In vitro toxicity of isolated strains and cyanobacterial bloom biomasses over Paramecium caudatum (ciliophora): Lessons from a non-metazoan model organism.

Cyanobacteria have been considered a major global threat because of their widespread ability to proliferate and contaminate inland and marine waters with toxic metabolites. For this reason, to avoid risks to humans and environmental health, regulatory legislation and guidelines have been established based on extensive toxicological data. However, most of what is known in this field come from works on microcystin (MC) variants, which effects were almost exclusively tested in metazoan models. In this work, we used acute end-point toxicological assays and high-resolution hybrid quadrupole time-of-flight mass spectrometer coupled with electrospray ionization source (ESI-Q-TOF-MS) analyses to evaluate the deleterious impact of aqueous extracts prepared from cultures of cyanobacteria and environmental bloom biomasses over a non-metazoan model organism, the cosmopolitan fresh/brackish water unicellular microeukaryote, Paramecium caudatum (Ciliophora). Our data suggest that all extracts produced time-dependent effects on P. caudatum survival, irrespective of their metabolite profile; and that this ciliate is more sensitive to extracts containing microginins than to extracts with only MCs, stressing that more toxicological investigations should be performed on the environmental impact of neglected cyanotoxins. Further, our data provide evidence that P. caudatum may be more sensitive to cyanotoxins than vertebrates, indicating that guidelines values, set on metazoans are likely to be inaccurate to protect organisms from basal food web positions. Thus, we highly recommend the widespread use of microeukaryotes, such as ciliates in environmental risk assessment frameworks for the establishment of more reliable cyanotoxin monitoring guideline values.

Bioglass/PLGA associated to photobiomodulation: effects on the healing process in an experimental model of calvarial bone defect.

Bioactive glasses (BG) are known for their ability to bond to bone tissue. However, in critical situations, even the osteogenic properties of BG may be not enough to induce bone consolidation. Thus, the enrichment of BG with polymers such as Poly (D, L-lactic-co-glycolic) acid (PLGA) and associated to photobiomodulation (PBM) may be a promising strategy to promote bone tissue healing. The aim of the present study was to investigate the in vivo performance of PLGA supplemented BG, associated to PBM therapy, using an experimental model of cranial bone defect in rats. Rats were distributed in 4 different groups (Bioglass, Bioglass/PBM, Bioglas/PLGA and BG/PLGA/PBM). After the surgical procedure to induce cranial bone defects, the pre-set samples were implanted and PBM treatment (low-level laser therapy) started (808 nm, 100 mW, 30 J/cm2). After 2 and 6 weeks, animals were euthanized, and the samples were retrieved for the histopathological, histomorphometric, picrosirius red staining and immunohistochemistry analysis. At 2 weeks post-surgery, it was observed granulation tissue and areas of newly formed bone in all experimental groups. At 6 weeks post-surgery, BG/PLGA (with or without PBM) more mature tissue around the biomaterial particles. Furthermore, there was a higher deposition of collagen for BG/PLGA in comparison with BG/PLGA/PBM, at second time-point. Histomorphometric analysis demonstrated higher values of BM.V/TV for BG compared to BG/PLGA (2 weeks post-surgery) and N.Ob/T.Ar for BG/PLGA compared to BG and BG/PBM (6 weeks post-surgery). This current study concluded that the use of BG/PLGA composites, associated or not to PBM, is a promising strategy for bone tissue engineering.

Neurotoxic effects of sublethal concentrations of cyanobacterial extract containing anatoxin-a(s) on Nauphoeta cinerea cockroaches.

