Aim To investigate the mechanism for the inhibitory effect of hydrogen sulfide on the expression of tissue factor(TF)induced by oxidative low-density lipoprotein(ox-LDL)in endothelial cells.Methods Human umbilical vein endothelial cells (HUVECs ) were treated with 50 mg·L-1 ox-LDL in the absence or presence of different concentrations of NaHS (25 , 50,100 and 200 μmol·L-1 )for 24 h.The mRNA expression and protein content of TF in HUVECs were determined by reverse transcription PCR and ELISA, respectively.The content of intracellular reactive oxy-gen species (ROS)was determined by DCFH,an oxi-dative sensitive fluorescent indicator.The activation of nuclear factor-kappaB (NF-κB)was estimated by its expression in nuclear extracts analyzed by Western blot.Results Ox-LDL induced TF mRNA expression and increased TF protein content in HUVECs.The in-crease in intracellular ROS production and the activa-tion of NF-κB were observed in HUVECs treated with ox-LDL.However,NaHS could markedly inhibit the increases in TF mRNA and protein levels induced by ox-LDL.Also the elevation of intracellular ROS pro-duction and the activation of NF-κB elicited by ox-LDL were significantly suppressed by pretreatment with NaHS.In addition,pretreatment with BAY 1 1-7082 (10 μmol·L-1 ),the inhibitor of NF-κB or N-acetyl-L-cysteine(1 mmol·L-1 ),an antioxidant,could also decrease the TF mRNA and protein level as well as ROS production and NF-κB activation induced by ox-LDL in HUVECs,similar to the effects of 200 μmol· L-1 NaHS.Conclusion The mechanism for the in-hibitory effect of H2 S on the ox-LDL- induced TF ex-pression in endothelial cells may be related to inhibi-ting intracellular ROS production and subsequently NF-κB activation.

OBJECTIVE Epileptic networks are char-acterized as two states,seizures or more prolonged inter-ictal periods.However,cellular mechanisms underlying the contribution of interictal periods to ictal events remain unclear.METHODS Here,we present the procedure for labeling seizure-activated and interictal-activated neuro-nal ensembles in mouse hippocampal kindling model using an enhanced-synaptic-activity-responsive element.This technique is combined with genetically encoded effectors to characterize and manipulate neuronal ensembles recruited by focal seizures(FS-Ens)and interictal periods(IP-Ens)in piriform cortex,a region that plays a key role in seizure generation.RESULTS Ca2+ activities and histo-logical evidence reveal a disjointed correlation between the two ensembles during FS dynamics.Optogenetic acti-vation of FS-Ens promotes further seizure development,while IP-Ens protects against it.Interestingly,both ensem-bles are functionally involved in generalized seizures(GS)due to circuit rearrangement.IP-Ens bidirectionally modulates FS but not GS by controlling coherence with hippocampus.CONCLUSION This study indicates that the interictal state may represent a seizure-preventing environment,and the interictal-activated ensemble may serve as a potential therapeutic target for epilepsy.

Objective:To investigate the effect of 1 064-nm Q-switched Nd:YAG laser at different energy settings on cell viability, protease activity and structures of Malassezia furfur. Methods:Cultured standard strains of Malassezia furfur were divided into several groups to be irradiated with 1 064-nm Q-switched Nd:YAG laser at different energies of 0 (control group) , 500, 600, 700, 800 and 900 mJ, respectively. Then, fungal suspensions in the above groups were inoculated onto the Leeming & Notman medium separately. After 7-day culture, the diameter and number of colonies were measured to evaluate the fungal cell viability, the protease activity was measured by using the whole-milk plate medium, and the ultrastructure of Malassezia furfur in each group was observed by transmission electron microscopy. One-way analysis of variance was used for comparisons among multiple groups, least significant difference- t test for multiple comparisons, and Pearson correlation analysis for analyzing correlations of laser energy with colony diameter, number and protease activity. Results:The colony diameter and number both significantly differed among the control group, 500-, 600-, 700-, 800- and 900-mJ groups (colony diameter: 4.05 ± 0.69, 3.76 ± 0.51, 3.28 ± 0.41, 3.09 ± 0.72, 2.54 ± 0.64 and 2.43 ± 0.41 mm, respectively; colony number: 4 787 ± 597, 4 287 ± 761, 1 879 ± 275, 1 082 ± 248 and 209 ± 42, 72 ± 31 colony-forming units, respectively; F = 14.83, 231.85, respectively, both P < 0.05) , and were significantly decreased in the 600-, 700-, 800- and 900-mJ groups compared with the control group (all P < 0.05) . The laser energy was negatively correlated with the colony diameter and number ( r = -0.67, -0.91, respectively, both P < 0.05) . The protease activity significantly differed among the control group, 500-, 700- and 900-mJ groups ( F = 346.60, P < 0.05) , and was significantly lower in the 700- and 900-mJ groups than in the control group (both P < 0.05) . There was a negative correlation between the laser energy and protease activity ( r = -0.94, P < 0.05) . Transmission electron microscopy showed intact fungal structures in the control group, relatively intact fungal structures in the 500-mJ group, and obviously damaged fungal structures in the 600- to 900-mJ groups, and the greater the laser energy, the more severely the fungal structures were damaged. Conclusion:The 1 064-nm Q-switched Nd:YAG laser could affect the cell viability of and protease activity in Malassezia furfur, and damage its structures.

