ABSTRACT
The selection of housekeeping genes is critical for gene expression studies. To address this issue, four candidate housekeeping genes, including several commonly used ones, were investigated in oral squamous cell carcinoma cell lines. A simple quantitative RT-PCR approach was employed by comparing relative expression of the four candidate genes within two cancerous cell lines (HN6 and HN31) and one noncancerous cell line (HaCaT) treated or not with EGF and TGF-beta1. Data were analyzed using ANOVA followed by the NormFinder software program. On this basis, stability of the candidate housekeeping genes was ranked and non statistical differences were found using ANOVA test. On the other hand, the NormFinder was able to show that GAPD and TUBB presented the less variable results, representing appropriated housekeeping genes for the samples and conditions analyzed. In conclusion, this study suggests that the GAPD and the TUBB represent adequate normalizers for gene profiling studies in OSCC cell lines, covering, respectively, high and low expression levels genes.
Subject(s)
Carcinoma, Squamous Cell/genetics , Gene Expression Profiling/methods , Glyceraldehyde-3-Phosphate Dehydrogenases/genetics , Mouth Neoplasms/genetics , Reverse Transcriptase Polymerase Chain Reaction/methods , Tubulin/genetics , Cell Line, Tumor , Humans , Reference Standards , Reverse Transcriptase Polymerase Chain Reaction/standardsABSTRACT
OBJECTIVE AND METHODS: Among the cells involved in immune and inflammatory responses in periodontal disease, mast cells have been shown to be capable of generating a large number of biologically active substances. The present study was undertaken to identify and quantify the presence of mast cells in different stages of human periodontal disease using histochemical (toluidine blue) and immunohistochemical (tryptase-positive mast cells) techniques. RESULTS: Mast cell densities (cells per mm(2)) were significantly increased in chronic periodontitis/gingivitis lesions compared with clinically healthy gingival tissues (Health) uniquely by immunohistochemical technique. Interestingly, mast cells were distributed specially in close apposition to mononuclear cells. CONCLUSIONS: In human periodontal disease there is an increase in the number of mast cells that may be participating either in the destructive events or in the defense mechanism of periodontal disease via secretion of cytokines, including perpetuation of the Th2 response, and cellular migration and healing processes.
Subject(s)
Gingivitis/immunology , Mast Cells , Periodontitis/immunology , Analysis of Variance , Cell Count , Chronic Disease , Gingivitis/pathology , Histocytochemistry , Humans , Immunoenzyme Techniques , Periodontitis/pathology , Staining and Labeling/methodsABSTRACT
In the periapex, the interaction among inflammatory cells and microorganisms and their products results both in specific and nonspecific immune responses. Many studies have reported quantitative analysis of the immunocompetent cells in periapical lesions, but the exact ratio of these cells in cysts and granulomas remains unknown. In the present study, we undertook a quantitative analysis of CD68+ cells and CD8+ T cells in human periapical granulomas and cysts. Immunoperoxidase staining revealed that CD68+ cells were present in both lesions, with no statistically significant difference, mainly distributed in the inner portion of the lesion, where the inflammation site is more active. On the other hand, CD8+ lymphocytes were more numerous in cysts. Thus, it appears that CD8+ T cells may play a more important role in a later phase of periapical lesion progression, probably exerting regulatory or cytotoxic functions in cellular immune response, which may lead to the stabilization of these lesions.
