ObjectiveTo investigate the effect of Yaoshu Zhuyu Fang on the regulation of the microRNA-17-5P (miR-17-5P)/murine double minute 2 (MDM2)/p53 axis in the proliferation and apoptosis of rat intervertebral disc annulus fibrosus cells, and its potential molecular mechanism. MethodsIntervertebral disc annulus fibrosus tissues were obtained from 8-week-old SPF-grade male SD rats, and annulus fibrosus cells were isolated and obtained by enzyme digestion and mechanical dispersion. Annulus fibrosus cells were divided into 6 groups: Group C was the blank control group, in which annulus fibrosus cells were not treated with interleukin-1β (IL-1β) but were cultured in RPMI 1640 complete medium. Group β was the degeneration model group constructed by treating annulus fibrosus cells with 10 ng/mL IL-1β for 24 h. Group β+B was the IL-1β + blank serum group, in which annulus fibrosus cells were first treated with IL-1β to construct the degeneration model, then treated with RPMI 1640 medium containing 5% blank serum for 24 h. Group β+W was the IL-1β + Yaoshu Zhuyu Fang-containing serum group, in which annulus fibrosus cells were first treated with IL-1β to construct the degeneration model, then treated with RPMI 1640 medium containing 5% Yaoshu Zhuyu Fang-containing serum for 24 h. Group β+I was the IL-1β + miR-17-5P inhibitor group, in which annulus fibrosus cells were first treated with IL-1β to construct the degeneration model, then transfected with miR-17-5P inhibitor. Group β+I+W was the IL-1β + miR-17-5P inhibitor + Yaoshu Zhuyu Fang-containing serum group, in which annulus fibrosus cells were first treated with IL-1β to construct the degeneration model, then transfected with miR-17-5P inhibitor, and finally treated with RPMI 1640 medium containing 5% Yaoshu Zhuyu Fang-containing serum for 24 h. CCK-8 assay was used to detect cell survival rate. Flow cytometry was used to detect cell apoptosis. Real-time quantitative PCR was used to detect the expression levels of miR-17-5P, MDM2 mRNA, and p53 mRNA in cells. Western blotting was used to detect the protein expression levels of MDM2 and p53 in cells. Dual-luciferase reporter system was used to analyze the targeting relationship between miR-17-5P and MDM2. ResultsCompared with Group C, Group β showed a significant decrease in cell survival rate (P<0.001), a significant increase in cell apoptosis rate (P<0.001), significantly increased expression of miR-17-5P, p53 mRNA, and p53 protein (P<0.001), and significantly decreased expression of MDM2 mRNA and protein (P<0.001). Compared with Group β, Group β+W, Group β+I, and Group β+I+W showed significantly increased cell survival rate, significantly decreased apoptosis rate, significantly decreased expression of miR-17-5P, p53 mRNA, and p53 protein, and significantly increased expression of MDM2 mRNA and protein (P<0.001). Moreover, changes in the above indicators were greater in Group β+I+W (P<0.001). Circular RNA Interactome predicted that miR-17-5P had specific binding sites with the 3' untranslated region (3'UTR) of MDM2. Transfection of miR-17-5P mimic significantly reduced the luciferase expression level of co-transfected luciferase reporter plasmid containing wild-type MDM2 3'UTR (P<0.05), but had no significant effect on luciferase expression in cells co-transfected with luciferase reporter plasmid containing mutant MDM2 3'UTR (P>0.05). ConclusionYaoshu Zhuyu Fang down-regulates the expression of miR-17-5P, promotes the synthesis of MDM2 protein, thereby down-regulates p53, promotes proliferation, and inhibits the apoptosis of rat intervertebral disc annulus fibrosus cells.

