BACKGROUND:Rev-erbα is involved in the regulation of inflammation,but pharmacological activation of Rev-erbα increases the risk for cardiovascular diseases.To reduce the relevant risk,an exploration on SR9009,a Rev-erbα agonist,combined with other drugs to relieve inflammation in skeletal myoblasts was conducted,laying the theoretical foundation for the treatment of inflammation-associated skeletal muscle atrophy. OBJECTIVE:To investigate the relationship of SR9009,indolepropionic acid and nuclear factor-κB signaling pathways in lipopolysaccharide-induced C2C12 myoblasts. METHODS:(1)C2C12 myoblasts were induced to differentiate in the presence of lipopolysaccharide(1 μg/mL).RNA-seq and KEGG pathway analysis were used to study signaling pathways.(2)C2C12 myoblast viability was assessed using the cell counting kit-8 assay to determine optimal concentrations of indolepropionic acid.Subsequently,cells were categorized into control group,lipopolysaccharide(1 μg/mL)group,SR9009(10 μmol/L)+lipopolysaccharide group,indolepropionic acid(80μmol/L)+lipopolysaccharide group,and SR9009+indolepropionic acid+lipopolysaccharide group.ELISA was employed to measure protein expression levels of interleukin-6 in the cultured supernatant.Real-time quantitative PCR were employed to measure mRNA expression levels of interleukin-6,tumor necrosis factor α,TLR4 and CD14.Western blot assay were employed to measure protein expression levels of NF-κB p65 and p-NF-κB p65.(3)After Rev-erbα was knocked down by siRNA,knockdown efficiency was assessed by RT-qPCR.And mRNA levels of interleukin-6 and tumor necrosis factor α were also measured. RESULTS AND CONCLUSION:Compared with the blank control group,lipopolysaccharide time-dependently inhibited myofibroblast fusion to form myotubes,the mRNA expression levels of interleukin-6 and tumor necrosis factor α were elevated,and the level of interleukin-6 in the cell supernatant was significantly increased.The results of KEGG pathway showed that the nuclear factor-κB signaling pathway was activated by lipopolysaccharide.Indolepropionic acid exhibited significant suppression of C2C12 myoblasts viability when its concentration exceeded 80 μmol/L.Indolepropionic acid and SR9009 inhibited the activation of NF-κB signaling pathway,thereby played an anti-inflammatory role,and suppressed the mRNA expression levels of interleukin-6,tumor necrosis factor α,TLR4 and CD14.Compared with the lipopolysaccharide group,the ratio of p-NF-κB p65/NF-κB p65 protein expression were downregulated.SR9009 combined with indolepropionic acid notably reduced lipopolysaccharide-induced inflammation,further downregulated the mRNA expression levels of interleukin-6,tumor necrosis factor α,TLR4 and CD14.The ratio of p-NF-κB p65/NF-κB p65 protein expression was significantly lower than that in the SR9009+lipopolysaccharide group or indolepropionic acid+lipopolysaccharide group.Rev-erbα increases time-dependently with lipopolysaccharide induction.The knockdown efficiency of Rev-erbα by siRNA reached over 58%,and lipopolysaccharide was added after Rev-erbα was successfully knocked down.Compared with the lipopolysaccharide group,the mRNA expression levels of interleukin-6 and tumor necrosis factor α were significantly up-regulated.These results conclude that Rev-erbα may act as a promising pharmacological target to reduce inflammation.SR9009 targeted activation of Rev-erbα combined with indolepropionic acid significantly inhibits the nuclear factor-κB signaling pathway and attenuates the inflammatory response of C2C12 myofibroblasts.Moreover,the combined anti-inflammatory effect is superior to that of the intervention alone.