The detection of cyanotoxins, such as the anatoxin-a(s), is essential to ensure the biological safety of water environments. Here, we propose the use of Nauphoeta cinerea cockroaches as an alternative biological model for the biomonitoring of the activity of anatoxin-a(s) in aquatic systems. In order to validate our proposed model, we compared the effects of a cyanobacterial extract containing anatoxin-a(s) (CECA) with those of the organophosphate trichlorfon (Tn) on biochemical and physiological parameters of the nervous system of Nauphoeta cinerea cockroaches. In brain homogenates from cockroaches, CECA (5 and 50 µg/g) inhibited acetylcholinesterase (AChE) activity by 53 ±â€¯2% and 51 ±â€¯7%, respectively, while Tn (5 and 50 µg/g) inhibited AChE activity by 35 ±â€¯4% and 80 ±â€¯9%, respectively (p < 0.05; n = 6). Moreover, CECA at concentrations of 5, 25, and 50 µg/g decreased the locomotor activity of the cockroaches, diminishing the distance travelled and increasing the frequency and duration of immobile episodes similarly to Tn (0.3 µg/g) (p < 0.05, n = 40, respectively). CECA (5, 25 and 50 µg/g) induced an increase in the leg grooming behavior, but not in the movement of antennae, similarly to the effect of Tn (0.3 µg/g). In addition, both CECA (50 µg/200 µl) and Tn (0.3 µg/200 µl) induced a negative chronotropism in the insect heart (37 ±â€¯1 and 47 ±â€¯8 beats/min in 30 min, respectively) (n = 9, p > 0.05). Finally, CECA (50 µg/g), Tn (0.3 µg/g) and neostigmine (50 µg/g) caused significant neuromuscular failure, as indicated by the monitoring of the in vivo neuromuscular function of the cockroaches, during 100 min (n = 6, p < 0.05, respectively). In conclusion, sublethal doses of CECA provoked entomotoxicity. The Tn-like effects of CECA on Nauphoeta cinerea cockroaches encompass both the central and peripheral nervous systems in our insect model. The inhibitory activity of CECA on AChE boosts a cascade of signaling events involving octopaminergic/dopaminergic neurotransmission. Therefore, this study indicates that this insect model could potentially be used as a powerful, practical, and inexpensive tool to understand the impacts of eutrophication and for orientating decontamination processes.

Interleukin-10 and collagen type II immunoexpression are modulated by photobiomodulation associated to aerobic and aquatic exercises in an experimental model of osteoarthritis.

The aim of this study was to compare the effects of photobiomodulation (PBM) associated with an aerobic and an aquatic exercise training on the degenerative process related to osteoarthritis (OA) in the articular cartilage in rats. Fifty male Wistar rats were randomly divided into 5 groups: OA control group (CG), OA plus aerobic training group (AET), OA plus aquatic training group (AQT), OA plus aerobic training associated with PBM group (AETL), OA plus aquatic training associated with PBM group (AQTL). The aerobic training (treadmill; 16 m/min; 50 min/day) and the aquatic training (water jumping; 50-80% of their body mass) started 4 weeks after the surgery and they were performed 3 days/week for 8 weeks. Moreover, PBM was performed after the physical exercise trainings on the left joint. Morphological characteristics and immunoexpression of IL-10, TGF-ß, and collagen type I (Col I) and II (Col II) of the articular cartilage were evaluated. The results showed that all the treated groups (exercise and PBM) presented less intense signs of degradation (measured by histopathological analysis and OARSI grade system). Additionally, aerobic and aquatic exercise training rats (associated or not with PBM) showed increased IL-10 (AET p = 0.0452; AETL p = 0.03; AQTL p = 0.0193) and Col II (AET p = 0.012; AQT p = 0.0437; AETL p = 0.0001; AQTL p = 0.0001) protein expression compared to CG. Furthermore, a statistically higher TGF-ß expression was observed in AET (p = 0.0084) and AETL (p = 0.0076) compared to CG. These results suggest that PBM associated with aerobic and aquatic exercise training were effective in mediating chondroprotective effects and maintaining the integrity of the articular tissue in the knees of OA rats.

The cytotoxic activity of miltefosine against Leishmania and macrophages is associated with dynamic changes in plasma membrane proteins.

