A novel technique of antimicrobial photodynamic therapy - aPDT using 1,9-dimethyl-methylene blue zinc chloride double salt-DMMB and polarized light on Staphylococcus aureus.

Antimicrobial Photodynamic Therapy (aPDT) is an alternative to conventional treatments of local infections such as the use of antibiotics, which may lead to the development of resistance. aPDT besides requiring the use of a photosensitiser also needs a light source do be carried out. In the search for efficient and low-cost procedure the use of multispectral polarized light (λ400-2000 nm) emerges as a possibility for the execution of aPDT. The use of a highly effective photosensitizer is also of great importance. 1,9-Dimethyl-Methylene Blue Zinc Chloride Double Salt - DMMB is a potent phenothiazine derivative that presents high photodynamic action due to its high lipophilicity as well as a greater quantum yield of Singlet oxygen and phototoxicity when compared to other Photosensitizers. The aim of this study was to assess, In Vitro, the efficacy of aPDT on Staphylococcus aureus (ATCC 25923) using different concentrations of DMMB associated to a Polarized light source (Bioptron®, 40 mW, ᴓ = 15.8 cm2) using different energy densities. Based on the IC50, 150 and 300 ng/mL of DMMB concentrations were chosen for this study. Twelve experimental groups were used: (Control, PLs, PSs and aPDTs). Serial dilutions (up to 10-8) of the bacterial inoculum were used and the DMMB was added using the two previously determined concentrations. After 5 min of preincubation the dilutions of the inoculum were illuminated by the polarized light source. Subsequently, 100 µL of each dilution, in triplicate, were inoculated into Petri dishes containing TSA medium and incubated in a bacteriological oven at 37 °C for 24-h and quantification of UFCs was done. The results showed significant exponential reduction (p < .0001) of 99.93% (150 ng/mL + LP 10 J/cm2) and 99.97% (300 ng/mL + LP 5 J/cm2) the CFU counts in comparison to non-illuminated control. The results of this study allow to conclude that aPDT carried out with 1,9-Dimethyl-Methylene Blue Zinc Chloride Double Salt-DMMB and a PL souce was efficacious on the reduction (99.97%), in vitro, of the bacterial counts of S. aureus.

Influence of laser therapy on the dynamic formation of extracellular matrix in standard second degree burns treated with bacterial cellulose membrane.

The present study aims to assess the influence of Aluminum-Gallium-Indium-Phosphide laser (AlGaInP laser, λ = 660 nm), whether or not in association with the application of a membrane of bacterial cellulose (Nexfill™), during recovery from induced second-degree burns at the dorsum of Wistar rats. (Rattus norvegicus, Wistar). Forty-eight animals have been distributed into four groups: Control (burns remained untreated), Group I (laser-treated), Group II (treated with Nexfill), and Group III (laser + Nexfill™). In addition to a morphological analysis, immunohistochemical analysis has been performed for type I collagen, type III collagen, fibronectin, and laminin. The Fisher's Test was used to assess differences among groups (p < 0,05). A larger amount of collagen type III was observed in Control, Group II and Group III when compared with Group I (p < 0,05). Group I and Group III have shown a greater collagen deposition when compared with Group II (p < 0,05), but the amount of collagen was similar in Group I, Group III, and Control. Group III has shown larger fibronectin amounts in comparison with Group II (p < 0,05). As regards laminin, Group I has shown a predominant discontinuity pattern on the basal lamina in comparison with Control, Group II, and Group III (p < 0,05). It is concluded that in this current study the laser when used alone (Group I) hasn't influenced collagen deposition neither has it acted on fiber pattern (fibril and/or reticular). Moreover, laser application hasn't accelerated the repair of wounds caused by inflicted second-degree burns.

Repair of surgical bone defects grafted with hydroxylapatite + ß-TCP and irradiated with λ=850 nm LED light.

The treatment of bone loss due to different etiologic factors is difficult and many techniques aim to improve the repair, including a wide range of biomaterials and recently, photobioengineering. This work aimed to assess by histological analysis the repair of bone defects grafted with biphasic synthetic micro-granular HA + ß-TCP associated with LED phototherapy. Forty rats were divided into 4 groups (Clot, LED, Biomaterial and LED + Biomaterial) each subdivided into 2 subgroups according to the time of animal death (15 and 30 days). Surgical bone defects were prepared on the femur of each animal with a trephine drill. In animals of the Clot group the defect was filled only by blood clot, in the LED group the defect filled with the clot was further irradiated. In the animals of Biomaterial and LED + Biomaterial groups the defect was filled by biomaterial and the last one was further irradiated (λ = 850 ± 10 nm, 150 mW, Φ ~ 0.5 cm2, 20 J/cm2 - session, 140 J/cm2- treatment) at 48-h intervals for 2 weeks. Following animal death, samples were taken and analyzed by light microscopy. Using the degree of maturation of the bone by assessment of the deposition/organization of the basophilic lines in the newly formed bone tissue, the LED + Biomaterial group was the one in a more advanced stage of bone repair process at the end of the experiment. It may be concluded that the use of LED phototherapy was effective in positively modulating the process of bone repair of bone defects in the femur of rats submitted or not to biomaterial grafting.