In traditional clinical teaching of impacted tooth extraction, there are problems including limited observation fields for students, insufficient doctor-patient communication training, inadequate clinical thinking development, and low levels of clinical participation. Based on the core elements of visualizing jaw structure with perspective, formulating clinical strategy, immersing in clinical participation, and cultivating the spirit of caregiving, we developed a virtual reality simulation teaching platform for the clinical decision analysis of impacted tooth extraction. The virtual simulation-based platform can create three different impacted tooth scenarios in 3D, which demonstrates the process of clinical decision analysis of impacted tooth extraction through virtuality-reality interaction, allowing students to immerse in the discovery, analysis, and resolution of the medical and humanistic problems in the process of impacted tooth extraction. The questionnaire survey showed that 81.36% (48/59) of the students believed that the software could improve their clinical thinking ability; 76.27% (45/59) of the students believed that it could help them master the basic process of impacted tooth extraction; and 62.71% (37/59) of the students believed that it could improve their skills of impacted tooth extraction. By providing immersive learning experience, constructive teaching design, and multi-dimensional teaching evaluation, the software achieved the goals of cultivating students' clinical thinking ability and professional literacy in oral and maxillofacial surgery.

Parthenogenetic embryonic stem (pES) cells isolated from parthenogenetic activation of oocytes and embryos, also called parthenogenetically induced pluripotent stem cells, exhibit pluripotency evidenced by both in vitro and in vivo differentiation potential. Differential proteomic analysis was performed using differential in-gel electrophoresis and isotope-coded affinity tag-based quantitative proteomics to investigate the molecular mechanisms underlying the developmental pluripotency of pES cells and to compare the protein expression of pES cells generated from either the in vivo-matured ovulated (IVO) oocytes or from the in vitro-matured (IVM) oocytes with that of fertilized embryonic stem (fES) cells derived from fertilized embryos. A total of 76 proteins were upregulated and 16 proteins were downregulated in the IVM pES cells, whereas 91 proteins were upregulated and 9 were downregulated in the IVO pES cells based on a minimal 1.5-fold change as the cutoff value. No distinct pathways were found in the differentially expressed proteins except for those involved in metabolism and physiological processes. Notably, no differences were found in the protein expression of imprinted genes between the pES and fES cells, suggesting that genomic imprinting can be corrected in the pES cells at least at the early passages. The germline competent IVM pES cells may be applicable for germ cell renewal in aging ovaries if oocytes are retrieved at a younger age.
Animals ; Cell Line ; Electrophoresis, Gel, Two-Dimensional ; Mice ; Parthenogenesis ; physiology ; Pluripotent Stem Cells ; metabolism ; Proteomics ; methods

Objective:To evaluate the role of exosomal long non-coding RNA (lncRNA) colon cancer associated transcript-2(CCAT2) in angiogenesis in nasopharyngeal carcinoma(NPC).Methods:Human umbilical vein endothelial cells (HUVECs) were divided into CNE2 supernatant coculture group and NP69 supernatant coculture group. The proliferation ability of HUVEC in each group was examined by CCK8. The lncRNA CCAT2 level in HUVEC was detected using real-time fluorescent quantitative PCR (qRT-PCR). HUVECs were divided into coculture group of serum in patients with NPC and coculture group of serum in healthy donors. The proliferation ability of HUVEC in each group was examined by CCK8. The lncRNA CCAT2 level in each group was detected using qRT-PCR. Exosomes from CNE2,NP69 supernatant were obtained by ultracentrifugation. HUVECs were divided into coculture group of CNE2′s supernatant exosomes and coculture group of NP69′s supernatant exosomes. Biological experiments like CCK8 were conducted to compare the angiogenesis ability of HUVEC in each group. shRNA transfection was used in HUVEC to suppress the expression of lncRNA CCAT2. Biological experiments like CCK8were conducted to compare the HUVEC′s function with different expression of lncRNA CCAT2. CNE2 was transfected with shRNA, the total supernatant exosomal RNA was extracted, and the difference of exosomal lncRNA CCAT2 in CNE2 supernatant with low expression and normal expression of lncRNA CCAT2 was detected by qRT-PCR. The above mentioned two groups of exosomes were cocultured with HUVEC respectively. The difference of lncRNA CCAT2 expression in each group′s HUVEC was detected by qRT-PCR and the function of HUVEC was examined with migration assay. GraphPad Prism 8 software was used for graphing and statistical analysis.Results:Compared with those in the NP69 supernatant coculture group, HUVECs in the CNE2 supernatant coculture group showed a higher proliferation ability and higher expression of lncRNA CCAT2(1 vs. 1.40±0.01, t=42.23, P=0.000). Compared with those in serum of healthy donors, HUVECs in serum of patients with NPC showed a higher proliferation ability and the expression of lncRNA CCAT2 increased (1 vs. 1.25±0.03, t=14.43, P=0.001). Compared with those in the coculture group of NP69 cells′ supernatant exosomes, HUVEC in the coculture group of CNE2 cells′ supernatant exosomes showed a higher proliferation ability and there were more cells that had passed the well during the migration experiment (53.12±2.13 vs. 154.74±4.17, t=37.73, P=0.000) and more knots formed in the tube-formation experiment(10.72±1.02 vs. 53.65±3.21, t=22.63, P=0.000). Compared with those in the sh-NC group, HUVEC in the sh-CCAT2 group showed a lower proliferation ability, and there were less cells that had passed the cell during the migration experiment(401.34±22.15 vs.138.25±6.85, t=23.19, P=0.000) and less MVC in the experiment of subcutaneous stroma embolization in nude mice(41.00±0.32 vs. 27.15±0.23, t=61.53, P=0.000). Compared with the supernatant exosomes in the sh-NC exosome group, the expression of lncRNA CCAT2 was lower in the supernatant exosomes of sh-CCAT2 group (1 vs. 0.65±0.02, t=30.31, P=0.000). Compared with those of the exosomes of sh-NC exosome group, the expression of lncRNA CCAT2 was lower in the HUVEC cocultured with exosomes of the sh-CCAT2 exosome group (1 vs. 0.73±0.01, t=46.77, P=0.000) and the exosomes of sh-CCAT2 exosome group showed a lower migration ability (389.73±26.34 vs. 190.54±8.36, t=12.47, P=0.001) after cocultured with HUVEC. Conclusion:Nasopharyngeal carcinoma-derived exosomal lncRNA CCAT2 could promote angiogenesis.

