Resposta celular Th1 /Th2 na doença periodontal experimental, em ratos: um estudo imuno-histoquímico

Host response plays a major role in the pathogenesis of periodontal disease. Mediators such as inflammatory cytokines which are secreted during the immune response to bacterial challenges have ambiguous functions that may or may not lead to protection of the attacked tissue. In this context, experimental evidence suggests that T-helper 1 (Th-1) and T-helper 2 (Th-2) mediated responses are potentially important during the disease process. The aims of this study therefore were to further clarify the role played by Th2 cells during different time points of the active phase of periodontal disease, as well as, to investigate whether there was any evidence of a Th1 response in the periodontal disease microenvironment. Experimental periodontitis was induced in 30 Wistar male rats by placing cotton ligatures around the mandibular first molars. The rats were then randomly divided into two groups. Group1 (G1=15) and Group 2 (G2=15). In G1 the ligatures were maintained for 2 days, whereas in G2 the ligatures were left for 15 days, a time point that corresponds to the advanced stage of periodontal disease The contra-lateral teeth served as controls (no ligatures). Immunohistochemical investigation for the presence in gingival tissue of Th2 specific transcription factor (GATA3) and the subunit of the IFN-γ receptor was carried out after the disease induction period. Light microscopy analysis revealed a decrease in the expression of GATA-3 as bone loss progressed. On the other hand, although IFN-γ R1 was detected at an early stage of the active phase of disease its expression remained unaltered during the remaining period of the study. These results indicate that the Th2 response have a protective role during the pathogenesis of periodontal disease and that the progression of the periodontal disease is related with the unbalance of the responses Th1/Th2

Análise imuno-histoquímica das metaloproteinases da matriz ( MMP-1,MMP-2 e MMP-9) na doença periodontal

The tissular destruction found in periodontal diseases is caused mainly by components of the host that have its production stimulated by the products of the microorganisms present on the plaque. Matrix Metalloproteinases (MMPs), a class of enzymes involved both in physiologic and pathologic extracellular matrix degradation are considered the main responsible for the characteristic tissular loss in periodontal disease, and the understanding of how this happens can have a series of beneficial implications for prevention, diagnosis and treatment of this illness. The aim of this work was to study the immunohistochemical expression of MMP-1, MMP-2, and MMP-9 in fragments of gingival biopsies with clinical diagnose of periodontal disease. MMP-1 has expressed significantly more than the others MMPs in gingivitis both in the epithelium (p=0,0008) and connective tissue (p=0,0049). In periodontitis, both MMP-1 and MMP-9 has expressed significantly more than MMP-2 in the epithelium (p<0,0001) and in the connective tissue (p=0,0002). MMP-1 and MMP-9 presented more expression in periodontitis than in gingivitis but MMP-1 only in the connective tissue (p=0,03) and MMP-9 in the epithelium (p=0,003) and in the connective tissue (p=0,04). In conclusion, these results indicate that the MMP-1 presents high expression in every stages of the periodontal diseases, and increases its expression in the connective tissue when the gingivitis evolves to periodontitis. Therefore, it may have an important role in connective tissue degradation and bone loss observed in disease, since early, in gingivitis, until late stages, in periodontitis, of the periodontal disease. MMP-9 has expressed more in periodontitis than in gingivitis, both in epithelium and in connective tissue. It means that this enzyme may have some importance in the progression of gingivitis to periodontitis by acting in bone resorption observed in this desease

Imunoexpressão de fatores reguladores da osteoclastogênese na doença periodontal em humanos e sua relação com os parâmetros clínicos

