A biocomplex to repair experimental critical size defects associated with photobiomodulation therapy

The association of scaffolds to repair extensive bone defects can contribute to their evolution and morphophysiological recomposition. The incorporation of particulate biomaterials into three-dimensional fibrin bioproducts together with photobiomodulation therapy (PBM) has potential and can improve regenerative medicine procedures. The objective of this experiment was to evaluate the effects of PBM therapy on critical size defects filled with xenogenic bone substitute associated with fibrin biopolymer. Methods: A critical defect of 8 mm was performed in 36 Wistar male adult rats that were divided into four groups. Groups BC and BC-PBM were defined as controls with defects filled by a clot (without or with PBM, respectively) and groups XS and XS-PBM that comprised those filled with biocomplex Bio-OssTM in association with fibrin biopolymer. PBM was applied immediately after the surgery and three times a week every other day, with the parameters: wavelength of 830 nm, energy density 6.2 J/cm2, output power 30 mW, beam area of 0.116 cm2, irradiance 0.258,62 W/cm2, energy/point 0.72 J, total energy 2.88 J. Fourteen and 42 days after the surgery, animals were euthanatized and subjected to microtomography, qualitative and quantitative histological analysis. Results: The BC-PBM and XS-PBM groups had a similar evolution in the tissue repair process, with a higher density of the volume of new formed bone in relation to the groups without PBM (p = 0.04086; p = 0.07093, respectively). Intense vascular proliferation and bone deposition around the biomaterial particles were observed in the animals of the groups in which biocomplex was applied (XS and XS-PBM). Conclusion: PBM therapy allowed an improvement in the formation of new bone, with a more organized deposition of collagen fibers in the defect area. Biocomplex favored the insertion and permanence of the particulate material in bone defects, creating a favorable microenvironment for accelerate repair process.(AU)

Effect of S53P4 bioactive glass and low-level laser therapy on calvarial bone repair in rats submitted to zoledronic acid therapy

ABSTRACT Purpose To evaluate the influence of bioactive glass and photobiomodulation therapy (PBMT) in calvarial bone repair process in rats submitted to zoledronic acid therapy. Methods Twenty-four rats were selected and treated with the dose of 0.035 mg/kg of zoledronic acid every two weeks, totalizing eight weeks, to induce osteonecrosis. After the drug therapy, surgical procedure was performed to create 5-mm diameter parietal bone defects in the calvarial region. The rats were then randomly assigned to groups according to the following treatments: AZC: control group, treated with blood clot; AZBIO: bone defect filled with bioactive glass; AZL: treated with blood clot and submitted to PBMT; and AZBIOL: treated with bioactive glass S53P4 and submitted to PBMT. Tissue samples were collected and submitted to histomorphometric analysis after 14 and 28 days. Results At 14 days, bone neoformation in the AZBIO (52.15 ± 9.77) and AZBIOL (49.77 ± 13.58) groups presented higher values (p ≤ 0.001) compared to the AZC (23.35 ± 10.15) and AZL groups (23.32 ± 8.75). At 28 days, AZBIO (80.24 ± 5.41)still presented significant higher bone recovery values when compared to AZC (59.59 ± 16.92)and AZL (45.25 ± 5.41) groups (p = 0.048). In the 28-day period, the AZBIOL group didn't show statistically significant difference with the other groups (71.79 ± 29.38). Conclusions The bioactive glass is an effective protocol to stimulate bone neoformation in critical defects surgically created in rats with drug induced osteonecrosis, in the studied periods of 14 and 28 days.

Effect of low-level laser therapy on the membrane induced by the Masquelet technique at an orthotopic site in rabbits

ABSTRACT Purpose: To evaluate the low-level laser therapy (LLLT) on the membrane induced by the Masquelet technique in rabbits. Methods: Twelve Norfolk rabbits at approximately 3 months of age were used. A 1-cm segmental defect was induced in both radii, which were filled with polymethylmethacrylate cylinder. LLLT was used postoperatively in the bone defect of one of the forelimbs every 48 hours for 15 days. Six rabbits were euthanatized on third and sixth postoperative weeks. Results: In both forelimbs, radiographs showed new bone growth from radius cut ends on the third postoperative week and more advanced stage on the sixth postoperative week. Ultrasound showed induced membrane one week after the surgery. Histologically, there were no significant differences in the semi-quantitative score of inflammation intensity, total number of blood vessels, bone metaplasia, and collagen. The average thicknesses were 2,050.17 and 1,451.96 μm for control membranes and 2,724.26 and 2,081.03 μm for irradiated membranes, respectively, on third and sixth postoperative weeks. Vascular endothelial growth factor A (VEGF-A) and platelet derived growth factor (PDGF) expression were present in the induced membranes of control and irradiated forelimbs, but there was no significant difference. Conclusions: Based on assessment methods, it was not possible to demonstrate the effect of LLLT on the induced membrane.

Antimicrobial photodinamic therapy combined with laser photobiomodulation in the treatment of skin wounds in a dog (Canis lupus familiaris)

