BACKGROUND:Individuals with chronic ankle instability are prone to inversion ankle sprains during landing.Moderately increasing the foot toe-out angle during landing may reduce the occurrence of inversion ankle sprains,but no studies have directly demonstrated this effect. OBJECTIVE:To explore the effect of increased toe-out angle during landing on the peak inversion angle,peak angular velocity,and the time to peak inversion among individuals with and without chronic ankle instability. METHODS:A total of 60 participants were recruited for this study,including 30 individuals with chronic ankle instability and 30 without chronic ankle instability.The study utilized a simulated sprain apparatus for drop-landing tests,featuring a platform that could tilt forward by 24° and inward by 15°,thus simulating the foot position during an ankle inversion sprain.Participants were required to perform drop-landing tests under two landing conditions:natural landing and toe-out landing,with the latter involving a greater foot toe-out angle,over 150%more than the former.Kinematic data of participants were recorded using a 12-camera three-dimensional motion capture system.Data analysis was conducted using two-way repeated measures analysis of variance and Spearman correlation analysis. RESULTS AND CONCLUSION:(1)Significant main effects of condition were found for peak inversion angle during drop-landing(P<0.001,η2 p=0.270),peak inversion velocity(P=0.015,η2 p=0.098),and peak inversion time(P<0.001,η2 p=0.260);a significant main effect of group was found for peak inversion velocity(P=0.029,η2 p=0.080).(2)There were significant negative correlations between the foot toe-out angle at landing and the peak ankle inversion angle(P=0.021,r=-0.310;P=0.042,r=-0.278)as well as the peak inversion time(P=0.018,r=-0.312;P=0.021,r=-0.309)in both chronic ankle instability and non-chronic ankle instability groups.Moreover,a significant negative correlation was also found between the foot toe-out angle and peak inversion velocity in the chronic ankle instability group(P=0.021,r=-0.312).(3)It is indicated that increasing the foot toe-out angle at landing can reduce the peak inversion angle,peak inversion velocity,and the peak inversion time during landing in patients with chronic ankle instability and non-chronic ankle instability,thereby decreasing the risk of ankle inversion sprains.

Aim To investigate the regulatory mecha-nisms of donepezil on the expression and enzymatic ac-tivity of cytochrome P450 3A4(CYP3A4),elucidate the synergistic impact of hypoxia on CYP3A4 function,and reveal its potential association with drug-induced cardiotoxicity,particularly QT interval prolongation.Methods Western blot,co-immunoprecipitation,and gene knockdown techniques were employed to evaluate the effects of donepezil and hypoxia on CYP3A4 pro-tein expression.CYP3A4 enzymatic activity was as-sessed using an in vitro incubation system with rat liver microsomes combined with high-performance liquid chromatography(HPLC),and the half-maximal inhib-itory concentration(IC50)was determined.Results Donepezil(10 μmol·L-1)and hypoxia reduced CYP3A4 protein expression to 31.75％and 45.90％of the control levels,respectively.Both interventions activated the gp78-mediated ubiquitin-proteasome path-way,significantly increasing CYP3A4 ubiquitination levels by 2.1-fold compared to the control group,thereby promoting proteasomal degradation.Donepezil inhibited CYP3A4 enzyme activity with an IC50 of 83.4μmol·L-1,and hypoxia synergistically enhanced this inhibitory effect,reducing the IC50 to 20.79 μmol·L-1.Conclusion Donepezil downregulates CYP3A4 function through dual mechanisms involving ubiquitin-mediated proteasomal degradation and direct enzymatic inhibition.Hypoxia potentiates this effect,leading to impaired metabolism of CYP3A4 substrate drugs,ele-vated plasma drug concentrations(1.6-2.3-fold in-crease compared to normal metabolic conditions),and an increased risk of QT interval prolongation and other forms of cardiotoxicity.