Periodontal disease is an infection initiated by oral periodontal pathogens that trigger an immune response culminating in tissue destruction. This destruction is mediated by the host by inducing the production and activation of lytic enzymes, cytokines and the stimulation of osteoclastogenesis. The aim of this study was to compare the immunohistochemical expression of factors involved in bone resorption, RANKL (Ligand Receptor Activator of Nuclear Factor kappa B), OPG (Osteoprotegerin) and TNF-α (tumor necrosis factor alpha) between the gingival healthy, gingivitis and chronic periodontitis and correlate them with clinical parameters. The sample consisted of 83 cases and 12 clinically healthy gums, 42 gingivitis and 29 periodontitis, from 74 adolescent and adult patients with a mean age of 35 years, without systemic changes and non-smokers, predominantly female and race brown. There was no statistically significant difference for the expression of anti-RANKL (p = 0.581) and RANKL / OPG ratio (p = 0.334) when comparing the three conditions, but the anti-OPG and anti-TNF-α showed statistically significant between the types of injury (p = 0.001 and p <0.001, respectively), showing greatest expression in periodontitis. In cases of periodontitis, the variable clinical attachment loss (PIC) was statistically significant and positive correlation, respectively, with immunostaining of anti-RANKL (p = 0.002, p = 0.001 and r = 0.642), anti-OPG (p = 0.018, p = 0.014 and r = 0.451), anti-TNF-α (p = 0.032, p = 0.014 and r = 0.453) and the percentage ratio of RANKL / OPG (p = 0.018, p = 0.002 and r = 0.544). The tooth mobility (MB) showed a statistically significant difference only with immunohistochemical anti-RANKL (p = 0.026), and probing depth (PD) was positively correlated with anti-RANKL (p = 0.028 and r = 0.409), both in cases of periodontitis. Only in cases of gingivitis TNF-α was positively correlated with RANKL (p = 0.012 and r = 0.384) and the RANKL / OPG ratio (p = 0.027 and r = 0.341). Given these results, we conclude that the greatest expression of TNF-α in periodontitis demonstrates a relationship with the progression and severity of periodontal disease and the correlation between all antibodies and clinical attachment loss demonstrates their involvement in periodontal bone resorption

Estudo in vitro dos efeitos da BMP-2 e do seu antagonista Noggin sobre a proliferação e migração celulares em carcinoma epidermóide de língua

Oral squamous cell carcinoma (OSCC) is the most prevalent malignancy in the oral cavity and reach a large number of individuals, has become an important public health problem. Studies have demonstrated changes in pathway components BMP in various types of cancers as prostate, colon, breast, gastric and OSCCs. Is the current knowledge that these proteins may exert pro-tumor effect in more advanced stages of neoplastic development coming to favor progression and invasion tumor. The inhibition of the signaling pathway BMP-2 through its antagonists, have shown positive results of antitumor activity and use of Noggin may be a novel therapeutic target for cancer. Given this evidence and the few studies with BMP-2, Noggin and OSCC, the objective of this research was to evaluate the effect of BMP-2 and its antagonist Noggin on proliferation and migration cell in line of cell cultures of human tongue squamous cell carcinoma (SCC25). The study was divided in three groups, a control group, where SCC25 cells suffered no treatment, a BMP-2 group, in which cells were treated with 100ng/ml of BMP-2 and a group of cells that were treated with 100ng/ml of Noggin. For the proliferation assay and cell cycle were established three time intervals (24, 48 and 72 hours). Proliferative activity was investigated by trypan blue and cell cycle analysis by staining with propidium iodide flow cytometry. The potential for migration / invasion of SCC25 cells was performing by a cell invasion assay using Matrigel in a 48-hour interval. The proliferation curve showed a higher proliferation in cells treated with BMP-2 in 72 hours (p < 0.05), and lower overgrowth and cell viability in Noggin group. Recombinant proteins favored a greater percentage of cells in cell cycle phase Go/G1 with a statistically significant difference in the interval of 24 hours (p < 0.05). BMP- 2 produced a greater invasion of cells studied as well as its antagonist Noggin inhibits invasion of cells (p < 0.05). Thus, these results indicate that BMP-2 promotes malignant phenotype, dues stimulates proliferation and invasion of SCC25 cells and, its antagonist Noggin may be an alternative treatment, due to inhibit the tumor progression

Aspectos anatômicos do olho e neuroquímicos da retina do mocó (Kerodon rupestris)