Repair of Surgical Bone Defects Grafted with Hydroxylapatite + ß-TCP and Irradiated with λ=850 nm LED Light

The treatment of bone loss due to different etiologic factors is difficult and many techniques aim to improve the repair, including a wide range of biomaterials and recently, photobioengineering. This work aimed to assess by histological analysis the repair of bone defects grafted with biphasic synthetic micro-granular HA + β-TCP associated with LED phototherapy. Forty rats were divided into 4 groups (Clot, LED, Biomaterial and LED + Biomaterial) each subdivided into 2 subgroups according to the time of animal death (15 and 30 days). Surgical bone defects were prepared on the femur of each animal with a trephine drill. In animals of the Clot group the defect was filled only by blood clot, in the LED group the defect filled with the clot was further irradiated. In the animals of Biomaterial and LED + Biomaterial groups the defect was filled by biomaterial and the last one was further irradiated (λ=850±10 nm, 150 mW, Φ ~ 0.5 cm2, 20 J/cm2 - session, 140 J/cm2- treatment) at 48-h intervals for 2 weeks. Following animal death, samples were taken and analyzed by light microscopy. Using the degree of maturation of the bone by assessment of the deposition/organization of the basophilic lines in the newly formed bone tissue, the LED + Biomaterial group was the one in a more advanced stage of bone repair process at the end of the experiment. It may be concluded that the use of LED phototherapy was effective in positively modulating the process of bone repair of bone defects in the femur of rats submitted or not to biomaterial grafting.

O tratamento de perdas ósseas devido a diferentes fatores etiológicos é difícil e muitas técnicas têm por objetivo melhorar o reparo incluindo o uso de uma ampla gama de biomateriais e, recentemente, a fotobioengenharia. Este trabalho teve como objetivo avaliar, por meio de análise histológica, o reparo de defeitos ósseos enxertados com HA bifásica micro-granular sintética + β -TCP associada à fototerapia LED. Quarenta ratos foram divididos em quatro grupos (Clot, LED, Biomaterial e LED + Biomaterial), subdivididos no dois subgrupos de acordo com o momento da morte (15 e 30 dias). Defeitos ósseos cirúrgicos foram criados em um fêmur de cada animal com uma broca trefina. Em animais do grupo coágulo, o defeito foi preenchido apenas pelo coágulo sanguíneo, no grupo de LED o defeito foi preenchido pelo coágulo e irradiado. Nos animais dos grupos do biomaterial e LED + biomaterial, os defeitos foram preenchidos com biomaterial e o último foi adicionalmente irradiado (λ=850±10 nm, 150 mW, Φ ~ 0.5 cm2, 20 J/cm2 - session, 140 J/cm2 -tratamento) a cada 48 h por duas semanas. Após a morte dos animais, amostras foram colhidas e analisadas por microscopia de luz, usando o grau de maturação do osso como marcador (deposição/organização das linhas basofílicas) no tecido ósseo neoformado. O grupo de LED + biomaterial apresentou processo de reparação mais avançado ao fim do experimento. Pode-se concluir que o uso da fototerapia LED foi eficaz na modulação positiva do processo de reparo ósseo de defeitos ósseos no fêmur de ratos submetidos ou não a enxerto com biomaterial.

Raman spectroscopic study of the repair of surgical bone defects grafted or not with biphasic synthetic micro-granular HA + ß-calcium triphosphate irradiated or not with λ850 nm LED light.

The handling of bone losses due to different etiologic factors is difficult and many techniques are aim to improve repair, including a wide range of biomaterials and, recently, photobioengineering. This work aimed to assess, through Raman spectroscopy, the level of bone mineralization using the intensities of the Raman peaks of both inorganic (~960, ~1,070, and 1,077 cm(-1)) and organic (~1,454 and ~1,666 cm(-1)) contents of bone tissue. Forty rats were divided into four groups each subdivided into two subgroups according to the time of sacrifice (15 and 30 days). Surgical bone defects were made on the femur of each animal with a trephine drill. On animals of group clot, the defect was filled only by blood clot, on group LED, the defect filled with the clot was further irradiated. On animals of groups biomaterial and LED + biomaterial, the defect was filled by biomaterial and the last one was further irradiated (λ850 ± 10 nm, 150 mW, Φ ~ 0.5 cm(2), 20 J/cm(2)-session, 140 J/cm(2)-treatment) at 48-h intervals and repeated for 2 weeks. At both 15th and 30th days following sacrifice, samples were taken and analyzed by Raman spectroscopy. At the end of the experimental time, the intensity of hydroxyapatite (HA) (~960 cm(-1)) were higher on group LED + biomaterial and the peaks of both organic content (~1,454 and ~1,666 cm(-1)) and transitional HA (~1,070 and ~1,077 cm(-1)) were lower on the same group. It is concluded that the use of LED phototherapy associated to biomaterial was effective in improving bone healing on bone defects as a result of the increasing deposition of HA measured by Raman spectroscopy.

