Angiogenesis and mast cells as predictive factors in behavior of Odontogenic cysts

Purpose: The aim of this study was to verify the density of mast cells (MCs) and microvessels in odontogenic cysts. Furthermore, the correlation between MCs and microvessels was evaluated to assess the contribution of MCs to angiogenesis and growth of odontogenic cysts. This approach may be a basis for the development of future pharmaceuticals addressed to MCs performance to manage odontogenic cysts. Methods:45 cases of odontogenic cysts consisting of 15 radicular cysts (RCs), 15 dentigerous cysts (DCs) and 15 odontogenickeratocysts (OKCs) were included in this study. Five high power fields in connective tissue wall were counted for each sample. Moreover, a total mean of five fields was calculated to calculate MVD and MCD of the samples. Results: RC showed the highest mean numbers of microvessels (p<0.05) and OKC showed the highest mean number of mast cells. A positive correlation between the numbers of MCs and microvessels was observed. Conclusion: Increase in angiogenesis of OKC confirms that aggressive lesions require a larger number of blood vessels and highlights the concept of unique clinical behavior of this cyst. In OKCs and DCs cysts, the MVD is closely related to of the inflammatory infiltrate. Mast cells are imperative in the pathogenesis of odontogenic cysts. Mast cells have a role in promoting the growth of odontogenic cysts, particularly OKC. Further studies on the in vivo functions of MCs will make the concept more clear. Angiogenesis and mast cells as predictive factors in behavior of Odontogenic cysts Mansoura Journal of Dentistry2020;7(28):43-49. 44 Omnia Mohamed Recently,mast cells were recognized in the pathogenesis of more aggressive pathologic lesions . Mast cells have an inhibitory role on the development of pathological lesions. However, stimulatory role of mast cells in the growth of pathological lesions is more prevalent and obvious than their inhibitory effect . With respect to several roles of mast cells such as participation in inflammation, degradation of extracellular matrix and bone resorption, previous studies have identified mast cells in odontogenic cysts, but there were limited studies about the role of mast cells in the pathogenesis of odontogenic cysts . There is a hypothesis that the more aggressive behavior of odontogenickeratocysts is related at least, partly, to distribution of mast cells. However, their pathogenesis and mechanism of expansion and enlargement have not been evaluated . The aim of this study was to determine the density of microvessels and MCs in odontogenic cysts. Correlate the microvessel density with their corresponding mast cells density in the three types of cysts, in order to detect their possible role in the variable behavior of these odontogenic cysts. Material and method : In this work, the study samples included 45 paraffin blocks of odontogenic cysts (15RC, 15 DC, 15 OKC) that were collected from archival files of Oral Pathology Departments, Faculties of Dentistry, Mansoura University. Four micrometer thick tissue sections were stained through standard immunohistochemical (CD 105) and histochemical (Toulidine blue) staining procedures according to the instructions of the manufacturer. The slides were deparaffinized in xylene and then hydrated in graded alcohol series. For histochemical evaluation, the sections were stained with freshly prepared 1% toluidine blue (adjusted at ph 2.0 – 2.5) for 2-3 minutes, using Toluidine blue staining protocol for mast cells . Following dehydration and clearing. For CD105 marker, the endogenous peroxidase activity was blocked by incubating the slides with 3% hydrogen peroxide in methanol for 30 minutes. The antigen retrieval for CD105 was carried out by treating sections with proteins-K for 5 minutes. To prevent nonspecific reactions, sections were incubated with 10% serum for 10 minutes. Mouse Monoclonal Anti-Human CD105 antibody (Catalog No. AM441-5M: Ready-to-Use Antibody for Use with BioGenex Super Sensitive Detection Systems, Fremont, CA 94538 USA) were used as primary antibodies in this study. CD105 antibody was incubated at room temperature for 90 minutes in a humidifying chamber, followed by incubation with secondary biotinylated antibodies and streptavidin for 15 min each. Diaminobenzidine was applied to produce brown staining followed by counterstaining with Mayer‟s hematoxylin. After each step, the slides were put in phosphate-buffered solution (PBS). For the negative control, the primary antibody was eliminated and replaced with PBS. For immunohistochemical and histochemical counting using the microvascular count technique, according to the method suggested by Weidner et al. . MVD and MCD was assessed as the mean number of microvessels and MCs per high power field. The field size for 400 magnification (40 objectives and 10 ocular) was approximately 0.18 mm. Scores of overall CD105 and Touilidine blue expression were represented as mean density/mm2 ± SD for quantitative variables using SPSS (Statistical package for Social Siences) software. Comparisons among the experimental groups were done using One-way analysis of variance test (ANOVA) (p˂0.05). To compare the number of microvessels and MCs between inflamed and non-inflamed DCs and OKCs, independent t-test was used. Pearson correlation coefficient test was used to determine the correlation between MVD and MCD. A p-value of <0.05 was considered statistically significant