Objective: To investigate the effect of specific deletion of SET-domain-containing 2 (Setd2) in dental germ mesenchymal cells on early tooth development and provide a research basis for exploring the etiology of dental developmental disorders. Methods: This study was approved by the Animal Care and Ethical Committee of Wuhan University. Wnt1Cre mice with a C57BL/6J background were crossed with Setd2flox/flox mice to generate the Setd2 knockout experimental group (Wnt1Cre; Setd2flox/flox) and the control group (Setd2flox/flox). Tail tissues of mouse embryos were collected at embryonic day (E) 13.5, E15.5, and E18.5 for polymerase chain reaction (PCR)-based genotyping, and embryonic heads were collected for paraffin sectioning to observe the development of the first mandibular molar germs. Immunohistochemical (IHC) staining was performed to verify the specific knockout of Setd2 in dental germ mesenchymal cells; hematoxylin and eosin (HE) staining and subrenal culture were used to evaluate the effect of Setd2 knockout in dental germ mesenchymal cells on tooth development; Ki67 staining and a terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay were employed to evaluate the effects of specific Setd2 knockout on the proliferation and apoptosis of dental germ mesenchymal cells; immunofluorescence (IF) staining was conducted to observe the impact of Setd2 knockout on the trimethylation of lysine 36 on histone 3 (H3K36me3). Results: PCR-based genotyping showed that the experimental group presented a single band (266 bp) for the Setd2flox/flox homozygote and a characteristic band for Wnt1Cre, while the control group only showed a single band (266 bp) for the Setd2flox/flox homozygote. IHC staining confirmed the successful knockout of Setd2 in dental germ mesenchymal cells of the experimental group; HE staining and subrenal culture demonstrated that specific knockout of Setd2 in the experimental group led to reduced size of tooth germs and increased condensation of mesenchymal cells at E15.5 and E18.5 (E15.5, P < 0.05; E18.5, P < 0.01); Ki67 staining showed no statistically significant effect on the proportion of division of dental germ mesenchymal cells between the experimental and control groups at E13.5 and E15.5 (E13.5, P = 0.694; E15.5, P = 0.503); the TUNEL assay demonstrated increased apoptosis of dental germ mesenchymal cells in the experimental group at both E13.5 and E15.5 (P < 0.001); IF staining revealed the absence of H3K36me3 modification in the dental germ mesenchymal cells of the experimental group across E13.5, E15.5, and E18.5 (P < 0.000 1). Conclusion Specific knockout of Setd2 in dental germ mesenchymal cells leads to increased cell apoptosis by impairing H3K36me3 modification, resulting in early developmental defects in mouse teeth characterized by reduced tooth germ volume.

Head and neck squamous cell carcinoma(HNSCC)is an immunosuppressive tumor with high morbidity and mortal-ity,and easy to develop treatment tolerance.In recent years,it has been found that the tumor microenvironment,especially the chemo-kine-chemokine receptor network,plays an important role in the occurrence,development,prognosis and treatment tolerance of HN-SCC.In this review,the research progress on the relationship between chemokine-chemokine receptor network and HNSCC is summa-rized,focusing on the regulatory role and mechanism of HNSCC progression,so as to provide new ideas for the diagnosis and treatment of HNSCC.