Raman ratios on the repair of grafted surgical bone defects irradiated or not with laser (λ780 nm) or LED (λ850 nm).

This work aimed to assess biochemical changes associated to mineralization and remodeling of bone defects filled with Hydroxyapatite+Beta-Beta-tricalcium phosphate irradiated or not with 2 light sources. Ratios of intensities, band position and bandwidth of selected Raman peaks of collagen and apatites were used. Sixty male Wistar rats were divided into 6 groups subdivided into 2 subgroups (15th and 30th days). A standard surgical defect was created on one femur of each animal. In 3 groups the defects were filled with blood clot (Clot, Clot+Laser and Clot+LED groups) and in the remaining 3 groups the defects were filled with biomaterial (Biomaterial, Biomaterial+Laser and Biomaterial+LED groups). When indicated, the defects were irradiated with either Laser (λ780 nm, 70 mW, Φ∼0.4 cm(2)) or LED (λ850±10 nm, 150 mW, Φ∼0.5 cm(2)), 20 J/cm(2) each session, at 48 h intervals/2 weeks (140 J/cm(2) treatment). Following sacrifice, bone fragments were analyzed by Raman spectroscopy. Statistical analysis (ANOVA General Linear Model, p<0.05) showed that both grafting and time were the variables that presented significance for the ratios of ∼1660/∼1670 cm(-1) (collagen maturation), ∼1077/∼854 cm(-1) (mineralization), ∼1077/∼1070 cm(-1) (carbonate substitution) and the position of the ∼960 cm(-1) (bone maturation). At 30th day, the ratios indicated an increased deposition of immature collagen for both Clot and Biomaterial groups. Biomaterial group showed increased collagen maturation. Only collagen deposition was significantly dependent upon irradiation independently of the light source, being the amount of collagen I increased in the Clot group at the end of the experimental time. On the other hand, collagen I deposition was reduced in biomaterial irradiated groups. Raman ratios of selected protein matrix and phosphate and carbonate HA indicated that the use of biphasic synthetic micro-granular HA+Beta-TCP graft improved the repair of bone defects, associated or not with Laser or LED light, because of the increasing deposition of HA.

Influence of the λ780nm laser light on the repair of surgical bone defects grafted or not with biphasic synthetic micro-granular hydroxylapatite+Beta-Calcium triphosphate.

The treatment of bone loss due to different etiologic factors is difficult and many techniques aim to improve repair, including a wide range of biomaterials and, recently, photobioengineering. This work aimed to assess, through histological analysis The aim of this study was to assess, by light microscopy, the repair of bone defects grafted or not with biphasic synthetic micro-granular Calcium hydroxyapatite (HA)+Beta-TCP associated or not with Laser phototherapy - LPT (λ780nm). Forty rats were divided into 4 groups each subdivided into 2 subgroups according to the time of sacrifice (15 and 30days). Surgical bone defects were made on femur of each animal with a trephine drill. On animals of Clot group the defect was filled only by blood clot, on Laser group the defect filled with the clot was further irradiated. On animals of Biomaterial and Laser+Biomaterial groups the defect was filled by biomaterial and the last one was further irradiated (λ780nm, 70mW, spot size∼0.4cm(2), 20J/cm(2)-session, 140J/cm(2)-treatment) in four points around the defect at 48-h intervals and repeated for 2weeks. At both 15th and 30th days following sacrifice, samples were taken and analyzed by light microscopy. Many similarities were observed histologically between groups on regards bone reabsorption and neoformation, inflammatory infiltrate and collagen deposition. The criterion degree of maturation, marked by the presence of basophilic lines, indicated that the use of LPT associated with HA+Beta TCP graft, resulted in more advanced stage of bone repair at the end of the experiment.

Raman study of the repair of surgical bone defects grafted with biphasic synthetic microgranular HA + ß-calcium triphosphate and irradiated or not with λ780 nm laser.