Background: Laser photobiomodulation has been used in the treatment of various injuries and diseases. This promotes modulation of the inflammatory process, edema reduction and devitalized tissue regeneration. The advantages of Antimicrobial photodynamic therapy are its easy application and the absence of side effects. Other advantages are the cost of the therapy, minimal damage to animal tissue, the broad spectrum of action, and efficiency against strains resistant to antimicrobials. The aim of this study was to report the clinical and their resolution in a female dog with a traumatic, infected wound treated with laser phototherapy as an alternative therapy method. Case: A 3-year-old bitch Border Collie, weighing 18 kg, from the municipality of Ilhéus, Bahia, Brazil, waspresented for examination with a history of traumatic laceration of the left thoracic member. On the anamnesis, it was reported that the patient presented with laceration of the left thoracic member. The wound was cleaned and an antibiotic [30 mg/kg of 12/12 h] and anti-inflammatory [0.1 mg/kg every 24 h were prescribed, both for 5 days]. Twenty-four h after the surgical procedure, there was dehiscence of the sutures, with daily topical therapy based on zinc oxide for secondary intention healing. Upon physical examination, the wound was found to be contaminated with swollen and erythematous edges, an ulcerated area with devitalized tissue, serous exudate, and 8.8 cm2 of wounded area. Given the macroscopic characteristics of the lesion, phototherapy was associated with conventional therapy until complete healing of the wound, with three weekly applications at 48 h intervals. Initially, the wound was cleaned with saline solution at 0.9% and a single treatment with aPDT was scheduled due to the high degree of contamination. The dosimetry parameters of irradiation were calculated according to the wounded area with a diode laser of 0.1W of power, continuous emission, spot area of 0.028 cm2, and energy of 9 J per application point. A gauze imbibed with 1 mL of methylene blue aqueous solution (300 µM), which was the photosensitizer was applied to the lesion, with a pre-irradiation time of 5 min, after which it was irradiated with red laser (RL) (λ = 660nm) for 90 s per point, using the sweeping technique. The edge of the lesion was irradiated with infrared laser (IRL) (λ = 808 nm), total energy of 5 J, using the technique of specific points and 1 J of energy/point. After aPDT, low-intensity laser therapy (LILT) sessions were set up with RL and IRL, with energies of 0.5 J/point and 1 J/point, respectively. The wound was cleaned daily, protected with a bandage, and clinically evaluated until complete regression. Discussion: In the present case, methylene blue was used as a photosensitizer, but it is noteworthy that, apparently, each microorganism responds differently to photosensitizers. Thus, the therapy becomes specific for each application, for example: the type of photosensitizer, its concentration, pre-irradiation time, type of light used in photosensitization, wavelength, energy, power, mode delivery of light. Thus, for the best result, the specific protocol in each application should be used Low-intensity laser therapy is an easily executed technique with effective results. The use of PDT associated with photobiomodulation therapy enabled rapid healing of the cutaneous wound, in addition to an improvement in clinical signs and pain caused by the lesion. The technique proved to be an efficient alternative in the treatment of wounds, whether used in isolation or associated with conventional therapy.

Staphylococcus aureus growth delay after exposure to low fluencies of blue light (470 nm) / Atraso no crescimento de Staphylococcus aureus após exposição à baixas fluências de luz azul (470 nm)

Antibiotic resistance is one of the greatest challenges to treat bacterial infections worldwide, leading to increase in medical expenses, prolonged hospital stay and increased mortality. The use of blue light has been suggested as an innovative alternative to overcome this problem. In this study we analyzed the antibacterial effect of blue light using low emission parameters on Staphylococcus aureus cultures. In vitro bacterial cultures were used in two experimental approaches. The first approach included single or fractionated blue light application provided by LED emitters (470 nm), with the following fluencies: 16.29, 27.16 and 54.32 J/cm2. For the second approach a power LED (470 nm) was used to deliver 54.32 J/cm2 fractionated in 3 applications. Our results demonstrated that bacterial cultures exposed to fractionated blue light radiation exhibited significantly smaller sizes colonies than the control group after 24 h incubation, however the affected bacteria were able to adapt and continue to proliferate after prolonged incubation time. We could conclude that the hypothetical clinical use of low fluencies of blue light as an antibacterial treatment is risky, since its action is not definitive and proves to be ineffective at least for the strain used in this study.

A resistência a antibióticos é um dos maiores desafios para o tratamento de infecções bacterianas em todo o mundo, levando ao aumento de despesas médicas, prolongamento da internação hospitalar e aumento da mortalidade. O uso da luz azul tem sido sugerido como uma alternativa inovadora para superar esse problema. Neste estudo, analisamos o efeito antibacteriano da luz azul usando parâmetros de baixa emissão em culturas de Staphylococcus aureus. Culturas bacterianas foram usadas em duas abordagens experimentais in vitro. A primeira abordagem incluiu o uso da aplicação única ou fracionada de luz azul fornecida por emissores de LED (470 nm), com as seguintes fluências: 16,29, 27,16 e 54,32 J/cm2. Para a segunda abordagem, um LED de potência (470 nm) foi usado para fornecer 54,32 J/cm2 fracionado em 3 aplicações. Nossos resultados demonstraram que as culturas bacterianas expostas à radiação de luz azul fracionada exibiram colônias de tamanhos significativamente menores do que o grupo controle após 24 h de incubação, no entanto, as bactérias afetadas foram capazes de se adaptar e continuar a proliferar após um tempo prolongado de incubação. Podemos concluir que o uso clínico hipotético de baixas fluências de luz azul como tratamento antibacteriano é arriscado, pois sua ação não é definitiva e mostra-se ineficaz, pelo menos para a cepa utilizada neste estudo.

A utilização do laser de baixa intensidade na cicatrização de ferida em cão / The use of low intensity laser in wound healing in dog

Background: Low-intensity laser is effective in cellular metabolism, analgesia, and tissue repair. The bioelectric, bioenergetic, and biochemical effects of laser therapy stimulate local circulation, collagen formation, and epithelization. The objective of this study was to report the use of gallium-aluminum arsenide (GaAlAs) laser (830 nm) in healing two woundscaused by necrosis in a dog. Case: An 8-year-old bitch, a rescued victim of animal abuse, had tibiotarsal dislocation in the left posterior limb. Orthopedic surgery was performed with the placement of an external fixation device with six pins in the distal third of the left posterior limb. A radiographic examination performed 42 days after the surgery revealed the loss of the tibiotarsal ratio, decreased radiopacity of the carpal bones, edema, inflammation, muscle atrophy, and the rotation of the anatomical axis of the metatarsals and phalanges. It was then decided to perform a second orthopedic surgery, in which a bone graft and arthrodesis with an 8-screw titanium plate were performed. Four of the plate screws were placed proximally to the tibia and four distally to the tarsus in addition to a screw and a pin transfixed through the distal metaphyseal region of the tibia and calcaneus. Traction was felt during suturing in the dermis and epidermis of the limb, suggesting the possibility of dehiscence of the stitches. After 3 h of surgery, the limb was cold and edematous. After 5 days, the tissue of the medial region of the tarsus developed necrosis, exposing a wound 6 cm long, 1.5 cm wide, and 2 mm deep (wound 1). Another necrotic lesion was observed in the dorsal region of the tarsos - 6.5 cm long, 2 cm wide, 3 mm deep, and exposing 5 cm of the titanium plate (wound 2). Therapy with GaAlAs laser was then performed over the entire length of the wounds. In each therapeutic session, the laser was first used in punctual mode at 10 J/cm², 830 nm, 20 s at each point of the length of the lesion and then in scan mode at 10 J/cm², 830 nm, 1000 Hz, and 40 s continuously. Laser therapy sessions were conducted at 2-4 day intervals, with a 12-day interval between the eighth and ninth sessions. During the entire treatment, cleansing and debridement of the wounds were performed every 48 h with saline and chlorhexidine digluconate, using a compression bandage, as described earlier. Wound 1 healed completely after two laser therapy sessions. In wound 2, tissue repair stagnated after the ninth and last laser session, leaving 4 cm of the titanium plate still exposed. The limb could not support the body weight of the patient, and radiographic examination revealed that the anatomical axis of the metatarsals and the phalanges was rotated and bone conformation was poor. Given the anatomical conditions associated with the patient's clinical picture, it was decided to amputate the limb between the femur and tibia. Discussion: Therapeutic lasers act on mitochondrial respiration, increasing respiratory metabolism and stimulating DNA synthesis and tissue proliferation. In the present case, there was a decrease in inflammatory cells, edema, and the size of the wounds. The pathological conditions of the affected site, influence of trauma, and degree of tissue damage affected the results of the laser therapy. Despite the size difference between wounds 1 and 2, the exposed titanium plate was a determining factor for the partial healing of wound 2.