In this study, we combined electron paramagnetic resonance (EPR) spectroscopy with an analysis of biophysical cellular parameters to study the mechanisms underlying the in vitro anti-leishmanial activity of miltefosine (MT). A thiol-specific spin label attached to membrane-bound proteins of Leishmania amazonensis and peritoneal macrophages indicated that MT may bind to plasma membrane proteins in large quantities via a detergent-like action and cause structural changes associated with a marked increase in dynamics and exposure to an aqueous environment. EPR spectra of a spin-labeled stearic acid indicated strong interactions between the probe and membrane proteins and a marked increase in the membrane fluidity of MT-treated cells. The cytotoxicity of MT was found to depend on the cell concentration used in the assay. This dependence was described by an equation involving the 50% inhibitory concentrations of MT in the aqueous medium (cw50) and the cell membrane (cm50) and the membrane-aqueous medium partition coefficient of MT (K). With a cw50 of 8.7µM, macrophages were less sensitive to MT than amastigotes and promastigotes of Leishmania, which had cw50 values of 2.4-3.1µM. The estimated cm50 of MT for Leishmania was 1.8M, which appears sufficient to cause ruptures or formation of pores in the plasma membrane. Additionally, we demonstrated that the changes in the plasma membrane detected by EPR spectroscopy occurred at cytotoxic concentrations of MT, as assessed through in vitro assays.

Photobiomodulation and bone healing in diabetic rats: evaluation of bone response using a tibial defect experimental model.

Diabetes mellitus (DM) leads to a delay in bone healing. Thus, some therapeutic approaches have been used to accelerate the process of bone repair such as photobiomodulation (PBM). Therefore, the present study aimed to evaluate the effects of PBM, in different fluences, in bone repair in an experimental model of tibial bone defects in diabetic rats. Sixty-four Wistar rats were submitted to a surgical procedure to perform bone defect and distributed in four groups: diabetic control group (DCG), diabetic laser group 30 J/cm(2) (L30), diabetic laser group 60 J/cm(2) (L60), and diabetic laser group 120 J/cm(2) (L120). A 808 nm Ga-Al-As (DMC Equipment, São Carlos, SP, Brazil) laser, 100 mW; 0.028 cm(2); 3.57 W/cm(2); 30, 60, and 120 J/cm(2); 0.84, 1.68, and 3.36 J; 8, 16, and 33 s was used. Animals were euthanized 15 and 30 days after the surgery. Histological, morphometric, immunohistochemistry, and biomechanical analyses were performed. In the histological and morphometric evaluation, all laser-treated groups showed a better histological pattern and a higher amount of newly formed bone compared to DCG. An intense RUNX2 immunoexpression was observed in the laser-treated groups, 15 days after the surgery. Receptor activator of nuclear factor κ-ß ligand (RANK-L) immunohistochemistry analysis showed a significant decrease in the immunoreactivity for L30 and L120, 30 days after surgery. There was no statistical difference in the biomechanical analysis among the groups. In conclusion, PBM, in all fluences used, showed an osteogenic potential in bone healing of diabetic rats.

Miltefosine increases lipid and protein dynamics in Leishmania amazonensis membranes at concentrations similar to those needed for cytotoxicity activity.

Miltefosine (MT) is a membrane-active alkylphospholipid licensed for the topical treatment of breast cancer skin metastases and the oral treatment of leishmaniasis, although its mechanism of action remains unclear. Electron paramagnetic resonance (EPR) spectroscopy of a spin-labeled lipid and a thiol-specific spin label in the plasma membrane of Leishmania promastigotes showed that MT causes dramatic increases in membrane dynamics. Although these alterations can be detected using a spin-labeled lipid, our experimental results indicated that MT interacts predominantly with the protein component of the membrane. Cell lysis was also detected by analyzing the supernatants of centrifuged samples for the presence of spin-labeled membrane fragments and cytoplasmic proteins. Using a method for the rapid incorporation of MT into the membrane, these effects were measured immediately after treatment under the same range of MT concentrations that cause cell growth inhibition. Cytotoxicity, estimated via microscopic counting of living and dead cells, indicated ∼70% cell death at the concentration of MT at which EPR spectroscopy detected a significant change in membrane dynamics. After this initial impact on the number of viable parasites, the processes of cell death and growth continued during the first 4 h of incubation. The EPR spectra of spin-labeled membrane-bound proteins were consistent with more expanded and solvent-exposed protein conformations, suggesting a detergent-like action. Thus, MT may form micelle-like structures around polypeptide chains, and proteins with a higher hydrophobicity may induce the penetration of hydrophilic groups of MT into the membrane, causing its rupture.

