Photobiomodulation effects on synovial morphology, iNOS gene, and protein expression in a model of acute inflammation.

PURPOSE: To evaluate morphological aspects and inducible nitric oxide synthase (iNOS) gene and protein expression in a model of acute inflammation. METHODS: Thirty-six female Wistar rats were assigned into three groups: control (saline, n = 12), sham (arthritis, n = 12), and PBM (arthritis and photobiomodulation, n = 12). Arthritis induction was performed with 200 µg of intra-articular Zymosan in sham and PBM animals. PBM was performed 24 h after induction with a laser device (λ = 808 nm, 25 mW of nominal power, fluence of 20 J/cm2, beam area of 0.02 mm2, time of 33 s, total energy of 0.825 J) with punctual and single dose application. Morphological analysis of joint structure (HE) and immunohistochemistry (anti-iNOS antibody) were performed on knee samples, and synovial tissue was submitted to RNA extraction, cDNA synthesis and gene expression analysis by quantitative polymerase chain reaction. Statistical analyses were performed with p < 0.05. RESULTS: It was observed an increase in the thickness of the synovial lining epithelium and inflammatory infiltrate in sham compared to PBM. Gene expression analysis showed higher iNOS expression in PBM, and iNOS protein expression decreased in PBM compared to sham. CONCLUSIONS: Photobiomodulation decreased inflammation in PBM animals, upregulated iNOS gene expression, however down egulated protein expression compared to sham.

Histomorphometric Changes of the Masseter Muscle of Rats After a Single Injection of Botulinum Toxin Type A.

BACKGROUND: It has been reported that botulinum toxin type A (BoNT-A) produces structural changes in masticatory muscles. However, not all histomorphometric parameters affected by BoNT-A parameters have been assessed. This study investigated the histomorphometric changes in the masseter muscle of rats after a single injection of BoNT-A. METHODS: Forty-four adult animals were randomly divided into control group (n = 22) and BoNT-A group (n = 22). Controls received a single dose of 0.14 mL/kg of saline in masseter muscles, and the BoNT-A group received a 7 U/Kg of BoNT-A. The groups received the same volume of injected substances. Animals were sacrificed on 7th (n = 5), 14th (n = 5), 21st (n = 5), 28th (n = 4) and 90th (n = 3) days post-treatment. Histological masseter tissue slides were obtained from hematoxylin-eosin treatment and analyzed in optical microscopy regarding muscle cross-sectional area, amount of connective tissue and quantity and diameter of myocytes. For statistical analysis, generalized linear models were used to compare the data (ANOVA). In all test, the significance level of 5% was set. RESULTS: BoNT-A values of cross-sectional area of the masseter muscle were significantly lower than controls (p < 0.01) throughout the study. Regarding myocytes quantity, BoNT-A subgroups presented higher values than controls (p < 0.0001) since the 14th day until the end of the study; however, the diameter of myocytes was smaller in all BoNT-A subgroups (p < 0.0001) in all assessment points. The amount of connective tissue was higher in BoNT-A subgroups (p < 0.0001) throughout the study. CONCLUSION: A single injection of BoNT-A altered the structure of masseter muscle of rats, regarding its histomorphometric parameters. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Arnica montana L. associated with microcurrent accelerates the dermis reorganisation of skin lesions.

The aim of this study was to test the effect of electrical stimulation in association with topical Arnica montana gel on organisational changes in the dermis during tissue repair. An experimental rat incisional skin lesion was used for the study. This involved making an incisional lesion on the dorsum of the animals using a scalpel. Ninety-six animals were used divided into the following groups: control (C), microcurrent (MC); topical treatment with Arnica montana gel (ARN); the ARN + microcurrent (ARN + MC). Treatments were administered daily, and injured tissue samples were collected and processed on Days 2, 6 and 10 for dermis analyses. Myeloperoxidase levels were greater in control than in treatment groups on Days 2 and 6. F4/80 expression was similar among all treatment groups and greater than that in control on Day 2. On Day 6, the expression of vascular endothelial growth factor was higher in the MC group than that in other groups, whereas transforming growth factor-ß expression increased in the MC and ARN + MC groups on Day 10. The expression of matrix metalloproteinase-2 was higher in the ARN + MC group when compared with other groups on Day 10. Expression levels of collagen I were increased in the ARN and ARN + MC groups when compared with control and MC groups on Day 6, while expression of collagen III was enhanced in MC, ARN, and ARN + MC groups when compared with the control. The protocol combining microcurrent with topical application of ARN reduces the inflammatory process, increases myofibroblasts proliferation and decreases the presence of macrophages in the dermis during skin repair in rats.

