Hsa_circRNA_101036 aggravates hypoxic-induced endoplasmic reticulum stress via the miR-21-3p/TMTC1 axis in oral squamous cell carcinoma.

Objective: Circular RNAs (circRNAs) have been identified as potential biomarkers and therapeutic targets for various types of cancer, including Oral squamous cell carcinoma (OSCC). Hsa_circRNA_101036 was found to function as a cancer suppressor gene in OSCC; however, the underlying regulatory mechanism remains unclear. We investigated the role of hsa_circRNA_101036 in OSCC development and progression and explored its potential as a therapeutic target. Methods: We performed a bioinformatics analysis and used experimental approaches to investigate the regulatory mechanism of hsa_circRNA_101036. The database StarBase v.2.0 was used to predict potential target-miRNAs of hsa_circRNA_101036. The levels of hsa_circRNA_101036, miR-21-3p, and TMTC2 expression in samples of OSCC cancer tissue (n = 15) and adjacent tissue (n = 15) were determined. We also examined the effects of hsa_circRNA_101036 overexpression on OSCC cell lines by using cell viability, migration, and invasion assays. The proportions of apoptotic cells and the reactive oxygen species (ROS) levels were analyzed by flow cytometry. We also investigated how hsa_circRNA_101036 overexpression affected the levels of miR-21-3p and TMTC2, and endoplasmic reticulum (ER) stress in OSCC cells. Results: The levels of hsa_circRNA_101036 and TMTC2 expression were significantly lower, while miR-21-3p expression was higher in tumor tissues and OSCC cells when compared to adjacent tissues and normal oral fibroblasts, respectively. The levels of HIF-1α and miR-21-3p expression were significantly increased under conditions of hypoxia, while the levels of hsa_circRNA_101036 and TMTC2 were decreased. The expression levels of proteins associated with ER stress, the proportions of apoptotic cells, and the levels of ROS were all increased by hypoxia stimulation. In addition, overexpression of hsa_circRNA_101036, but not mutant hsa_circRNA_101036, was found to enhance the effect of hypoxia on HSC3 and OECM-1 cells. Hsa_circRNA_101036 overexpression suppressed tumor growth and induced ER stress. Finally, knockdown of miR-21-3p had the same effect as overexpression of hsa_circRNA_101036. Conclusion: Our findings suggest that hsa_circRNA_101036 plays a critical role in the development and progression of OSCC. Overexpression of hsa_circRNA_101036 aggravated ER stress, and increased cell apoptosis and ROS production in OSCC under hypoxic conditions. Hsa_circRNA_101036 up-regulated TMTC2 expression by sponging miR-21-3p in OSCC.

Gene mutation analysis of oral submucous fibrosis cancerization in Hainan Island.

Objective: The sequencing panel composed of 61 target genes was used to explore the related mutation genes of oral squamous cell carcinoma (OSCC) and oral submucous fibrosis (OSF) cancerization, so as to provide a theoretical basis for the early diagnosis of oral submucous fibrosis cancerization, find the most important mutations in OSF cancerization, and more targeted prevention of OSF cancerization. Methods: A total of 74 clinically diagnosed samples were included, including 36 cases of OSCC and 38 cases of OSF cancer patients. DNA was extracted, and targeted gene panel sequencing technology was used to analyze the gene frequency of pathogenic mutation sites in clinical samples. Results: Gene panel sequencing analysis showed that there were 69 mutations in 18 genes in OSCC and OSF cancerous specimens. The results of gene panel sequencing were screened, and 18 mutant genes were finally screened out and their mutation frequencies in the samples were analyzed. According to the frequency of gene mutations from high to low, they were TP53, FLT4, PIK3CA, CDKN2A, FGFR4, HRAS, BRCA1, PTPN11, NF1, KMT2A, RB1, PTEN, MSH2, MLH1, KMT2D, FLCN, BRCA2, APC. The mutation frequency of FLT4 gene was significantly higher than that of OSCC group (P < 0.05). Conclusion: FLT4 gene may be related to OSF cancerization and is expected to be an early diagnostic biomarker for OSF cancerization.

CircFKBP5 Suppresses Apoptosis and Inflammation and Promotes Osteogenic Differentiation.

OBJECTIVES: Dental pulp stem cells (DPSCs) are a type of mesenchymal stem cell possessing self-renewal and multilineage differentiation capabilities. The dysfunction of DPSCs is related to the pathologic process of pulpitis. The participation of circular RNAs (circRNAs) in DPSC differentiation has been identified. This work focussed on exploring the functions and mechanism of circFKBP5 in DPSC dysfunction evoked by lipopolysaccharide (LPS). MATERIALS AND METHODS: The viability and apoptosis of human DPSCs (hDPSCs) were determined using Cell Counting Kit-8 assay and flow cytometry. Inflammation was analysed by measuring the release of inflammatory cytokines. The osteogenic differentiation of hDPSCs was investigated by performing alkaline phosphatase (ALP) staining and alizarin red S staining and detecting the changes of ALP and runt-related transcription factor 2 (RUNX2) proteins. The dual-luciferase reporter, RNA immunoprecipitation (RIP), and pull-down assays were used to confirm the binding between miR-708-5p and circFKBP5 or G-protein-coupled receptor (GPCR)-kinase interacting protein 2 (GIT2). RESULTS: CircFKBP5 expression was decreased in hDPSCs and, functionally, reexpression of circFKBP5 attenuated LPS-induced apoptosis, inflammation, and inhibition of proliferation ability and osteogenic differentiation in hDPSCs. Mechanistically, circFKBP5 acted as a sponge for miR-708-5p, which was verified to target GIT2. LPS induced miR-708-5p expression in hDPSCs, and knockdown of miR-708-5p protected against LPS-evoked hDPSC dysfunction. Besides, GIT2 expression was decreased in hDPSCs after LPS treatment. Rescue experiments showed that GIT2 could mediate the protective functions of circFKBP5 on hDPSCs under LPS treatment. CONCLUSIONS: CircFKBP5 could protect against LPS-induced apoptosis, inflammation, and osteogenic differentiation inhibition in hDPSCs via the miR-708-5p/GIT2 axis.