The catalytic activity of 3-mercaptopyruvate (3MP) sulfurtransferase (MPST) converts 3MP to hydrogen sulfide (H₂S). However, the regulatory mechanisms governing MPST and its impact on the brain remain largely unexplored. Our study reveals the neuroprotective role of endothelial MPST-generated H₂S, regulated by protein phosphatase 2A (PP2A). Bioinformatics analysis and RNA sequencing demonstrated that endothelial PP2A is associated with neurodegenerative disease pathways. Cerebral ischemic mice exhibited significant inactivation of endothelial PP2A, evidenced by the reduction of PP2Acα in the brain endothelium. Mice with endothelium-specific null PP2A (PP2A^EC-cKO) exhibited neuronal loss, cognitive dysfunction, and long-term potentiation deficits. Postnatal inactivation of endothelial PP2A also contributes to cognitive dysfunction and neuronal loss. However, regaining endothelial PP2A activity by overexpressing Ppp2ca rescued neuronal dysfunction. Mechanistically, PP2A deficiency is intricately linked to the MPST-H₂S signaling pathway. A robust reduction in endothelial MPST-dependent H₂S production followed PP2A deficiency. Exogenous H₂S treatment and AAV-mediated overexpression of MPST in brain endothelial cells significantly mitigated neuronal dysfunction in PP2A^EC-cKO mice. Furthermore, PP2A deficiency promotes an increase in calcium influx and calpain2 phosphorylation, subsequently leading to MPST degradation. The PP2A activator (FTY720) and MPST activator (3MP sodium) both remarkably restored endothelial MPST-dependent H₂S production, subsequently rescuing ischemia-induced neurological deficits. In conclusion, our study demonstrates that endothelial PP2A deficiency leads to MPST degradation by activating calpain2, thus damaging neuronal function.

Objective To explore the application effect of health education based on gain and loss message framing on the treatment behavior intention and self-management of patients with high-risk diabetic foot.Methods From July to September 2024,convenience sampling was used to select patients with high-risk diabetic foot who were hospitalized in the endocrinology department of a tertiary general hospital in Guiyang as the study subjects.They were divided into 3 groups according to the admission time,with 30 patients in each group.The experimental group adopted health education based on gain message framing or framing loss message,while in the control group,health education was provided in a conventional manner.Before and after intervention,the differences of intervention effects among the 3 groups were compared by using diabetic foot pre-hospital delay intention questionnaire,diabetic foot care knowledge questionnaire and Chinese version of Nottingham foot care assessment scale(CNAFF).Results Ultimately,29 cases in the gain framing group,29 cases in the loss framing group,and 29 cases in the control group completed the study.After intervention,the score of pre-hospital delay intention questionnaire of diabetic foot in the gain framing group was(21.48±4.32),and it was(24.31±2.49)in the loss framing group,and(17.76±5.03)in the control group.The difference among the 3 groups was statistically significant(F=18.725,P＜0.001);the loss framing group was superior to the gain framing group(P=0.01)and the control group(P＜0.001).After the intervention,the score of the CNAFF in the gain framing group was(55.83±3.06),and it was(59.14±2.90)in the loss framing group,and(48.66±2.58)in the control group.The difference between the 3 groups was statistically significant(F=102.245,P＜0.001).The loss framing group was superior to the gain framing group and the control group(all P＜0.001).Conclusion Health education based on the loss message framing is more conducive to improving patients' intention to delay diabetic foot visits,leading to good foot care behaviors,and may provide an effective means of pre-hospital prevention and control of diabetic foot.

Objective To investigate the intervention mechanism of Bushen Zhupai Decoction on polycystic ovary syndrome rats with insulin resistance based on IL-6/PI3K/AKT signaling pathway Methods A rat model of polycystic ovary syndrome with insulin resistance was established by feeding high-fat diet combined with oral administration of letrozole,and the blank group was fed with ordinary diet.The modeled rats were randomly divided into model group,metformin group and Bushen Zhupai Decoction low-,medium-and high-dosage groups.The corresponding intervention were given for 28 days.Fasting blood glucose(FBG)content was detected,and insulin resistance index(HOMA-IR)was calculated.ELISA was used to detect serum fasting insulin(FINS),testosterone(T),estradiol(E2),luteinizing hormone(LH)and follicle stimulating hormone(FSH)contents.HE staining was used to observe the morphology of ovarian tissue.Immunohistochemistry was used to detected the expressions of interleukin(IL)-6,p-PI3K,p-AKT and GLUT4 in ovarian tissue.Western blot was used to detect the protein expressions of IL-6,PI3K,p-PI3K,AKT,p-AKT and GLUT4 in ovarian tissue,while RT-PCR was used to detect the mRNA expressions of IL-6,PI3K,AKT and GLUT4.Results Compared with the blank group,the body mass,FBG,FINS and serum T and LH contents of the model group rats significantly increased(P<0.01),while the serum E2 and FSH contents significantly decreased(P<0.01);ovarian tissue showed polycystic changes,with reduced layers and loose arrangement of granulosa cells,the expression of IL-6 protein and mRNA in ovarian tissue significantly increased(P<0.01),the expressions of p-PI3K,p-AKT and GLUT4 proteins significantly decreased(P<0.01),and the expressions of PI3K,AKT and GLUT4 mRNA significantly decreased(P<0.01).Compared with the model group,the body mass of rats in each administration group decreased,and the contents of FBG,FINS and serum T and LH decreased(P<0.01,P<0.05),while the contents of E2 and FSH increased;polycystic changes in ovarian tissue were reduced,mature follicles increased,and granulosa cells were tightly arranged,the expression of IL-6 protein and mRNA in ovarian tissue decreased,the expressions of p-PI3K,p-AKT,GLUT4 protein and PI3K,AKT,GLUT4 mRNA increased.Bushen Zhupai Decoction high-dosage group showed statistical significance(P<0.01).Conclusion Bushen Zhupai Decoction can improve reproductive and metabolic disorders in polycystic ovary syndrome with insulin resistance rats,and its mechanism may be related to inhibiting IL-6 expression,activating the PI3K/AKT signaling pathway,and alleviating insulin resistance.