BACKGROUND:Ankle inversion injuries frequently occur during landing,injuring the anterior talofibular and calcaneofibular ligaments.Previous studies usually used indirect indicators,such as inversion angle,as an injury risk indicator,but epidemiological support is lacking.OBJECTIVE:To calculate anterior talofibular and calcaneofibular ligament strains using a three-dimensional multi-body foot model during a drop-landing and to investigate whether increasing the foot toe-out angle for landing would reduce the risk of inversion sprains.METHODS:Thirty-five participants with high sports demands[15 males and 20 females,age:(21.0±0.9)years,height:(176.2±8.8)cm,body mass:(71.6±12.8)kg]were recruited to perform a drop-landing test using a trapdoor device to simulate ankle inversion sprains.Two landing conditions were tested,i.e.,natural landing and toe-out landing.Kinematic data were collected using a 12-camera motion analysis system,the strains of the anterior talofibular and calcaneofibular ligaments were calculated using a three-dimensional rigid-body foot model.RESULTS AND CONCLUSION:From natural landing to toe-out landing conditions,the anterior talofibular ligament strain decreased[natural landing=(3.57±1.92)％,toe-out landing=(0.36±1.18)％,P＜0.001,Cohen's d=2.01),as was the calcaneofibular ligament strain[natural landing=(1.38±1.80)％,toe-out landing=(0.28±2.29)％,P=0.003,Cohen's d=0.81).It could be concluded that increasing foot toe-out angle reduces anterior talofibular and calcaneofibular ligament strains during drop-landing with ankle inversion,thereby reducing the potential of ankle inversion sprains.

Objective To investigate the effect of miR-7975 on the malignant phenotype of oral squamous cell carcinoma(OSCC)and its potential mechanisms.Methods This study compared the expression levels of miR-7975 in different oral cell lines by qRT-PCR.miR-7975 mimic and miR-7975 inhibitor were transfected into OSCC cell lines HSC3 and HN4 respectively.Colony formation as-say,CCK8 assay,Transwell assay,and wound healing assay were conducted to evaluate the effects of miR-7975 on the malignant phe-notype of OSCC cells.Western blot was employed to analyze changes in the expression of EMT related proteins and proteins associated with the RAS/ERK signaling pathway.Subcutaneous tumor model of nude mice was used to further validate the tumorigenic effect of miR-7975 in vivo.Results The expression of miR-7975 was downregulated in OSCC cells.Overexpression of miR-7975 reduced the proliferation,migration,and invasion abilities of OSCC cells,whereas downregulation of miR-7975 enhanced these abilities.After miR-7975 overexpression,the expression of the EMT-related protein E-cadherin was upregulated,while N-cadherin,Vimentin,β-catenin,Snail,and Slug were downregulated.Additionally,the expression of proteins related to the RAS/ERK signaling pathway increased.Conversely,the expression of EMT and RAS/ERK signaling pathway-related proteins showed opposite changes when miR-7975 was downregulated.Compared to the control group,the volume and weight of tumors formed in nude mice were significantly smaller after miR-7975 overexpression,while they were significantly larger when miR-7975 expression was reduced.Conclusion miR-7975 exerts its tumor-suppressive effects by inhibiting the proliferation,migration,and invasion of OSCC through the regulation of EMT and the RAS/ERK signaling pathway.