Maxillary and dental development in the offspring of protein-restricted female rats.

Nutritional restriction during developmental periods impairs organ physiology. Female rats were subjected to protein restriction during pregnancy and lactation to analyze dental and maxillary development. Four exposure groups were considered: normal-protein diet during pregnancy and lactation (NP, 17% casein), low-protein diet during lactation (LP-L, 6% casein), low-protein diet during pregnancy and lactation (LP), and low-protein diet during pregnancy (LP-G). Maxillae from 15-day-old male pups were collected. All protein-restricted groups presented increased dentin thickness and reduced alveolar bone area. When protein restriction was applied during both gestation and lactation (LP), harmful effects were observed in the form of loss of protective OPG (osteoprotegerin) in tooth epithelium-mesenchyme, due to higher RANKL expression, delay in odontoblast maturation, less dental pulp vascularity, reduction in amount of alveolar bone, and less matrix mineralization. In the LP-L group, effects of protein restriction seemed less harmful, and despite less alveolar bone, the enhancement in BMP-7, VEGF, and RANKL seems a compensatory signal to maintain maxillary osteogenesis. In LP-G animals, Dspp expression was higher, suggesting a delay in odontoblast maturation or expression recuperation. In conclusion, maternal protein restriction affects dental and maxillary development. A low-protein diet only in gestation allows for normal development. A low-protein diet during gestation-lactation results in impaired odontogenesis that may increase susceptibility of dental anomalies.

Morphofunctional and immunological cardiac evaluation of protein restriction on rat offspring.

Cardiovascular diseases affect a large part of the population during adulthood. Several epidemiological and experimental studies have demonstrated an association between maternal protein restriction and a marked risk of developing heart disease during early intrauterine and postnatal life. Maternal nutritional conditions act by modulating the microenvironment, thus favoring adaptive processes of the fetal organism for development to occur. However, the physiological profile is established in this period and its effects can be observed in the long term. In the present study, the cardiac muscles of 15-day-old offspring of Wistar rats subjected to maternal protein restriction during pregnancy and/or lactation were evaluated to identify possible cardiac changes relevant for heart disease in adulthood. The offspring of restricted female rats during pregnancy had a lower birth weight. Male offspring subjected to restriction during pregnancy and lactation showed an increase in the concentration of H2O2, a reduction in the expression of the Mn-SOD enzyme, and a greater expression of ß-MHC and Connexin 43. There was also an increase in the MPO enzyme activity in the tissue. It was observed that the effects of protein restriction are sex-specific, since the cardiac muscle of male animals showed alterations suggestive of oxidative stress, hypertrophy, signs of tissue inflammation, and increased expression of important proteins in intercellular communication. These changes characterize the ongoing cardiac remodeling process. Finally, the data revealed that the lactation phase accentuated harmful effects on the cardiac tissue of the offspring.

Pulpal outcomes in orthodontic tooth movement in diabetes mellitus.

