ObjectiveTo integrate network pharmacology prediction with multi-level experimental verification methods， and to explore in depth the therapeutic efficacy and potential mechanism of luteolin in treating gout. MethodsDatabases were used to obtain potential pharmacodynamic targets of luteolin. Protein-protein interaction （PPI） network construction and network pharmacology analysis techniques were used to screen key core targets of luteolin in gout treatment. Further biological function enrichment analysis and signaling pathway analysis were performed on these targets. Molecular docking simulation was used to calculate the binding energy between luteolin and potential core targets， clarifying the strength of their interactions. In the in vivo experiment for hyperuricemia， 48 mice were randomly divided into a blank group， a model group， an allopurinol group （5 mg·kg^-1）， and low-dose （10 mg·kg^-1）， medium-dose （30 mg·kg^-1）， and high-dose （90 mg·kg^-1） luteolin groups. For the first three days， the blank and model groups were gavaged with an equal volume of normal saline， while the allopurinol group and luteolin groups were gavaged with corresponding drugs. From day 4 onwards， modeling was performed by intraperitoneal injection at 12：00 daily （normal saline for the blank group， and oxonic acid potassium-hypoxanthine mixture for other groups， with 300 mg·kg^-1 for each group）. Gavage intervention was administered at 18：00 daily （normal saline for the blank/model groups， and corresponding drugs for the treatment groups） until day 7. After sampling， levels of serum uric acid （UA）， alanine aminotransferase （ALT）， and aspartate aminotransferase （AST） were measured. Levels of xanthine oxidase （XO） in the liver and kidney， ATP-binding cassette transporter G2 （ABCG2） and malondialdehyde （MDA） in the kidney， and superoxide dismutase （SOD） in the liver were determined. Renal HE staining was also performed. In the pharmacodynamic study of gouty arthritis， 36 rats were randomly divided into a blank group， a model group， a colchicine group （0.315 mg·kg^-1）， and low-dose （7 mg·kg^-1）， medium-dose （21 mg·kg^-1）， and high-dose （63 mg·kg^-1） luteolin groups. The model was established by vertically injecting 100 µL of 25 g·L^-1 monosodium urate suspension into the posterior lateral aspect of the right ankle joint （the blank group was injected with an equal volume of normal saline）， with repeated injections every two days for reinforcement. From day 2 after modeling， daily gavage administration was performed （normal saline for the blank/model groups， and corresponding drugs for the treatment groups） for a total of 16 days. During the experiment， ankle swelling and pain threshold were measured regularly. After sampling， levels of serum tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and interleukin-1β （IL-1β） were determined. Ankle joints were subjected to HE， Masson， and safranin O-fast green staining， and HE staining was also performed on ankle synovial tissue and various organs. Western blot was used to determine the expression levels of key proteins in gout-related signaling pathways. ResultsNetwork pharmacology analysis predicted that luteolin may regulate over 20 core targets， such as XO， ABCG2， nuclear factor erythroid 2-related factor 2 （Nrf2）， and SOD， through acting on signaling pathways including NF-κB， phosphoinositide 3-kinase/protein kinase B （PI3K/Akt）， and ABC transporters， thereby affecting uric acid metabolism and inflammatory responses. In the hyperuricemia model， compared with the blank group， the model group showed significantly increased serum UA level， liver and kidney XO activity， renal ABCG2 expression， and liver SOD activity （P<0.01）. Compared with the model group， the high-dose luteolin group significantly reduced serum UA level （P<0.01）， inhibited liver and kidney XO activity （P<0.01）， and significantly increased renal ABCG2 expression and liver SOD activity （P<0.01）， effectively alleviating renal oxidative stress damage and improving renal histopathological status. In the gouty arthritis model， compared with the blank group， the model group showed significant ankle swelling， decreased pain threshold， and significantly increased levels of IL-6， IL-1β， and TNF-α in serum and synovial tissue （P<0.01）. The high-dose luteolin group significantly reduced ankle swelling， prolonged hot plate pain threshold， effectively decreased the levels of the above inflammatory factors in serum and synovial tissue （P<0.01）， and significantly improved ankle pathological damage， showing good analgesic and anti-inflammatory effects. Western blot results further confirmed that luteolin significantly upregulated Nrf2 protein expression and downregulated XO and nucleotide-binding oligomerization domain （NOD）-like receptor protein 3 （NLRP3） expression in animals. ConclusionLuteolin can improve symptoms of hyperuricemia and gouty arthritis， and its potential mechanism may be related to inhibiting XO activity， increasing ABCG2 and SOD levels， and regulating Nrf2-mediated oxidative stress-related pathways.