Healing of skin wounds in rats: influence of low-level laser therapy, alone or combined with natural products

Background: A great variety of natural products have been evaluated for the skin wound healing due to anti-inflammatory,antioxidant and antibacterial activities, and procollagen synthesis, of which may be mentioned the honey and propolis. Inturn, low-level laser therapy (LLLT) is considered an important tool in the treatment of skin wounds, because of ability toraise the ATP production, to stimulate the microcirculation, and formation of new blood vessels. Therefore, this study aimedto assess the influence of LLLT, alone or combined with natural products, in the healing of excisional skin wounds in rats.Materials, Methods & Results: One hundred twenty-six male Wistar rats, aged 14 weeks, were randomly divided intoseven groups (n = 18 per group) according to wound treatments: G1 (negative control): 0.9% saline solution; G2 (positive control): allantoin + zinc oxide ointment; G3: LLLT; G4: mixture of honey + 5% propolis hydroalcoholic solution;G5: LLLT + mixture of honey + hydroalcoholic solution of 5% propolis; G6: LLLT + honey in natura; G7: LLLT + 5%propolis hydroalcoholic solution. Six rats from each group were euthanized on the 7th, 14th, and 21th postoperative days.Macroscopic and histological evaluations of wound healing were performed. A decrease in wound area occurred in allgroups, but in general G4 had the higher reduction and G1 and G3 had the lower reductions. These findings correspondedto the percentage of wound contraction, since the largest contraction was G4 and the lowest contractions were G1 andG3. Histological analysis showed no significant difference among groups...

Healing of skin wounds in rats: influence of low-level laser therapy, alone or combined with natural products

Background: A great variety of natural products have been evaluated for the skin wound healing due to anti-inflammatory,antioxidant and antibacterial activities, and procollagen synthesis, of which may be mentioned the honey and propolis. Inturn, low-level laser therapy (LLLT) is considered an important tool in the treatment of skin wounds, because of ability toraise the ATP production, to stimulate the microcirculation, and formation of new blood vessels. Therefore, this study aimedto assess the influence of LLLT, alone or combined with natural products, in the healing of excisional skin wounds in rats.Materials, Methods & Results: One hundred twenty-six male Wistar rats, aged 14 weeks, were randomly divided intoseven groups (n = 18 per group) according to wound treatments: G1 (negative control): 0.9% saline solution; G2 (positive control): allantoin + zinc oxide ointment; G3: LLLT; G4: mixture of honey + 5% propolis hydroalcoholic solution;G5: LLLT + mixture of honey + hydroalcoholic solution of 5% propolis; G6: LLLT + honey in natura; G7: LLLT + 5%propolis hydroalcoholic solution. Six rats from each group were euthanized on the 7th, 14th, and 21th postoperative days.Macroscopic and histological evaluations of wound healing were performed. A decrease in wound area occurred in allgroups, but in general G4 had the higher reduction and G1 and G3 had the lower reductions. These findings correspondedto the percentage of wound contraction, since the largest contraction was G4 and the lowest contractions were G1 andG3. Histological analysis showed no significant difference among groups...(AU)

Use of phytotherapics, low power laser and ozone for biting wound in dog

Background: Skin wounds are among the most common cases in small animals, highlighting the importance of alternativetherapies as an aid to the healing process. These include ozone, which has a bactericidal and immunomodulatory action; thelow power laser that determines photodynamic changes triggering metabolism elevation; and the use of medicinal plants,in which several herbal medicines has been mentioned in research relating their benefits to wound healing in humans andanimals. Thus, it aims to report the case of a dog submitted to these treatments due to chronic wound due to bite.Case: A 6-year-old canine, female, mixed-breed, 8 kg, with a perforating wound in the left anterior limb in the scapularand humeral region, caused by trauma from the bite of another dog, previously treated, without clinical improvement,has been attended presenting apathy, increased body temperature (40.1°C), difficulty in walking and inability to supportthe injured limb, which was swollen, hyperemic, with elevated temperature, absence of hair, elevated pain sensitivity andpresence of 2 circular lesions, approximately 2 cm in diameter, caused by trauma, and others of size between 0.5 and 1 cmin diameter, which appeared during the course of the disease, characterized as fistulas, through which there was drainageof characteristic fluid exudative and purulent discharge. It was collected blood sample for blood count and purulent material through sterile swab for culture and antibiogram. Systemic treatment with ceftiofur, dipyrone, meloxican and topicalcleaning of the wounds with hydrogen peroxide and physiological solution; dimethyl sulfoxide ointment, and hot compresses was chosen. After seven days, the animal showed significant general clinical...