Diabetes mellitus impairs angiogenesis and tissue reorganization during orthodontic tooth movement (OTM). Thus, this study evaluated pulpal outcomes in orthodontic tooth movement through metabolic changes in diabetes. Male Wistar rats were used, and the in vivo study design consisted of four groups (n = 10/group): C-non-diabetic animals not subjected to orthodontic tooth movement; D-diabetic animals not subjected to orthodontic tooth movement; OTM-non-diabetic animals subjected to orthodontic tooth movement; and D + OTM-diabetic animals subjected to orthodontic tooth movement. In addition, the pulps of the distovestibular root (DV) and mesiovestibular root (MV) were assessed by histomorphometric analyses and immunoexpression of the RANKL/OPG system. Pulpal analysis of the MV root showed an increase in blood vessels in diabetic animals. Inflammatory infiltrate and fibroblastic cells were elevated in diabetic animals with tooth movement in the DV and MV roots. In the DV and MV roots, diabetic rats with OTM showed a reduction in birefringent collagen fibers. The immunostaining for RANKL was higher in the pulp tissue of OTM in diabetic and non-diabetic animals. It was concluded that the pulp tissue has less adaptive and repair capacity during OTM in diabetes. Orthodontic strength can alter the inflammatory processes in the pulp.

Influence of microcurrent on the modulation of remodelling genes in a wound healing assay.

The literature has shown the beneficial effects of microcurrent (MC) therapy on tissue repair. We investigated if the application of MC at 10 µA/90 s could modulate the expression of remodeling genes transforming growth factor beta (Tgfb), connective tissue growth factor (Ctgf), insulin-like growth factor 1 (Igf1), tenascin C (Tnc), Fibronectin (Fn1), Scleraxis (Scx), Fibromodulin (Fmod) and tenomodulin in NIH/3T3 fibroblasts in a wound healing assay. The cell migration was analyzed between days 0 and 4 in both fibroblasts (F) and fibroblasts + MC (F+MC) groups. On the 4th day, cell viability and gene expression were also analyzed after daily MC application. Higher expression of Ctgf and lower expression of Tnc and Fmod, respectively, were observed in the F+MC group in relation to F group (p < 0.05), and no difference was observed between the groups for the genes Tgfb, Fn1 and Scx. In cell migration, a higher number of cells in the scratch region was observed in group F+MC (p < 0.05) compared to group F on the 4th day, and the cell viability assay showed no difference between the groups. In conclusion, MC therapy at an intensity/time of 10 µA/90 s with 4 daily applications did not affect cell viability, stimulated fibroblasts migration with the involvement of Ctgf, and reduced the Tnc and Fmod expression.

Photobiomodulation (λ=808nm) and Platelet-Rich Plasma (PRP) for the Treatment of Acute Rheumatoid Arthritis in Wistar Rats.

Introduction: Rheumatoid arthritis (RA) causes inflammation, pain, edema, and articular degradation and its treatment can be based on anti-inflammatory drugs, photobiomodulation (PBM) and/or platelet-rich plasma (PRP) that can decrease cell flow and promote local healing. In the present study, we evaluate the effects of PBM and PRP on acute arthritis in Wistar rats through inflammatory and oxidative stress parameters. Methods: Thirty female Wistar rats were assigned to five groups (n=6, each group): Control, Sham, PRP, Laser, and PRP+Laser. For arthritis induction, all animals of groups Sham, PRP, Laser and PRP+Laser received an intraarticular injection of Zymosan® (200µg) in the right knee. Twenty-four hours post-arthritis induction, PRP was prepared and injected (8 × 105 of platelets) in animals of PRP and PRP+Laser groups. PBM was performed in Laser and PRP+Laser groups by single-dose therapy with the GaAlAs laser (λ=808 nm, P=25 mW, fluence=30 J/cm2, beam area=0.02 mm2, t=33 seconds, E=0.825 J, punctual application). After seven days of induction, serum samples were collected and thiobarbituric acid reactive substances (TBARS), nitric oxide (NO) and catalase activity were analysed. Morphological parameters were measured for inflammation areas, cartilage thickness, and C3 protein expression in knee samples. Statistical analysis was performed with an ANOVA test and Tukey's post-hoc test with a significance level of 5% (P<0.05). Results: NO was lower in the treated groups compared to the Sham group, and TBARS did not show any differences, while catalase showed greater activity between PRP+Laser versus PRP (P<0.05). Inflammatory areas and cartilage thickness were lower in the treated groups compared to Sham (P<0.05), while no differences in C3 protein expression was observed. Conclusion: PBM associated with PRP is better for anti-inflammatory and joint preservation by morphological aspects and NO levels that concern a potential clinical application.