The treatment of bone loss due to different etiologic factors is difficult, and many techniques aim to improve repair, including a wide range of biomaterials and, recently, photobioengineering. This work aimed to assess, through Raman spectroscopy, the level of bone mineralization using the intensities of the Raman peaks of both inorganic (∼ 960, ∼ 1,070, and ∼ 1,077 cm(-1)) and organic (∼ 1,454 and ∼ 1,666 cm(-1)) contents of bone tissue. Forty rats were divided into four groups each subdivided into two subgroups according to the time of killing (15 and 30 days). Surgical bone defects were made on femur of each animal with a trephine drill. On animals of group Clot, the defect was filled only by blood clot; on group Laser, the defect filled with the clot was further irradiated. On animals of groups Biomaterial and Laser + Biomaterial, the defect was filled by biomaterial and the last one was further irradiated (λ780 nm, 70 mW, Φ âˆ¼ 0.4 cm(2), 20 J/cm(2) session, 140 J/cm(2) treatment) in four points around the defect at 48-h intervals and repeated for 2 weeks. At both 15th and 30th day following killing, samples were taken and analyzed by Raman spectroscopy. At the end of the experimental time, the intensities of both inorganic and organic contents were higher on group Laser + Biomaterial. It is concluded that the use of laser phototherapy associated to biomaterial was effective in improving bone healing on bone defects as a result of the increasing deposition of calcium hydroxyapatite measured by Raman spectroscopy.

New bone formation around implants inserted on autologous and xenografts irradiated or not with IR laser light: a histomorphometric study in rabbits.

Use of biomaterials and light on bone grafts has been widely reported. This work assessed the influence of low-level laser therapy (LLLT) on bone volume (BV) and bone implant contact (BIC) interface around implants inserted in blocks of bovine or autologous bone grafts (autografts), irradiated or not, in rabbit femurs. Twenty-four adult rabbits were divided in 8 groups: AG: autograft; XG: xenograft; AG/L: autograft + laser; XG/L: xenograft + laser; AG/I: autograft + titanium (Ti) implant; XG/I: xenograft + Ti implant; AG/I/L: autograft + Ti implant + laser; and XG/I/L: xenograft + Ti implant + laser. The animals received the Ti implant after incorporation of the grafts. The laser parameters in the groups AG/L and XG/L were λ=780 nm, 70 mW, CW, 21.5 J/cm 2 , while in the groups AG/I/L and XG/I/L the following parameters were used: λ=780 nm, 70 mW, 0.5 cm 2 (spot), 4 J/cm 2 per point (4), 16 J/cm 2 per session, 48 h interval × 12 sessions, CW, contact mode. LLLT was repeated every other day during 2 weeks. To avoid systemic effect, only one limb of each rabbit was double grafted. All animals were sacrificed 9 weeks after implantation. Specimens were routinely stained and histomorphometry carried out. Comparison of non-irradiated and irradiated grafts (AG/L versus AG and XG/L versus XG) showed that irradiation increased significantly BV on both grafts (p=0.05, p=0.001). Comparison between irradiated and non-irradiated grafts (AG/I/L versus AG/I and XG/I/L versus XG/I) showed a significant (p=0.02) increase of the BIC in autografts. The same was seen when xenografts were used, without significant difference. The results of this investigation suggest that the use of LLLT is effective for enhancing new bone formation with consequent increase of bone-implant interface in both autologous grafts and xenografts.

New Bone Formation around Implants Inserted on Autologous and Xenografts Irradiated or not with IR Laser Light: A Histomorphometric Study in Rabbits

Use of biomaterials and light on bone grafts has been widely reported. This work assessed the influence of low-level laser therapy (LLLT) on bone volume (BV) and bone implant contact (BIC) interface around implants inserted in blocks of bovine or autologous bone grafts (autografts), irradiated or not, in rabbit femurs. Twenty-four adult rabbits were divided in 8 groups: AG: autograft; XG: xenograft; AG/L: autograft + laser; XG/L: xenograft + laser; AG/I: autograft + titanium (Ti) implant; XG/I: xenograft + Ti implant; AG/I/L: autograft + Ti implant + laser; and XG/I/L: xenograft + Ti implant + laser. The animals received the Ti implant after incorporation of the grafts. The laser parameters in the groups AG/L and XG/L were λ=780 nm, 70 mW, CW, 21.5 J/cm 2 , while in the groups AG/I/L and XG/I/L the following parameters were used: λ=780 nm, 70 mW, 0.5 cm 2 (spot), 4 J/cm 2 per point (4), 16 J/cm 2 per session, 48 h interval × 12 sessions, CW, contact mode. LLLT was repeated every other day during 2 weeks. To avoid systemic effect, only one limb of each rabbit was double grafted. All animals were sacrificed 9 weeks after implantation. Specimens were routinely stained and histomorphometry carried out. Comparison of non-irradiated and irradiated grafts (AG/L versus AG and XG/L versus XG) showed that irradiation increased significantly BV on both grafts (p=0.05, p=0.001). Comparison between irradiated and non-irradiated grafts (AG/I/L versus AG/I and XG/I/L versus XG/I) showed a significant (p=0.02) increase of the BIC in autografts. The same was seen when xenografts were used, without significant difference. The results of this investigation suggest that the use of LLLT is effective for enhancing new bone formation with consequent increase of bone-implant interface in both autologous grafts and xenografts.