BACKGROUND:Primary cortical neurons and microglial cells play a crucial role in exploring cell therapies for neurological disorders,and most of the current methods for obtaining the two types of cells are cumbersome and require separate extraction.It is therefore crucial to find a convenient and rapid method to extract both types of cells simultaneously. OBJECTIVE:To explore a novel method for simultaneous extraction of primary cortical neurons and microglial cells. METHODS:Newborn suckling SD rats were taken within 24 hours.The brain was removed and placed in a dish with DMEM,and the pia mater was removed for later use.Primary neurons were extracted from the same brain tissue,and then the remaining brain tissue was used to extract microglial cells.The whole process was performed on ice.Extraction and culture steps of primary cortical neurons:The cerebral cortex was taken 2.0-3.0 mm with forceps,and the tissue was digested with papain for 20 minutes.After aborting digestion,the blown tissue presented an adherent tissue suspension.The supernatant cell suspension was obtained,filtered,and dispensed into 15 mL centrifuge tubes.After centrifugation and re-suspension,the cells were inoculated onto 6-well plate crawls coated with L-polylysine.Neuronal morphology was observed at 1-day intervals,and staining could be performed for identification using immunofluorescence staining of MAP2 and β-Tubulin by day 7.Microglia extraction and culture steps:The remaining brain tissue at 8-10 mm thick was subjected to microglial cell extraction,digested by trypsin for 20 minutes.After digestion was stopped,the tissue was blown to a homogenate,and then the homogenate was transferred to the culture bottle for culture.On day 14,the culture flasks were sealed and subjected to constant temperature horizontal shaking for 2 hours.Microglial cells were shed in the supernatant.Purified microglial cells were taken and continued to be cultured for 3 days for identification by Iba1 immunofluorescence staining. RESULTS AND CONCLUSION:(1)After 24 hours of culture,the neurons were adherent to the wall,the cytosol was enlarged,and some neurons developed synapses.After 3 and 5 days of culture,the cytosol was further enlarged,and most of the neurons were in the form of synapses,and some neurons were growing in clusters.On day 7,neuronal synapses were prolonged and thickened,and they were connected with each other to form a network.The neurons were identified by β-Tubulin and MAP2 immunofluorescence staining.(2)The cells grew close to the wall on day 1 of culture.On days 3,5,and 7,the density of microglial cells was small,and the cell morphology was bright oval or round,but the cells basically grew in clumps on the upper layer of other cells.On day 10,the density of microglial cells increased significantly.On day 14,microglial cells grew in dense clumps on the upper layer of other cells,and then they could be isolated and purified.The isolated and purified cells were taken and re-cultured to day 3 and identified as microglial cells by Iba1 immunofluorescence;their purity was greater than 95%.(3)The results show that primary cortical neurons and microglial cells obtained by this method after extraction and culture are of high purity,good morphology,and high viability.

Objective To assess the effect of early nutritional intervention on the patients with respiratory diseases at nutritional risk. Methods A total of 130 patients with respiratory disease who were hospitalized in Shanghai Fifth People’s Hospital, Fudan University between May 2023 and December 2024 and had a nutritional risk screening 2002 score ≥3 points. Based on the initiation time of nutritional intervention, patients were divided into an early group (≤5 days, n＝65) and a late group (＞5 days, n＝65). Results In the early group, prealbumin (P-ALB) and retinol-binding protein (RBP) levels were significantly higher (P＜0.01), C-reactive protein (CRP), procalcitonin (PCT) levels were significantly lower after intervention (P＜0.05). Compared with the late group, the hospital costs were lower and hospital stays were shorter in the early group (P＜0.001). Spearman analysis showed ALB, P-ALB, and total protein (TP) were negatively correlated with hospital costs (r＝－0.37, －0.20, and－0.22, P＜0.05). RBP, ALB, P-ALB, and lymphocyte count (LYM) were negatively correlated with CRP (r＝－0.30, －0.26, －0.37, －0.18, P＜0.01), RBP, ALB, P-ALB, hemoglobin (HB), and TP were negatively correlated with PCT (r＝－0.23,－0.36, －0.40, －0.30, －0.19, P＜0.05). Conclusions For patients with respiratory diseases, early nutritional assessment should be underwent, and for patients with nutritional risk screening 2002 score ≥3 points, early nutritional intervention could improve the nutritional status and alleviate inflammatory response, promote recovery, shorten the hospital stays.