Laser treatment of synovial inflammatory process in experimentally induced microcrystalline arthritis in Wistar rats.

The presence of intra-articular crystals is detected in different articular pathologies of acute or chronic nature. The aim of this work was to analyze the action of the indium gallium aluminum and phosphorus (InGaAlP) (λ = 670 nm) laser on the synovial membrane present in the knee joint in experimentally induced microcrystalline arthritis in male adult Wistar rats. The animals were divided into three experimental groups (n = 24): control (A), experimentally induced arthritis (B), experimentally induced arthritis+InGaAlP laser therapy (C). The laser treatment was made daily in the patellar region of the right knee after 48 h of the experimental induction. After 7, 14, and 21 days of therapy, the rats were euthanized and the right knees were removed and processed for histomorphometric, immunohistochemical, ultrastructural, and biochemical investigation of the synovium. The number of granulocytes on the 14th and 21st days was higher in B and lower in C and, lastly, in A. The number of fibroblasts on the 14th and 21st days was similar between A and C and below B. The number of blood vessels on the 21st day was higher in B than in the other groups. The positive number of cells for the TUNEL test was higher on the 14th and 21st days in B compared to the others. The percentage of tissue area occupied by birefringent collagen fibers was higher in B on the 21st day than in the others. The ultrastructure of cells showed fibroblast-like morphology in all groups and periods evaluated. The quantification of glycosaminoglycans did not present significant differences between the groups in all the experimental periods. The amount of hydroxyproline was higher in B compared to the other groups on the 14th and 21st days. The content of non-collagen proteins was higher in B on the 21st day in relation to the other groups. Quantification of TNF-α on the 21st day was higher in A and B than in C. For TGF-ß on the 21st day, groups B and C presented similar and higher values than A. For MMP-13, groups A and B presented data similar to and above C. In relation to ADAMT-S4, on the 21st day, groups B and C presented data similar to and lower than A. InGaAlP-670 nm therapy reduced the inflammatory process and tissue injuries of the synovial membrane in comparison to the untreated group, indicating its potential utilization in clinical studies aiming in the recovery of acute arthritis in patients.

Gestational protein restriction alters the RANKL/OPG system in the dental germ of offsprings.

Evaluate the dentinogenesis in the offspring of rats submitted to gestational protein restriction (GPR). DESIGN: The offspring were evaluated at the 21st day of gestation (21 dG). Assessments were made of morphological parameters and the RANKL/OPG system - bone tissue maturation markers - in the upper incisor tooth germ. Pregnant 10-week-old female Wistar rats were divided into normal protein (NP, 17% casein, n = 5) and low protein (LP, 6% casein, n = 5) diet groups. At 21 dG, the offspring maxillae were collected for histomorphometric and immunohistochemical analyses. RESULTS: The LP group showed decreased thickness of the dentin and odontoblast cell layers on the tooth germ. GPR led to decreased OPG expression and increased RANKL expression in the incisor germ. CONCLUSION: The results suggested that gestational protein restriction altered odontoblast RANKL/OPG expression and decreased dentin matrix deposition and thickness in tooth development.

Argon Atmospheric Plasma Treatment Promotes Burn Healing by Stimulating Inflammation and Controlling the Redox State.