Successful cloning through somatic cell nuclear transfer (SCNT) faces significant challenges due to epigenetic obstacles. Recent studies have highlighted the roles of H3K4me3 and H3K27me3 as potential contributors to these obstacles. However, the underlying mechanisms remain largely unclear. In this study, we generated genome-wide maps of H3K4me3 and H3K27me3 in mouse pre-implantation NT embryos. Our analysis revealed that aberrantly over-represented broad H3K4me3 domain and H3K27me3 signal lead to increased bivalent marks at gene promoters in NT embryos compared with naturally fertilized (NF) embryos at the 2-cell stage, which may link to relatively low levels of H3K36me3 in NT 2-cell embryos. Notably, the overexpression of Setd2, a H3K36me3 methyltransferase, successfully restored multiple epigenetic marks, including H3K36me3, H3K4me3, and H3K27me3. In addition, it reinstated the expression levels of ZGA-related genes by reestablishing H3K36me3 at gene body regions, which excluded H3K27me3 from bivalent promoters, ultimately improving cloning efficiency. These findings highlight the excessive bivalent state at gene promoters as a potent barrier and emphasize the removal of these barriers as a promising approach for achieving higher cloning efficiency.
Animals ; Mice ; Histone-Lysine N-Methyltransferase/biosynthesis* ; Histones/genetics* ; Nuclear Transfer Techniques ; Female ; Gene Expression Regulation, Developmental ; Promoter Regions, Genetic ; Epigenesis, Genetic ; Embryo, Mammalian/metabolism*

OBJECTIVE: To evaluate the impact of critical care warning platform (CWP) on clinical outcomes of patients transferred from internal medical ward to intensive care unit (ICU) based on real-world data. METHODS: A retrospective cohort study was conducted. The patients transferred from internal medical ward to ICU of Zhongda Hospital, Southeast University, between January 2022 and October 2024, were enrolled. They were divided into critical care warning group and conventional treatment group based on whether they were connected to the CWP. The patients in the critical care warning group were connected to the CWP, which collected real-time vital signs and treatment data. The platform automatically calculated severity scores, generated individualized risk assessments, and triggered warning alerts, allowing clinicians to adjust treatment plans accordingly. The patients in the conventional treatment group were not connected to the CWP and relied on conventional clinical judgment and nursing measures for treatment management. Baseline characteristics [gender, age, body mass index (BMI), admission type, severity score of illness, underlying diseases, and disease type at ICU admission], primary clinical outcome (in-hospital mortality), and secondary clinical outcomes [ICU mortality, length of ICU stay, total length of hospital stay, and mechanical ventilation and continuous renal replacement therapy (CRRT) status] were collected. Multivariate Logistic regression was used to analyze the impact of CWP on in-hospital death, and subgroup analyses were performed based on different patient characteristics. RESULTS: A total of 1 281 patients were enrolled, with 768 in the critical care warning group and 513 in the conventional treatment group. Compared with the conventional treatment group, the proportion of patients in the critical care warning group with underlying diseases of diabetes and malignancy and transferred to ICU due to sepsis was lowered, however, there were no statistically significant differences in other baseline characteristics between the two groups. Regarding the primary clinical outcome, the in-hospital mortality in the critical care warning group was significantly lower than that in the conventional treatment group [17.6% (135/768) vs. 25.7% (132/513), P < 0.01]. For secondary clinical outcomes, compared with the conventional treatment group, the patients in the critical care warning group had significantly fewer days of mechanical ventilation within 28 days [days: 2 (1, 6) vs. 2 (1, 8), P < 0.05], significantly shorter length of ICU stay [days: 3 (2, 8) vs. 4 (2, 10), P < 0.01], and significantly lower ICU mortality [15.1% (116/768) vs. 21.4% (110/513), P < 0.01]. Multivariate Logistic regression analysis showed that, after adjusting for age and underlying diseases, the use of CWP was significantly associated with a reduction of in-hospital mortality among patients transferred from internal medical ward to ICU [odds ratio (OR) = 0.670, 95% confidence interval (95%CI) was 0.502-0.894, P = 0.006]. Further subgroup analysis revealed that, among patients transferred to ICU due to sepsis, the use of CWP significantly reduced in-hospital mortality (OR = 0.514, 95%CI was 0.367-0.722, P < 0.001). In patients aged ≥ 70 years old (OR = 0.587, 95%CI was 0.415-0.831, P = 0.003) and those with underlying diseases of malignancy (OR = 0.124, 95%CI was 0.046-0.330, P < 0.001), CWP also showed significant protective effects on in-hospital prognosis. CONCLUSION The use of CWP is significantly associated with a reduction in in-hospital mortality among patients transferred from internal medical ward to ICU, demonstrating its potential in assessing the deterioration of hospitalized patients.
