Biomechanical and histological characterization of MPS I mice femurs.

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder characterized by alpha-L-iduronidase (IDUA) deficiency, an enzyme responsible for glycosaminoglycan degradation. Musculoskeletal impairment is an important component of the morbidity related to the disease, as it has a major impact on patients' quality of life. To understand how this disease affects bone structure, morphological, biomechanical and histological analyses of femurs from 3- and 6-month-old wild type (Idua +/+) and MPS I knockout mice (Idua -/-) were performed. Femurs from 3-month-old Idua -/- mice were found to be smaller and less resistant to fracture when compared to their age matched controls. In addition, at this age, the femurs presented important alterations in articular cartilage, trabecular bone architecture, and deposition of type I and III collagen. At 6 months of age, femurs from Idua -/- mice were more resistant to fracture than those from Idua +/+. Our results suggest that the abnormalities observed in bone matrix and articular cartilage in 3-month-old Idua -/- animals caused bone tissue to be less flexible and more likely to fracture, whereas in 6-month-old Idua -/- group the ability to withstand more load before fracturing than wild type animals is possibly due to changes in the bone matrix.

Low-Level Laser Therapy and Spinal Cord Injury: Effects of 3 Different Fluences in the Intermediate Period of Repair in an Experimental Model in Rats

Abstract The aim of this study was to evaluate the effectiveness of 3 different fluences of low-level laser therapy (LLLT) in intermediate period of spinal cord repair using an experimental model in rats. Thirty two rats were randomly divided into four experimental groups: Control Group (CG); Laser treated group 500 J/cm² (L-500); Laser treated group 750 J/cm² (L-750) and Laser treated group 1000 J/cm² (L-1000). Spinal cord injury (SCI) was performed by an impactor equipment (between 9th and 10th thoracic vertebrae), with a pressure of 150 kdyn. Afterwards, the injured region was irradiated daily for 14 consecutive sessions, using an 808 nm laser, at the respective fluence of each experimental groups. Locomotor function and tactile sensitivity were performed on days 1 and 15 post-surgery. Animals were euthanized 15 days post-surgery and samples were retrieved for histological and immunohistochemistry analysis. Functional behavior and tactile sensitivity were improved after laser irradiation. Moreover, higher fluencies of LLLT reduced the volume of injury. Additionally, LLLT produced a decreased CD-68 expression. These results demonstrated that, for an intermediate period of SCI repair, LLLT at higher fluences, was effective in promoting functional recovery and modulating the inflammatory process in the spinal cord of rats after traumatic SCI.

Effects of different fluences of low-level laser therapy in an experimental model of spinal cord injury in rats.

The aim of this study was to evaluate the in vivo response of different fluences of low-level laser therapy (LLLT) on the area of the injury, inflammatory markers, and functional recovery using an experimental model of traumatic spinal cord injury (SCI). Thirty two rats were randomly divided into four experimental groups: control group (CG), laser-treated group 500 J/cm2 (L-500), laser-treated group 750 J/cm2 (L-750), and laser-treated group 1000 J/cm2 (L-1000). SCI was performed by an impactor equipment (between the ninth and tenth thoracic vertebrae), with a pressure of 150 kdyn. Afterwards, the injured region was irradiated daily for seven consecutive sessions, using an 808-nm laser, at the respective fluence of each experimental groups. Motor function and tactile sensitivity were performed on days 1 and 7 post-surgery. Animals were euthanized on the eighth day after injury, and the samples were retrieved for histological and immunohistochemistry analyses. Functional evaluation and tactile sensitivity were improved after LLLT, at the higher fluence. Additionally, LLLT, at 750 and 1000 J/cm2, reduces the lesion volume and modulates the inflammatory process with decrease of CD-68 protein expression. These results suggest that LLLT at higher doses was effective in promoting functional recovery and modulating inflammatory process in the spinal cord of rats after SCI.

Low level laser therapy accelerates bone healing in spinal cord injured rats.

Bone loss occurs rapidly and consistently after the occurrence of a spinal cord injury (SCI), leading to a decrease in bone mineral density (BMD) and a higher risk of fractures. In this context, the stimulatory effects of low level laser therapy (LLLT) also known as photobiomodulation (PBM) have been highlighted, mainly due to its osteogenic potential. The aim of the present study was to evaluate the effects of LLLT on bone healing using an experimental model of tibial bone defect in SCI rats. Twenty-four female Wistar rats were randomly divided into 3 groups: Sham group (SG), SCI control group (SC) and SCI laser treated group (SL). Two weeks after the induction of the SCI, animals were submitted to surgery to induce a tibial bone defect. Treatment was performed 3days a week, for 2weeks, at a single point over the area of the injury, using an 808nm laser (30mW, 100J/cm(2); 0.028cm(2), 1.7W/cm², 2.8J). The results of the histological and morphometric evaluation demonstrated that the SL group showed a larger amount of newly formed bone compared to the SC group. Moreover, a significant immunoexpression of runt-related transcription factor 2 (RUNX2) was observed in the SL group. There was no statistical difference in the biomechanical evaluation. In conclusion, the results suggest that LLLT accelerated the process of bone repair in rats with complete SCI.