Objective To evaluate drug-related problems(DRPs)and to analyze the influencing factors of patients un-dergoing video-assisted thoracoscopic surgery(VATS)before operation in thoracic surgery.Methods Clinical pharmacists used the Pharmaceutical Care Network Europe(PCNE)classification system(version 9.1)to analyze DRPs and influencing fac-tors of patients who received VATS from March 1 to May 31,2023,and had at least one comorbidity.Results Out of 300 pa-tients,174 were involved in a total of 200 DRPs.The most common category of DRPs is treatment safety(47.50％),followed by treatment effectiveness(46.00％)and others(6.50％).The most common cause of the problem is drug selection(33.83％),fol-lowed by other(33.33％)and patient cause(19.90％).367 interventions were conducted for DRPs,with the most interventions being at the drug level(55.86％),followed by the doctor level(39.24％)and the patient level(3.54％).In the end,96.00％of the intervention plan was accepted,and 86.50％of the problems were resolved.There were significant differences(P＜0.05)in the number of underlying diseases,medication varieties,body mass index(BMI),and length of hospital stay between the group with and without DRPs.The results of multivariate analysis showed that comorbidities,number of medication types,and BMI were independent risk factors for preoperative occurrence(or potential)of DRPs in VATS patients in thoracic surgery(OR＞1,P＜0.05).Conclusions Clinical pharmacists can effectively evaluate preoperative DRPs in patients undergoing VATS in thoracic surgery through the PCNE classification system.Comorbidities,number of medications,and BMI are influential factors for the oc-currence of preoperative DRPs.Future clinical practice should focus on these risk factors to optimize treatment strategies and re-duce the occurrence of DRPs.

Objective: To explore the associations of moderate to vigorous intensity physical activity (MVPA) and sedentary behavior (SB) among primary and secondary school students and their parents, so as to provide a scientific basis for formulating targeted physical activity promotion strategies for children and adolescents. Methods: From 2021 to 2022, basic information and 24 h movement behaviors of 2 484 pairs of students and their parents were collected from five primary and secondary schools in Haizhu District, Guangzhou City, with a convenient sampling combining with cluster sampling method. Component regression models were constructed to analyze the relationship between parental MVPA, SB and primary and secondary school students MVPA and SB, and a component isochronous substitution model was used to explore the effects of mutual substitution between parental MVPA, residual components (time use components other than SB during the 24 h period), and SB on the behavioral activities of MVPA and SB in primary and secondary school students. Results: Parental MVPA and SB of students in grade 1 to 3 were positively correlated with both students MVPA and SB ( β=0.06, 0.12, P <0.01). The component isochronous substitution model showed that substituting 10 and 20 minutes of MVPA for SB by parents in grade 1 to 3 was associated with an increase in MVPA of students, and substituting 10 and 20 minutes of residual ingredients for SB was associated with a decrease in SB of students, with mean changes of 0.8 (95% CI =0.4-1.2) and 1.4 (95% CI =0.7-2.2) and -1.4 (95% CI =-1.7 to -1.1) and -2.9 (95% CI =-3.4 to -2.3)( P <0.05). No statistically significant associations were observed between parents of students in grades 4 to 6 and 7 to 9 and students physical activity and sedentary behaviour ( P >0.05). Conclusions Parents of students in grades 1 to 3 increases MVPA and decrease SB are beneficial to increase MVPA and decrease SB of students. Parents could promote physical activity among primary and secondary school students, and the intervention gateway should be advanced, with the low grades as the optimal intervention period.