Objective: To analyze the spatio-temporal distribution of pulmonary tuberculosis (PTB) among students in Suzhou City, Jiangsu Province from 2015 to 2023, so as to provide the evidence for the prevention and control of PTB in schools. Methods: Data of PTB cases among students in Suzhou City from 2015 to 2023 were collected from Chinese Disease Prevention and Control Information System and Suzhou Report of Investigation and Disposal of Tuberculosis in Schools. The seasonal incidence of PTB among students was analyzed using seasonal index (SI). The spatio-temporal clustering characteristics of PTB among students were analyzed using spatial autocorrelation and retrospective spatio-temporal permutation scanning. Results: Totally 1 374 PTB cases among students were reported in Suzhou City from 2015 to 2023. PTB cases were reported in each month, and the SIs were 100.69%, 124.38%, 108.98%, 135.04%, 106.61% and 106.61% in April, May, July, September, October and November, respectively, indicating the prevalence of PTB among students. Spatial autocorrelation analysis showed there was a positive spatial correlation of PTB among students in 2019 and 2020 (Moran's I=0.053 and 0.089, both P<0.05). From 2015 to 2023, there were high-high clustering sites mainly in Hengtang Street and Shishan Street. Retrospective spatio-temporal permutation scanning showed a primary cluster in Hengtang Street, with aggregation time in 2017, and 6 secondary clusters covering 25 towns (streets). Conclusion From 2015 to 2023, the PTB cases among students in Suzhou City were mainly concentrated in summer and autumn, and were predominantly clustered in Hengtang Street and Shishan Street.

BackgroundCurrently， the problem of depressed mood in college students is becoming more prominent. The experience of childhood maltreatment is a significant contributor to depression among college students. Although the association between the two has been confirmed， the specific psychosocial mechanisms underlying how childhood maltreatment affects college students' mental health remain insufficiently evidenced. ObjectiveTo explore the mediating role of emotion regulation difficulties in the relationship between childhood maltreatment and depression among college students， and to investigate the moderated effects of psychological resilience and family socioeconomic status， aiming to provide references for improving depressive symptoms in college students. MethodsOn 14 March 2024， a cluster sampling method was employed to recruit 751 college students from a university in Heilongjiang Province. Participants were assessed with Childhood Trauma Questionnaire （CTQ）， Difficulties in Emotion Regulation Scale （DERS）， Patients' Health Questionnaire Depression Scale-9 item （PHQ-9）， 10-item Connor-Davidson Resilience Scale （CD-RISC-10） and Family Socioeconomic Status Questionnaire. Pearson correlation analysis was adopted to examine the correlation between the scores of scales. Model 4 and model 7 in Process 4.2 were used to test the mediating effects of emotional regulation difficulties and the moderated effects of psychological resilience and family socioeconomic status. Results① A total of 712 （94.81%） valid questionnaires were collected. ② College students' CTQ score was positively correlated with DERS score and PHQ-9 score （r=0.296， 0.507， P<0.01）， and negatively correlated with CD-RISC-10 score and Family Socioeconomic Status Questionnaire score （r=-0.148， -0.229， P<0.01）. ③ The indirect effect value of difficulties in emotion regulation on the relationship between childhood maltreatment and depression was 0.091 （95% CI： 0.018~0.046）， accounting for 17.95% of the total effect. ④ The first half of the mediation model "childhood maltreatment → difficulties in emotion regulation → depression" （childhood maltreatment → difficulties in emotion regulation） was moderated by psychological resilience （β=-0.030， t=-6.147， 95% CI： -0.040~-0.020） and family socioeconomic status （β=-0.051， t=-3.929， 95% CI： -0.077~-0.026）. ConclusionChildhood maltreatment exerts both a direct effect on college students' depression and an indirect effect through emotion regulation difficulties. The childhood maltreatment → emotion regulation difficulties pathway in this mediation model is moderated by psychological resilience and family socioeconomic status. ［Funded by Qiqihar Medical University Graduate Student Innovation Fund Project （number， QYYCX2023-48）； Special Research Fund Project for Young Doctors of Qiqihar Academy of Medical Sciences （number， QMSI2021B-08）］