Behavioural changes observed in demyelination model shares similarities with white matter abnormalities in humans.

Multiple sclerosis (MS) is a chronic, inflammatory, demyelinating disease of the central nervous system (CNS). Further to the symptoms resulting from demyelination, new studies point to the involvement of neuroinflammation and white matter abnormalities in psychiatric disorders and neurodegenerative diseases. Cuprizone, a model of MS, produces consistent demyelination and elicits behavioural, morphological and inflammatory changes in animals that share some similarities with those observed in humans. In this study, we used the cuprizone model in Lewis rats to evaluate clinical signs triggered by the demyelination process which could be comparable with the symptoms seen in white matter abnormalities in human beings. To induce the demyelination process, 0.6% cuprizone was added to the Lewis rats' diet for 4 weeks. We proceeded with behavioural, morphological and immunological analyses. Animals fed with cuprizone exhibited behavioural changes: higher scores in the neurotoxicity test, reduced exploratory and locomotion behaviour, and also an increase of permanency in the closed arm of the elevated plus maze test, were observed. In these analyses, the animals showed motor coordination impairment and anxiety-like behaviour. Demyelination also triggered changes in discrimination of objects identified by an increase in the time spent close to a familiar object. These behavioural alterations were associated with a significant increase in the levels of TNF-alpha and corticosterone, consistent with the activation of microglia and astrocytes. Taken together, the results of this work show the cuprizone/Lewis rat model demyelination as an attractive paradigm for studying the correlation between white matter abnormalities and behaviour.

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.

The effects of low level laser therapy on injured skeletal muscle

The main purpose of the present study was to investigate the effects of low-level laser therapy (LLLT) used in two different fluencies on injured skeletal muscle after cryolesion by means of histopathological analysis and immunohistochemistry for COX-2. A total of sixty male Wistar rats were randomly distributed into three groups: injured animals without any treatment; 808 nm laser treated group, at 10 J/cm² and 808 nm laser treated group, at 50 J/cm². Each group was divided into two different subgroups (n=10) on days 6 and 13 post-injury. The results showed that the animals irradiated with laser at 10 J/cm² or 50 J/cm² presented the areas with cell infiltrate and pointed out to minor and mild areas with destroyed zones compared with the control group. Also, a COX-2 downregulation was noticed in the groups exposed to laser at two fluences evaluated in this study. Significant statistically differences (p<0.05) were noticed to collagen deposition in the laser treated animals, with the fluence of 50 J/cm² when compared to the other groups on day 13 post-surgery. Taken together, these results suggested that laser therapy could have positive effects on muscle repair in the rats after cryolesion.

Transplanting neural progenitors into a complete transection model of spinal cord injury.

Neural progenitor cell (NPC) transplantation is a promising therapeutic strategy for spinal cord injury (SCI) because of the potential for cell replacement and restoration of connectivity. Our previous studies have shown that transplants of NPC, composed of neuron- and glia-restricted progenitors derived from the embryonic spinal cord, survived well in partial lesion models and generated graft-derived neurons, which could be used to form a functional relay. We have now examined the properties of a similar NPC transplant using a complete transection model in juvenile and adult rats. We found poor survival of grafted cells despite using a variety of lesion methods, matrices, and delays of transplantation. If, instead of cultured progenitor cells, the transplants were composed of segmental or dissociated segments of fetal spinal cord (FSC) derived from similar-staged embryos, grafted cells survived and integrated well with host tissue in juvenile and adult rats. FSC transplants differentiated into neurons and glial cells, including astrocytes and oligodendrocytes. Graft-derived neurons expressed glutaminergic and GABAergic markers. Grafted cells also migrated and extended processes into host tissue. Analysis of axon growth from the host spinal cord showed serotonin-positive fibers and biotinylated dextran amine-traced propriospinal axons growing into the transplants. These results suggest that in treating severe SCI, such as complete transection, NPC grafting faces major challenges related to cell survival and formation of a functional relay. Lessons learned from the efficacy of FSC transplants could be used to develop a therapeutic strategy based on neural progenitor cells for severe SCI.

