Exploration of the underlying mechanisms of tumor occurrence and development, as well as evaluation of the efficacy of anticancer drug treatments, relies on various research models both in vivo and in vitro. Over the past few decades, with the rapid advancement of biomedical technology, significant achievements have been made in this field. Gene detection technology has progressed from a single-gene perspective to multi-gene approaches, resulting in rapid development of bioinformatics and transformation of the conceptual understanding of malignant tumors. Moreover, in vitro cell research models have evolved from monolayer two-dimensional and primary cultures to three-dimensional configurations, which better imitate the cellular interactions and functions within tumor tissues. Furthermore, in vivo animal research models have transitioned from traditional carcinogen induction and cell or tissue xenografts to genetically engineered animal models or xenograft models, enabling targeted investigation into the roles of relevant genes in the occurrence and development of tumors. Clinical research has shifted from simple retrospective to prospective studies, including phase Ⅰ/Ⅱ/Ⅲ clinical trials, investigator-initiated clinical trials, and real-world clinical trials. The major shortcomings of current malignant tumor research models include their singularity, insufficient simulation of the tumor microenvironment, disparities between animal models and human tumors, and the lack of consideration for personalized medicine. Further research and optimization of the models are still needed in the future, along with more effective integration of different models to form an optimized comprehensive experimental model system. This review systematically examines and comprehensively overviews the evolution of malignant tumor research models with the aim of providing more references to researchers engaged in oncology research.

Objective To investigate the impact of metabolic labeling on Porphyromonas gingivalis (Pg) and compare the imaging effects of two fluorescent probes. Methods This study was reviewed by the unit Ethics Committee and was approved by the Experimental Animal Welfare Ethics Branch of the Unit Experimental Biomedical Ethics Committee. Pg integrated N-azidoacetylgalactosamine (Ac₄GalNAz) via a bioorthogonal reaction and was labeled with Cy5-DBCO or Cy7-DBCO via a click chemistry reaction. The bacteria were divided into Pg group (control, not fluorescently labeled), Cy5-Pg group (tagged by Cy5-DBCO), and Cy7-Pg group (tagged by Cy7-DBCO). A live/dead staining kit was applied to test the viability of Pg, Cy5-Pg, and Cy7-Pg. The mRNA levels of interleukin-6 (IL-6) and IL-8 and cell proliferation were examined in human gingival fibroblasts (HGFs) after the challenge of Cy5-Pg, Cy7-Pg, or Pg. To investigate the stability of metabolic labeling, Cy5-Pg or Cy7-Pg was cocultured with Escherichia coli (E. coli). Cy5-Pg and Cy7-Pg signal intensity with serial dilutions were examined using an in vivo imaging system (IVIS). Finally, C57BL/6J mice were orally administered Cy5-Pg or Cy7-Pg for IVIS detection, and the signal-to-background ratios were calculated. Results Metabolic labeling could be applied to label live Pg in vitro. The optimal labeling concentrations for Cy5 and Cy7 were 20 μmol/L and 30 μmol/L, respectively. The area ratios of live to dead bacteria were approximately 2.0 in the three groups (F = 0.318, P>0.05). After a 6-h challenge with Cy5-Pg, Cy7-Pg, or Pg, the mRNA levels of HGFs increased by 7.86-, 7.46-, and 6.56-fold for IL-6, respectively (F = 40.886, P<0.001) and 12.43-, 13.03-, and 13.71-fold for IL-8 (F = 18.781, P<0.01), were spectively, compared to that of the Ctrl group, which was not challenged by bacteria, where no significant differences were observed among the three groups (P>0.05). HGFs were further challenged by Cy5-Pg, Cy7-Pg, or Pg at different MOIs, and cell proliferation was significantly inhibited (MOI = 10⁴∶1, F = 153.52, P<0.001; MOI = 10⁵∶1, F = 331.21, P<0.001; MOI = 10⁶∶1, F = 533.65, P<0.001), with no significant differences among the three groups (P>0.05). Within 24 h of co-culturing Cy5-Pg or Cy7-Pg with E. coli, minimal E. coli was detected. The intensities of Cy5 and Cy7 remained stable for 3 h. Additionally, the fluorescence signal intensities of Cy5 and Cy7 were linearly correlated with the concentration (R² = 0.97). After oral gavage of Cy5-Pg or Cy7-Pg in mice for the abdominal region at 1 h and 3 h, the signal-to-background ratios of Cy7-Pg were approximately 4.24-fold (t = 6.893, P<0.01) and 3.77-fold (t = 4.407, P<0.05) higher, respectively, than those of Cy5-Pg. For the isolated gastrointestinal tracts at 3 h, the signal-to-background ratio of Cy7-Pg was 5.19-fold higher than that of Cy5-Pg (t = 4.418, P<0.05). Conclusions Metabolic labeling did not significantly affect viability, immunomodulatory ability, and toxicity. The imaging effect of Cy7 on IVIS was better than that of Cy5. Our study provided experimental evidence for the correlation between periodontitis and overall health.