O uso de biomateriais e luz em enxertos ósseos têm sido relatados. Esse trabalho avaliou a influência do laser baixa potência - LBP no volume ósseo (VO) e superfície de contato osso-implante (COI) ao redor de implantes dentários inseridos em blocos de enxerto bovino ou autólogos incorporados, irradiados ou não, em fêmures de coelho. Vinte e quatro coelhos adultos foram divididos em 8 grupos: EA: enxerto autólogo; EX: enxerto xenógeno; EA/L: enxerto autólogo + laser; EX/L: enxerto xenógeno + laser; EA/I: enxerto autólogo + implante; EX/I: enxerto xenógeno + implante; EA/I/L: enxerto autólogo + implante de titânio + laser; EX/I/L: enxerto xenógeno + implante de titânio + laser. Os animais receberam um implante de titânio após a incorporação dos enxertos. Os parâmetros de laser nos grupos EA/L e EX/L foram λ =780 nm, 70 mW, CW, 21,5 J/cm 2 ), enquanto que nos grupos EA/I/L e EX/I/L os seguintes parâmetros de laser foram utilizados: λ =780 nm, 70 mW, 0,5 cm 2 (spot), 4 J/cm 2 por ponto (4), 16 J/cm 2 por sessão, intervalo de 48 h × 12 sessões, CW, modo contato. O LBP foi repetido a cada 48 h (2 semanas). Para evitar efeito sistêmico apenas um membro de cada coelho foi duplamente enxertado. Todos os animais foram sacrificados 9 semanas após o implante. Os espécimes foram corados rotineiramente e histomorfometria foi realizada. A comparação dos enxertos não-irradiados e irradiados (EA/L versus EA e EX/L versus EX) mostrou que a irradiação aumentou significantemente (p=0,02) o VO para ambos os tipos de enxertos (p=0,05, p=0,001). A comparação dos enxertos não-irradiados e irradiados (EA/I/L versus EA/I e EX/I/L versus EX/I) mostrou um aumento significante (p=0,02) do COI nos enxertos autólogos e xenógenos sem diferença estatística. Os resultados desta investigação sugerem que o uso de LBP é efetivo para aumentar a neoformação óssea com consequente aumento do COI em enxertos autólogos e xenógenos.

Differential diagnosis between experimental endophthalmitis and uveitis in vitreous with Raman spectroscopy and principal components analysis.

Raman spectroscopy has been used for the diagnosis of various eye diseases. A diagnostic tool based on Raman spectroscopy has been developed to discriminate endophthalmitis from uveitis in vitreous tissues of rabbits' eyes in vitro. Twenty-two New Zealand rabbits suffering from endophthalmitis induced by Staphylococcus aureus (n=10), non-infectious uveitis induced by lipopolysaccharide from Escherichia coli (LPS) (n=10 animals) and control (n=2) were included in the study. After eye inoculation, vitreous tissues were dissected and a fragment was submitted to dispersive Raman spectroscopy using near-infrared laser excitation (830 nm, 100 mW) and spectrograph/CCD camera for detection of Raman signal with integration time of 50 s. A routine was developed to classify the spectra of endophthalmitis and uveitis using principal components analysis (PCA) and Mahalanobis distance. The mean Raman spectra of tissues with uveitis and endophthalmitis showed several bands in the region of 800-1800 cm(-1), which have been attributed to nucleic acids, amino acids and proteins from inflamed tissue and proliferating bacteria. The bands at 1004, 1258, 1339, 1451 and 1635 cm(-1) showed statistically significant differences between both diseases. It was observed that principal components PC1, PC3 and PC4 showed statistically significant differences for the two tissue types, indicating that these PCs can be used to discriminate between the two groups. The diagnostic model showed 94% sensitivity, 95% specificity and 95% accuracy using PC3×PC4. The Raman spectroscopy technique has been shown to be useful in differentiating uveitis and endophthalmitis in vitreous tissues in vitro, and these results may be clinically relevant for differentiating vitreous tissues to optimise the diagnosis of inflammatory and infectious vitreoretinal diseases.

Tooth movement after infrared laser phototherapy: clinical study in rodents.