Dual role of cyclooxygenase-2 during tissue repair induced by low level laser therapy: an intriguing issue.

Tissue repair is an excellent example of pathophysiological model for studying the role of cyclooxygenase-2 (COX-2) on eukaryotic cells. It has been established that two COX isoforms are expressed in human tissues: constitutive or induced. COX-1 activity is constitutive, present in nearly all cell types at a constant level; COX-2 activity is normally absent from cells, and when induced, the protein levels increase and decrease in a matter of hours after a single stimulus. Thus, the purpose of this review was to describe the role of COX-2 during tissue repair induced by low level laser therapy (LLLT) in humans and experimental models. COX-2 expression has been implicated in the onset or the exacerbation of inflammation during tissue repair induced by LLLT in a number of studies, Many studies are conducted to investigate the role of COX-2 during tissue repair induced by LLLT using different experimental protocols and dosages. Therefore, this is an area that warrants investigation, since the estimation of COX-2 expression from using such important techniques in therapeutics with respect to tissue repair will be added to those already established in the literature as a way to improve health status and prevention of side effects.

Acute spinal cord injury induces genetic damage in multiple organs of rats.

Spinal cord injury (SCI) is a devastating condition with important functional and psychological consequences. However, the underlying mechanisms by which these alterations occur are still not fully understood. The aim of this study was to analyze genomic instability in multiple organs in the acute phase of SCI by means of single cell gel (comet) assay. Rats were randomly distributed into two groups (n = 5): a SHAM and a SCI group killed 24 h after cord transection surgery. The results pointed out genetic damage in blood cells as depicted by the tail moment results. DNA breakage was also detected in liver and kidney cells after SCI. Taken together, our results suggest that SCI induces genomic damage in multiple organs of Wistar rats.

Low-level laser therapy improves repair following complete resection of the sciatic nerve in rats.

The aim of this study is to analyze the effects of low-level laser therapy (LLLT) on the regeneration of the sciatic nerve in rats following a complete nerve resection. Male Wistar rats were divided into a control injury group, injury groups irradiated with a 660-nm laser at 10 or 50 J/cm(2), and injury groups irradiated with an 808-nm laser at 10 or 50 J/cm(2). Treatment began 24 h following nerve resection and continued for 15 days. Using the sciatic functional index (SFI), we show that the injured animals treated with 660 nm at 10 and 50 J/cm(2) had better SFI values compared with the control injury and the 808-nm groups. Animals irradiated with the 808-nm laser at 50 J/cm(2) show higher values for fiber density than do control animals. In addition, axon and fiber diameters were larger in animals irradiated with 660 nm at 50 J/cm(2) compared to the control group. These findings indicate that 660-nm LLLT is able to provide functional gait recovery and leads to increases in fiber diameter following sciatic nerve resection.

Temporal modifications in bone following spinal cord injury in rats.

INTRODUCTION: The aim of this study was to investigate the temporal modifications in bone mass, bone biomechanical properties and bone morphology in spinal cord injured rats 2, 4 and 6 weeks after a transection. MATERIAL AND METHODS: Control animals were randomly distributed into four groups (n = 10 each group): control group (CG) - control animals sacrificed immediately after surgery; spinal cord-injured 2 weeks (2W) - spinal cord-injured animals sacrificed 2 weeks after surgery; spinal cord-injured 4 weeks (4W) - spinal cord-injured animals sacrificed 4 weeks after surgery; spinal cord-injured 6 weeks (6W) - spinal cord-injured animals sacrificed 6 weeks after surgery. RESULTS: Biomechanical properties of the right tibia were determined by a three-point bending test and injured animals showed a statistically significant decrease in maximal load compared to control animals. The right femur was used for densitometric analysis and bone mineral content of the animals sacrificed 4 and 6 weeks after surgery was significantly higher compared to the control animals and animals sacrificed 2 weeks after surgery. Histopathological and morphological analysis of tibiae revealed intense resorptive areas in the group 2 weeks after injury only. CONCLUSIONS: The results of this study show that this rat model is a valuable tool to investigate bone remodeling processes specifically associated with SCI. Taken together, our results suggest that spinal cord injury induced bone loss within 2 weeks after injury in rats.

Comparison of the effects of electrical field stimulation and low-level laser therapy on bone loss in spinal cord-injured rats.

