present a cDNA sequence encoding a full-length precursor form of a new member (ACLD) of the metalloproteinase-disintegrin-like protein family from the venom glands of Agkistrodon contortrix laticinctus (broad-banded copperhead) snake. Comparison of the deduced amino acid sequence of ACLD with those of other members of the metalloproteinase-disintegrin protein family from both mammalian and snake venom origin suggests that some conserved residues may be involved in processing of the disintegrin domain.

LOW-INTENSITY PULSED ULTRASOUND PRODUCED AN INCREASE OF OSTEOGENIC GENES EXPRESSION DURING THE PROCESS OF BONE HEALING IN RATS

The aim of this study was to measure the temporal expression of osteogenic genes during the process of bone healing in low-intensity pulsed ultrasound (LIPUS) treated bone defects by means of histopathologic and real-time polymerase chain reaction (PCR) analysis. Animals were randomly distributed into two groups (n = 30): control group (bone defect without treatment) and LIPUS treated (bone defect treated with LIPUS). On days 7, 13 and 25 postinjury, 10 rats per group were sacrificed. Rats were treated with a 30 mW/cm(2) LIPUS. The results pointed out intense new bone formation surrounded by highly vascularized connective tissue presenting a slight osteogenic activity, with primary bone deposition was observed in the group exposed to LIPUS in the intermediary (13 days) and late stages of repair (25 days) in the treated animals. In addition, quantitative real-time polymerase chain reaction (RT-qPCR) showed an upregulation of bone morphogenetic protein 4 (BMP4), osteocalcin and Runx2 genes 7 days after the surgery. In the intermediary period, there was no increase in the expression. The expression of alkaline phosphatase, BMP4 and Runx2 was significantly increased at the last period. Our results indicate that LIPUS therapy improves bone repair in rats and upregulated osteogenic genes, mainly at the late stages of recovery.

Resistance training improves body composition and increases matrix metalloproteinase 2 activity in biceps and gastrocnemius muscles of diet-induced obese rats

OBJECTIVE: We investigated the influence of resistance training on body composition and matrix metalloproteinase 2 activity in skeletal muscles of rats fed a high-fat diet. METHODS: Thirty-two Wistar rats were divided into four experimental groups (n = 8/each) according to diet and exercise status: Control (standard diet), Obese Control (high-fat diet), Resistance Training (standard diet) and Obese Resistance Training (high-fat diet) groups. Animals were fed a high-fat diet for 12 weeks to promote excessive weight gain. Resistance Training groups performed 12 weeks of training periods after this period in a vertical ladder three times/week. Fat percentage, fat-free mass and fat mass were assessed using dual-energy X-ray absorptiometry, and matrix metalloproteinase 2 activity in biceps and gastrocnemius muscles was analyzed using zymography. RESULTS: Resistance training significantly reduced body and fat masses and fat percentages in both trained groups (p<0.05). The maximal carrying load between trained groups was not different, but relative force was higher in the Resistance Training group (p<0.05). Of note, increased matrix metalloproteinase 2 activity was noted in the tested muscles of both trained groups (p<0.05). CONCLUSION: In conclusion, altered body composition and muscle matrix metalloproteinase 2 activity promoted by excessive weight gain were positively modified by resistance training.

Low-level Laser Therapy Associated With High Intensity Resistance Training on Cardiac Autonomic Control of Heart Rate and Skeletal Muscle Remodeling in Wistar Rats

Phototherapy plus dynamic exercise can enhance physical performance and improve health. The aim of our study was to evaluate the effect of low‐level laser therapy (LLLT) associated with high intensity resistance training (HIT) on cardiac autonomic and muscle metabolic responses in rats.