Objective:To investigate the role of WW domain containing E3 ubiquitin protein ligase 1 (WWP1) in enamel development of mice.Methods:Single-cell RNA sequencing data of incisor tissues of postnatal day 7 (P7) mice and mandibular first molar tooth germs of P3.5 mice were used to analyze the expression of Wwp1 in dental epithelial cells. Immunohistochemistry was performed to observe the distribution and expression levels of WWP1 in the epithelium of mouse incisors and mandibular first molar tooth germs. Wwp1 knockout (Wwp1 KO) mice were generated and collected with their control littermates at P1, P7, three mice per group, as well as at P14, P28, 2 months (2M), and 3M, six mice per group. The enamel volumes of molars and incisors were analyzed using micro-CT. Scanning electron microscopy was employed to examine the enamel cross-sections of Wwp1 KO and control mice. Energy dispersive spectroscopy (EDS) was used to analyze the calcium and phosphorus content of the enamel rod of incisors. Immunofluorescence was performed to detect the expression of amelogenin (AMELX) in the ameloblasts of Wwp1 KO and control mice. Additionally, LS-8 ameloblast-like epithelial cells were cultured, and Wwp1 siRNA or overexpression plasmids were transfected to knock down or overexpress WWP1. The protein levels of AMELX were then assessed by Western blotting.Results:Single-cell sequencing result showed a high Wwp1 mRNA expression level in the epithelial cells of mouse incisors and mandibular molar tooth germs. Immunohistochemistry revealed the expression of WWP1 in presecretory, secretory, transitional, and mature ameloblasts. Wwp1 KO mice exhibited enamel developmental defects. The enamel volumes of molars and incisors in Wwp1 KO mice [(0.155±0.016), (0.300±0.017) μm 3] were reduced by 23.95% ( P<0.001) and 28.31% ( P<0.001) compared with the control group [(0.203±0.062), (0.418±0.023) μm 3] respectively. Scanning electron microscopy showed disorganized enamel structures in Wwp1 KO incisors and molars. EDS results showed the weight percent of calcium in the enamel rod of incisors decreased in Wwp1 KO mice [(20.74±0.91)%] compared with the control group [(30.30±3.83)%] ( P<0.001), and the calcium-to-phosphorus ratio decreased in Wwp1 KO mice (1.93±0.01) compared with the control group (2.02±0.01) ( P<0.001). Immunofluorescence showed weaker AMELX expression in ameloblasts of mandibular first molar tooth germs from P1 and P7 Wwp1 KO mice compared with the control group ( P<0.001, P<0.001). In LS-8 cells, Wwp1 knocked-down led to a decrease of AMELX protein expression, while WWP1 overexpression resulted in an increased AMELX protein level. Conclusions:WWP1 promotes ameloblast differentiation and enamel matrix mineralization, playing a critical role in enamel formation.

Head and neck squamous cell carcinoma(HNSCC)is the sixth most common malignant tumor in the world.Due to its high incidence,poor prognosis,relapse-prone characteristics,and the existence of treatment resist-ance,HNSCC has a serious impact on patients'daily lives.As a bridge between innate immunity and adaptive immu-nity,dendritic cell(DC)plays a crucial role in tumor progression.In recent years,numerous studies have found that DC has complex and diverse functions in the tumor microenvironment(TME),and has a significant impact on the oc-currence and development of HNSCC.In this article,we provide a review of the mechanism of DC in HNSCC TME.

Objective To explore the value of pulmonary perfusion SPECT/CT imaging for diagnosing pulmonary embolism(PE)and combining with clinical and laboratory indicators for predicting prognosis.Methods Totally 260 patients with suspected PE were retrospectively collected,and PE was clinically confirmed in 237 cases.The diagnostic efficacy of pulmonary perfusion SPECT/CT imaging and CT pulmonary angiography(CTPA)were compared.Among 237 patients with clinically confirmed PE,160 cases were initially diagnosed who underwent standard anticoagulant therapy and were regularly followed up to 3 months after discharge,and their prognosis were analyzed.Logistic regression was used to analyze clinical data,laboratory indicators and pulmonary perfusion defect scores based on pulmonary perfusion SPECT/CT imaging to screen the independent predictors of poor prognosis of PE of the above patients,and a prediction model was then constructed.Receiver operating characteristic(ROC)curve was plotted,and the area under the curve(AUC)was calculated to evaluate the predictive efficacy of this model for predicting prognosis of PE.Results The positive rate of PE diagnosed with pulmonary perfusion SPECT/CT was higher than that with CTPA(66.54％[173/260]vs.59.62％[155/260],x2=4.765,P=0.038).Among 160 cases of clinically confirmed PE,poor prognosis was observed in 51 cases,while good prognosis was noticed in 109 cases.Patients' age,blood urea nitrogen,serum creatinine,uric acid,D-dimer level and pulmonary perfusion defect score were all independent predictors for poor prognosis of PE(all P＜0.05),and AUC of the established combined model was 0.731.Conclusion Pulmonary perfusion SPECT/CT imaging could be used for diagnosing PE,and combining with clinical and laboratory indicators could effectively predict prognosis of PE.