Humans ; Intensive Care Units ; Retrospective Studies ; Hospital Mortality ; Prognosis ; Critical Care ; Male ; Female ; Patient Transfer ; Middle Aged ; Aged ; Cohort Studies

To summarize Professor WANG Xixing's clinical experience in treating advanced breast cancer with anxiety and depression from the perspective of shaoyang pivot. It is believed that the core pathogenesis of advanced breast cancer with anxiety and depression lies in the dysfunction of shaoyang pivot （referring to the imbalanced regulatory function of the shaoyang meridian system that governs the transportation and transformation of qi， blood， and body fluids）. This dysfunction can lead to abnormal circulation of qi， blood， and body fluids， as well as the intermingling of phlegm and blood stasis， which further promotes the spread and diffusion of cancer toxin. Meanwhile， it disturbs mental activity， resulting in a condition characterized by stagnation of cancer toxin and concurrent disorders of both the physical body and the spirit. Based on this pathogenesis， the basic therapeutic principles of harmonizing shaoyang， regulating the pivot to calm the spirit， and dissipating masses and resolving toxins are proposed. Clinically， the disease is classified into three syndromes for differentiation and treatment. For shaoyang pivot dysfunction syndrome， treatment should use self-prescribed Chaiqin Hengshu Ningxin Decoction （柴芩衡枢宁神汤）； for sanjiao pivot dysfunction syndrome， treatment should prescribe Chaigui Tongshu Dashen Decoction （柴归通枢达神饮）； for gallbladder function disorder syndrome， treatment should apply Wendan Qishu Shoushen Decoction （温胆启枢守神汤）. Throughout the treatment process， the concept of "simultaneous treatment of cancer and depression" is implemented to smooth the shaoyang pivot， block the vicious cycle where cancer toxin and emotional abnormalities mutually reinforce each other.

Objective To construct an in vitro research model for studying human liver regeneration based on human pluripotent stem cells(hPSCs)-derived hepatocyte-like organoid(HLO).Methods The hPSCs-derived HLO was obtained by inducing differentiation and the regeneration model after liver injury was constructed by adding acetaminophen(APAP)at fixed time points in HLO culture conditions to simulate acute liver injury.Subse-quently,HLO with catenin/cadherin-associated protein beta 1(CTNNB1)knockout,a key gene regulating liver re-generation,was constructed using CRISPR/Cas9 gene editing technology,and regeneration experiments with APAP injury were performed.HLO as a model for liver regeneration studies was further evaluated by morphological observation,RT-qPCR,Western blot and pathological analysis.Results Morphology evidence as well as expres-sion of marker genes showed that hPSCs-derived HLO was able to initiate a post-injury regeneration response after APAP treatment.CTNNB1-deficient HLO showed delayed recovery in dimension and down-regulated or delayed ex-pression of related genes during post-injury regeneration as compared to control HLO.Conclusions A HLO-based hPSCs-derived human liver regeneration model is successfully constructed,which can be used for gene function studies during liver regeneration.