Use of phytotherapics, low power laser and ozone for biting wound in dog

Background: Skin wounds are among the most common cases in small animals, highlighting the importance of alternativetherapies as an aid to the healing process. These include ozone, which has a bactericidal and immunomodulatory action; thelow power laser that determines photodynamic changes triggering metabolism elevation; and the use of medicinal plants,in which several herbal medicines has been mentioned in research relating their benefits to wound healing in humans andanimals. Thus, it aims to report the case of a dog submitted to these treatments due to chronic wound due to bite.Case: A 6-year-old canine, female, mixed-breed, 8 kg, with a perforating wound in the left anterior limb in the scapularand humeral region, caused by trauma from the bite of another dog, previously treated, without clinical improvement,has been attended presenting apathy, increased body temperature (40.1°C), difficulty in walking and inability to supportthe injured limb, which was swollen, hyperemic, with elevated temperature, absence of hair, elevated pain sensitivity andpresence of 2 circular lesions, approximately 2 cm in diameter, caused by trauma, and others of size between 0.5 and 1 cmin diameter, which appeared during the course of the disease, characterized as fistulas, through which there was drainageof characteristic fluid exudative and purulent discharge. It was collected blood sample for blood count and purulent material through sterile swab for culture and antibiogram. Systemic treatment with ceftiofur, dipyrone, meloxican and topicalcleaning of the wounds with hydrogen peroxide and physiological solution; dimethyl sulfoxide ointment, and hot compresses was chosen. After seven days, the animal showed significant general clinical...(AU)

Effect of low power laser in biomodulation of cultured osteoblastic cells of Wistar rats

Purpose:To analyze aspects of the biomodulating effect of light in biological tissues, bone cells from surgical explants of the femur of rats were irradiated with low intensity laser.Methods:Bone cells were cultured and irradiated with LASER light (GaAlAs). Growth, cell viability, mineralized matrix formation, total protein dosage, immunostimulatory properties, cytochemical analysis, gene expression of bone proteins were examined using live cell imaging and cell counting by colorimetric assay. The gene expression of: alkaline phosphatase (ALP), type 1 collagen, osteocalcin and osteopontin through the real-time polymerase chain reaction.Results:At 8 days, the viability of the irradiated culture was 82.3% and 72.4% in non-irradiated cells. At 18 days, the cellular viability (with laser) was 77.42% and 47.62% without laser. At 8 days, the total protein concentration was 21.622 mg / mol in the irradiated group and 16, 604 mg / mol in the non-irradiated group and at 18 days the concentration was 37.25 mg / mol in the irradiated group and 24, 95 mg / mol in the non-irradiated group.Conclusion:The laser interfered in the histochemical reaction, cell viability, matrix mineralization, and maintained the cellular expression of proteins.(AU)

Influence of low-level laser irradiation on osteocalcin protein and gene expression in bone tissue1

Purpose: To evaluate osteocalcin gene and protein expression in vitro and in an in vivo model of ostectomy. Methods: Twenty Wistar rats were assigned into two groups A (n=10, laser) and B (n=10, control). Ostectomy was performed in the femur diaphysis; the twenty fragments removed, composed in vitro groups named as in vivo (A and B) and cultivated in CO2 atmosphere for thirteen days. Low-level laser irradiation was performed in groups A (in vivo and in vitro) by an GaAlAs device (λ=808 nm, dose of 2J/cm2, power of 200mW, power density of 0.2W/cm2, total energy of 1.25J, spot diameter of 0.02mm) for 5 seconds, at one point, daily. It was performed immunocytochemistry assays in vivo and in vitro groups. In vitro groups were also submitted to RNA extraction, cDNA synthesis and gene expression by quantitative PCR. Statistical analysis was realized with p<0.05. Results: Immunocytochemistry scores showed no significant differences between control and laser groups either in vivo and in vitro. Gene expression also showed no statistical differences. Conclusion: Low-level laser irradiation did not alter osteocalcin protein and gene expression in vivo and in vitro in the studied period but it may have been expressed them in an earlier period.(AU)

Laserterapia Para Tratamento De Tendinopatia Induzida Em Ratos Wistar: Estudo Histomorfométrico / Laser Therapy For Treatment Of Tendinopathy Induced In Wistar Rats: An Histomorfometric Study / Terapia Con Láser Para Tratamiento De Tendinopatia Inducida En Ratas Wistar: Estudio Histomorfométrico

O objetivo desse estudo foi avaliar os efeitos da laserterapia na lesão do tendão calcanear comum de ratos Wistar. Foram utilizados 12 ratos machos adultos, distribuídos aleatoriamente em dois grupos (L=laser e C=controle). Todos foram submetidos à tendinopatia unilateral (escolha aleatória) mediante compressão transversal do tendão (10 segundos) com pinça Halstead mosquito, assim como escarificações (com bisturi). Após 24 horas da indução da lesão os animais do grupo L receberam laser (904 nm/3 J/cm²/9s) por 20 dias. Os do grupo C foram manipulados como se fossem receber a radiação. Após 3, 7, 14 e 21 dias da realização da lesão, três ratos de cada grupo foram submetidos à eutanásia, e os tendões obtidos para análise histomorfométrica. As amostras foram processadas como de rotina e os fragmentos corados com hematoxilina-eosina, tricrômico de Masson e Picrosirius Red. Os dados foram analisados por ANOVA, a 5% de probabilidade e análise de regressão. Não houve diferença entre grupos nem entre tempos para as características hemorragia, angiogênese, espessamento do epitendão. Independente do tratamento ocorreu diminuição (p=0,0129) da formação de aderência fibrinosa (do 3o ao 21o dias). Por outro lado, a avaliação morfométrica revelou maiores (p=0,0120) quantidades de fibroblastos no grupo que recebeu laserterapia, não havendo efeito de tempo. Avaliação semiquantitativa, revelou maiores (p=0,0000) quantidades de fibroblastos no grupo tratado, porém nessa análise, a quantidade dessas células aumentou com o tempo (p=0,0001) em ambos os grupos. Diferentemente, ANOVA revelou redução do infiltrado inflamatório do 3o ao 21o dia em ambos os grupos (histologia: p=0,0003; morfometria: p=0,0000).(AU)