BACKGROUND:Prostaglandin E1(PGE1)has been shown to play a regulatory role in vasodilatation,inflammation,and leukocyte migration and adhesion,but its effects on spinal cord microcirculation after traumatic spinal cord injury(SCI)remain poorly understood. OBJECTIVE:To investigate the mechanism underlying the protective effects of PGE1 administered during the acute phase of traumatic SCI in rats on the regulation of vascular-related factors and microcirculatory function. METHODS:Seventy-two female Sprague-Dawley rats were divided into three groups(n=24 per group):control group,SCI group,and PGE1 group.An in vivo SCI model was established using Allen's blow method.Rats in the PGE1 group were injected with PGE1(10 μg/kg)via the tail vein immediately after SCI.Spinal cord microcirculatory blood flow and oxygen saturation,spinal cord microvessel diameter and area,spinal cord water content,vascular function regulators(von Willebrand factor,thromboxane A2,prostacyclin,endothelin-1),and inflammatory factors(tumor necrosis factor-α,interleukin-1β)were measured at 2 and 24 hours after SCI. RESULTS AND CONCLUSION:At 2 hours after SCI,the diameter and area of spinal cord microvessels,spinal cord microcirculatory blood flow,and oxygen saturation in the PGE1 group were higher than those in the SCI group(P<0.05),the water content of the spinal cord was lower than that in the SCI group(P<0.05),and the level of plasma von Willebrand Factor,the ratio of thromboxane A2/prostacyclin of the spinal cord and the level of endothelin-1 were lower than those in the SCI group(P<0.05).At 24 hours after SCI,the spinal cord microvessel area,blood flow,and oxygen saturation of rats in the PGE1 group were higher than those in the SCI group(P<0.05),the spinal cord water content was lower than that in the SCI group(P<0.05),and the levels of plasma von Willebrand factor,spinal cord tissue thromboxane A2/prostacyclin ratio and the levels of endothelin-1,tumor necrosis factor-α and interleukin-1β were lower than those in the SCI group(P<0.05).The diameter and area of spinal cord microvessels,spinal cord microcirculatory blood flow and blood oxygen saturation of rats in the SCI group were higher than those in the SCI group at 24 hours post-injury(P<0.05),and the levels of plasma von Willebrand factor,spinal tissue thromboxane A2/prostacyclin ratio,tumor necrosis factor-α and interleukin-1β were higher than those at 2 hours post-injury(P<0.05),but the level of endothelin-1 in spinal cord tissue was lower than that at 2 hours(P<0.05).The blood flow and oxygen saturation of spinal cord microcirculation in the PGE1 group rats at 24 hours post-injury were lower than those at 2 hours post-injury(P<0.05),and the diameter and area of spinal cord microvessels and water content of the spinal cord were higher than those at 2 hours post-injury(P<0.05).The above results indicate that intravenous administration of PGE1 in SCI rats immediately after injury can regulate vascular function regulators,inflammatory factors and improve microcirculation of the spinal cord after SCI,which provides a potential basis for the search of drugs for the treatment of acute SCI.