Objective To study the effect of the combinational use of miR-34a-functionalized Bio-Oss^® bone powder with transglutaminase crosslinked gelatin (Col-Tgel) on the osteoblastic differentiation of bone marrow mesenchymal stem cells (BMSCs) and bone defect healing after irradiation. Methods The experiment was approved by the Animal Ethics Committee. BMSCs were isolated from the bone marrow of 2-week-old Sprague-Dawley (SD) rats and identified. After reaching 80% confluence, BMSCs were irradiated with 2 Gy of X-ray radiation to establish a radiation-damaged BMSC model for further experimentation. 2.5 μL or 5 μL of Col-Tgel was mixed with 10 mg of Bio-Oss^® (P) to prepare PG-2.5 and PG-5. The optimal proportion of Bio-Oss^® (P) and Col-Tgel was determined through in vitro and in vivo experiments. Cy3-labeled agomiR-34a, agomiR-34a, or agomiR NC was mixed with lipofectamine 2000 and added to 10 mg of Bio-Oss^® (P). The mixtures were lyophilized, and 2.5 μL Col-Tgel was added to each group of lyophilized Bio-Oss^®/lipofectamine/miRNA complexes or to 10 mg of Bio-Oss^® to obtain PG-Cy3-miR-34a, PG-miR-34a, PG-miR NC, and PG. Irradiated BMSCs were cocultured with PG-Cy3-miR-34a to evaluate cellular uptake of Cy3-agomiR-34a using confocal microscopy. Then, irradiated BMSCs were cocultured with PG-miR-34a, PG-miR NC, and PG. The expression of miR-34a was tested by RT-qPCR and cell proliferation was tested by CCK-8 assay. After 14 days of osteogenic induction, the mRNA expression of Runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), and osteocalcin (OCN) was tested by RT-qPCR. The bilateral tibias of 8-week-old SD rats were irradiated with a single dose of 15 Gy of X-ray radiation. Three weeks later, tibial defects with a diameter of 3 mm and a depth of 2 mm were created 2-3 mm below the epiphyseal line in the tibial metaphysis. The composite bone substitute materials of PG-miR-34a, PG-miR NC, and PG were implanted into the defect area. Eight weeks after implantation, the tibias were harvested and evaluated for bone regeneration using micro-CT analysis and HE staining. Results The results demonstrated that 2 Gy irradiation adversely affected the osteogenic differentiation capacity of BMSCs, evidenced by the decreased ALP staining and number of mineralized nodules stained with Alizarin red in the irradiated group compared to the non-irradiated group. The composite material consisting of 10 mg Bio-Oss^® and 2.5 μL Col-Tgel exhibited good osteogenic induction capability and handling properties and was used for subsequent experiments. The PG-Cy3-miR-34a could deliver the loaded Cy3-agomiR-34a into irradiated BMSCs. PG-miR-34a enhanced the expression of miR-34a in irradiated BMSCs without affecting cell proliferation. PG-miR-34a significantly upregulated the expression of osteogenic-related genes, including Runx2, ALP, and OCN. In the experiment of bone defect healing in irradiated tibias, micro-CT analysis showed that PG-miR-34a group had a higher bone volume in the bone defect area compared to other groups. The HE staining results also confirmed that implantation of PG-miR-34a can promote the healing of bone defects in irradiated tibias. Conclusion The combinational use of miR-34a-functionalized Bio-Oss^® bone powder with Col-Tgel could promote the osteogenic differentiation of irradiated BMSCs and enhance bone regeneration in irradiated bone defects.