The aim of this study was to evaluate the effects of laser therapy in tendinopathy of Achilles tendon of Wistar rats. Thus, 12 adult male rats which were divided into two groups (L=laser and C=control) were used. All animals were submitted to unilateral tendinopathy (random selection) by transverse compression of the tendon (10 seconds) with Halsted forceps, as well as 10 scarifications (using a scalpel). After 24 hours of induction of lesion all animals of Group L received laser (904 nm/3 J/cm²/ 9 s) for 20 days, while Group C were handled as if they would receive radiation. After 3, 7, 14 and 21 days lesion induction, three rats from each group were euthanized, and the tendons obtained for histomorphometric analysis. The samples were processed as routine and the sections stained with hematoxylin-eosin, Masson's trichrome and Picrosirius Red. Data were analyzed by ANOVA, at 5% probability and regression analysis. No differences between groups neither between time for hemorrhage characteristics, angiogenesis, thickening of epitendon. Independently of the treatment there was a decrease (p=0.0129) of fibrinous adhesion (3rd to 21st day). On the other hand, morphometric analysis revealed higher (p=0.0120) amounts of fibroblasts cells in the group receiving laser therapy, with no effect of time. Semiquantitative assessment also showed higher (p=0.0000) amounts of fibroblasts cells in the treated group, but in this analysis, the number of fibroblasts increased with time (p=0.0001) in both group. In contrast, ANOVA revealed a reduction of the inflammatory cells from 3rd to 21st day in both groups (histology: p=0.0003; morphometry: p=0.0000).(AU)

La finalidad de ese estudio fue evaluar los efectos del láser en la lesión del tendón calcáneo común de ratas Wistar. Fueron utilizadas 12 ratas machos adultos, distribuidos al azar en dos grupos (L=láser y C=control). Todos fueron sometidos a tendinopatía unilateral (selección aleatoria) por la compresión transversal del tendón (10 segundos) con una pinza Halstead-Mosquito, y escarificación (con bisturí). Después de 24 horas de la inducción de la lesión los animales del grupo L recibieron láser (904 nm/3J/ cm²/9s) durante 20 días. Los tendones del grupo C fueran manejados como si recibera la radiación. Después de 3, 7, 14 y 21 días de realización de la lesión, tres ratas de cada grupo fueron sacrificadas, y los tendones obtenidos para análisis histomorfométrico. Las muestras fueron procesadas como de rutina y los fragmentos se tiñeron con hematoxilina-eosina, tricrómico de Masson y Picrosirius Red. Los datos fueron analizados por ANOVA, con una probabilidad de 5%, además de análisis de regresión. No hubo diferencias entre los grupos o tiempos para las variables hemorragia, angiogénesis, engrosamiento del epitendón. Independientemente del tratamiento se redujo (p=0,0129) la formación de adherencia fibrinosa (del 3o al 21o días). Por otra parte, la evaluación morfométrica mostró cantidades más altas (p=0,0120) de los fibroblastos en el grupo que recibió el tratamiento con láser, sin efecto del tiempo. Evaluación semicuantitativa reveló mayor (p=0,0000) cantidad de fibroblastos en el grupo tratado, pero en esa análisis la cantidad de esas células aumentó con el tiempo (p=0,0001) en ambos grupos. En contraste, ANOVA reveló reducción del infiltrado inflamatorio entre el 3o y el 21o días, en ambos grupos (histología: p=0,0003; morfometría: p=0,0000). Aunque no hubo diferencias entre grupos en la cantidad de fibras de colágeno (I y III), la morfometría reveló que las ratas del grupo L presentaram mayores cantidades (p=0,0096) de fibras de colágeno.(AU)

Laserterapia Para Tratamento De Tendinopatia Induzida Em Ratos Wistar: Estudo Histomorfométrico / Laser Therapy For Treatment Of Tendinopathy Induced In Wistar Rats: An Histomorfometric Study / Terapia Con Láser Para Tratamiento De Tendinopatia Inducida En Ratas Wistar: Estudio Histomorfométrico

O objetivo desse estudo foi avaliar os efeitos da laserterapia na lesão do tendão calcanear comum de ratos Wistar. Foram utilizados 12 ratos machos adultos, distribuídos aleatoriamente em dois grupos (L=laser e C=controle). Todos foram submetidos à tendinopatia unilateral (escolha aleatória) mediante compressão transversal do tendão (10 segundos) com pinça Halstead mosquito, assim como escarificações (com bisturi). Após 24 horas da indução da lesão os animais do grupo L receberam laser (904 nm/3 J/cm²/9s) por 20 dias. Os do grupo C foram manipulados como se fossem receber a radiação. Após 3, 7, 14 e 21 dias da realização da lesão, três ratos de cada grupo foram submetidos à eutanásia, e os tendões obtidos para análise histomorfométrica. As amostras foram processadas como de rotina e os fragmentos corados com hematoxilina-eosina, tricrômico de Masson e Picrosirius Red. Os dados foram analisados por ANOVA, a 5% de probabilidade e análise de regressão. Não houve diferença entre grupos nem entre tempos para as características hemorragia, angiogênese, espessamento do epitendão. Independente do tratamento ocorreu diminuição (p=0,0129) da formação de aderência fibrinosa (do 3o ao 21o dias). Por outro lado, a avaliação morfométrica revelou maiores (p=0,0120) quantidades de fibroblastos no grupo que recebeu laserterapia, não havendo efeito de tempo. Avaliação semiquantitativa, revelou maiores (p=0,0000) quantidades de fibroblastos no grupo tratado, porém nessa análise, a quantidade dessas células aumentou com o tempo (p=0,0001) em ambos os grupos. Diferentemente, ANOVA revelou redução do infiltrado inflamatório do 3o ao 21o dia em ambos os grupos (histologia: p=0,0003; morfometria: p=0,0000).

The aim of this study was to evaluate the effects of laser therapy in tendinopathy of Achilles tendon of Wistar rats. Thus, 12 adult male rats which were divided into two groups (L=laser and C=control) were used. All animals were submitted to unilateral tendinopathy (random selection) by transverse compression of the tendon (10 seconds) with Halsted forceps, as well as 10 scarifications (using a scalpel). After 24 hours of induction of lesion all animals of Group L received laser (904 nm/3 J/cm²/ 9 s) for 20 days, while Group C were handled as if they would receive radiation. After 3, 7, 14 and 21 days lesion induction, three rats from each group were euthanized, and the tendons obtained for histomorphometric analysis. The samples were processed as routine and the sections stained with hematoxylin-eosin, Masson's trichrome and Picrosirius Red. Data were analyzed by ANOVA, at 5% probability and regression analysis. No differences between groups neither between time for hemorrhage characteristics, angiogenesis, thickening of epitendon. Independently of the treatment there was a decrease (p=0.0129) of fibrinous adhesion (3rd to 21st day). On the other hand, morphometric analysis revealed higher (p=0.0120) amounts of fibroblasts cells in the group receiving laser therapy, with no effect of time. Semiquantitative assessment also showed higher (p=0.0000) amounts of fibroblasts cells in the treated group, but in this analysis, the number of fibroblasts increased with time (p=0.0001) in both group. In contrast, ANOVA revealed a reduction of the inflammatory cells from 3rd to 21st day in both groups (histology: p=0.0003; morphometry: p=0.0000).