Burns are a public health problem, with second-degree burns as one of the most common types. Although intense inflammation worsens burn healing, effective therapies are scarce. Thus, infections and hypertrophic scars may occur, which compromise patient quality of life and may delay healing. Argon atmospheric plasma (AP) has been shown to positively influence wound healing. In the context of identifying effective and alternative therapies for the treatment of second-degree burns, the present study evaluated AP in the treatment of second-degree burns in rats compared to that for sham treatment on the 2nd, 7th, 14th, and 21st days post-injury. Our results revealed proinflammatory effect for AP by recruiting predominantly neutrophils on the 7th day and macrophages on the 21st day compared to sham treatment, allowing a greater production of interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, and IL-17, and also controlled the inflammation by IL-10 and transforming growth factor (TGF)-ß1. AP also showed antioxidant activity important for controlling oxidative damage on the 2nd day. This favored the induction of angiogenesis from the 2nd day and induction fibroplasia and fibrillogenesis after the 14th day, which enhanced burn healing with the formation of a thinner burn eschar before the 21st day post-burn. Thus, AP effectively modulated the inflammatory phase of second-degree burn healing through the control of oxidative damage that favored the following phases. Therefore, AP is a relevant alternative in the treatment of second-degree burns.

Inhibitory effect of red LED irradiation on fibroblasts and co-culture of adipose-derived mesenchymal stem cells.

The objective of this study was to evaluate the effects of red Light Emiting Diode (red LED) irradiation on fibroblasts in adipose-derived mesenchymal stem cells (ASC) co-culture on the scratch assay. We hypothesized that red LED irradiation could stimulate paracrine secretion of ASC, contributing to the activation of genes and molecules involved in cell migration and tissue repair. ASC were co-cultured with NIH/3T3 fibroblasts through direct contact and subjected to red LED irradiation (1.45 J/cm2/5min6s) after the scratch assay, during 4 days. Four groups were established: fibroblasts (F), fibroblasts + LED (FL), fibroblasts + ASC (FC) and fibroblasts + LED + ASC (FLC). The analyzes were based on Ctgf and Reck expression, quantification of collagen types I and III, tenomodulin, VEGF, TGF-ß1, MMP-2 and MMP-9, as well as viability analysis and cell migration. Higher Ctgf expression was observed in FC compared to F. Group FC presented higher amount of tenomodulin and VEGF in relation to the other groups. In the cell migration analysis, a higher number of cells was observed in the scratched area of the FC group on the 4th day. There were no differences between groups considering cell viability, Reck expression, amount of collagen types I and III, MMP-2 and TGF-ß1, whereas TGF-ß1 was not detected in the FC group and the MMP-9 in none of the groups. Our hypothesis was not supported by the results because the red LED irradiation decreased the healing response of ASC. An inhibitory effect of the LED irradiation associated with ASC co-culture was observed with reduction of the amount of TGF-ß1, VEGF and tenomodulin, possibly involved in the reduced cell migration. In turn, the ASC alone seem to have modulated fibroblast behavior by increasing Ctgf, VEGF and tenomodulin, leading to greater cell migration. In conclusion, red LED and ASC therapy can have independent effects on fibroblast wound healing, but the combination of both does not have a synergistic effect. Therefore, future studies with other parameters of red LED associated with ASC should be tested aiming clinical application for tissue repair.

Diabetes mellitus and periodontitis: Inflammatory response in orthodontic tooth movement.

OBJECTIVES: This study evaluated, in experimental model, the inflammatory alterations in gingival tissue and alveolar bone during the orthodontic tooth movement (OTM) in diabetes mellitus (D) and periodontitis (P). SETTING AND SAMPLE POPULATION: Forty male Wistar rats, 90 days old and weighing 300 g. MATERIALS AND METHODS: The sample was divided into four groups (n = 10). OTM: orthodontic movement (10 days, 0.4 N force); P + OTM: periodontitis (ligature-induced periodontitis, 3-0 silk suture thread) and orthodontic movement; D + OTM: diabetes (Alloxan-induced diabetes, 150 mg/kg) and orthodontic movement; and D + P + OTM: diabetes, periodontitis and orthodontic movement. Tooth displacement was measured; fibroblast, inflammatory cells, osteoclast and blood vessels were quantified by histomorphometric analysis. Inflammatory markers, interleukin-6 (IL-6) and tumour necrosis factor (TNF-α) were quantified by ELISA (Enzyme-Linked Immunosorbent Assay) in gingival tissue. The fibroblastic growth factor (bFGF), transforming growth factor (TGF-ß1) and the vascular endothelial growth factor (VEGF) were measured via Western blotting in the alveolar bone. The results were analysed by ANOVA and Tukey's test at a 5% significance level. RESULTS: The quantification of inflammatory cells and the expression of IL-6, TNF-α, TGF-ß1 and bFGF were increased in diabetes and periodontitis. However, the number of fibroblasts and blood vessels and the percentage of birefringent collagen fibres were higher in healthy animals. There was greater tooth displacement in the OTM group. CONCLUSION: Diabetes Mellitus modifies the inflammatory response. The increased expression of inflammatory markers IL-6, TNF-α and TGF-ß1 in diabetic animals impairs neovasculogenesis and tissue reorganization during orthodontic tooth movement, which may be aggravated by periodontitis.