Objective To employ next-generation sequencing (NGS) to analyze differentially expressed mRNAs in the gingival tissue of hypertensive rats with or without periodontitis to provide a theoretical basis for the prevention and treatment of hypertension with periodontitis. Methods After obtaining approval from the Animal Experiment Ethics Committee, a hypertensive rat model was established by administering high-salt feed containing 8%（w/w） NaCl, and a periodontitis rat model was established by ligating the first molar of the mandibular region using 3-0 sterile silk thread. Rat models of the normal control (N), hypertension (H), and hypertension with periodontitis (PH) groups were established. The blood pressure, heart rate, alveolar bone resorption, and number of osteoclasts in the alveolar bone were measured, before harvesting the gingival tissues from the three groups for NGS to analyze the expression of significantly different genes. Gene ontology (GO) enrichment analysis was performed for all significantly differentially expressed genes between the H and PH groups. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was performed. Key genes were screened by protein-protein interaction (PPI) networks, and the key gene expression in each group was verified using immunohistochemistry (IHC). The expression of key genes in the systemic circulation of each group was analyzed using enzyme-linked immunosorbent assay (ELISA). Results At the end of the experiment (11th week), the blood pressure was higher in both the H and PH groups than that in the N group (P<0.001), but there was no statistically significant difference in blood pressure between the H and PH groups. There was no statistical difference in heart rate among the 3 groups. Micro-CT showed that the distance from the cemento-enamel junction to the alveolar bone crest (CEJ-ABC) of the mandibular first molar in the PH group was significantly higher than that in the N and H groups (P<0.016 7). The number of osteoclasts in the alveolar bone of the PH group was significantly higher than that of the N and H groups (P<0.0167). No common differentially expressed genes were found among the 3 groups. There were 235 significantly differentially expressed genes in the gingival tissue between the H and PH groups, and 137 upregulated genes (e.g., P-selectin, keratin 16, and S100 calcium binding protein A) and 98 downregulated genes (e.g., FK506 binding protein 5, mediator complex subunit 22, zinc finger and BTB domain containing 16) in the PH group compared to the H group. GO analysis showed that the major enriched biological processes (BP) were leukocyte migration, the major cellular component (CC) was complex of collagen trimers, and the significant molecular function (MF) was extracellular matrix structural constituent in the H and PH groups. KEGG pathway analysis showed that signaling pathways such as cytokine-cytokine receptor interaction, IL-17 signaling pathway, and TNF-α signaling pathway were significantly enriched in the H and PH groups. PPI analysis identified four key genes affecting periodontitis in hypertensive conditions, including interleukin-1β (IL-1β), matrix metalloproteinase-9 (MMP-9), collagen type I alpha1 (COL1α1), and chemokine ligand 1 (CXCL1). Compared to the N and H groups, the expressions of IL-1β and TNF-α were all upregulated in the gingival tissue and systemic serum in the PH group (P<0.016 7). Conclusion The differentially expressed mRNAs in hypertension with or without periodontitis included IL-1β and MMP-9, while the differentially expressed signaling pathways were IL-17 and TNF-α. These results provide a theoretical reference for further investigation of the molecular regulatory mechanism of hypertension with periodontitis in the future.

