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Introduction: Dental age estimation has always been a very important and very susceptible subject in forensic science, as it is an essential requirement in living, dead human individual’s remains. Aspartic acid racemization is considered to be one of the advanced, reliable, accurate, and complex biochemical methods. There will be an amendment of Levo aspartic acid to dexto aspartic acid when the age advances. To estimate the dental age of single-rooted teeth using aspartic acid racemization. Material and Methods: Thirty anterior teeth were collected from maxillary and mandibular arches distributed into four groups. All teeth were analyzed for Levo (l) and Dextro (d) form of aspartic acid using a high-performance liquid chromatography technique and the ratio of amino acid racemization for each sample was determined. The obtained data were subjected to step-wise linear regression analysis for the ratio of d and l aspartic acid against actual age with estimated age. Results: Distribution of teeth about age maximum samples was collected at 51 to 60 years. l-aspartic acid ratio was more in the younger age group but when the age increases d-aspartic acid ratio was increased. Linear regression analysis showed maximum samples showed very close to the actual age (30%). Conclusion: By analyzing the levels of aspartic acid using amino acid racemization, we can estimate an individual with maximum accuracy. Accretion of d-aspartic acid in dentin is synchronous with the aging of an individual and this method can be used as one of the precise methods for dental age estimation.

This study aimed to develop an in vitro three-dimensional (3D) cell culture model of oral carcinogenesis for the rapid, scalable testing of chemotherapeutic agents. Spheroids of normal (HOK) and dysplastic (DOK) human oral keratinocytes were cultured and treated with 4-nitroquinoline-1-oxide (4NQO). A 3D invasion assay using Matrigel was performed to validate the model. RNA was extracted and subjected to transcriptomic analysis to validate the model and assess carcinogen-induced changes. The VEGF inhibitors pazopanib and lenvatinib were tested in the model and were validated by a 3D invasion assay, which demonstrated that changes induced by the carcinogen in spheroids were consistent with a malignant phenotype. Further validation was obtained by bioinformatic analyses, which showed the enrichment of pathways associated with hallmarks of cancer and VEGF signalling. Overexpression of common genes associated with tobacco-induced oral squamous cell carcinoma (OSCC), such as MMP1, MMP3, MMP9, YAP1, CYP1A1, and CYP1B1, was also observed. Pazopanib and lenvatinib inhibited the invasion of transformed spheroids. In summary, we successfully established a 3D spheroid model of oral carcinogenesis for biomarker discovery and drug testing. This model is a validated preclinical model for OSCC development and would be suitable for testing a range of chemotherapeutic agents.

Three-dimensional (3-D) cell culture models, such as spheroids, organoids, and organotypic cultures, are more physiologically representative of the human tumor microenvironment (TME) than traditional two-dimensional (2-D) cell culture models. They have been used as in vitro models to investigate various aspects of oral cancer but, to date, have not be widely used in investigations of the process of oral carcinogenesis. The aim of this scoping review was to evaluate the use of 3-D cell cultures in oral squamous cell carcinoma (OSCC) research, with a particular emphasis on oral carcinogenesis studies. Databases (PubMed, Scopus, and Web of Science) were systematically searched to identify research applying 3-D cell culture techniques to cells from normal, dysplastic, and malignant oral mucosae. A total of 119 studies were included for qualitative analysis including 53 studies utilizing spheroids, 62 utilizing organotypic cultures, and 4 using organoids. We found that 3-D oral carcinogenesis studies had been limited to just two organotypic culture models and that to date, spheroids and organoids had not been utilized for this purpose. Spheroid culture was most frequently used as a tumorosphere forming assay and the organoids cultured from human OSCCs most often used in drug sensitivity testing. These results indicate that there are significant opportunities to utilize 3-D cell culture to explore the development of oral cancer, particularly as the physiological relevance of these models continues to improve.