Objective To investigate the protective effect of dapagliflozin on kidney and the expression of AMP-activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR)signaling pathway in diabetic nephropathy(DN)rats.Methods A total of 40 SPF Wistar male rats were randomly divided into normal group,model group,low-dose group and high-dose group,with 10 rats in each group.After the DN model was successfully prepared,the rats in normal group were given normal diet + normal saline by gavage,the rats in model group was given high sugar and high fat feed + normal saline by gavage,the rats in low-dose group was given high sugar and high fat feed+1mg/(kg·d)of dapagliflozin by gavage,the rats in high-dose group was given high sugar and high fat feed+10mg/(kg·d)of dapagliflozin by gavage.Rats in each group were continuously gavaged for 12 weeks.Renal function indexes,renal pathological changes,p-AMPK and p-mTOR protein expression,collagen type Ⅰ(COL Ⅰ),collagen type Ⅳ(COL Ⅳ)and fibronectin(FN)of all groups were compared.Results Blood urea nitrogen(BUN),serum creatinine(SCr),24h urinary protein quantity,p-mTOR protein expression,COL Ⅰ,COL Ⅳ and FN levels of rats in model group,low-dose group and high-dose group were significantly higher than those in normal group,and p-AMPK protein expression was significantly lower than that of normal group(P<0.05).BUN,SCr,24h urinary protein quantity,p-mTOR protein expression,COL Ⅰ,COL Ⅳ and FN levels of rats in low-dose group and high-dose group were significantly lower than those in model group,while p-AMPK protein expression was significantly higher than that in model group(P<0.05).BUN,SCr,24h urinary protein quantity,p-mTOR protein expression,COL Ⅰ,COL Ⅳ and FN levels in high-dose group were significantly lower than those in low-dose group,and p-AMPK protein expression was significantly higher than that in low-dose group(P<0.05).Conclusion Dapagliflozin has a good kidney protection effect on DN rats,and its mechanism may be related to the regulation of AMPK/mTOR signaling pathway.

Objective:To evaluate the relationship between gluten-free diet and rheumatoid arthritis (RA).Methods:Data were obtained from large-scale genome-wide association studies (GWAS), and genetic loci that are independent of gluten-free diet and RA of people of Europe2 were selected as instrumental variables. The gluten-free diet GWAS data included 64 949 individuals and 9 851 867 controls. Data were obtained from GWAS of 58 284 RA patients and 13 108 512 controls. The inverse variance weighted (IVW), MR Egger, weighted median method and weighted model were used to conduct two sample Mendelian randomization (MR) analysis. Cochran Q test and mendelian randomness pleiotropy residual sum and outlier (MR-PRESSO) were used to assess SNP heterogeneity. Applying the MR Egger intercept to test the level pleiotropy of SNP. The sensitivity analysis of the "leave one method" that evaluates whether MR studies were influenced by a single SNP. Results:After matching GFD and RA data, three SNPs were included as instrumental variables in the study. IVW showed that GFD could significantly reduce the risk of RA ( β=-60.83, s x=3.82, P<0.001). The weighted median method and weighted pattern also showed that the gluten free diet could reduce the risk of RA ( β=-57.97, s x=4.41, P<0.001; β=-55.81, s x=5.10, P=0.008). Sensitivity analysis of the correlation between GFD and RA showed that there might be heterogeneity between SNPs (Cochran Q test, Q=12.80, P=0.002). The MR-PRESSO results showed that no abnormal SNP was detected ( P=0.174). The forest map showed that SNPs was closely related to GFD and RA stability. The method comparison chart showed that the results of multiple testing methods were basically consistent. The funnel plot showed that SNPs were basically symmetrical, indicating that there was no pleiotropy in MR analysis. The MR Egger intercept test showed no horizontal pleiotropy in MR analysis (intercept value was-0.24, P=0.174). The sensitivity analysis of the "leave one method" is suggested that no single SNP had a significant impact on the overall results. Conclusion:Gluten free diet is related to the risk reduction of RA.

Objective To evaluate the protective effect and underlying mechanism of ulinastatin on hepatic ischemia-reperfusion injury.Methods Twenty-four male SD rats were divided into three groups:sham operation group(Sham group),hepatic ischemia-reperfusion injury group(HIRI group)and hepatic ischemia-reperfusion injury+ulinastatin pretreatment group(HIRI+UTI group),with 8 rats in each group.The HIRI rat models were established by occluding hepatic portal vein and hepatic artery for 1 h.In the HIRI+UTI group,the rats were intraperitoneally injected with ulinastatin at 30 min before model establishment,and an equivalent amount of normal saline was given in the Sham and HIRI groups.The rats were sacrificed at 6 h after model establishment.Serum samples were collected to detect alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels.Pathological changes of liver tissues were observed by hematoxylin-eosin(HE)staining.Ultrastructural changes of mitochondria in liver tissues were observed by transmission electron microscopy.The expression of glutathione peroxidase 4(GPX4)was determined by immunofluorescent staining.The contents of malondialdehyde(MDA),glutathione(GSH),Fe,reactive oxygen species(ROS)and GPX4 were detected.The expression levels of GPX4 and acyl-CoA synthetase long-chain family 4(ACSL4)messenger RNA(mRNA)and proteins in liver tissue were measured.Results Compared with the Sham group,serum ALT and AST levels were up-regulated,pathological changes such as congestion,hepatocyte necrosis and abnormal hepatic lobule structure were observed,pathological score was increased,the mitochondria shrank,the membrane density was increased,the mitochondrial crest was damaged or even absent,the contents of ROS,MDA and Fe were elevated,the GSH content was decreased,the fluorescent intensity of GPX4 was weakened,the relative expression levels of ACSL4 mRNA and protein were up-regulated,and the relative expression levels of GPX4 mRNA and protein were down-regulated in the HIRI group(all P＜0.05).Compared with the HIRI group,serum ALT and AST levels were down-regulated,liver tissue injury was alleviated,pathological score was decreased,mitochondrial shrinkage and crest breakage were mitigated,the contents of ROS,MDA and Fe were down-regulated,the GSH content was up-regulated,the fluorescent intensity of GPX4 was enhanced,the relative expression levels of ACSL4 mRNA and protein were down-regulated,and the relative expression levels of GPX4 mRNA and protein were up-regulated in the HIRI+UTI group(all P＜0.05).Conclusions Ulinastatin may alleviate hepatic ischemia-reperfusion injury in rats probably through inhibiting ferroptosis.