Activators of SIRT1 in wound repair: an animal model study.

Caloric restriction (CR) and resveratrol activate SIRT1 and induce anti-inflammatory and antioxidant properties. We perform excisional lesion on the dorsum of four groups anesthetized animals: ad libitum-AL diet, AL diet with topical application of 2% resveratrol-Rv, 30% calorie restricted, and finally 30% calorie restricted with 2% resveratrol and we examine CR and Rv effects in wound repair. Restricted animals remained with CR for 31 days. The lesion was performed on day 18 of CR, and resveratrol application was started on day 19. Lesion samples were then collected on days 3 and 10 of treatment for structural, morphometric, and protein analyses. Our results showed that CR and Rv group as well as R group had enhanced numbers of blood vessels, VEGF, fibroblast, birefringent collagen fiber areas in the lesion. We conclude that effects in wound repair suggests that both CR and resveratrol may modulate angiogenesis, fibroplasia, and collagenesis, which could be ascribed to the action of SIRT1.

Effects of low-level laser therapy on the organization of articular cartilage in an experimental microcrystalline arthritis model.

The aim of this study was to evaluate the effects of low-level laser therapy using the gallium arsenide laser (λ = 830 nm) on the articular cartilage (AC) organization from knee joint in an experimental model of microcrystalline arthritis in adult male Wistar rats. Seventy-two animals were divided into three groups: A (control), B (induced arthritis), and C (induced arthritis + laser therapy). The arthritis was induced in the right knee using 2 mg of Na4P2O7 in 0.5 mL of saline solution. The treatments were daily applied in the patellar region of the right knee after 48 h of induction. On the 7th, 14th, and 21st days of treatment, the animals were euthanized and their right knees were removed and processed for structural and biochemical analysis of the AC. The chondrocytes positively labeled for the TUNEL reaction were lower in C than in B on the 14th and 21st days. The content of glycosaminoglycans and hydroxyproline in A and C was higher than B on the 21st day. The amount of tibial TNF-α in B and C was lower than in A. The amount of tibial BMP-7 in B and C was higher than in A. The femoral MMP-13 was lower in B and C than for A. The tibial TGF-ß for C was higher than the others. The femoral ADAMT-S4 content of A and C presented similar and inferior data to B on the 21st day. The AsGa-830 nm therapy preserved the content of glycosaminoglycans, reduced the cellular changes and the inflammatory process compared to the untreated group.

Electrical stimulation: Complementary therapy to improve the performance of grafts in bone defects?

The limitations of bone reconstruction techniques have stimulated the tissue engineering for the repair of large bone defects using osteoconductive materials and osteoinductive agents. This study evaluated the effects of low intensity electric current on the inorganic bovine graft in calvaria defects. Bone defects were performed with piezoelectric system in the calvaria of Wistar rats divided into four groups (n = 24): (C) without grafting and without electrical stimulation; (E) with grafting; (MC) without grafting and submitted to electrical stimulation; (MC + E) with grafting and submitted to electrical stimulation. Inflammatory, angiogenic and osteogenic events during bone repair at the 10th, 30th, 60th, and 90th days were considered. Several inflammatory markers demonstrated the efficacy of grafting in reducing inflammation, particularly when subjected to electrical stimulation. Angiogenesis and collagen organization were more evident by electrical stimulation application on the grafts. Moreover, the osteogenic cell differentiation process indicated that the application of microcurrent on grafting modulated the homeostasis of bone remodeling. It is concluded that microcurrent favored the performance of grafts in calvarial rat model. Low-intensity electrical current might improve the osteoconductive property of grafting in bone defects. Therefore, electrical current becomes an option as complementary therapy in clinical trials involving bone surgeries and injuries. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 924-932, 2019.