AIM:To investigate the potential impact of mitoNEET[mitochondrial protein containing Asn-Glu-Glu-Thr(NEET)sequence]on mitochondrial metabolism in brown adipocytes,and to elucidate its underlying mecha-nism.METHODS:An in vitro model of primary mouse brown adipocytes was established.Western blot were utilized to detect relevant proteins,and iron ion and ATP content was measured using kits.Mitochondrial membrane potential and re-active oxygen species(ROS)were assessed by fluorescence microscopy and flow cytometry.RESULTS:The expression of the ferroptosis-related protein ACSL4 increased by 1.13 times in ferroptosis inducer erastin treatment group,whereas the expression of SLC7A11 and GPX4 decreased by 27.33%and 25.33%,respectively,compared with control group(P<0.05).The expression of Nrf1,PGC-1α,MFN2 and UCP1 proteins,related to mitochondrial energy metabolism,de-creased by 20.98%,15.17%,15.03%and 34.22%,respectively(P<0.05).Additionally,the mitoNEET protein con-tent was significantly reduced by 42.14%(P<0.05).The iron ion content in erastin group was substantially increased by 1.80 times compared with control group.However,a notable decrease in ATP content of 14.95%was seen(P<0.05).The results obtained from fluorescence microscopy and flow cytometry demonstrated a significant decrease in the mitochon-drial membrane potential of brown adipocytes in erastin group,with reductions of 52.18%and 61.31%(P<0.05),re-spectively.A substantial increase in mitochondrial ROS content of 80.97%was seen(P<0.05).Western blot analysis of overexpressed stable strains revealed a significant elevation in mitoNEET levels in brown adipocytes following lentivirus transfection,exhibiting an increase of 11.19 times(P<0.05),thus confirming successful transfection.The LV-mitoNEET group exhibited a significant decrease of 37.95%in the expression of ferroptosis-related protein ACSL4 in brown adipose cells compared with control group.Additionally,there was a notable increase of 77.82%and 66.3%in the expression of SLC7A11 and GPX4,respectively(P<0.05).Up-regulation was observed in the expression of MFN2(79.06%),PGC-1α(72.89%),Nrf1(40.14%),and UCP1(31.68%)(P<0.05).The test results demonstrated that the LV-mitoNEET group experienced a reduction of 43.5%in iron ion content compared with control group while exhibiting an increase of 33.5%in ATP content(P<0.05).The results obtained from fluorescence microscopy and flow cytometry demonstrated that mitoNEET overexpression led to a significant increase in the mitochondrial membrane potential of erastin-induced brown adipocytes,with increments of 17.61%and 96.05%,respectively.Additionally,mitoNEET overexpression effec-tively reduced the production of mitochondrial ROS by 24.48%(P<0.05).CONCLUSION:Our findings suggest that mitoNEET overexpression can effectively inhibit the disruption of mitochondrial energy metabolism caused by ferroptosis-induced death of brown adipocytes.

Anti-seizure medications (ASMs) are the main therapy for epilepsy.There are many kinds of ASMs with complex mechanism of action, so it is difficult for pharmacists to examine prescriptions.This paper put forward some suggestions on the indications, dosage forms/routes of administration, appropriateness of usage and dosage, combined medication and drug interaction, long-term prescription review, individual differences in pathophysiology of children, and drug selection when complicated with common epilepsy, for the reference of doctors and pharmacists.