OBJECTIVES: The aim of this research was to investigate the influence of low-power laser on tooth movement in rats. BACKGROUND: Tooth movement is closely related to the process of bone remodeling. The biologic result, with the application of a force to the tooth, is bone absorption on the pressure side and neoformation on the traction side of the alveolar bone. The laser photobiomodulation is capable of providing an increase in cellular metabolism, blood flow, and lymphatic drainage. METHODS: Thirty young-adult male Wistar rats weighing between 250 and 300 g were divided into two groups, control and experimental, containing 15 animals each. The animals received orthodontic devices calibrated to release a force of 40 g/F, with the purpose of moving the first upper molar mesially. Low-intensity laser, wavelength 790 nm, was used in the experimental group; the dose was 4.5 J/cm(2) per point, mesial and distal, on the palatal side, 11 J/cm(2) on the buccal side, and this procedure was repeated every 48 h, totaling nine applications. The active movement was clinically evaluated after 7, 13, and 19 days. RESULTS AND CONCLUSION: The results showed no statistically significant difference, p = 0.079 (T0-T7), p = 0.597 (T7-T13), and p = 0.550 (T13-T19) between the laser and control groups on the amount of tooth movement in the different times evaluated. It may be concluded that laser phototherapy, with the parameters in the present study, did not significantly increase the amount of tooth displacement during induced orthodontic movement in rodents.

Influence of the use of laser phototherapy (lambda660 or 790 nm) on the survival of cutaneous flaps on diabetic rats.

OBJECTIVE: The aim of this study was to assess and compare the effects of laser phototherapy (LPT) on cutaneous flaps on diabetic rats. BACKGROUND: Diabetes mellitus is characterized by high blood glucose levels. Its main complications are delayed wound healing, an impaired blood supply, and a decrease in collagen production. Cutaneous flaps are routinely used in several surgical procedures, and most failures are related to poor blood supply. LPT has been studied using several healing models. ANIMALS AND METHODS: Twelve Wistar rats were randomized into three groups: group 1 (G1; diabetic animals without treatment), group 2 (G2; diabetic animals irradiated with lambda680 nm), and group 3 (G3; diabetic animals irradiated with lambda790 nm). Diabetes was induced with streptozotocin. A 2- x 8-cm cutaneous flap was raised on the dorsum of each animal, and a plastic sheet was introduced between the flap and the bed to cause poor blood supply. Nonirradiated animals acted as controls. The dose per session was 40 J/cm(2). Laser light was applied transcutaneously and fractioned on 16 contact points at the wound margins (16 x 2.5 J/cm(2)). Animal death occurred on day 8 after surgery. Specimens were taken, processed, cut, stained with eosin (HE) and sirius red, and underwent histological analysis. RESULTS: It is shown that accute inflammation was mostly discrete for G3. Chronic inflammation was more evident for G2. Fibroblast number was higher for G3. Angiogenesis was more evident for G3. Necrosis was more evident for G2. Statistical analysis among all groups showed significant differences (p = 0.04) on the level of acute inflammation between G1 and G3, tissue necrosis between G1 and G2 (p = 0.03), chronic inflammation between (p = 0.04), fibroblastic proliferation between G2 and G3 (p = 0.05), and neovascularization between G2 and G3 (p = 0.04). CONCLUSION: LPT was effective in increasing angiogenesis as seen on irradiated subjects and was more pronounced when IR laser light was used.

Bone repair following bone grafting hydroxyapatite guided bone regeneration and infra-red laser photobiomodulation: a histological study in a rodent model.

The aim of the investigation was to assess histologically the effect of laser photobiomodulation (LPBM) on a repair of defects surgically created in the femurs of rats. Forty-five Wistar rats were divided into four groups: group I (control); group II (LPBM); group III (hydroxyapatite guided bone regeneration; HA GBR); group IV (HA GBR LPBM). The animals in the irradiated groups were subjected to the first irradiation immediately after surgery, and it was repeated every day for 2 weeks. The animals were killed 15 days, 21 days and 30 days after surgery. When the groups irradiated with implant and membrane were compared, it was observed that the repair of the defects submitted to LPBM was also processed faster, starting from the 15th day. At the 30th day, the level of repair of the defects was similar in the irradiated groups and those not irradiated. New bone formation was seen inside the cavity, probably by the osteoconduction of the implant, and, in the irradiated groups, this new bone formation was incremental. The present preliminary data seem to suggest that LPMB therapy might have a positive effect upon early wound healing of bone defects treated with a combination of HA and GBR.

Infrared laser light further improves bone healing when associated with bone morphogenetic proteins and guided bone regeneration: an in vivo study in a rodent model.