La finalidad de ese estudio fue evaluar los efectos del láser en la lesión del tendón calcáneo común de ratas Wistar. Fueron utilizadas 12 ratas machos adultos, distribuidos al azar en dos grupos (L=láser y C=control). Todos fueron sometidos a tendinopatía unilateral (selección aleatoria) por la compresión transversal del tendón (10 segundos) con una pinza Halstead-Mosquito, y escarificación (con bisturí). Después de 24 horas de la inducción de la lesión los animales del grupo L recibieron láser (904 nm/3J/ cm²/9s) durante 20 días. Los tendones del grupo C fueran manejados como si recibera la radiación. Después de 3, 7, 14 y 21 días de realización de la lesión, tres ratas de cada grupo fueron sacrificadas, y los tendones obtenidos para análisis histomorfométrico. Las muestras fueron procesadas como de rutina y los fragmentos se tiñeron con hematoxilina-eosina, tricrómico de Masson y Picrosirius Red. Los datos fueron analizados por ANOVA, con una probabilidad de 5%, además de análisis de regresión. No hubo diferencias entre los grupos o tiempos para las variables hemorragia, angiogénesis, engrosamiento del epitendón. Independientemente del tratamiento se redujo (p=0,0129) la formación de adherencia fibrinosa (del 3o al 21o días). Por otra parte, la evaluación morfométrica mostró cantidades más altas (p=0,0120) de los fibroblastos en el grupo que recibió el tratamiento con láser, sin efecto del tiempo. Evaluación semicuantitativa reveló mayor (p=0,0000) cantidad de fibroblastos en el grupo tratado, pero en esa análisis la cantidad de esas células aumentó con el tiempo (p=0,0001) en ambos grupos. En contraste, ANOVA reveló reducción del infiltrado inflamatorio entre el 3o y el 21o días, en ambos grupos (histología: p=0,0003; morfometría: p=0,0000). Aunque no hubo diferencias entre grupos en la cantidad de fibras de colágeno (I y III), la morfometría reveló que las ratas del grupo L presentaram mayores cantidades (p=0,0096) de fibras de colágeno.

Epitelização de enxertos cutâneos em feridas recentes de coelhos tratados com membrana amniótica canina e/ou laserterapia / Skin graft epithelialization in rabbit fresh wounds treated with amniotic membrane and/or laser therapy

A enxertia cutânea é uma técnica cirúrgica simples e bastante útil para o reparo de feridas, principalmente aquelas onde existe dificuldade da aplicação do fechamento primário ou de outras técnicas reconstrutivas. Entretanto, para a sobrevivência do enxerto, é necessário que o leito da ferida esteja saudável e com presença de tecido de granulação exuberante. O objetivo do presente trabalho foi avaliar a aplicação da membrana amniótica e da laserterapia como potenciais estimulantes da cicatrização em enxertos aplicados em feridas sem tecido de granulação. Foram utilizados 42 coelhos, divididos em quatro grupos: grupo controle (GC), grupo membrana (GM), grupo laser (GL) e grupo membrana e laser (GML), submetidos a avaliações macro e microscópicas. Na avaliação macroscópica, foi possível notar que os pacientes dos grupos nos quais a membrana amniótica foi utilizada (GM e GML) apresentaram evidências associadas à acentuada reação inflamatória, à falha de integração do enxerto e à consequente necrose dele. Já os pacientes do GL apresentaram melhor aspecto do enxerto no último dia de avaliação. Na análise microscópica, observou-se intensa integração do enxerto à derme, reepitelização acentuada e escassas células inflamatórias no local do enxerto no GL. O contrário foi observado nos pacientes do GM e GML, nos quais aparentemente houve rejeição da membrana. A formação de colágeno não se correlacionou com outros fatores, como inflamação e necrose, em nenhum dos grupos de tratamento. Dessa forma, é possível afirmar que a laserterapia mostrou ser efetiva, contribuindo para o processo cicatricial e a integração do enxerto. Já a membrana amniótica canina não deve ser utilizada para esse fim, pois provoca intensa reação inflamatória, além de impedir a nutrição do enxerto.(AU)

Skin grafting is a simple surgical technique and useful to repair wounds, especially those where there is a difficulty to apply primary closure skin or other reconstructive techniques. However, for graft survival a healthy wound bed and the presence of an exuberant granulation tissue are necessary. This study aimed to evaluate the application of amniotic membrane and laser therapy as potential healing stimulants in grafts applied in wounds without granulation tissue. For this, we used 42 rabbits divided into four treatment groups, control group (CG), membrane group (MG), laser group (LG) and membrane and laser group (MLG), submitted to macroscopic and microscopic evaluation. The macroscopic examination showed that the patients of the groups where the amniotic membrane was used in order to stimulate re-epithelialization (MG and MLG) presented evidences associated with severe inflammatory reaction, graft integration failure and consequent necrosis. LG patients apparently had the best graph aspect in the last valuation date. Microscopic examination showed intense integration of the graft to the dermis, high re-epithelialization level, and scarce inflammatory cells in the graft site of LG patients. The opposite was observed in patients in the MG and MLG groups, where a rejection of the membrane was observed. Finally, collagen formation was not correlated with other factors such as inflammation and necrosis in any of the treatment groups. We can conclude that laser therapy was effective, contributing to the healing process and integration of the graft. Thus, canine amniotic membrane should not be used for this purpose because it causes intense inflammatory reaction besides avoiding graft nutrition.(AU)

Epitelização de enxertos cutâneos em feridas recentes de coelhos tratados com membrana amniótica canina e/ou laserterapia / Skin graft epithelialization in rabbit fresh wounds treated with amniotic membrane and/or laser therapy