Evolution of Periodontal Disease: Immune Response and RANK/RANKL/OPG System.

The aim of this study was to evaluate markers of bone loss and immune response present in evolution of periodontal disease. One hundred and two Wistar rats were divided into three animals groups: PD0, without ligation and PD15 days and PD60 days, submitted to ligation placement with a sterile 3-0 silk cord in the cervical region of the upper first molar on both sides. Samples were obtained from the gingival tissue for histomorphometric analysis, immunohistochemical analysis of RANK, RANKL, OPG, characterization of the inflammatory infiltrate, quantification of nitric oxide, MCP-1, RANTES, IP10 chemokines, and expression of the TGF-b1, VEG, and bFGF. The number of inflammatory cells in gingival tissue was higher in PD60 samples. The collagen content and the area occupied by birefringent collagen fibers were lower for PD60. Differential leukocyte counting showed that there was a significantly higher polymorphonuclear influx in group PD15, while PD60 showed a greater number of lymphocytes. PD60 showed higher RANTES, IP-10, MCP-1 gene transcripts, as well as a higher nitric oxide concentration. Clinical evaluation revealed that the PD60 group presented an increase in furcal area. In conclusion, in this animal model the increase of RANK/RANKL and HGF markers is related to a specific immune response, and probably contributed to the evolution of periodontal disease. Investigating the effect of these biomarkers can help in targeted therapy for bone resorption, since blocking these can inhibit bone loss.

Evolution of Periodontal Disease: Immune Response and RANK/RANKL/OPG System

Abstract The aim of this study was to evaluate markers of bone loss and immune response present in evolution of periodontal disease. One hundred and two Wistar rats were divided into three animals groups: PD0, without ligation and PD15 days and PD60 days, submitted to ligation placement with a sterile 3-0 silk cord in the cervical region of the upper first molar on both sides. Samples were obtained from the gingival tissue for histomorphometric analysis, immunohistochemical analysis of RANK, RANKL, OPG, characterization of the inflammatory infiltrate, quantification of nitric oxide, MCP-1, RANTES, IP10 chemokines, and expression of the TGF-b1, VEG, and bFGF. The number of inflammatory cells in gingival tissue was higher in PD60 samples. The collagen content and the area occupied by birefringent collagen fibers were lower for PD60. Differential leukocyte counting showed that there was a significantly higher polymorphonuclear influx in group PD15, while PD60 showed a greater number of lymphocytes. PD60 showed higher RANTES, IP-10, MCP-1 gene transcripts, as well as a higher nitric oxide concentration. Clinical evaluation revealed that the PD60 group presented an increase in furcal area. In conclusion, in this animal model the increase of RANK/RANKL and HGF markers is related to a specific immune response, and probably contributed to the evolution of periodontal disease. Investigating the effect of these biomarkers can help in targeted therapy for bone resorption, since blocking these can inhibit bone loss.