OBJECTIVE To investigate the potential mechanism of epigallocatechin gallate （EGCG） enhancing the sensitivity of hepatocellular carcinoma （HCC） cells to lenvatinib based on the phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway. METHODS Five human HCC cell lines （HepG2， Huh-7， SMMC-7721， SNU-368 and SNU-739） were used to evaluate the effects of lenvatinib alone and in combination with EGCG on survival rates， clone number， proliferation rate， invasion number and the expressions of mRNAs and proteins related to the PI3K/Akt signaling pathway. The PI3K activator insulin-like growth factor-1 （IGF-1） was introduced to investigate the effect of activating the PI3K/Akt signaling pathway on the sensitization effect of EGCG. RESULTS Compared with the control group， lenvatinib （10 μmol/L） and different concentrations of EGCG+ lenvatinib （1， 5 and 10 μg/mL EGCG+10 μmol/L lenvatinib） significantly reduced the survival rates and clone numbers of all five HCC cell lines in a dose-dependent manner （P＜0.05）. Lenvatinib （10 μmol/L） and EGCG+lenvatinib （10 μg/mL EGCG+10 μmol/L lenvatinib） also markedly inhibited the proliferation rate and invasion numbers of these cells， and decreased the mRNA expressions of PI3K， Akt， mammalian target of rapamycin （mTOR）， P70S6K and 4EBP， and the phosphorylation levels of PI3K and Akt， as well as the protein expressions of mTOR and B cell lymphoma-2 （Bcl-2） in HepG2 cells or all five HCC cells； conversely， the mRNA and protein expressions of phosphatase and tensin homologue deleted on chromosome 10（PTEN）， and the protein expressions of caspase-3 and cleaved caspase-3 were significantly upregulated， with more pronounced effects observed in the EGCG+lenvatinib group than in the lenvatinib group （P＜0.05）. Compared with the lenvatinib group and the EGCG+lenvatinib group， the clone number， proliferation rate and invasion number of HepG2 cells in the EGCG+lenvatinib+IGF-1 group （10 μg/mL EGCG+10 μmol/L lenvatinib+50 ng/mL IGF-1） were significantly increased （P＜0.05）. CONCLUSIONS EGCG can enhance the sensitivity of HCC cells to lenvatinib， and its underlying mechanism may be related to the inhibition of the activation of PI3K/Akt signaling pathway activation.

OBJECTIVE To provide a reference for optimizing and promoting the quality standards of Shechuan naolitong granules. METHODS Fifteen batches of Shechuan naolitong granules were used as samples to establish HPLC fingerprints using the Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine （2012 edition）. Similarity evaluation and common peak identification were performed， and orthogonal partial least squares discriminant analysis （OPLS-DA） was used to assess quality differences among different batches and to screen quality differential components. Using salvianolic acid B（SAB） as the internal reference， quantitative analysis of multi-components by single-marker （QAMS） was developed to simultaneously determine geniposidic acid （GA）， chlorogenic acid （CA）， vaccarin （VA）， ferulic acid （FA） and senkyunolide I （SI）. The results were compared with those obtained by the external standard method. RESULTS A total of 13 common peaks were identified in the HPLC fingerprints of 15 batches of samples， and the similarities of the spectra were all above 0.96. Seven chromatographic peaks were identified as GA （peak 3）， CA （peak 6）， VA （peak 8）， FA （peak 9）， SI （peak 11）， SAB（peak 12） and TA（peak 13）. OPLS-DA indicated that the differential quality markers among 15 batches were peaks 5， 11 （SI）， and 12 （SAB）.Using SAB as the internal reference， the relative correction factors for GA， CA， VA， FA and SI were calculated as 1.058 4， 0.594 3， 0.643 3， 0.342 7 and 0.262 8， respectively. The mean content of GA， CA， VA， FA， SI and SAB across the 15 batches of samples were 0.155 0， 0.085 4， 0.140 3， 0.071 8， 0.072 7， 1.276 3 mg/g， respectively， showing no significant difference compared with the ESM （P＞0.05）. CONCLUSIONS The established HPLC fingerprint and QAMS are simple， efficient and economical， providing a reference for the quality control and further development of Shechuan naolitong granules.