Objective To evaluate the caries excavation efficacy and minimally invasive potential of three dentine caries excavation methods including traditional excavation, chemomechanical excavation, and fluorescence-aided caries excavation using micro-computerized tomography (micro-CT). Methods This study was approved by the Biomedical Research Ethics Committee, and all patients provided informed consent. Thirty molars and premolars with dentin caries were collected and randomly divided into three groups. The samples were obtained by traditional excavation (traditional excavation group), chemomechanical excavation (chemomechanical excavation group), and fluorescence-aided caries excavation (fluorescence-aided caries excavation group), and the operation time for each sample was recorded. Micro-CT was used to scan and record the caries volume and healthy tooth volume of each tooth before and after caries excavation. The caries excavation efficacy and minimally invasive potential of the three caries excavation methods were evaluated based on the caries volume and the healthy tooth volume before and after caries excavation. Results In terms of caries excavation operation time, the chemomechanical excavation group (501.7 s ± 143.6 s) had a longer operation time than the traditional excavation group (263.9 s ± 121.2 s) and the fluorescence-aided caries excavation group (284.2 s ± 135.6 s), with a statistically significant difference (P<0.01); there was no significant difference between the traditional excavation group and the fluorescence-aided caries excavation group. In terms of caries excavation efficacy, the ratio of residual caries volume to initial caries volume was determined in the traditional excavation group (0.087 ± 0.04), followed by the fluorescence-aided caries excavation group (0.36 ± 0.10), and the chemomechanical excavation group was the highest (0.51 ± 0.10); the observed disparity between the groups exhibited statistical significance (P<0.01). In terms of minimally invasive potential, the ratio of the traditional excavation group (0.87 ± 0.05) was lower than the chemomechanical excavation group (0.99 ± 0.01) and fluorescence-aided caries excavation group (0.98 ± 0.01), with statistically significant differences (P<0.01); the difference between the ratio of the chemomechanical excavation group and the fluorescence-aided caries excavation group was not statistically significant. Conclusion The traditional excavation group had the shortest operation time, but the traditional excavation removed too much healthy dentin and demineralized dentin. The chemomechanical excavation group retained demineralized dentin and healthy dentin and had the best minimally invasive potential, but the caries excavation efficacy was poor and the operation time was long. The fluorescence-aided caries excavation preserved part of the demineralized dentin and healthy dentin, had certain minimally invasive potential, and the clinical operation time was moderate.

Objective To compare the accuracy of the original-mirror alignment algorithm and a landmark-independent method in constructing the midsagittal plane (MSP) of the cone beam computed tomography in patients with facial deformities, so as to provide a theoretical basis for symmetric analysis. Methods The study was approved by the hospital ethics committee. Cone beam computed tomography data of 30 patients with facial deformities were obtained, and the output was saved in DICOM format. The scan data were imported into Mimics 21.0; after segmentation, three-dimensional (3D) skull models were reconstructed. Furthermore, the 3D scan data of skulls were transformed into mirror skull models using Geomagic Studio 2014 reverse engineering software. The MSP of each skull was generated using both the original-mirror alignment algorithm and the landmark-independent method. Original-mirror alignment algorithm: the original skull model and its mirror model were combined, and the new data to calculate the MSP (S1) of the original data in Geomagic Studio 2014 were obtained. Landmark-independent method: the following anatomical landmarks were determined using Mimics 21.0: nasion (N), crista galli (CG), sella (S), basion (Ba), vomer (V), posterior nasal spine (PNS), incisive foramen (IF), and anterior nasal spine (ANS). The MSP (S2) of best fit was then found by minimizing the mean square distance of these eight anatomical landmarks to a plane in Geomagic Studio 2014. The results of the S1 and S2 models constructed using the original-mirror alignment algorithm and the landmark-independent method, respectively, were scored subjectively by five senior maxillofacial surgeons, and a paired t-test was performed for the two groups. The internal consistency analysis was performed based on secondary experiments to verify the repeatability of the expert evaluation method. Results The average scores of the S1 and S2 models were 65.73 and 75.90, respectively. The average score of the model constructed using the landmark-independent method was significantly higher than that of the model constructed using the original-mirror alignment algorithm (P<0.01). Furthermore, the results of the internal consistency analysis showed that the expert evaluation method had good reliability and validity. Conclusion In patients with facial deformities, the MSP constructed using the landmark-independent method is superior to that constructed using the original-mirror alignment algorithm. This study provides a theoretical basis for maxillofacial symmetry analysis in clinical settings and is clinically feasible.