Aim To investigate the regulatory mecha-nisms of donepezil on the expression and enzymatic ac-tivity of cytochrome P450 3A4(CYP3A4),elucidate the synergistic impact of hypoxia on CYP3A4 function,and reveal its potential association with drug-induced cardiotoxicity,particularly QT interval prolongation.Methods Western blot,co-immunoprecipitation,and gene knockdown techniques were employed to evaluate the effects of donepezil and hypoxia on CYP3A4 pro-tein expression.CYP3A4 enzymatic activity was as-sessed using an in vitro incubation system with rat liver microsomes combined with high-performance liquid chromatography(HPLC),and the half-maximal inhib-itory concentration(IC50)was determined.Results Donepezil(10 μmol·L-1)and hypoxia reduced CYP3A4 protein expression to 31.75％and 45.90％of the control levels,respectively.Both interventions activated the gp78-mediated ubiquitin-proteasome path-way,significantly increasing CYP3A4 ubiquitination levels by 2.1-fold compared to the control group,thereby promoting proteasomal degradation.Donepezil inhibited CYP3A4 enzyme activity with an IC50 of 83.4μmol·L-1,and hypoxia synergistically enhanced this inhibitory effect,reducing the IC50 to 20.79 μmol·L-1.Conclusion Donepezil downregulates CYP3A4 function through dual mechanisms involving ubiquitin-mediated proteasomal degradation and direct enzymatic inhibition.Hypoxia potentiates this effect,leading to impaired metabolism of CYP3A4 substrate drugs,ele-vated plasma drug concentrations(1.6-2.3-fold in-crease compared to normal metabolic conditions),and an increased risk of QT interval prolongation and other forms of cardiotoxicity.

Objective To investigate the effect of miR-7975 on the malignant phenotype of oral squamous cell carcinoma(OSCC)and its potential mechanisms.Methods This study compared the expression levels of miR-7975 in different oral cell lines by qRT-PCR.miR-7975 mimic and miR-7975 inhibitor were transfected into OSCC cell lines HSC3 and HN4 respectively.Colony formation as-say,CCK8 assay,Transwell assay,and wound healing assay were conducted to evaluate the effects of miR-7975 on the malignant phe-notype of OSCC cells.Western blot was employed to analyze changes in the expression of EMT related proteins and proteins associated with the RAS/ERK signaling pathway.Subcutaneous tumor model of nude mice was used to further validate the tumorigenic effect of miR-7975 in vivo.Results The expression of miR-7975 was downregulated in OSCC cells.Overexpression of miR-7975 reduced the proliferation,migration,and invasion abilities of OSCC cells,whereas downregulation of miR-7975 enhanced these abilities.After miR-7975 overexpression,the expression of the EMT-related protein E-cadherin was upregulated,while N-cadherin,Vimentin,β-catenin,Snail,and Slug were downregulated.Additionally,the expression of proteins related to the RAS/ERK signaling pathway increased.Conversely,the expression of EMT and RAS/ERK signaling pathway-related proteins showed opposite changes when miR-7975 was downregulated.Compared to the control group,the volume and weight of tumors formed in nude mice were significantly smaller after miR-7975 overexpression,while they were significantly larger when miR-7975 expression was reduced.Conclusion miR-7975 exerts its tumor-suppressive effects by inhibiting the proliferation,migration,and invasion of OSCC through the regulation of EMT and the RAS/ERK signaling pathway.

Objective:To investigate the effects of long non coding RNA KCNQ1OT1(LncRNA KCNQ1OT1)on the migration and invasion of oral squamous cell carcinoma(OSCC)cells by regulating the microRNA-875-5p(miR-875-5p)/ETS like transcription factor 4(ELK4)axis.Methods:QRT-PCR was applied to detect the mRNA levels of LncRNA KCNQ1OT1,miR-875-5p,and ELK4 in OSCC cell lines(HSC-3,PE/CA-PJ15,HN13)and tissues.The dual luciferase assay was applied to detect the targeting relationship between LncRNA KCNQ1OT1 and miR-875-5p,and target relationship between miR-875-5p and ELK4.HSC-3 cells were used in control group,sh-NC group,sh-KCNQ1OT1 group,sh-KCNQ1OT1+anti-NC group,sh-KCNQ1OT1+anti-miR-875-5p group,miR-NC group,miR-875-5p mimic group,miR-875-5p mimic+pcDNA-NC group,and miR-875-5p mimic+ELK4 group.The migration and invasion abilities of HSC-3 cells were