Background::While type 2 diabetes mellitus (T2DM) is considered a putative causal risk factor for coronary artery disease (CAD), the intrinsic link underlying T2DM and CAD is not fully understood. We aimed to highlight the importance of integrated care targeting both diseases by investigating the phenotypic and genetic relationships between T2DM and CAD.Methods::We evaluated phenotypic associations using data from the United Kingdom Biobank ( N = 472,050). We investigated genetic relationships by leveraging genomic data conducted in European ancestry for T2DM, with and without adjustment for body mass index (BMI) (T2DM: Ncase/ Ncontrol = 74,124/824,006; T2DM adjusted for BMI [T2DM adjBMI]: Ncase/ Ncontrol = 50,409/523,897) and for CAD ( Ncase/ Ncontrol = 181,522/984,168). We performed additional analyses using genomic data conducted in multiancestry individuals for T2DM ( Ncase/ Ncontrol = 180,834/1,159,055). Results::Observational analysis suggested a bidirectional relationship between T2DM and CAD (T2DM→CAD: hazard ratio [HR] = 2.12, 95% confidence interval [CI]: 2.01–2.24; CAD→T2DM: HR = 1.72, 95% CI: 1.63–1.81). A positive overall genetic correlation between T2DM and CAD was observed ( rg = 0.39, P = 1.43 × 10 -75), which was largely independent of BMI (T2DM adjBMI–CAD: rg = 0.31, P = 1.20 × 10 –36). This was corroborated by six local signals, among which 9p21.3 showed the strongest genetic correlation. Cross-trait meta-analysis replicated 101 previously reported loci and discovered six novel pleiotropic loci. Mendelian randomization analysis supported a bidirectional causal relationship (T2DM→CAD: odds ratio [OR] = 1.13, 95% CI: 1.11-1.16; CAD→T2DM: OR = 1.12, 95% CI: 1.07-1.18), which was confirmed in multiancestry individuals (T2DM→CAD: OR = 1.13, 95% CI: 1.10-1.16; CAD→T2DM: OR = 1.08, 95% CI: 1.04-1.13). This bidirectional relationship was significantly mediated by systolic blood pressure and intake of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors, with mediation proportions of 54.1% (95% CI: 24.9-83.4%) and 90.4% (95% CI: 29.3-151.5%), respectively. Conclusion::Our observational and genetic analyses demonstrated an intrinsic bidirectional relationship between T2DM and CAD and clarified the biological mechanisms underlying this relationship.

Background::Homoharringtonine (HHT) is an effective anti-inflammatory, anti-viral, and anti-tumor protein synthesis inhibitor that has been applied clinically. Here, we explored the therapeutic effects of HHT in a mouse heart transplant model.Methods::Healthy C57BL/6 mice were used to observe the toxicity of HHT in the liver, kidney, and hematology. A mouse heart transplantation model was constructed, and the potential mechanism of HHT prolonging allograft survival was evaluated using Kaplan–Meier analysis, immunostaining, and bulk RNA sequencing analysis. The HHT-T cell crosstalk was modeled ex vivo to further verify the molecular mechanism of HHT-induced regulatory T cells (Tregs) differentiation. Results::HHT inhibited the activation and proliferation of T cells and promoted their apoptosis ex vivo. Treatment of 0.5 mg/kg HHT for 10 days significantly prolonged the mean graft survival time of the allografts from 7 days to 48 days ( P <0.001) without non-immune toxicity. The allografts had long-term survival after continuous HHT treatment for 28 days. HHT significantly reduced lymphocyte infiltration in the graft, and interferon-γ-secreting CD4 + and CD8 + T cells in the spleen ( P <0.01). HHT significantly increased the number of peripheral Tregs (about 20%, P <0.001) and serum interleukin (IL)-10 levels. HHT downregulated the expression of T cell receptor (TCR) signaling pathway-related genes ( CD4, H2-Eb1, TRAT1, and CD74) and upregulated the expression of IL-10 and transforming growth factor (TGF) -β pathway-related genes and Treg signature genes ( CTLA4, Foxp3, CD74, and ICOS). HHT increased CD4 + Foxp3 + cells and Foxp3 expression ex vivo, and it enhanced the inhibitory function of inducible Tregs. Conclusions::HHT promotes Treg cell differentiation and enhances Treg suppressive function by attenuating the TCR signaling pathway and upregulating the expression of Treg signature genes and IL-10 levels, thereby promoting mouse heart allograft acceptance. These findings may have therapeutic implications for organ transplant recipients, particularly those with viral infections and malignancies, which require a more suitable anti-rejection medication.

This article reviews the concept and research tools of social distancing, the current situation and influencing factors of social distancing in elderly diabetic patients, in order to provide a basis for the development of targeted intervention strategies for elderly diabetic patients, reduce the sense of social alienation in elderly diabetic patients, and promote their social participation and sense of meaning in life.