The source and process of mandible development are significantly different from those of other bones in the body, and abnormal development can lead to various bone-related diseases, seriously affecting the quality of life of patients. In recent years, the role of the Hedgehog signaling pathway in bone development has received increasing attention. The Hedgehog gene includes three subtypes: Sonic Hedgehog (Shh), Indian Hedgehog (Ihh), and Desert Hedgehog (Dhh). Shh and Ihh can participate in bone metabolism regulation through various pathways, with Shh primarily involved in limb development and Ihh playing a key role in endochondral osteogenesis. The Hedgehog signaling pathway includes Hedgehog signaling protein ligands, Patched (Ptch) receptors, Smoothed (Smo) receptors, nuclear transcription factors, glioma-associated oncogene homologues (Gli), and downstream target genes. The activation of typical Hedgehog signaling pathways requires the involvement of Gli, whereas atypical Hedgehog signaling is mainly regulated by Ptch, Smo, and others. Shh regulates various biological behaviors during early vertebrate embryogenesis, such as organ differentiation, neural stem formation, stem cell differentiation and proliferation, limb bone development, and tooth germ development. During the process of bone cell differentiation, Shh, Ptch1, and Gli1 are expressed in osteoblasts, further promoting the differentiation of bone marrow mesenchymal stem cells into osteoblasts and chondrocytes. IHh plays an indispensable functional role in bone growth, development, and homeostasis and participates in the formation of intramembrane bone collars, proliferation, and maturation of chondrocytes. IHh is expressed in mature skull osteoblasts and can act as a promoter of bone factor regulation of Ptch and bone morphogenetic protein (BMP) expression to induce intramembrane ossification. Brain and muscle ARNT-like protein 1 (BMAL1) can regulate the Hedgehog signaling pathway by binding to Ptch1 and Ihh, playing a crucial role in cartilage formation and endochondral osteogenesis in the temporomandibular joint. Hedgehog signal activators can improve the reduction in mandibular bone mass caused by BMAL1 deficiency. Hedgehog signaling imbalance can have a significant impact on bone development and lead to a series of bone diseases, such as abnormal bone development, fractures, osteoporosis, and osteoarthritis. The mechanism of the Hedgehog signaling pathway in relation to mandibular diseases has not been fully elucidated, and future research should seek to further explore Hedgehog signaling as a potential target for treating mandibular developmental-related diseases.

Inflammatory bowel disease (IBD) is a group of chronic, non-specific inflammatory diseases of the gastrointestinal tract including primarily Crohn’s disease and ulcerative colitis, which are affected by multiple factors. Periodontitis is a type of disease characterized by plaque biofilm as the initiating factor and chronic destruction of alveolar bone via resorption. An increasing number of studies have reported a correlation between periodontitis and IBD, but the relationship between the two remains unclear. In this study, we explore the internal relationships between the two diseases from three dimensions, including epidemiological, biological, and associated treatment evidence. Based on epidemiological evidence, periodontitis was found to be associated with an increased risk of IBD, which also affects periodontal health, although the bidirectional correlation needs to be further studied by expanding the number of data sources. From the biological evidence, both clinical studies and animal experiments show that IBD and periodontitis are interconnected. Based on evidence from association therapy, drugs that are beneficial for the treatment of IBD are also effective in the prevention and treatment of periodontitis. In addition, drugs that are good for improving periodontitis can also significantly alleviate IBD. The interaction mechanism between IBD and periodontitis includes the microbial pathway and the immunization route. The microbial pathway refers to the increase in the probability of intestinal tract ectopic colonization by oral bacteria transmitted through the mouth-gut axis or blood, resulting from the increase in the proportion of opportunistic pathogens in the oral cavity of patients with periodontitis and the influence of IBD on the secretion of gastric juice and the balance of intestinal flora. These microorganisms further aggravate IBD inflammation by releasing virulence factors, destroying the intestinal mucosal barrier, and triggering inflammatory responses. In periodontitis, adaptive immunity is activated in the mouth, leading to the production of a large number of immune cells, including Th17 containing the intestinal homing marker α4β7 integrin on their surface. Increased ligand expression of α4β7 integrin in the intestinal mucosa of patients with IBD accelerates oral Th17 cell transfer to the intestine, thereby worsening intestinal inflammation. In parallel, the abnormal expression of cytokines, such as TNF-α, IL-1β, IL-10, IL-6, IL-21, soluble CD40 ligand (sCD40L), IL-23, and INF-γ, in the oral cavity of patients with IBD was observed, suggesting that IBD may affect periodontitis through immunity. These cytokines represent targets for the treatment of both diseases and provide a research direction for their prevention and treatment in the future.