The visual system is an important link between the animal and the environment, com profound influences on the habits and lifestyle in various habitats. Adaptive mechanismsto the temporal niche are present in the visual system of many vertebrates, involving changins in ocular dimensios and design, retinal cell distribution and organization of neurochemical circuits related to the retinal resolution or sensitivity. The sensory system of the eye is represented by the retina, whose organization is responsible by receipty, initial analysis, and transmission of the information to the brain. The knowledge of the position of the eyes in the head and the distribution of retinal cells allow to identify adaptive aspects of each species to its visual field, which is characteristic to the ecological niche it occupies. In this research, we study eye anatomical characteristics and retina neurochemical features of the rock cavy (Kerodon rupestris), a tipical Brazilian rodent from the suborder Hystricomorpha, family Caviidae. The rock cavy has lateral eyes well constitute bony orbit and well differentiated extrinsic muscle. The study of the descriptive and morphometric anatomy of the showed mean values of axial diameter 10.7±0,5mm and equatorial diameter 11.6±0.7mm. The pupil is slit shaped and the lens has mean axial diameter 5.4±0.03 mm, corresponding to ~45% of the axial diameter of the eye. The posterior nodal distance and the retinal magnification factor were estimated at 6.74 mm e 118 μm/grau, respectively. Flat mounts were processed for Nissl stain, and the topographic distribution of ganglion cells showed a moderate visual band, just below the optic disc, with higher density in the ventral retina. Retinal vertical sections and flat mounts were processed for immunohistochemistry to visualize tyrosine hydroxilase (TH) and thus two types of TH+ cells were detected. Type 1 cells had strong TH-immunoreactivity, the body cell varied from 120.047 to 269.373 μm2 stratifying in the sublamina 1 of the IPL. Type 2 cells were weakly TH-imunoreactive, had cell body located mostly in the IPL, varying from 54.848 to 177.142 μm2, constituting ~10% of the TH+ cells. Both cell types exhibited similar topographic distribution with higher density found in a horizontal band along of the naso-temporal axis in the dorsal retina. The total population of dopaminergic cells was 2,156±469,4 cells, occupying an average area of 198,164 μm2. The presence of cones and rods was detected by immunohistochemistry in vertical sections and flat mounts. S cones density is around 10 times smaller than L cones, with different degree of spatial organization. Other retinal neuronal populations of the rock cavy were also detected in vertical sections with specific markers. Comparative analysis of the anatomical characteristics of the rock cavy eye 12 suggest that it was designed to acquire higher sensitivity to light, at expense of image sharpness, compatible with a vision at mesopic conditions. Additionally, the distribution of the 2 subtypes of dopaminergic cells in a naso-temporal band in the dorsal retina seems suitable to a gain in sensitivity, coherent with an animal with predominantly crepuscular activity pattern

Condição peri-implantar e fatores associados às doenças peri-implantares em pacientes reabilitados no serviço odontológico da faculdade de odontologia da UFRN