Background::B-cell maturation antigen (BCMA)-directed chimeric antigen receptor T (CAR-T) therapy yield remarkable responses in patients with relapsed/refractory multiple myeloma (R/RMM). Circulating tumor DNA (ctDNA) reportedly exhibits distinct advantages in addressing the challenges posed by tumor heterogeneity in the distribution and genetic variations in R/RMM.Methods::Herein, the ctDNA of 108 peripheral blood plasma samples from patients with R/RMM at the First Affiliated Hospital, School of Medicine, Zhejiang University was thoroughly investigated before administration of anti-BCMA CAR-T therapy to establish its predictive potential. Flow cytometry is used primarily to detect subgroups of T cells or CAR-T cells.Results::In this study, several tumor and T cell effector-mediated factors were considered to be related to treatment failure by an integrat analysis, including higher percentages of multiple myeloma (MM) cells in the bone marrow ( P = 0.0125), lower percentages of CAR-T cells in the peripheral blood at peak ( P = 0.0375), and higher percentages of CD8 + T cells ( P = 0.0340). Furthermore, there is a substantial correlation between high ctDNA level (>143 ng/mL) and shorter progression-free survival (PFS) ( P = 0.007). Multivariate Cox regression analysis showed that high levels of ctDNA (>143 ng/mL), MM-driven high-risk mutations (including IGLL5 [ P = 0.004], IRF4 [ P = 0.024], and CREBBP [ P = 0.041]), number of multisite mutations, and resistance-related mutation ( ERBB4, P = 0.040) were independent risk factors for PFS. Conclusion::Finally, a ctDNA-based risk model was built based on the above independent risk factors, which serves as an adjunct non-invasive measure of substantial tumor burden and a prognostic genetic feature that can assist in predicting the response to anti-BCMA CAR-T therapy.

Objective To construct a model combining with traditional Chinese medicine(TCM)syndrome for predicting the risk of disease progression in patients with idiopathic membranous nephropathy(IMN)by machine learning methods,thus to quantitatively evaluating the value of TCM syndrome in the prediction of the risk of disease progression in IMN.Methods Monofactor analysis,recursive feature elimination(RFE)and multivariate binary Logistic regression analysis were used to screen the independent related factors affecting the risk of disease progression of IMN,and then a risk prediction model was constructed.A total of 102 patients with IMN were randomly assigned to the training set and the test set in a ratio of 65∶35,and then the comparison was conducted in the performance indicators of accuracy,sensitivity,specificity,F1 value,and area under the receiver operating characteristic(ROC)area under the curve(AUC)of the risk prediction model with or without the inclusion of the TCM syndrome information.Results Before the inclusion of TCM syndrome information,12 clinical characteristic variables for patients with MN were obtained after monofactor analysis combined with RFE screening,and they were age,hemoglobin quantification,urinary occult blood,24-hour urine protein quantification,urine protein-creatinine ratio,estimated glomerular filtration rate(eGFR),creatinine,uric acid,alanine transaminase,anti-phospholipase A2 receptor antibody(PLA2R-Ab),total cholesterol,and low-density lipoprotein cholesterd.A risk cholesterol prediction model containing the above variables was constructed.The multivariate binary Logistic regression analysis showed that the differences of the clinical variables mentioned above between the training-set group and test-set group were statistically significant,and the risk prediction model presented good sensitivity and predictability.Monofactor analysis combined with RFE screening was performed again after the inclusion of TCM syndrome information,and then 14 variables were obtained,which included blood stasis syndrome and dampness obstruction syndrome.The sensitivity and specificity of the model with the inclusion of the TCM syndrome information were significantly improved when compared with those without the inclusion of TCM syndrome information.Conclusion The results of the study initially indicate that TCM syndrome can be used as an important supplementary variable for predicting the risk of disease progression in IMN,and will provide a reference for intelligent diagnosis through the integration of traditional Chinese and western medicine information,and will supply the guidance for the treatment of IMN with TCM.

Objective To explore the effect of acyl-CoA synthetase short-chain family member 3(ACSS3)gene on the proliferation of human large cell lung cancer cells(NCI-H460)using CRISPR/Cas 9 gene editing technology.Methods The expression of ACSS3 was detected by Western blot.ACSS3-targeting sgRNAs were designed,and a CRISPR/Cas 9 knockout vector was constructed and transfected into NCI-H460 cells.The transfected cells were selected with puromycin based on vector-carried resistance.ACSS3-knockout monoclonal cell strains were established by limited dilution method and then expanded in culture.Knockout efficiency was confirmed by Western blot.Cell proliferation was assessed using MTT and colony formation assays.Results The expression of ACSS3 was significantly elevated in NCI-H460 cells as compared with human normal lung epithelial cells BEAS-2B(P＜0.05).No ACSS3 protein was detected in the knockout monoclonal strain,indicating successful generation of ACSS3-knockout NCI-H460 cells.Compared with the control cells transfected with empty vector,the proliferation and colo-ny formation ability were inhibited in NCI-H460 cells with ACSS3 knockout(P＜0.05).Conclusions The ACSS3-knockout NCI-H460 cell strain was successfully established,which provides a foundation for further study on the role of ACSS3 in lung cancer.