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Abstract Aims Different regions of oral squamous cell carcinoma (OSCC) have particular histopathological characteristics, and the individual histological characteristics of the tumors are poorly understood. Therefore, calculating the proportion of tumor cells in different regions that allow assessment of the prognostic outcomes for OSCC patients would be of great clinical significance. Methods and Results We established an open‐source software‐based analytic pipeline that defines the inner tumor and invasive tumor front (ITF) in pancytokeratin‐stained whole slide images (WSIs) and quantifies the tumor‐stroma ratio (TSR) within the two regions. We applied this method to 114 patients with OSCC and predicted patient prognosis by the TSR. The proportion of tumor area in the inner tumor was generally higher than that in the ITF (p

Head and neck cancer ranks as the sixth most prevalent malignancy, constituting 5 % of all cancer cases. Its inconspicuous onset often leads to advanced stage diagnoses, prompting the need for early detection to enhance patient prognosis. Currently, research into early diagnostic markers relies predominantly on genomics, proteomics, transcriptomics, and other methods, which, unfortunately, necessitate tumor tissue homogenization, resulting in the loss of temporal and spatial information. Emerging as a recent addition to the omics toolkit, spatial metabolomics stands out. This method conducts in situ mass spectrometry analyses on fresh tissue specimens while effectively preserving their spatiotemporal information. The utilization of spatial metabolomics in life science research offers distinct advantages. This article comprehensively reviews the progress of spatial metabolomics in head and neck cancer research, encompassing insights into cancer cell metabolic reprogramming. Various mass spectrometry imaging techniques, such as secondary ion mass spectrometry, stroma-assisted laser desorption/ionization, and desorption electrospray ionization, enable in situ metabolite analysis for head and neck cancer. Finally, significant emphasis is placed on the application of presently available techniques for early diagnosis, margin assessment, and prognosis of head and neck cancer.

Objective: To clarify the effects on the development, position and morphology of the permanent successors of primary molars affected by apical periodontitis (AP). Method: A total of 132 panoramic radiographs of children aged from 4 to 10 were screened out and a total of 159 mandibular second primary molars with chronic apical periodontitis（AP）(93 males and 66 females) were analyzed. The maturation values of permanent successors were interpreted and scored according to Nolla's method and compared with normal ones'. The proportion of abnormalities in the morphology and orientation of permanent successors were counted, and the differences between men and women was analyzed. The distribution of various abnormalities in different age groups was also analyzed. Result: There were significant differences in development of permanent successors in this study compared with the normal ones' in all age groups, among which the differences were statistically significant in males aged in 4,5,7 groups and females aged in 4,6 (P

Abstract The survival rate of oral squamous cell carcinoma (OSCC) can be greatly improved if intervention could be initiated as early as possible. This poses a technical demand for developing a sensitive screening and specific in vitro diagnosis method for OSCC. Herewith, a large cohort consisting of 241 healthy contrast (HC) and 578 OSCC patients were recruited for conducting the rapid metabolic profiling of trace volume serum using conductive polymer spray ionization mass spectrometry (CPSI‐MS). Statistical analysis picked out 65 metabolite ions as potential characteristic markers for differentiating OSCC from HC. With the aid of a supporting vector machine (SVM), OSCC can be distinguished from HC with an accuracy of 98.0% by cross‐validation in the discovery cohort and 89.6% accuracy in the validation cohort. Furthermore, orthogonal partial least square‐discriminant analysis (OPLS‐DA) also initially showed the potential for OSCC staging, especially between T1/T2 and T3/T4 stages with an accuracy of 90.1%. CPSI‐MS combined with SVM or OPLS‐DA can not only quickly distinguish OSCC from HC but also predict the OSCC progression from T1/2 to T3/4 stages in a few minutes, making it a promising tool for both screening and diagnosing high‐risk population. Key points Sixty‐five characteristic metabolite ions significantly changed in OSCC serum metabolic profile compared to that in the HC group. CPSI‐MS combined with SVM achieved 89.6% accuracy on the validation cohort for OSCC prediction. CPSI‐MS/OPLS‐DA can distinguish T1/T2 from T3/T4 stages with an accuracy of 90.1% by cross‐validation.