The oral cavity harbors a diverse population of microorganisms, making it one of the most heavily colonized sites in the human body. Maintaining a balanced microecology is crucial for oral health. Ligilactobacillus salivarius as a species of Ligilactobacillus, has good oral colonization ability and potential to improve oral microecology for disease prevention and control. Currently, the application and mechanism of Ligilactobacillus salivarius in oral diseases include several aspects. First, by directly inhibiting the growth of Streptococcus mutans and downregulating the expression of its cariogenic virulence factor, gtfs, the aim is to reduce the number of adherent Streptococcus mutans on the tooth surface, thereby preventing dental caries. Second, reducing the number of keystone taxa in periodontitis, and the virulence factors of Aggregatibacter actinomycetemcomitans, including CdtB and LtxA, can alleviate local stimulation in patients with periodontitis. Additionally, directly inhibiting macrophage MAPK and NF-κB pathway activation suppresses osteoclastogenesis and reduces periodontal bone absorption. In mucosal inflammation, Ligilactobacillus salivarius competes with Candida albicans, inhibits the formation of pathogenic hyphae or germ tubes, and prevents monilial stomatitis. Ligilactobacillus salivarius can also reduce the amount of Staphylococcus aureus and mitigate the activation of the macrophage TLR/PI3K/Akt/mTOR and TLR/PI3K/Akt/IκB/NF-κB pathways induced by S. aureus infections, thus alleviating inflammation in the oral and pharyngeal regions. In vitro studies on oral tumors have revealed that Ligilactobacillus salivarius can downregulate the expression of cancer cell Akt/Cyclin D1, induce direct apoptosis of tumor cells, reduce COX-2 expression, and improve the tumor immune-suppressive microenvironment. Previous studies have revealed considerable variability in Ligilactobacillus salivarius, necessitating more detailed research to clarify its clinical effects, safety, and mechanisms. Despite the emergence of novel microbiological research techniques, their application to Ligilactobacillus salivarius remains relatively limited. One crucial direction for future research is to better utilize these methods to investigate the effects of Ligilactobacillus salivarius on oral diseases. Considering these factors, this study provides a comprehensive review of existing research studies on Ligilactobacillus salivarius in the fields of oral medicine and dentistry, with the aim to serve as a reference and guide for future studies.

Bacterial overproliferation and virulence factors in plaque biofilms can cause inflammation of soft and hard tissues around the implant, resulting in peri-implantitis. If not well controlled, severe peri-implantitis can lead to failure of implant osseointegration and implant loosening and loss. Currently, peri-implantitis can be treated by surgical and non-surgical treatment such as mechanical debridement and chemotherapy, but there remain problems related to the unpredictable therapeutic effect and high recurrence rate. Therefore, gaining a comprehensive understanding of the relationship between plaque biofilm formation and peri-implantitis is crucial for the prevention and treatment of peri-implantitis. In this article, we comprehensively review current research on the specific composition and formation process of biofilms and the influence of implant material characteristics on biofilm formation. The results of the research review indicated that peri-implantitis biofilms are composed of extracellular matrix, with a predominant population of anaerobic Gram-negative bacteria embedded within. The formation process includes the acquisition of an acquired membrane, microbial adhesion, and biofilm detachment and dispersion. Biofilm formation is primarily influenced by the implant surface roughness, surface free energy (SFE), and material properties. Current strategies for biofilm removal around implants mainly involve implant surface coating techniques, mechanical debridement, chemical agents, laser therapy, and photodynamic therapy; however, the therapeutic outcomes remain uncertain. The future research direction will be based on the characteristics of the plaque biofilm around the implant, combined with cutting-edge methods, such as nanotechnology, immunotherapy, and gene therapy, to continuously prevent the formation of plaque biofilm on the surface of the implant to prevent and treat peri-implantitis.