Cristiane Miranda França

LOW-INTENSITY RED LASER ON THE PREVENTION AND TREATMENT OF INDUCED-ORAL MUCOSITIS IN HAMSTERS

OBJECTIVE The purpose of this study was to investigate the effects of laser phototherapy as preventive and therapeutic regime on induced-oral mucositis in hamsters. DESIGN The animals were divided into four groups: preventive cryotherapy, preventive laser, therapeutic laser and therapeutic control group. Mucositis was induced in hamsters by intraperitoneal injection of 5-fluorouracil (5-FU) and superficial scratching. All preventive treatment was performed on the right cheek pouch mucosa. The left pouch mucosa was used for a spontaneous development of mucositis and did not receive any preventive therapy. Laser parameters were: lambda=660nm, P= 30mW, D=1.2J/cm(2), Deltat=40s, spot size 3mm(2), I=1W/cm(2). Cryotherapy was done positioning ice packs in the hamster mucosa 5min before 5-FU infusion and 10min afterward. To study the healing of mucositis, the left pouch mucosa of each of the hamsters in the TLG received laser irradiation on the injured area. Irradiation parameters were kept the same as abovementioned. The control hamsters in the TCG did not receive any treatment. The mucositis degree and the animals body mass were evaluated. An assessment of blood vessels was made based on immunohistochemical staining. RESULTS The CG animals lost 15.16% of theirs initial body mass while the LG animals lost 8.97% during the first 5 days. The laser treated animals had a better clinical outcome with a faster healing, and more granulation tissue. The quantity of blood vessels at both LG and CG were higher than in healthy mucosa. Regarding the therapeutic analysis, the severity of the mucositis in the TLG was always lower than TCG. TLG presented higher organization of the granulation tissue, parallel collagen fibrils, and increased angiogenesis. CONCLUSION The results suggest that laser phototherapy had a positive effect in reducing mucositis severity, and a more pronounced effect in treating established mucositis.

Low-Level Laser Therapy Restores the Oxidative Stress Balance in Acute Lung Injury Induced by Gut Ischemia and Reperfusion

It remains unknown if the oxidative stress can be regulated by low‐level laser therapy (LLLT) in lung inflammation induced by intestinal reperfusion (i‐I/R). A study was developed in which rats were irradiated (660 nm, 30 mW, 5.4 J) on the skin over the bronchus and euthanized 2 h after the initial of intestinal reperfusion. Lung edema and bronchoalveolar lavage fluid neutrophils were measured by the Evans blue extravasation and myeloperoxidase (MPO) activity respectively. Lung histology was used for analyzing the injury score. Reactive oxygen species (ROS) was measured by fluorescence. Both expression intercellular adhesion molecule 1 (ICAM‐1) and peroxisome proliferator‐activated receptor‐y (PPARy) were measured by RT‐PCR. The lung immunohistochemical localization of ICAM‐1 was visualized as a brown stain. Both lung HSP70 and glutathione protein were evaluated by ELISA. LLLT reduced neatly the edema, neutrophils influx, MPO activity and ICAM‐1 mRNA expression. LLLT also reduced the ROS formation and oppositely increased GSH concentration in lung from i‐I/R groups. Both HSP70 and PPARy expression also were elevated after laser irradiation. Results indicate that laser effect in attenuating the acute lung inflammation is driven to restore the balance between the pro‐ and antioxidants mediators rising of PPARy expression and consequently the HSP70 production.

Histomorphometric and Microbiological Assessment of Photodynamic Therapy as an Adjuvant Treatment for Periodontitis: A Short-Term Evaluation of Inflammatory Periodontal Conditions and Bacterial Reduction in a Rat Model

OBJECTIVE The aim of this study was to investigate the short-term effects of photodynamic therapy (PDT) in periodontal tissue when it is used as an adjuvant treatment for periodontitis. BACKGROUND DATA PDT has been used as an adjuvant in the combat of local infections, such as periodontitis, and combines a photosensitizer (PS) with a light source to induce reactive oxygen species (ROS) and kill microbial cells. METHODS Fifty healthy male rats were used in this study. Periodontitis was induced by placing a cotton ligature around the upper left second molar in a subgingival position. Posterior maxillas were removed and histologically prepared with hematoxylin & eosin (H&E) staining techniques. PDT was performed with a diode laser (λ=660 nm) with an output power of 100 mW. Methylene blue aqueous solution (100 μM) was used as the PS while control group used phosphate buffered saline (PBS). Collagen organization, inflammatory infiltrate, and bone loss were evaluated. Bacterial samples were collected before and immediately after treatment to determine bacterial reduction. RESULTS The experimental group that was treated with PDT presented better periodontal healing, as measured by collagen organization, inflammatory infiltrate, and bone loss. Significant bacterial reduction was achieved following treatment with or without PDT compared to control, with a higher microbial reduction observed in the PDT group. CONCLUSIONS PDT used as an adjuvant treatment showed effective short-term control of periodontitis infection.