detected.Immunoblotting was applied to detect the protein expression of ELK4,MMP-2,MMP-9 and epithelial mesenchymal transition(EMT)(E-Cadherin,N-Cadherin,Vimentin).The nude mouse transplant tumor was applied to verify the effect of LncRNA KCNQ1OT1 on OSCC transplant tumors.Results:LncRNA KCNQ1OT1 and ELK4 mRNA expression increased in OSCC tissues and cancer cell lines,while miR-875-5p expression decreased(P＜0.05).Database predictions show that miR-875-5p specifically bound to LncRNAs KCNQ1OT1 and ELK4,respectively.Compared with the sh-NC group,the numbers of cell migration and cell invasion,the expression of LncRNA KCNQ1OT1,ELK4,MMP-2,MMP-9,N-Cadherin,and Vimentin in the sh-KC-NQ1OT1 group were lower,while the expression of miR-875-5p and E-Cadherin was higher(P＜0.05).Compared with the sh-KC-NQ1OT1+anti-NC group,the expression of miR-875-5p and E-Cadherin in the sh-KCNQ1OT1+anti-miR-875-5p group was lower,while the numbers of cell migration and cell invasion,the expression of ELK4,MMP-2,MMP-9,N-Cadherin,and Vimentin were higher(P＜0.05).Compared with the miR-NC group,the expression of miR-875-5p and E-Cadherin in the miR-875-5p mimic group was higher,while the numbers of cell migration and cell invasion,the expression of ELK4,MMP-2,MMP-9,N-Cadherin,and Vim-entin were lower(P＜0.05).Compared with the miR-875-5p mimic+pcDNA-NC group,the numbers of cell migration and cell inva-sion,the expression of ELK4,MMP-2,MMP-9,N-Cadherin,and Vimentin in the miR-875-5p mimic+ELK4 group were higher,while the expression of E-Cadherin was lower(P＜0.05).The transplant tumor volume and weight of the sh-KCNQ1OT1 group were smaller than those of the sh-NC group,the mRNA and protein expression levels of LncRNA KCNQ1OT1,ELK4 were lower than those of the sh-NC group,and the expression level of miR-875-5p was higher than that of the sh-NC group(P＜0.05).Conclusion:Inhibition of LncRNA KCNQ1OT1 can target the miR-875-5p/ELK4 axis to inhibit migration,invasion,and EMT of OSCC cells.

Objective:To improve the isolation and culture method of seasonal influenza virus in embryonated chicken eggs (ECEs), and evaluate their isolation efficiency.Methods:We randomly selected 80 positive samples of H1N1 (H1N1pdm09) and seasonal H3N2 (H3N2snl) influenza virus nucleic acid, and inoculated them into the amniotic and urinary sac cavities of 10-day-old (traditional method) and 14-day-old (improved method) ECEs respectively to adapt the virus to the ECEs (E1-E2). Both method were used to inoculate 10-day-old urinary sac amplification virus (E2-E3), and the final virus isolation positive rates of the two method were compared; using fluorescence quantitative PCR method to detect viral nucleic acids in the improved amniotic and urinary sac cultures, and evaluate the viral proliferation at different inoculation sites; we analyzed the correlation between virus content and isolation positivity rate in the original specimen based on the CT value of nucleic acid testing and the final virus isolation positivity rate using the improved method.Results:The improved method obtained 42 strains of H1N1pdm09 strain, with a positive rate of 52.5% ( χ2=38.571, P<0.01); obtained 54 strains of H3N2snl strain, with a positive rate of 67.5% ( χ2=40.921, P<0.01). Significant differences were observed in the isolation efficiency of H1N1pdm09 samples when the improved method was applied to different inoculation sites of chicken embryos ( χ2=30.476, P<0.01), and similar differences were noted for H3N2snl samples ( χ2=4.928, P=0.026). There was no significant difference in the isolation rate of different CT value intervals of the original samples ( χH1N1pdm092=10.226, χH3N2snl2=3.764, P>0.05). Conclusions:The improved method of inoculating 14-day old ECEs adapted the virus, and the final number of strains obtained was significantly higher than the traditional method of inoculating 10 day old ECEs, which can significantly improve the positive isolation rate of H1N1pdm09 and H3N2snl influenza virus in ECEs. The amniotic cavity is more sensitive to H1N1pdm09 and H3N2snl influenza viruses, which helps the virus adapt in ECEs. There was no significant difference in the sample isolation rate and total positive rate of virus isolation among different CT value ranges, and further verification is needed.