VEGF gene transfection restores the angiogenesis of oral submucous fibrosis in mice.

BACKGROUND: To explore the effectiveness of adenovirus-enhanced green fluorescent protein-vascular endothelial growth factor165 (AD-EGFP-VEGF165) transfection on fibroblasts from mice, and we assessed whether VEGF165 restores the angiogenesis of oral submucous fibrosis (OSF) in mice. METHODS: AD-EGFP-VEGF165 and AD-EGFP were transfected into fibroblasts from mouse buccal tissues in vitro. The expression of VEGF before and after transfection was detected by RT-qPCR and ELISA in each group of fibroblasts. Fifteen OSF mice (pre-experimental construction) were randomly divided into 3 groups, and equal amounts of AD-EGFP-VEGF165 virus, AD-EGFP virus, and saline were injected into the buccal submucosal tissue of OSF mice. The expression of VEGF and local tissue angiogenesis were observed and measured in each group of animals. RESULTS: The Ad-EGFP-VEGF165-transfected fibroblasts increased human and mouse VEGF expression compared to the Ad-EGFP group and control group (P<0.05). The buccal submucosal tissue of mice was injected with Ad-EGFP-VEGF165 after the 6th day, and the expression of VEGF was effectively expressed in AD-EGFP-VEGF165 group (P<0.05), while no positive expression observed in other groups. and the number of microvessels in the AD-EGFP-VEGF165 group increased significantly compared to the other groups (P<0.05). CONCLUSIONS: Ad-EGFP-VEGF165 can be successfully transfected into fibroblasts from mice, and restored the angiogenesis of OSF in mice.

Development of a mouse model of arecoline-induced oral mucosal fibrosis.

OBJECTIVE: To develop a BALB/c mouse model of oral submucous fibrosis (OSF) induced by arecoline and to exhibit an accumulation of collagen and angiogenesis changes. METHODS: BALB/c mice were randomly assigned to either the control (distilled water) or experimental group (arecoline) (n = 40). Eight mice from each group were sacrificed every 4 weeks since 8 weeks post treatment. Changes in histopathologic features, levels of collagen type I and collagen type III, and angiogenesis were measured. RESULTS: In the 8th week, epithelium atrophy, collagen cumulation and micrangium pathologic changes in the lamina propria were observed in the oral mucosa. In the 20th week, hyaline degeneration of the connective tissues was observed on the tongue and palate mucosa. The angiogenesis and collagen type I changed significantly as the diseases advanced (P < 0.05); however, collagen type III was not statistically different. CONCLUSIONS: An OSF model involving mice can be rapidly induced by drinking a high-dose of arecoline. OSF angiogenic changes in mice primarily decrease and collagen accumulation is mainly collagen type I.

Improving Surgeons' Comfort With Prismatic Glasses During Cleft Palate Surgery: Preliminary Findings.

PURPOSE: We aimed to observe the effect of prismatic glasses on improving surgeons' comfort during cleft palate surgery. MATERIALS AND METHODS: A within-subjects design was used. We included 3 oral-maxillofacial surgeons and 6 patients with complete cleft palate in the study. One group of cleft palate patients (3 complete cleft palates) was allocated to each of the 3 surgeons not wearing prismatic glasses, and another similar group of cleft palate patients was allocated to the same 3 surgeons wearing prismatic glasses. The push-back method was performed in all cleft palate patients by all surgeons. The degree of neck flexion exhibited by all surgeons was digitally video recorded. Screen-capture images of the video recordings were collected, and neck flexion in all video recordings was analyzed. All surgeons completed a questionnaire based on a visual analog scale to assess their discomfort symptoms of the neck, shoulders, and back. Operative time and bleeding volume were recorded to assess operational efficiency. RESULTS: Use of prismatic glasses significantly reduced surgeons' working time spent in pronounced neck flexion during cleft palate surgery (P < .05), and there was a statistically significant reduction in the visual analog scale discomfort scores for the neck, back, and shoulders with the use of prismatic glasses (P < .05). However, no significant difference was found in operational time (P = .337) and bleeding volume (P = .183) attributable to the presence or absence of prismatic glasses. CONCLUSIONS: An ergonomic approach to cleft palate surgery in which surgeons wore prismatic glasses improved neck, back, and shoulder comfort.