Pulp capping materials exert an effect on the secretion of IL-1β and IL-8 by migrating human neutrophils

Pulp repair is a complex process whose mechanisms are not yet fully understood. The first immune cells to reach the damaged pulp are neutrophils that play an important role in releasing cytokines and in phagocytosis. The objective of this study was to analyze the effect of different pulp-capping materials on the secretion of interleukin-1 beta (IL-1β) and interleukin-8 (IL-8) by migrating human neutrophils. Neutrophils were obtained from the blood of three healthy donors. The experimental groups were calcium hydroxide [Ca(OH)(2)], an adhesive system (Single Bond), and mineral trioxide aggregate (MTA). Untreated cells were used as control. Transwell chambers were used in performing the assays to mimic an in vivo situation of neutrophil chemotaxis. The pulp-capping materials were placed in the lower chamber and the human neutrophils, in the upper chamber. The cells were counted and the culture medium was assayed using ELISA kits for detecting and quantifying IL-1β and IL8. The data were compared by ANOVA followed by Tukeys test (p < 0.05). The secretion of IL-8 was significantly higher in all groups in comparison to the control group (p < 0.05). The adhesive system group showed higher IL-8 than the MTA group (p < 0.05). The secretion of IL-1β was significantly greater only in the MTA group (p < 0.001). It was concluded that only MTA is able to improve the secretion of IL-1β, and all materials tested increased IL-8 secretion. These results combined with all the other biological advantages of MTA indicate that it could be considered the material of choice for dental pulp capping.

Tissue Responses to Postoperative Laser Therapy in Diabetic Rats Submitted to Excisional Wounds

In a previous study about low-level laser therapy biomodulation on a full-thickness burn model we showed that single and fractionated dose regimens increased wound healing and leukocyte influx similarly when compared with untreated control. In order to verify if this finding would be similar in an impaired wound model, we investigated the effect of single and multiple irradiations on wound closure rate, type of inflammatory infiltrate, myofibroblasts, collagen deposition, and optical retardation of collagen in diabetic rats. Female Wistar rats in the same estrous cycle had diabetes induced with streptozotocin and an 8-mm excisional wound performed with a punch. The experimental groups were: control group – untreated ulcer; single-dose group – ulcer submitted to single dose of diode laser therapy (λ = 660 ± 2 nm; P = 30 mW; energy density: 4 J/cm2) and fractionated-dose group – ulcer submitted to 1 J/cm2 laser therapy on Days 1, 3, 8, and 10. The ulcers were photographed on the experimental days and after euthanasia tissue samples were routinely processed for histological and immunohistochemistry analyses. Independently of the energy density, laser therapy accelerated wound closure by approximately 40% in the first three days in comparison to the control group. Laser therapy increased acute inflammatory infiltrate until Day 3. Both laser groups exhibited more myofibroblasts and better collagen organization than the control group. The findings demonstrate that low-level laser therapy in the immediate postoperative period can enhance the tissue repair process in a diabetes model. Similar effects were achieved with laser therapy applied a single time with an energy density of 4 J/cm2 and applied four times with an energy density of 1 J/cm2. The application of laser therapy in the inflammatory phase was the most important factor to the enhancement of the tissue repair process.

The effect of low‐level laser therapy (LLLT) applied prior to muscle injury

To evaluate the effect of LLLT (780 nm; 10 J/cm2, 40 mW, 3.2 J) prior to injury on the morphological analysis, collagen deposition, and activity of matrix metalloproteinase‐2 (MMP‐2).

Use of low-power laser to assist the healing of traumatic wounds in rats

PURPOSE To investigate the morphological aspects of the healing of traumatic wounds in rats using low-power laser. METHODS Twenty four non isogenic, young adult male Wistar rats (Rattus norvegicus) weighing between 200 and 300 g was used. The animals were randomly distributed into two groups: Control (GC) and Laser (GL), with 12 animals each. After shaving, anesthesia was performed in the dorsal region and then a surgical procedure using a scalpel was carried out to make the traumatic wound. GL received five sessions of laser therapy in consecutive days using the following laser parameters: wavelength 660 nm, power 100 mW, dose 10 J/cm2. The wounds were evaluated through measurement of the area and depth of the wound (MW) and histological analysis (HA). RESULTS When comparing the GC with the GL in MW there was a difference in area (p<0.001) and depth (p=0.003) measurement of the wounds in GL. The laser group presented more epithelization than GC (p=0.03). The other histological parameters were similar. CONCLUSION The healing of wounds in rats was improved with the use of the laser.