Resumo Este estudo avaliou marcadores de perda óssea e da resposta imune presentes na evolução da doença periodontal. Cento e dois ratos Wistar foram divididos em três grupos de animais: PD0, sem ligadura e PD15 dias e PD60 dias, submetidos a colocação de ligadura com um fio de seda estéril 3-0 na região cervical do primeiro molar superior em ambos os lados. Foram obtidas amostras de tecido gengival para análise histomorfométrica, análises imunohistoquímicas de RANK, RANKL, OPG, caracterização do infiltrado inflamatório, quantificação de óxido nítrico, expressão de quimiocinas MCP-1, RANTES, IP10 e do TGF-b1, VEGF e bFGF . O número de células inflamatórias no tecido gengival foi maior nas amostras PD60. O teor de colágeno na área ocupada pelas fibras de colágeno birrefringentes foram menores para PD60. A contagem diferencial de leucócitos mostrou que houve um influxo polimorfonuclear significativamente maior no grupo PD15, enquanto que PD60 mostrou número maior de linfócitos. PD60 apresentou transcritos de genes RANTES, IP-10, MCP-1 mais elevados, bem como uma maior concentração de óxido nítrico. A avaliação clínica revelou que o grupo PD60 apresentou aumento da área óssea exposta na região da furca. Em conclusão, neste modelo animal o aumento dos marcadores RANK/RANKL e HGF está relacionado a uma resposta imunológica específica e provavelmente contribuiu para a evolução da doença periodontal. Investigar o efeito destes biomarcadores pode ajudar na terapia dirigida para a reabsorção óssea, uma vez que bloquear estes pode inibir a perda óssea.

Differing energy densities with laser 670nm InGaP controls inflammation and collagen reorganization in burns.

PURPOSE: This study compared different energy densities of laser on second degrees burns in rats aiming to determine the most effective dosimetry in stimulation of the healing process. METHODS: Burns were induced in the dorsal skin of 54 animals divided into three groups (n: 18): 1-without treatment; 2-irradiated lesions by the Indium Gallium Phosphide (InGaP) 670nm (4.93J/cm2) laser; 3-irradiated lesions by the InGaP-670nm (9.86J/cm2) laser. Samples were collected on the 2, 10 and 18 days after injury for structural, morphometry, biochemical analysis and Western blotting. RESULTS: The energy densities examined were effective in significantly increasing the total number of fibroblasts and blood vessels and reduce the number of inflammatory cells particularly in irradiated lesions with 9.86J/cm2. This same energy density significantly increased the amount of GAGs (Glycosaminoglycans), decreased the TGF-ß1 (Transforming Growth Factor ß1) and increased the VEGF (Vascular and Endothelial Growth Factor) during the experimental period. This energy density also significantly increased the Collagen type I and decreased Collagen type III and the active isoform of metalloproteinase 9 (MMP-9). CONCLUSIONS: The energy density of 9.86J/cm2 was more effective in promoting cellular responses related to neoangiogenesis, decreasing inflammation and collagen fibers reorganization.

Electrical stimulation enhances tissue reorganization during orthodontic tooth movement in rats.

OBJECTIVE: This study evaluated the effects of a low-intensity electric current on tissue reorganization during experimental orthodontic tooth movement. MATERIALS AND METHODS: Thirty-two animals were divided into two groups evaluated on days 3 and 7: OTM-orthodontic tooth movement and OTM + MC-orthodontic tooth movement and microcurrent application (10 µA/5 min). The samples were processed for histological, morphometric, and Western blotting analysis. RESULTS: Analysis of the periodontal ligament (PL) showed a significantly smaller number of granulocytes in the OTM + MC group on day 7.The number of fibroblasts was significantly higher in the OTM + MC group on days 3 and 7. The area of birefringent collagen fibers was more organized in the OTM + MC group on days 3 and 7. The number of blood vessels was significantly higher in the OTM + MC group on day 7. Microcurrent application significantly increased the number of osteoclasts in the compression region of the PL. In the OTM + MC group on day 7 of tooth movement, the expression of TGF-ß1 and VEGF was significantly reduced whereas the expression of bFGF was increased in PL. CONCLUSIONS: Electrical stimulation enhances tissue responses, reducing the number of granulocytes and increasing the number of fibroblasts, blood vessels, and osteoclasts and modulates the expression of TGF-ß1, VEFG, and bFGF. CLINICAL RELEVANCE: This technique is used in many areas of medicine, but poorly explored in dentistry and orthodontics. This treatment is cheap and non-invasive and can be applied by own orthodontist, and it can improve the treatment with a faster and safe tooth movement, without pain.