Low-level laser therapy enhances the expression of osteogenic factors during bone repair in rats.

The aim of this study was to evaluate the effects of low-level laser therapy (LLLT) on bone formation, immunoexpression of osteogenic factors, and biomechanical properties in a tibial bone defect model in rats. Sixty male Wistar rats were distributed into bone defect control group (CG) and laser irradiated group (LG). Animals were euthanized on days 15, 30, and 45 post-injury. The histological and morphometric analysis showed that the treated animals presented no inflammatory infiltrate and a better tissue organization at 15 and 30 days postsurgery. Also, a higher amount of newly formed bone was observed at 15 days postsurgery. No statistically significant difference was observed in cyclooxygenase-2 immunoexpression among the groups at 15, 30, and 45 days in the immunohistochemical analysis. Considering RUNX-2, the immunoexpression was statistically higher in the LG compared to the CG at 45 days. BMP-9 immunoexpression was significantly higher in the LG in comparison to CG at day 30. However, there was no expressivity for this immunomarker, both in the CG and LG, at the day 45 postsurgery. No statistically significant difference was observed in the receptor activator of nuclear factor kappa-B ligand immunoexpression among the groups in all periods evaluated. No statistically significant difference among the groups was observed in the maximal load in any period of time. Our findings indicate that laser therapy improved bone healing by accelerating the development of newly formed bone and activating the osteogenic factors on tibial defects, but the biomechanical properties in LG were not improved.

The effects of low-level laser irradiation on bone tissue in diabetic rats.

Diabetes mellitus (DM) leads to a decrease in bone mass and increase the risk of osteoporosis and in this context, many treatments have shown to accelerate bone metabolism. It seems that low-level laser therapy (LLLT) is able of stimulating osteoblast activity and produced increased biomechanical properties. However, its effects on bone in diabetic rats are not fully elucidated. The aim of this study was to evaluate the effects of LLLT on bone formation, immunoexpression of osteogenic factors, biomechanical properties and densitometric parameters in diabetic rats. Thirty male Wistar rats were randomly distributed into three experimental groups: control group, diabetic group, and laser-treated diabetic group. DM was induced by streptozotocin (STZ) and after 1 week laser treatment started. An 830-nm laser was used, performed for 18 sessions, during 6 weeks. At the end of the experiment, animals were euthanized and tibias and femurs were defleshed for analysis. Extensive resorptive areas as a result of osteoclasts activity were noticed in DG when compared to control. Laser-treated animals showed an increased cortical area. The immunohistochemical analysis revealed that LLLT produced an increased RUNX-2 expression compared to other groups. Similar RANK-L immunoexpression was observed for all experimental groups. In addition, laser irradiation produced a statistically increase in fracture force, bone mineral content (BMC) and bone mineral density compared to DG. The results of this study indicate that the STZ model was efficient in inducing DM 1 and producing a decrease in cortical diameter, biomechanical properties and in densitometric variables. In addition, it seems that LLLT stimulated bone metabolism, decreased resorptive areas, increased RUNX-2 expression, cortical area, fracture force, BMD, and BMC. Further studies should be developed to provide additional information concerning the mechanisms of action of laser therapy in diabetic bone in experimental and clinical trials.

The effects of 780-nm low-level laser therapy on muscle healing process after cryolesion.

The objective of this study was to assess the effects of 780-nm low-level laser therapy at different periods of 7, 14 and 21 days after cryolesion, including the dose (10 or 50 J/cm(2)), to promote a better muscle repair evidenced by histopathological and immunohistochemical analyses. Fifty-four male rats were divided into three groups: injured control group (CG)-injured animals without any treatment; injured 780-nm laser-treated group, at 10 J/cm(2) (G10); and injured 780-nm laser-treated group, at 50 J/cm(2) (G50). Each group was divided into three subgroups (n = 6): 7, 14 and 21 days post-injury. Histopathological findings revealed better organised muscle fibres in the G10 and G50 during the periods of 7 and 14 days compared to the CG. The G10 and G50 during the 7 days showed a significant reduction (p < 0.05) of lesion area compared to the CG, without differences between groups treated for 14 and 21 days. The G10 showed an increase of the amount of vessels after 14 days compared to the G50, but not in relation to controls. With regard to the immunohistochemical analyses of the MyoD factor, the G10 and G50 during the 7 days showed higher concentrations of immunomarkers than controls. Myogenin immunomarkers were similarly observed at days 7 and 14 in all the three groups analysed, whereas immunomarkers were found in none of the groups after 21 days of laser therapy. The results showed that laser, regardless the applied dose, has positive effects on muscle repair.