OBJECTIVE: This study investigated the effects of low-level laser therapy (LLLT) and electrical stimulation (ES) on bone loss in spinal cord-injured rats. MATERIALS AND METHODS: Thirty-seven male Wistar rats were divided into four groups: standard control group (CG); spinal cord-injured control (SC); spinal cord-injured treated with laser (SCL; GaAlAs, 830 nm, CW, 30 mW/cm, 250 J/cm(2)); and spinal cord-injured treated with electrical field stimulation (SCE; 1.5 MHz, 1:4 duty cycles, 30 mW, 20 min). Biomechanical, densitometric, and morphometric analyses were performed. RESULTS: SC rats showed a significant decrease in bone mass, biomechanical properties, and morphometric parameters (versus CG). SCE rats showed significantly higher values of inner diameter and internal and external areas of tibia diaphyses; and the SCL group showed a trend toward the same result (versus SC). No increase was found in either mechanical or densitometric parameters. CONCLUSION: We conclude that the mentioned treatments were able to initiate a positive bone-tissue response, maybe through stimulation of osteoblasts, which was able to determine the observed morphometric modifications. However, the evoked tissue response could not determine either biomechanical or densitometric modifications.

Chromosome damage and cytotoxicity in oral mucosa cells after 2 months of exposure to anabolic steroids (decadurabolin and winstrol) in weight lifting.

The aim of the present study was to evaluate DNA damage (micronucleus) and cellular death (pyknosis, karyolysis and karyorrhexis) in exfoliated buccal mucosa cells from anabolic steroid users after 2 months of exposure. Two experimental groups consisting of 15 adult males who practise weight lifting and are anabolic steroid users or 15 adult males who practise weight lifting, but are non-anabolic steroid users, were recruited. In addition, 20 sedentary males, who do not practise any physical activity regularly, were matched by age with experimental groups. No significant statistical differences (p>0.05) were noticed in individuals who practise physical activity only. On the other hand, an increase of micronucleated cells (MNCs) in anabolic steroid (decadurabulin and Winstrol) users was observed. Regarding cytotoxic parameters, the same observation has occurred, that is, significant statistical differences (p<0.05) were noticed in the group exposed to anabolic steroids when compared with other controls, as depicted by high frequencies of pyknosis, karyolysis and karyorrhexis. Taken together, our results suggest that genomic instability and cytotoxicity are induced by anabolic steroid administration in oral mucosa cells as assessed by the micronucleus test.

Involvement of the histaminergic system on appetitive learning and its interaction with haloperidol in goldfish.

This study investigated the actions of the histaminergic system on appetitive learning and memory, and its interaction with the dopaminergic system in goldfish. It consisted of nine sessions, in which fish were tested in a four-arm tank. On day 1, the animals were habituated for 10 min. On day 2, they were placed in one arm and had to find food at the left or the right arm. Time to begin feeding was recorded, and the procedure repeated for more 3 days (training phase). On training day 4, seven groups were injected with saline, seven with haloperidol (2.0 mg/kg) and one with DMSO solution before training and after feeding, three groups received saline, six chlorpheniramine (CPA) (1.0, 4.0 and 8.0 mg/kg), and six l-histidine (LH) (25, 50 and 100 mg/kg). Saline groups were considered as control of CPA and LH treated groups and DMSO as control of haloperidol. A non-injected group was also included. Testing occurred after 24 h. A reversal procedure was conducted 24h after testing and repeated for 3 days. The groups receiving CPA at 1.0 and 8.0 mg/kg and LH at 25, 50 and 100 mg/kg differed between Test and Reversal day 1. Pre-treatment with haloperidol plus 8.0 mg/kg of CPA and 25 and 50 mg/kg of LH reverted the treatment effect. However, in the groups treated with 1.0 mg/kg of CPA and 100 mg/kg of LH, the difference remained. This study confirmed the interaction between the histaminergic and the dopaminergic systems on memory process in goldfish.

Neurochemical analysis of brain monoamines after L-histidine and chlorpheniramine administration in goldfish.

This study investigated the effects of chlorpheniramine (CPA) and L-histidine (LH) administration on catecholaminergic levels in goldfish brain using neurochemical analysis. Fifty-eight animals were used. After 20 min of i.p. administration of the drugs or saline the animals were decapitated, and the telencephalon and the diencephalon were dissected. We also measured catecholamines in a non-injected (NI) group. Results showed lower homovanillic acid (HVA) levels after treatment with 100 mg/kg of LH when compared to saline and 5-hydroxyindoleacetic acid levels were lower in the saline group when compared to the NI group. In the diencephalon the NI group and animals treated with CPA at 4.0 and 8.0 mg/kg had lower HVA levels. Results suggest that LH had an inhibitory effect on dopaminergic activity and an anxiolytic-like effect for CPA results is suggested.