A enxertia cutânea é uma técnica cirúrgica simples e bastante útil para o reparo de feridas, principalmente aquelas onde existe dificuldade da aplicação do fechamento primário ou de outras técnicas reconstrutivas. Entretanto, para a sobrevivência do enxerto, é necessário que o leito da ferida esteja saudável e com presença de tecido de granulação exuberante. O objetivo do presente trabalho foi avaliar a aplicação da membrana amniótica e da laserterapia como potenciais estimulantes da cicatrização em enxertos aplicados em feridas sem tecido de granulação. Foram utilizados 42 coelhos, divididos em quatro grupos: grupo controle (GC), grupo membrana (GM), grupo laser (GL) e grupo membrana e laser (GML), submetidos a avaliações macro e microscópicas. Na avaliação macroscópica, foi possível notar que os pacientes dos grupos nos quais a membrana amniótica foi utilizada (GM e GML) apresentaram evidências associadas à acentuada reação inflamatória, à falha de integração do enxerto e à consequente necrose dele. Já os pacientes do GL apresentaram melhor aspecto do enxerto no último dia de avaliação. Na análise microscópica, observou-se intensa integração do enxerto à derme, reepitelização acentuada e escassas células inflamatórias no local do enxerto no GL. O contrário foi observado nos pacientes do GM e GML, nos quais aparentemente houve rejeição da membrana. A formação de colágeno não se correlacionou com outros fatores, como inflamação e necrose, em nenhum dos grupos de tratamento. Dessa forma, é possível afirmar que a laserterapia mostrou ser efetiva, contribuindo para o processo cicatricial e a integração do enxerto. Já a membrana amniótica canina não deve ser utilizada para esse fim, pois provoca intensa reação inflamatória, além de impedir a nutrição do enxerto.(AU)

Skin grafting is a simple surgical technique and useful to repair wounds, especially those where there is a difficulty to apply primary closure skin or other reconstructive techniques. However, for graft survival a healthy wound bed and the presence of an exuberant granulation tissue are necessary. This study aimed to evaluate the application of amniotic membrane and laser therapy as potential healing stimulants in grafts applied in wounds without granulation tissue. For this, we used 42 rabbits divided into four treatment groups, control group (CG), membrane group (MG), laser group (LG) and membrane and laser group (MLG), submitted to macroscopic and microscopic evaluation. The macroscopic examination showed that the patients of the groups where the amniotic membrane was used in order to stimulate re-epithelialization (MG and MLG) presented evidences associated with severe inflammatory reaction, graft integration failure and consequent necrosis. LG patients apparently had the best graph aspect in the last valuation date. Microscopic examination showed intense integration of the graft to the dermis, high re-epithelialization level, and scarce inflammatory cells in the graft site of LG patients. The opposite was observed in patients in the MG and MLG groups, where a rejection of the membrane was observed. Finally, collagen formation was not correlated with other factors such as inflammation and necrosis in any of the treatment groups. We can conclude that laser therapy was effective, contributing to the healing process and integration of the graft. Thus, canine amniotic membrane should not be used for this purpose because it causes intense inflammatory reaction besides avoiding graft nutrition.(AU)

The effects of photobiomodulation therapy on Staphylococcus aureus infected surgical wounds in diabetic rats. A microbiological, histopathological, and biomechanical study

PURPOSE:To evaluate the effects of photobiomodulation therapy (PBMT) at 685 nm on diabetic wound healing in rats suffering from bacterial infection induced by Staphylococcus aureus (S. aureus).METHODS:Thirty streptozotocin-induced diabetic rats were allocated into two groups: control and PBMT. A 4-cm full-thickness linear-incision was made on the dorsal midline and was contaminated with S. aureus. The wounds in the PBMT group were irradiated daily for 5 consecutive days, starting 3 days after the induction and always in the mornings.RESULTS:The result revealed that PBMT resulted in a significant decrease in S. aureus CFU in the PBMT group in comparison to the control group (P<0.05). The length of wounds, in the 2nd and 3rd weeks, in the PBMT group were significantly shorter compared to the control group (P<0.05). PBMT caused a significant increase in the histological parameters in comparison to the control group (P<0.05). Moreover, PBMT significantly increased the breaking strength of the surgical scars produced in the skin of the PBMT group when compared to the control group (P<0.05).CONCLUSION:Photobiomodulation therapy may be useful in the management of wound infection through a significant bacterial growth inhibition and an acceleration of wound healing process.(AU)

Effect of irradiation with different laser wavelengths on oxidative stress of non-hepatectomized rats

PURPOSE: To assess the effect of two laser wavelengths, either separate or combined, on intact rat livers. METHOD: Nineteen male Wistar rats (200-300 g) were submitted to laser irradiation at 5 different sites on the liver surface.Wavelengths 660 and 780 nm were used, with a dose of irradiation of 60 J/cm2/site.The animals were divided into the groups:control (C) and animals irradiated with 660 nm laser (L1), with 780 nm laser (L2) or withboth wavelengths (L3).Mitochondrial function, mitochondrial swelling, and hepatocellular malondialdehyde (MDA) levels were determined.Data were analyzed by the Mann-Whitney test, with the level of significance set at 5%. RESULTS: There was a reduction of ADP-activated respiration (state 3) in group L1 compared to group C (p=0.0016), whereas the values of group L2 were similar to control.Group L3 also showed a reduction of state 3 (p=0.0159).There was a reduction of RCR in group L1 compared to control (p=0.0001) and to group L2 (p=0.0040).Mitochondrial swelling only differed between group L3 and control (p=0.0286).There was a increase in MDA levels in group L3 compared to control (p=0.0476) and to group L2 (p=0.0286) and in group L1 compared to group L2 (p=0.0132). CONCLUSION: Although laser irradiation reduced mitochondrial function,it did not interfere with the hepatocellular energy status.(AU)

The effects of low level laser therapy on Staphylococcus aureus infected third-degree burns in diabetic rats

PURPOSE:To characterize the effects of low-level laser (LLL) on third-degreeburn wounds which were infected with Staphylococcus aureus (S. aureus) in diabetic rats.METHODS:Thirty streptozotocin-induced diabetic rats were divided into two groups: the control and the LLL groups. Third-degree burns were induced using a heated metal rod, and then, were contaminated with S. aureus. The wounds in the LLL group were irradiated with a LLL (685nm) daily for five consecutive days, starting three days after the induction. The wound area was measured at 3, 5, 8, 14 and 21 days after burning. At the end of trial, the skin samples were harvested.RESULTS:Reduction in wound areas in the LLL and control groups were significantly different only on the 21st day (p<0.05). The mean bacterial numbers in the LLL group were significantly lower (p<0.05) than those in the control group.The number of macrophages, new blood vessels, fibroblast, and elevated collagen deposition in the LLL group significantly increased compared to the control group (p<0.05).The mean breaking strength of scars in the control group was significantly lower (p<0.05) than that of the LLL group.CONCLUSION:The low-level laser improved the healing of S. aureus third-degree burn infections in diabetic rats.(AU)