Low-level laser therapy prevents degenerative morphological changes in an experimental model of anterior cruciate ligament transection in rats.

The aim of this study was to analyze the effects of low-level laser therapy (LLLT) on the prevention of cartilage damage after the anterior cruciate ligament transection (ACLT) in knees of rats. Thirty male rats (Wistar) were distributed into three groups (n = 10 each): injured control group (CG); injured laser-treated group at 10 J/cm(2) (L10), and injured laser-treated group at 50 J/cm(2) (L50). Laser treatment started immediately after the surgery and it was performed for 15 sessions. An 808 nm laser, at 10 and 50 J/cm(2), was used. To evaluate the effects of LLLT, the qualitative and semi-quantitative histological, morphometric, and immunohistochemistry analysis were performed. Initial signs of tissue degradation were observed in CG. Interestingly, laser-treated animals presented a better tissue organization, especially at the fluence of 10 J/cm(2). Furthermore, laser phototherapy was able of modulating some of the aspects related to the degenerative process, such as the prevention of proteoglycans loss and the increase in cartilage area. However, LLLT was not able of modulating chondrocytes proliferation and the immunoexpression of markers related to inflammatory process (IL-1 and MMP-13). This study showed that 808 nm laser, at both fluences, prevented features related to the articular degenerative process in the knees of rats after ACLT.

A systematic review on guanitoxin: General characteristics and ecological risks.

Guanitoxin (GNT) is a potent cyanotoxin, with a relatively low number of publications (n = 51) compared to other cyanotoxins. Among the published studies, 35 % were on the effect of the toxin in animals, mainly in rodents and in vitro testing, followed by studies that identified species of cyanobacteria that produce GNT in aquatic systems and consequently accidental poisoning in wild and domestic animals (27 %). Studies that developed or tested methods for identifying the molecule, based on colorimetric and analytical techniques, represented 14 %, while 8 % were on GNT biosynthesis. Review articles and chemical isolation (6 %) and on the stability of the molecule (4 %) were the topics with the lowest number of publications. The results show the occurrence of GNT was identified mainly in eutrophic environments with a higher incidence in the American continent. Chemical characteristics of the molecule, such as short half-life in the environment, instability in solutions with alkaline pH values, temperature >23 °C, added to the lack of an analytical standard, are factors that make it difficult to identify and quantify it. However, GNT monitoring can be performed using LC-MS-MRM methods or genes specific to the newly discovered molecule.

Nandrolone decanoate induces genetic damage in multiple organs of rats.

To evaluate the impact potential of nandrolone decanoate on DNA damage in multiple organs of Wistar rats by means of single-cell gel (comet) assay and micronucleus test. A total of 15 animals were distributed into three groups of five animals each as follows: control group = animal not exposed to nandrolone decanoate; experimental group = animals exposed to nandrolone decanoate for 24 h at 5 mg/kg subcutaneously; and experimental group = animals exposed to nandrolone decanoate for 24 h at 15 mg/kg subcutaneously. Significant statistical differences (p < 0.05) were noted in peripheral blood, liver, and heart cells exposed to nandrolone decanoate at the two doses evaluated. A clear dose-response relationship was observed between groups. Kidney cells showed genetic damage at only the highest dose (15 mg/kg) used. However, micronucleus data did not show remarkable differences among groups. In conclusion, the present study indicates that nandrolone decanoate induces genetic damage in rat blood, liver, heart, and kidney cells as shown by single-cell gel (comet) assay results.

DNA damage, p53, Ki-67 and COX-2 expression in rat tongue cells exposed to nandrolone decanoate.