A introdução dos implantes dentários osseointegrados como uma ferramenta na reabilitação oral de pacientes edêntulos e parcialmente edêntulos é uma realidade no cotidiano do cirurgião-dentista. Estudos reportam uma alta taxa de sucesso da utilização de implantes no tratamento reabilitador. Entretanto, outras investigações têm mostrado a perda desses implantes devido a infecções peri-implantares, como a mucosite e a peri-implantite. O objetivo deste trabalho foi avaliar a frequência das doenças peri-implantares e seus fatores associados em pacientes com implantes dentais em função reabilitados no serviço odontológico da Faculdade de Odontologia da UFRN. Foram examinados 155 indivíduos portadores de 523 implantes e 2718 dentes. Dentes e implantes foram avaliados por meio de sondagem periodontal, observando-se a profundidade de sondagem, retração gengival, bem como foram avaliados índices de placa visível (IPV) e sangramento gengival (ISG) e presença de supuração. Os dados foram armazenados em fichas clínicas e avaliados estatisticamente por meio da estatística descritiva e inferencial. A idade média dos pacientes foi de 54,05 (± 12,61) anos, sendo 79,4% do sexo feminino. As frequências da mucosite, peri-implantite e periodontite em indivíduos foram 54%, 28% e 50%, respectivamente. Dos 523 implantes avaliados, 43% tinham mucosite, 14% peri-implantite e 43% saúde. Os testes Qui-quadrado de Pearson e Exato de Fisher mostraram que as doenças peri-implantares estão associadas as doenças periodontais, uso de medicação, alterações sistêmicas número de implantes, IPV, ISG, ao tempo de função das próteses, região do implante, número de roscas expostas e faixa de mucosa queratinizada (p<0,05). A análise de regressão múltipla, através da regressão binária logística, constatou que indivíduos que faziam uso de medicação (OR = 1,784), com um ISG > 10% (OR = 1,742), com implantes instalados na maxila (OR = 2,654), onde a prótese sobre o implante tinham mais de 2 anos em função (OR = 3,144) e que radiograficamente apresentavam uma perda óssea atingindo a terceira rosca do implante (OR = 4,701) mostram uma associação positiva com as doenças peri-implantares de maneira que esses indivíduos têm mais chances de ter essas doenças. Os resultados sugerem que a frequência das doenças peri-implantares na população em estudo foi de 82% dos pacientes e que estas doenças estão associadas a fatores relacionados aos indivíduos como: a presença da doença periodontal, piores IPV e IS, alterações sistêmicas, uso de medicação e maior número de implantes; e a fatores locais relacionados aos implantes como: ausência ou faixa de mucosa menor que 2mm, implantes na maxila e na região anterior, perda óssea atingindo a terceira rosca do implante e a um tempo de reabilitação prótetica maior que 2 anos

Estudo da descelularização tecidual na produção de arcabouços biológicos para enxertos

The regeneration of bone defects with loss of substance remains as a therapeutic challenge in the medical field. There are basically four types of grafts: autologous, allogenic, xenogenic and isogenic. It is a consensus that autologous bone is the most suitable material for this purpose, but there are limitations to its use, especially the insufficient amount in the donor. Surveys show that the components of the extracellular matrix (ECM) are generally conserved between different species and are well tolerated even in xenogenic recipient. Thus, several studies have been conducted in the search for a replacement for autogenous bone scaffold using the technique of decellularization. To obtain these scaffolds, tissue must undergo a process of cell removal that causes minimal adverse effects on the composition, biological activity and mechanical integrity of the remaining extracellular matrix. There is not, however, a conformity among researchers about the best protocol for decellularization, since each of these treatments interfere differently in biochemical composition, ultrastructure and mechanical properties of the extracellular matrix, affecting the type of immune response to the material. Further down the arsenal of research involving decellularization bone tissue represents another obstacle to the arrival of a consensus protocol. The present study aimed to evaluate the influence of decellularization methods in the production of biological scaffolds from skeletal organs of mice, for their use for grafting. This was a laboratory study, sequenced in two distinct stages. In the first phase 12 mice hemi-calvariae were evaluated, divided into three groups (n = 4) and submitted to three different decellularization protocols (SDS [group I], trypsin [Group II], Triton X-100 [Group III]). We tried to identify the one that promotes most efficient cell removal, simultaneously to the best structural preservation of the bone extracellular matrix. Therefore, we performed quantitative analysis of the number of remaining cells and descriptive analysis of the scaffolds, made possible by microscopy. In the second stage, a study was conducted to evaluate the in vitro adhesion of mice bone marrow mesenchymal cells, cultured on these scaffolds, previously decellularized. Through manual counting of cells on scaffolds there was a complete cell removal in Group II, Group I showed a practically complete cell removal, and Group III displayed cell remains. The findings allowed us to observe a significant difference only between Groups II and III (p = 0.042). Better maintenance of the collagen structure was obtained with Triton X-100, whereas the decellularization with Trypsin was responsible for the major structural changes in the scaffolds. After culture, the adhesion of mesenchymal cells was only observed in specimens deccelularized with Trypsin. Due to the potential for total removal of cells and the ability to allow adherence of these, the protocol based on the use of Trypsin (Group II) was considered the most suitable for use in future experiments involving bone grafting decellularized scaffolds