OBJECTIVE: This study assessed histologically the effect of laser photobiomodulation on the repair of surgical defects created in the femurs of Wistar rats treated or not treated with bone morphogenetic proteins (BMPs) and organic bovine bone graft. BACKGROUND DATA: This paper is part of an ongoing series of works in which biomaterials and/or guided bone regeneration (GBR) are used in association with laser photobiomodulation. Several previous reports from our group have shown that the use of laser photobiomodulation improves the treatment of bone defects. MATERIALS AND METHODS: Forty-eight adult male Wistar rats were divided into four randomized groups: group 1 (controls, n = 12); group 2 (laser photobiomodulation, n = 12); group 3 (BMPs + organic bovine bone graft + GBR, n = 12); and group 4 (BMPs + organic bovine bone graft + GBR + laser photobiomodulation, n = 12). The irradiated groups received seven irradiations every 48 h, the first immediately after the surgical procedure. Laser photobiomodulation (830 nm, 40 mW, CW, phi approximately 0.6 mm) consisted of a total of 16 J/cm2 per session at four points (4 J/cm2 each) equally spaced around the periphery of the defect. The animals were sacrificed after 15, 21, and 30 d, and the specimens were routinely embedded in wax and stained with hematoxylin and eosin and Sirius red stains and analyzed under light microscopy. RESULTS: The results showed histological evidence of increased deposition of collagen fibers (at 15 and 21 d), as well as an increased amount of well-organized bone trabeculi at the end of the experimental period (30 d) in irradiated animals compared to non-irradiated controls. CONCLUSION: We concluded that the use of laser photobiomodulation in association with BMPs, organic bovine bone grafts, and GBR increases the positive biomodulating effects of laser energy.

Infrared laser light further improves bone healing when associated with bone morphogenic proteins: an in vivo study in a rodent model.

OBJECTIVE: This study assessed histologically the effect of laser photobiomodulation (LPBM) on the repair of surgical defects created in the femurs of Wistar rats treated or not treated with bone morphogenic proteins (BMPs) and organic bovine bone graft. BACKGROUND DATA: This paper is part of an ongoing series of works in which biomaterials are used in association with LPBM. Several previous reports by our group have shown that the use of laser photobiomodulation improves the treatment of bone defects. MATERIALS AND METHODS: Forty-eight adult male Wistar rats were divided into four randomized groups: group I (control, n = 12); group II (LPBM, n = 12); group III (BMPs + organic bovine bone graft, n = 12); and group IV (BMPs + organic bovine bone graft + LPBM, n = 12). The irradiated groups received seven irradiations every 48 h, beginning immediately after the surgical procedure. The laser therapy (lambda = 830 nm, 40 mW CW, varphi = 0.6 mm) consisted of 16 J/cm(2) per session divided equally over four points (4 J/cm(2) each) around the defect. The subjects were sacrificed after 15, 21, and 30 d, and the specimens were routinely embedded in wax, stained with hematoxylin and eosin and sirius red, and analyzed under light microscopy. RESULTS: The results showed histological evidence of increased deposition of collagen fibers (at 15 and 21 d), as well as an increased amount of well-organized bone trabeculae at the end of the experimental period (30 d) in the irradiated animals versus the non-irradiated controls. CONCLUSION: The use of LPBM with BMPs and organic bovine bone grafts increases the positive biomodulating effects of laser light.

Use of the CO(2) laser on orthodontic patients suffering from gingival hyperplasia.

OBJECTIVE: The present study aimed to assess the effect of the use of the CO(2) laser on the treatment of gingival hyperplasia in orthodontic patients wearing fixed appliances. BACKGROUND DATA: Gingival hyperplasia is a condition very frequent in patients undergoing fixed orthodontic treatment. Amongst the treatments available for this is the use of surgical lasers. METHODS: Ten patients entered this study and signed an informed consent. Seventy-five anterior teeth with gingival hyperplasia were selected for laser surgery. Prior to surgery, the length of the crowns were measured using a digital caliper, and depth of the pocket was probed. The hyperplasic gingival was removed with a CO(2) laser under local anesthesia. Immediately after surgery, measurement of the length of the crowns and probing were carried out and were repeated. RESULTS: The results were statistically analyzed and significant differences were detected regarding the length of the crown (p = 0.000) and depth of the gingival sulcus (p = 0.000). CONCLUSION: It is concluded that the use of the CO(2) laser was effective in the treatment of gingival hyperplasia.

Photoengineering of bone repair processes.