The objective of this article was to evaluate the impact potential of nandrolone decanoate on DNA damage, cellular regulatory proteins and cyclooxygenase (COX)-2 in oral mucosa cells of Wistar rats. A total of 40 rats were distributed into four groups. Two experimental groups were treated with nandrolone decanoate, at 5 mg/kg doses, subcutaneously, three times a week in two periods: 15 and 30 days. The remaining groups received only 0.9% saline subcutaneously, three times a week. To evaluate genetic damage, nandrolone decanoate at 15 mg/kg dose was exposed to 24 h. In the histopathological analysis, no remarkable morphological changes were observed in tongue tissue in all groups. Significant increase in immunoexpression of Ki-67, p53, COX-2 proteins was detected in the groups treated with nandrolone decanoate during 15 and 30 days, when compared to their respective controls. A positive correlation between immunoexpression of p53 and COX-2 protein was detected following nandrolone decanoate exposure. DNA damage was induced by nandrolone decanoate in oral mucosa cells at 15 mg/kg dose. Our results suggest that nandrolone decanoate was able to alter the expression of cell cycle-related proteins, as well as to induce genetic damage and COX-2 immunoexpression in tongue cells of Wistar rats.

Radiographers' musculoskeletal health in Western Switzerland: WRMSDs symptoms prevalence and risk factors.

BACKGROUND: Work-related musculoskeletal disorders (WRMSDs) are a significant occupational health concern in radiographers. OBJECTIVE: This study aimed to describe radiographers' WRMSDs symptoms prevalence and severity, exploring associations with occupational risk factors. METHODS: A cross-sectional study was conducted to explore WRMSDs symptoms and occupational risk factors in radiographers of Western Switzerland using an online survey, based on the Nordic Musculoskeletal Questionnaire (NMQ). Descriptive statistics were conducted to analyze the collected data, and associative statistics to identify the risk factors related to symptoms. RESULTS: Participants (n = 359) presented a high prevalence of WRMSDs symptoms in the last 12 months (94.7%), with a related absenteeism rate of 15.6%. In the last 7 days, symptoms prevalence was 67.7%. The most affected anatomical regions, over both time periods, were the neck (73.0%, 36.8%) and low back (67.4%, 35.7%). Associative statistics underlined risk factors affecting significantly radiographers' health (OR >2) were the "awkward postures" (OR = 2.86; 95% CI 1.78-4.58) and "feeling anxiety/stress at work" both for low back (OR = 2.38; 95% CI 1.39-4.08), and being a woman for the neck (OR = 2.64; 95% CI 1.51-4.61). CONCLUSIONS: There is a high WRMSDs symptoms prevalence in Western Switzerland radiographers. Radiographers' work demands namely for awkward postures increases the odds for WRMSDs symptoms presence, affecting predominantly neck, upper and lower back. Our data suggest that further research is needed to implement adapted prevention to this specific context.

Toxicological effects of cyanobacterial metabolites on zebrafish larval development.

Freshwater cyanobacteria are known worldwide for their potential to produce toxins. However, these organisms are also found in marine, terrestrial and extreme environments and produce unique compounds, other than toxins. Nevertheless, their effects on biological systems are still barely known. This work tested extracts of different cyanobacterial strains against zebrafish (Danio rerio) larvae and analyzed their metabolomic profiles using liquid chromatography combined with mass spectrometry. Strains Desertifilum tharense, Anagnostidinema amphibium, and Nostoc sp. promoted morphological abnormalities such as pericardial edema, edema in the digestive system region, curvature of the tail and spine in zebrafish larvae in vivo. In contrast, Microcystis aeruginosa and Chlorogloeopsis sp. did not promote such changes. Metabolomics revealed unique compounds belonging to the classes of terpenoids, peptides, and linear lipopeptides/microginins in the nontoxic strains. The toxic strains were shown to contain unique compounds belonging to the classes of cyclic peptides, amino acids and other peptides, anabaenopeptins, lipopeptides, terpenoids, and alkaloids and derivatives. Other unknown compounds were also detected, highlighting the rich structural diversity of secondary metabolites produced by cyanobacteria. The effects of cyanobacterial metabolites on living organisms, mainly those related to potential human and ecotoxicological risks, are still poorly known. This work highlights the diverse, complex, and unique metabolomic profiles of cyanobacteria and the biotechnological potential and associated risks of exposure to their metabolites.