Objective:To investigate the role of WW domain containing E3 ubiquitin protein ligase 1 (WWP1) in enamel development of mice.Methods:Single-cell RNA sequencing data of incisor tissues of postnatal day 7 (P7) mice and mandibular first molar tooth germs of P3.5 mice were used to analyze the expression of Wwp1 in dental epithelial cells. Immunohistochemistry was performed to observe the distribution and expression levels of WWP1 in the epithelium of mouse incisors and mandibular first molar tooth germs. Wwp1 knockout (Wwp1 KO) mice were generated and collected with their control littermates at P1, P7, three mice per group, as well as at P14, P28, 2 months (2M), and 3M, six mice per group. The enamel volumes of molars and incisors were analyzed using micro-CT. Scanning electron microscopy was employed to examine the enamel cross-sections of Wwp1 KO and control mice. Energy dispersive spectroscopy (EDS) was used to analyze the calcium and phosphorus content of the enamel rod of incisors. Immunofluorescence was performed to detect the expression of amelogenin (AMELX) in the ameloblasts of Wwp1 KO and control mice. Additionally, LS-8 ameloblast-like epithelial cells were cultured, and Wwp1 siRNA or overexpression plasmids were transfected to knock down or overexpress WWP1. The protein levels of AMELX were then assessed by Western blotting.Results:Single-cell sequencing result showed a high Wwp1 mRNA expression level in the epithelial cells of mouse incisors and mandibular molar tooth germs. Immunohistochemistry revealed the expression of WWP1 in presecretory, secretory, transitional, and mature ameloblasts. Wwp1 KO mice exhibited enamel developmental defects. The enamel volumes of molars and incisors in Wwp1 KO mice [(0.155±0.016), (0.300±0.017) μm 3] were reduced by 23.95% ( P<0.001) and 28.31% ( P<0.001) compared with the control group [(0.203±0.062), (0.418±0.023) μm 3] respectively. Scanning electron microscopy showed disorganized enamel structures in Wwp1 KO incisors and molars. EDS results showed the weight percent of calcium in the enamel rod of incisors decreased in Wwp1 KO mice [(20.74±0.91)%] compared with the control group [(30.30±3.83)%] ( P<0.001), and the calcium-to-phosphorus ratio decreased in Wwp1 KO mice (1.93±0.01) compared with the control group (2.02±0.01) ( P<0.001). Immunofluorescence showed weaker AMELX expression in ameloblasts of mandibular first molar tooth germs from P1 and P7 Wwp1 KO mice compared with the control group ( P<0.001, P<0.001). In LS-8 cells, Wwp1 knocked-down led to a decrease of AMELX protein expression, while WWP1 overexpression resulted in an increased AMELX protein level. Conclusions:WWP1 promotes ameloblast differentiation and enamel matrix mineralization, playing a critical role in enamel formation.

Head and neck squamous cell carcinoma(HNSCC)is the sixth most common malignant tumor in the world.Due to its high incidence,poor prognosis,relapse-prone characteristics,and the existence of treatment resist-ance,HNSCC has a serious impact on patients'daily lives.As a bridge between innate immunity and adaptive immu-nity,dendritic cell(DC)plays a crucial role in tumor progression.In recent years,numerous studies have found that DC has complex and diverse functions in the tumor microenvironment(TME),and has a significant impact on the oc-currence and development of HNSCC.In this article,we provide a review of the mechanism of DC in HNSCC TME.