Background: Although there is consensus that the optimal safe margin is ≥ 5mm, obtaining clear margins (≥5 mm) intraoperatively seems to be the major challenge. We applied a molecular diagnostic method at the lipidomic level to determine the safe surgical resection margin of OSCC by desorption electrospray ionisation mass spectrometry imaging (DESI-MSI). Methods: By overlaying mass spectrometry images with hematoxylin-eosin staining (H&E) from 18 recruited OSCC participants, the mass spectra of all pixels across the diagnosed tumour and continuous mucosal margin regions were extracted to serve as the training and validation datasets. A Lasso regression model was used to evaluate the test performance. Findings: By leave-one-out validation, the Lasso model achieved 88.6% accuracy in distinguishing between tumour and normal regions. To determine the safe surgical resection distance and margin status of OSCC, a set of 14 lipid ions that gradually decreased from tumour to normal tissue was assigned higher weight coefficients in the Lasso model. The safe surgical resection distance of OSCC was measured using the developed 14 lipid ion molecular diagnostic model for clinical reference. The overall accuracy of predicting tumours, positive margins, and negative margins was 92.6%. Interpretation: The spatial segmentation results based on our diagnostic model not only clearly delineated the tumour and normal tissue, but also distinguished the different status of surgical margins. Meanwhile, the safe surgical resection margin of OSCC on frozen sections can also be accurately measured using the developed diagnostic model. Funding: This study was supported by Nanjing Municipal Key Medical Laboratory Constructional Project Funding (since 2016) and the Centre of Nanjing Clinical Medicine Tumour (since 2014).

Dysregulated amino acids metabolism reciprocally interplays with evolutionary phenotypic characteristics of cancer cells to enhance metastasis. The high metastasis potential of oral squamous cell carcinoma (OSCC) can manifest with perineural invasion (PNI). We here aimed to determine the role of amino acids metabolism in OSCCs with different PNI statuses. Targeted metabolomics was used to quantify 48 amino acids in 20 fresh OSCC samples and 25 amino acids were successfully detected, within which 9 were significantly up-regulated in PNI positive (PNI+) samples. As its highest area under the curve value (0.9063), l-asparagine was selected as the biomarker to distinguish PNI+ from PNI negative (PNI−). Then, the key enzyme of l-asparagine, asparagine synthetase (ASNS), was investigated using immunohistochemistry with 86 OSCC patients. The results showed that ASNS mainly expressed in tumor epitheliums and positively correlated with lymph node metastasis and PNI. Moreover, subgroup survival analysis revealed that ASNS expression combined with PNI status significantly improved their prognostic value, which was confirmed by the TCGA OSCC cohort (n = 279). To validate whether ASNS promotes PNI, we determined ASNS expression levels in five OSCC cell lines and one normal oral keratinocyte, and HSC3 showed the lowest ASNS level but CAL33 had the highest. Therefore, HSC3 and CAL33 (or PBS as control) were selected and injected separately into sciatic nerves to construct the in vivo PNI mouse models. Although both models eventually developed the hind-limb paralysis, nerve dysfunction in the CAL33 model progressed significantly earlier than HSC3 (Day 9 vs. Day 24). Besides, CAL33 migrated significantly farther than HSC3 in the nerve microenvironment (P = 0.0003), indicating high ASNS expression is indispensable for OSCC progression, especially PNI formation, through l-asparagine metabolism alteration. This study provides novel insights into how amino acids metabolism disorders alter tumor neurotropism which helps cancer metastasis.

The 5-year survival rate for oral squamous cell carcinoma (OSCC), one of the most common head and neck cancers, has not improved in the last 20 years. Poor prognosis of OSCC is the result of failure in early and precise diagnosis. Metabolic reprogramming, including the alteration of the uptake and utilisation of glucose, amino acids and lipids, is an important feature of OSCC and can be used to identify its biomarkers for early and precise diagnosis. In this review, we summarise how recent findings of rewired metabolic networks in OSCC have facilitated early and precise diagnosis of OSCC.

BACKGROUND: Cyclin D1 (CCND1) has been associated with chemotherapy resistance and poor prognosis. In this study, we tested the hypothesis that CCND1 expression determines response and clinical outcomes in locally advanced head and neck squamous cell carcinoma (HNSCC) patients treated with neoadjuvant chemotherapy followed by surgery and radiotherapy. METHODOLOGY AND FINDINGS: 224 patients with HNSCC were treated with either cisplatin-based chemotherapy followed by surgery and radiotherapy (neoadjuvant group, n = 100) or surgery and radiotherapy (non-neoadjuvant group, n = 124). CCND1 expression was assessed by immunohistochemistry. CCND1 levels were analyzed with chemotherapy response, disease-free survival (DFS) and overall survival (OS). There was no significant difference between the neoadjuvant group and non-neoadjuvant group in DFS and OS (p = 0.929 and p = 0.760) when patients treated with the indiscriminate administration of cisplatin-based chemotherapy. However, in the neoadjuvant group, patients whose tumors showed a low CCND1 expression more likely respond to chemotherapy (p