Different Species of Marine Sponges Diverge in Osteogenic Potential When Therapeutically Applied as Natural Scaffolds for Bone Regeneration in Rats.

A highly porous structure, and an inorganic (biosilica) and collagen-like organic content (spongin) makes marine sponges potential candidates to be used as natural scaffolds in bone tissue engineering. The aim of this study was to characterize (through SEM, FTIR, EDS, XRD, pH, mass degradation and porosity tests) scaffolds produced from two species of marine sponges, Dragmacidon reticulatum (DR) and Amphimedon viridis (AV), and to evaluate the osteogenic potential of these scaffolds by using a bone defect model in rats. First, it was shown that the same chemical composition and porosity (84 ± 5% for DR and 90 ± 2% for AV) occurs among scaffolds from the two species. Higher material degradation was observed in the scaffolds of the DR group, with a greater loss of organic matter after incubation. Later, scaffolds from both species were surgically introduced in rat tibial defects, and histopathological analysis after 15 days showed the presence of neo-formed bone and osteoid tissue within the bone defect in DR, always around the silica spicules. In turn, AV exhibited a fibrous capsule around the lesion (19.9 ± 17.1%), no formation of bone tissue and only a small amount of osteoid tissue. The results showed that scaffolds manufactured from Dragmacidon reticulatum presented a more suitable structure for stimulation of osteoid tissue formation when compared to Amphimedon viridis marine sponge species.

Effect of low-level laser therapy (660 nm) on the healing of second-degree skin burns in rats.

The aim of this study was to investigate the effects of 660 nm laser on the healing of burn wounds made on the backs of rats. Thirty-two Wistar male rats were used. The animals were randomly distributed into 2 groups of 16 animals each: control group (burned rats without treatment) and laser-treated group (burned rats treated with laser therapy). Each group was divided into two different subgroups, euthanized in different periods (subgroup A: 7 days post-surgery and subgroup B: 14 days post-surgery). Histopathological analysis revealed a significant decrease in the necrotic area in the laser-treated group compared to the controls at days 7 and 14 post-injury. COX-2 positive cells were found in a strong pattern in the group submitted to laser therapy after 7 days. Regarding VEGF immunomarker, a significant VEGF immunoexpression was detected in the laser-exposed group after 14 days when compared to the negative control group. Taken together, our results demonstrate that laser therapy is able to promote skin repair of burned rats as a result of decreasing necrotic area and an up-regulation of COX-2 and VEGF immunoexpression.

Availability of Guanitoxin in Water Samples Containing Sphaerospermopsis torques-reginae Cells Submitted to Dissolution Tests.

Guanitoxin (GNT) is a potent neurotoxin produced by freshwater cyanobacteria that can cause the deaths of wild and domestic animals. Through reports of animal intoxication by cyanobacteria cells that produce GNT, this study aimed to investigate the bio-accessibility of GNT in simulated solutions of the gastrointestinal content in order to understand the process of toxicosis promoted by GNT in vivo. Dissolution tests were conducted with a mixture of Sphaerospermopsis torques-reginae (Cyanobacteria; Nostocales) cultures (30%) and gastrointestinal solutions with and without proteolytic enzymes (70%) at a temperature of 37 °C and rotation at 100 rpm for 2 h. The identification of GNT was performed by LC-QqQ-MS/MS through the transitions [M + H]+m/z 253 > 58 and [M + H]+m/z 253 > 159, which showed high concentrations of GNT in simulated gastric fluid solutions (p-value < 0.001) in comparison to simulated solutions of intestinal content. The gastric solution with pepsin promoted the stability of GNT (p-value < 0.05) compared to the simulated solution of gastric fluid at the same pH without the enzyme. However, the results showed that GNT is also available in intestinal fluids for a period of 2 h, and solutions containing the pancreatin enzyme influenced the bio-accessibility of the toxin more compared to the intestinal medium without enzyme (p-value < 0.05). Therefore, the bio-accessibility of the toxin must be considered both in the stomach and in the intestine, and may help in the diagnosis and prediction of exposure and risk in vivo through the oral ingestion of GNT-producing cyanobacteria cells.