Increase in cholinergic modulation with pyridostigmine induces anti-inflammatory cell recruitment soon after acute myocardial infarction in rats

We tested the hypothesis that an increase in the anti-inflammatory cholinergic pathway, when induced by pyridostigmine (PY), may modulate subtypes of lymphocytes (CD4+, CD8+, FOXP3+) and macrophages (M1/M2) soon after myocardial infarction (MI) in rats. Wistar rats, randomly allocated to receive PY (40 mg·kg(-1)·day(-1)) in drinking water or to stay without treatment, were followed for 4 days and then were subjected to ligation of the left coronary artery. The groups-denominated as the pyridostigmine-treated infarcted (IP) and infarcted control (I) groups-were submitted to euthanasia 3 days after MI; the heart was removed for immunohistochemistry, and the peripheral blood and spleen were collected for flow cytometry analysis. Noninfarcted and untreated rats were used as controls (C Group). Echocardiographic measurements were registered on the second day after MI, and heart rate variability was measured on the third day after MI. The infarcted groups had similar MI areas, degrees of systolic dysfunction, blood pressures, and heart rates. Compared with the I Group, the IP Group showed a significant higher parasympathetic modulation and a lower sympathetic modulation, which were associated with a small, but significant, increase in diastolic function. The IP Group showed a significant increase in M2 macrophages and FOXP3(+)cells in the infarcted and peri-infarcted areas, a significantly higher frequency of circulating Treg cells (CD4(+)CD25(+)FOXP3(+)), and a less extreme decrease in conventional T cells (CD25(+)FOXP3(-)) compared with the I Group. Therefore, increasing cholinergic modulation with PY induces greater anti-inflammatory cell recruitment soon after MY in rats.

Effect of papain-based gel on type I collagen--spectroscopy applied for microstructural analysis.

Considering the improvement of biomaterials that facilitate atraumatic restorative techniques in dentistry, a papain-based gel can be used in the chemomechanical removal of decayed dental tissue. However, there is no information regarding the influence of this gel on the structure of sound collagen. The aim of the present study was to investigate the adsorption of a papain-based gel (PapacarieTM) to collagen and determine collagen integrity after treatment. A pilot study was first performed with 10 samples of type I collagen membrane obtained from bovine Achilles deep tendon to compare the influence of hydration (Milli-Q water) on infrared bands of collagen. In a further experiment, 10 samples of type I collagen membrane were used to evaluate the effects of PapacarieTM on the collagen microstructure. All analyses were performed using the attenuated total reflectance technique of Fourier transform infrared (ATR-FTIR). The results demonstrated that the application of PapacarieTM does not lead to the degradation of collagen and this product can be safely used in minimally invasive dentistry. As the integrity of sound collagen is preserved after the application of the papain-based gel, this product is indicated for the selective removal of infected dentin, leaving the affected dentin intact and capable of re-mineralization.

Macrophage migration inhibitory factor and oral cancer

BACKGROUND Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine with pro-inflammatory functions and involved in tumorigenesis. The aim of this study was to evaluate the expression and localization of the macrophage MIF in oral squamous carcinoma (OSC). In addition, the relationship between MIF expression and clinicopathological parameters such as survival data, tobacco use, alcohol habits, TNM stage, tumor graduation, and peritumoral inflammatory infiltrate were evaluated. METHODS Using immunohistochemistry, expression and localization of MIF was detected in 44 specimens of OSC. The absolute number and relative proportions of MIF-positive cells detected were also determined separately for tumor parenchyma vs. stroma. All counts were determined from 10 consecutive high-power fields using an integration graticule. Moreover, some parameters were analyzed separately for lip and intra-oral cancers. RESULTS Migration inhibitory factor-positive cells were observed in both the tumor parenchyma and in inflammatory cells of all specimens. In contrast, MIF expression was not detected in tumoral nests associated with poorly differentiated tumors. In specimens of lip cancer, a greater number of MIF-positive stromal immune cells were detected than in intra-oral cancer specimens (Mann-Whitney test, P = 0.049). CONCLUSIONS Oral squamous carcinoma cells consistently express MIF independent of their location. Lip tumors presented more MIF-positive peritumoral inflammatory cells, similar to control, suggesting that immunological differences in leukocyte activation exist between in lip and intra-oral cancers.