Histomorphometric analysis of the Achilles tendon of Wistar rats treated with laser therapy and eccentric exercise / Análise histomorfométrica do tendão calcanear comum de ratos Wistar tratado com laserterapia e exercício excêntrico

Low-level laser therapy is recommended for the treatment of tendinopathies despite the contradictory results related to the ideal dose of energy, wavelength and time of application. This study aimed to assess the effects of laser therapy and eccentric exercise on tendinopathy of the Achilles tendon of Wistar rats. Forty-eight adult male rats were randomly distributed into four groups (L= laser; E= eccentric exercise; LE = laser and eccentric exercise; and R= rest). Laser therapy (904nm/3J/cm2) and/or eccentric exercise (downhill walking; 15o incline treadmill; 12m/min; 50min/day) was started 24h after induction of unilateral tendinopathy and remained for 20 days. At 3, 7, 14 and 21 days after lesion induction, three rats from each group were euthanized and the tendons were collected for histological and morphometric analyses. There was no difference among groups or among times for the characteristics hemorrhage (p=0.4154), fibrinous adhesion formation (p=0.0712), and organization of collagen fibers (p=0.2583) and of the connective tissue (p=0.1046). For these groups, regardless of the time, eccentric exercise led to epitenon thickening (p=0.0204), which was lower in the group treated with laser therapy. Histological analysis revealed differences (p=0.0032) in the number of inflammatory cells over time. They were more numerous in the group that only exercised. This result was confirmed by morphometric analysis, which showed a significant interaction (groups x time) for this characteristic. Eccentric exercise increased (p=0.0014) the inflammatory infiltrate over time (3 and 21 days). However, association with laser therapy reduced inflammatory reaction. On the other hand, the combination of the treatments increased angiogenesis in morphometric (p=0.0000) and histological (p=0.0006) analyses compared with the other groups, while the isolated application of low-level laser reduced this characteristic over time. Animals maintained at rest presented the lowest amount (p=0.0000) of fibroblasts, according to the morphometric analysis. However, histological evaluation showed a significant group x time interaction (p=0.0024). Greater amounts of fibroblasts were observed in groups E, L and LE on the 7th, 14th and 21st days, respectively. The animals that received laser therapy and were exercised showed a greater (p=0.0000) amount of collagen fibers over time. Laser therapy at a dose of 3J and at a wavelength of 904nm, starting 24h after surgical induction of tendinopathy in Wistar rats, is suitable for angiogenesis and prevention of tendon thickening, which can be associated with the intensity of inflammatory process. When associated with eccentric exercise, the therapy has the advantage of increasing the amount of collagen fibers, reducing fibrinous adhesions and inflammatory infiltrate, despite prolonging angiogenesis. Therefore, eccentric exercise performed concomitantly with laser therapy improves the histological properties of the injured tendon.(AU)

O laser de baixa potência é recomendado para o tratamento das tendinopatias apesar dos resultados contraditórios no que se refere a dose de energia ideal, comprimento de onda e tempo de aplicação. O objetivo desse estudo foi avaliar os efeitos da laserterapia e do exercício excêntrico na tendinopatia do tendão calcanear comum de ratos Wistar. Foram utilizados 48 ratos machos adultos. Os animais foram distribuídos aleatoriamente em quatro grupos (L=laser; E=exercício excêntrico; LE=laser e exercício excêntrico, e R=repouso). Após 24h da indução de tendinopatia unilateral, foi iniciada laserterapia (904nm/3J/cm2) e/ou exercício excêntrico (caminhada em declive; esteira com 15o de inclinação; 12m/min; 50min/dia) que permaneceu por 20 dias. Aos 3, 7, 14 e 21 dias após indução da lesão, três ratos de cada grupo foram submetidos à eutanásia, e os tendões foram obtidos para análises histológica e morfométrica. Não houve diferença entre grupos nem entre tempos nas características hemorragia (p=0,4154), formação de aderência fibrinosa (p=0,0712) e organização das fibras colágenas (p=0,2583) e do tecido conjuntivo (p=0,1046). Considerando os grupos, independentemente do tempo, foi observado que o exercício excêntrico ocasionou espessamento do epitendão (p=0,0204), que foi menor no grupo submetido à laserterapia. A análise histológica revelou diferença (p=0,0032) na quantidade de células inflamatórias ao longo do tempo, sendo observado em maior quantidade no grupo apenas exercitado. Esse resultado foi confirmado pela análise morfométrica, que demonstrou haver interação significativa (grupos x tempo) nessa característica, na qual o exercício excêntrico aumentou (p=0,0014) o infiltrado inflamatório ao longo do tempo (3 e 21 dias), porém, quando associado à laserterapia, ocorreu redução da reação inflamatória. Por outro lado, a associação dos tratamentos ocasionou maior angiogênese observada nas análises morfométrica (p=0,0000) e histológica (p=0,0006), quando comparada com os demais grupos, enquanto a aplicação isolada do laser de baixa potência reduziu essa característica ao longo do tempo. Os animais mantidos em repouso foram os que apresentaram menor (p=0,0000) quantidade de fibroblastos, na análise morfométrica, porém na avaliação histológica foi observada interação significativa entre grupo e tempo (p=0,0024). Maior quantidade de fibroblastos foi observada nos grupos E, L e LE, no 7o, 14o e 21o dias, respectivamente. Os animais que receberam laserterapia e foram exercitados apresentaram maior (p=0,0000) quantidade de fibras colágenas ao longo do tempo. A laserterapia na dose de 3J e comprimento de onda de 904nm, iniciada 24h após indução cirúrgica de tendinopatia em ratos Wistar é adequada para angiogênese e para evitar o espessamento do tendão, que pode estar relacionado com a intensidade do processo inflamatório. Quando associada ao exercício excêntrico, a terapia apresenta a vantagem de aumentar a quantidade de fibras colágenas, reduzir a aderência fibrinosa e infiltrado inflamatório, apesar de prolongar a angiogênese. Portanto, o exercício excêntrico realizado concomitantemente com a laserterapia melhora as propriedades histológicas do tendão lesionado.(AU)