Objective To explore the value of pulmonary perfusion SPECT/CT imaging for diagnosing pulmonary embolism(PE)and combining with clinical and laboratory indicators for predicting prognosis.Methods Totally 260 patients with suspected PE were retrospectively collected,and PE was clinically confirmed in 237 cases.The diagnostic efficacy of pulmonary perfusion SPECT/CT imaging and CT pulmonary angiography(CTPA)were compared.Among 237 patients with clinically confirmed PE,160 cases were initially diagnosed who underwent standard anticoagulant therapy and were regularly followed up to 3 months after discharge,and their prognosis were analyzed.Logistic regression was used to analyze clinical data,laboratory indicators and pulmonary perfusion defect scores based on pulmonary perfusion SPECT/CT imaging to screen the independent predictors of poor prognosis of PE of the above patients,and a prediction model was then constructed.Receiver operating characteristic(ROC)curve was plotted,and the area under the curve(AUC)was calculated to evaluate the predictive efficacy of this model for predicting prognosis of PE.Results The positive rate of PE diagnosed with pulmonary perfusion SPECT/CT was higher than that with CTPA(66.54％[173/260]vs.59.62％[155/260],x2=4.765,P=0.038).Among 160 cases of clinically confirmed PE,poor prognosis was observed in 51 cases,while good prognosis was noticed in 109 cases.Patients' age,blood urea nitrogen,serum creatinine,uric acid,D-dimer level and pulmonary perfusion defect score were all independent predictors for poor prognosis of PE(all P＜0.05),and AUC of the established combined model was 0.731.Conclusion Pulmonary perfusion SPECT/CT imaging could be used for diagnosing PE,and combining with clinical and laboratory indicators could effectively predict prognosis of PE.

Head and neck squamous cell carcinoma(HNSCC)is an immunosuppressive tumor with high morbidity and mortal-ity,and easy to develop treatment tolerance.In recent years,it has been found that the tumor microenvironment,especially the chemo-kine-chemokine receptor network,plays an important role in the occurrence,development,prognosis and treatment tolerance of HN-SCC.In this review,the research progress on the relationship between chemokine-chemokine receptor network and HNSCC is summa-rized,focusing on the regulatory role and mechanism of HNSCC progression,so as to provide new ideas for the diagnosis and treatment of HNSCC.

Objective To investigate the relationship of miRNA gene polymorphisms with blood pressure(BP)responses to the sodium and potassium diet intervention.Methods In 2004,we recruited 514 participants from 124 families in seven villages of Baoji,Shaanxi Province,China.All subjects were given a three-day normal diet,followed by a seven-day low-salt diet,a seven-day high-salt diet,and finally a seven-day high-salt and potassium supplementation.A total of 19 miRNA single nucleotide polymorphisms(SNPs)were selected for analysis.Results Throughout the sodium-potassium dietary intervention,the BP of the subjects fluctuated across all phases,showing a decrease during the low-salt period and an increase during the high-salt period,followed by a reduction in BP subsequent to potassium supplementation during the high-salt diet.MiR-210-3p SNP rs 12364149 was significantly associated with systolic BP(SBP),diastolic BP(DBP)and mean arterial pressure(MAP)responses to low-salt diet.MiR-4638-3p SNP rs6601178 was significantly associated with SBP while miR-26b-3p SNP rs115254818 was significantly associated with MAP responses to low-salt intervention.In addition,miR-26b-3p SNP rs115254818 was significantly correlated with SBP,DBP and MAP responses to high-salt intervention.MiR-1307-5p SNPs rs1 1191676 and rs2292807 were associated with SBP and MAP responses to high-salt diet.MiR-4638-3p SNP rs6601178,miR-210-3p SNP rs12364149,miR-382-5p SNP rs4906032 and rs4143957 were significantly associated with SBP response to high-salt diet.In addition,miR-26b-3p SNP rs115254818 was significantly associated with SBP,DBP and MAP responses to potassium supplementation.MiR-1307-5p SNPs rs11191676,rs2292807,and miR-19a-3p SNP rs4284505 were significantly associated with SBP responses to high-salt and potassium supplementation.Conclusion miRNA gene polymorphisms are associated with BP response to sodium and potassium,suggesting that miRNA genes may be involved in the pathophysiological process of salt sensitivity and potassium sensitivity.