OBJECTIVE: This paper aims to report the state of the art with respect to photoengineering of bone repair using laser therapy. BACKGROUND DATA: Laser therapy has been reported as an important tool to positively stimulate bone both in vivo and in vitro. These results indicate that photophysical and photochemical properties of some wavelengths are primarily responsible for the tissue responses. The use of correct and appropriate parameters has been shown to be effective in the promotion of a positive biomodulative effect in healing bone. METHODS: A series of papers reporting the effects of laser therapy on bone cells and tissue are presented, and new and promising protocols developed by our group are presented. RESULTS: The results of our studies and others indicate that bone irradiated mostly with infrared (IR) wavelengths shows increased osteoblastic proliferation, collagen deposition, and bone neorformation when compared to nonirradiated bone. Further, the effect of laser therapy is more effective if the treatment is carried out at early stages when high cellular proliferation occurs. Vascular responses to laser therapy were also suggested as one of the possible mechanisms responsible for the positive clinical results observed following laser therapy. It still remains uncertain if bone stimulation by laser light is a general effect or if the isolate stimulation of osteoblasts is possible. CONCLUSION: It is possible that the laser therapy effect on bone regeneration depends not only on the total dose of irradiation, but also on the irradiation time and the irradiation mode. The threshold parameter energy density and intensity are biologically independent of one another. This independence accounts for the success and the failure of laser therapy achieved at low-energy density levels.

Laser therapy improves healing of bone defects submitted to autologous bone graft.

OBJECTIVE: The aim of the present study was to assess histologically the effect of low-level laser thrapy (LLLT) (lambda 830 nm) on the healing of bone defects associated with autologous bone graft. BACKGROUND DATA: LLLT has been used on the modulation of bone healing because of the photo-physical and photochemical properties of some wavelengths. The use of correct and appropriate parameters has been shown to be effective in the promotion of a positive biomodulative effect on the healing bone. METHODS: Sixty male Wistar rats were divided into four groups: G1 (control), G2 (LLLT on the surgical bed), G3 (LLLT on the graft), and G4 (LLLT on both the graft and the surgical bed). The dose per session was 10 J/cm(2), and it was applied to the surgical bed (G2/G4) and on the bone graft (G3/G4). LLLT was carried out every other day for 15 days (lambda 830 nm, phi = 0.5 cm(2), 50 Mw, 10 J/cm(2)). The dose was fractioned in four points. The animals were sacrificed 15, 21, and 30 days after surgery; specimens were taken and routinely processed (wax, cut, and stain with H&E and Sirius red stains). Light microscopic analysis was performed by a pathologist. RESULTS: In the groups in which the LLLT was used trans-operatively on the surgical bed (G2/G4), bone remodeling was both quantitatively and qualitatively more evident when compared to subjects of groups G1 and G3. CONCLUSION: The present study indicates that the use of LLLT trans-operatively resulted in a positive biomodulative effect on the healing of bone defects associated with autologous bone grafts.

Polarized light (400-2000 nm) and non-ablative laser (685 nm): a description of the wound healing process using immunohistochemical analysis.

OBJECTIVE: This study aimed to describe, through morphologic and cytochemical analysis, the healing process of wounds submitted (or not) to laser therapy (lambda685 nm) or polarized light (lambda400-2000 nm). BACKGROUND DATA: There are many reports on different effects of several types of phototherapies on the treatment of distinct conditions, amongst them, on wound healing. Laser therapy and the use of polarized light are still controversial despite successive reports on their positive effects on several biological processes. METHODS: Thirty male Wistar rats, approximately 4 months old, were used, and standardized excisional wounds were created on their dorsum. The wounds were irradiated in four equidistant points with laser light or illuminated with polarized light, both with doses of 20 or 40 J/cm2. Group 1 acted as untreated controls. Animals were irradiated every 48 h during 7 days, starting immediately after surgery, and were humanely killed on the 8th post-operative day. Specimens were taken and routinely processed and stained with H&E, and for descriptive analysis of myofibroblasts and collagen fibers, the specimens were imunnomarked by smooth muscle alpha-actin and picrosirius stain. RESULTS: Control specimens showed the presence of ulceration, hyperemia, discrete edema, intense, and diffuse inflammation, collagen deposition was irregular, and myofibroblasts were seen parallel to the wound margins. Wounds treated by laser therapy with a dose of 20 J/cm2 showed mild hyperemia, inflammation varied from moderate to intense, the number of fibroblasts was large, and the distribution of collagen fibers was more regular. Increasing the dose to 40 J/cm2 evidenced exuberant neovascularization, severe hyperemia, moderate to severe inflammation, large collagen deposition, and fewer myofibroblasts. On subjects illuminated with polarized light with a dose of 20 J/cm2, mild to moderate hyperemia was detectable, and collagen matrix was expressive and unevenly distributed; a larger number of myofibroblasts was present and no re-epithelialization was seen. Increasing the dose resulted in mild to moderate hyperemia, no re-epithelialization was seen, edema was discrete, and inflammation was moderate. CONCLUSION: The use of 685-nm laser light or polarized light with a dose of 20 J/cm2 resulted in increased collagen deposition and better organization on healing wounds, and the number of myofibroblast was increased when polarized light is used.