Objective:To investigate the effects of long non coding RNA KCNQ1OT1(LncRNA KCNQ1OT1)on the migration and invasion of oral squamous cell carcinoma(OSCC)cells by regulating the microRNA-875-5p(miR-875-5p)/ETS like transcription factor 4(ELK4)axis.Methods:QRT-PCR was applied to detect the mRNA levels of LncRNA KCNQ1OT1,miR-875-5p,and ELK4 in OSCC cell lines(HSC-3,PE/CA-PJ15,HN13)and tissues.The dual luciferase assay was applied to detect the targeting relationship between LncRNA KCNQ1OT1 and miR-875-5p,and target relationship between miR-875-5p and ELK4.HSC-3 cells were used in control group,sh-NC group,sh-KCNQ1OT1 group,sh-KCNQ1OT1+anti-NC group,sh-KCNQ1OT1+anti-miR-875-5p group,miR-NC group,miR-875-5p mimic group,miR-875-5p mimic+pcDNA-NC group,and miR-875-5p mimic+ELK4 group.The migration and invasion abilities of HSC-3 cells were detected.Immunoblotting was applied to detect the protein expression of ELK4,MMP-2,MMP-9 and epithelial mesenchymal transition(EMT)(E-Cadherin,N-Cadherin,Vimentin).The nude mouse transplant tumor was applied to verify the effect of LncRNA KCNQ1OT1 on OSCC transplant tumors.Results:LncRNA KCNQ1OT1 and ELK4 mRNA expression increased in OSCC tissues and cancer cell lines,while miR-875-5p expression decreased(P＜0.05).Database predictions show that miR-875-5p specifically bound to LncRNAs KCNQ1OT1 and ELK4,respectively.Compared with the sh-NC group,the numbers of cell migration and cell invasion,the expression of LncRNA KCNQ1OT1,ELK4,MMP-2,MMP-9,N-Cadherin,and Vimentin in the sh-KC-NQ1OT1 group were lower,while the expression of miR-875-5p and E-Cadherin was higher(P＜0.05).Compared with the sh-KC-NQ1OT1+anti-NC group,the expression of miR-875-5p and E-Cadherin in the sh-KCNQ1OT1+anti-miR-875-5p group was lower,while the numbers of cell migration and cell invasion,the expression of ELK4,MMP-2,MMP-9,N-Cadherin,and Vimentin were higher(P＜0.05).Compared with the miR-NC group,the expression of miR-875-5p and E-Cadherin in the miR-875-5p mimic group was higher,while the numbers of cell migration and cell invasion,the expression of ELK4,MMP-2,MMP-9,N-Cadherin,and Vim-entin were lower(P＜0.05).Compared with the miR-875-5p mimic+pcDNA-NC group,the numbers of cell migration and cell inva-sion,the expression of ELK4,MMP-2,MMP-9,N-Cadherin,and Vimentin in the miR-875-5p mimic+ELK4 group were higher,while the expression of E-Cadherin was lower(P＜0.05).The transplant tumor volume and weight of the sh-KCNQ1OT1 group were smaller than those of the sh-NC group,the mRNA and protein expression levels of LncRNA KCNQ1OT1,ELK4 were lower than those of the sh-NC group,and the expression level of miR-875-5p was higher than that of the sh-NC group(P＜0.05).Conclusion:Inhibition of LncRNA KCNQ1OT1 can target the miR-875-5p/ELK4 axis to inhibit migration,invasion,and EMT of OSCC cells.