Objective:To investigate the differences in programmed death-ligand 1 (PD-L1) expression and immune cell infiltration characteristics in different molecular subtypes of endometrial cancer.Methods:A retrospective case series study was conducted. Ninety primary treated EC patients who underwent surgery without preoperative neoadjuvant therapy at the Second Hospital of Tianjin Medical University from November 2016 to May 2022 were collected. The surgical paraffin-embedded tissues were selected, and the molecular subtypes of endometrial cancer were classified according to 2020 World Health Organization (WHO) molecular subtypes using POLE gene Sanger sequencing and immunohistochemical staining. The expression of PD-L1, CD3, CD4, CD8, CD68, and CD20 proteins were detected by immunohistochemistry. Stained slides were digitally scanned for quantitative analysis of PD-L1 and immune cell infiltration density. The PD-L1-related scores were evaluated, including tumor cell score (TCS, the percentage of PD-L1 positive tumor cells among total tumor cells ≥1% was TCS positive, <1% was TCS negative), immune cell score (ICS, the percentage of PD-L1 positive tumor-associated lymphocytes and macrophages among total tumor-associated lymphocytes and macrophages ≥1% was ICS positive, <1% was ICS negative) and combined positive score [CPS, PD-L1 positive stained cells (including tumor cells, lymphocytes and macrophages)/total number of viable tumor cells ×100 ≥ 1 was CPS positive, < 1 was CPS negative]. Clinicopathological characteristics, PD-L1 scores and immune cell infiltration densities among different molecular subtypes were analyzed. Kaplan-Meier method was used to plot disease-free survival (DFS) curves for molecular subtypes, PD-L1 scores and immune cell infiltration densities, with subgroup comparisons using log-rank test. Cox proportional hazards models were used for univariate and multivariate analyses of poor DFS in endometrial cancer patients.Results:The median age of 90 patients was 58 years old (range: 33-72 years old); endometrioid carcinoma was present in 78 cases (86.7%), and non-endometrioid carcinoma was present in 12 cases (13.3%). Molecular subtyping identified POLE-mutated subtype in 6 cases (6.7%), mismatch repair deficient (MMRd) subtype in 23 cases (25.6%), p53 abnormal subtype in 14 cases (15.6%), and non-specific molecular profile (NSMP) subtype in 47 cases (52.2%). Significant differences were observed among the 4 molecular subtypes in International Federation of Gynecology and Obstetrics (FIGO) stage, histological grade, morphological subtype, tertiary lymphoid structures, estrogen receptor expression, and progesterone receptor expression (all P < 0.05). Among the 90 cases, 18 cases (20.0%) were positive for TCS, 31 cases (34.4%) were positive for ICS, and 39 cases (43.3%) were positive for CPS. Significant differences were found among the 4 molecular subtypes in PD-L1 + cell density, distribution of patients with ICS positivity, and distribution of patients with CPS positivity (all P < 0.01), but not in distribution of patients with TCS positivity ( P = 0.090); compared to NSMP subtype, the proportions of ICS-positive patients in POLE-mutated and MMRd subtypes were higher, the proportion of CPS-positive patients and PD-L1 + cell density in MMRd and p53 abnormal subtypes were higher, and the differences were statistically significant (all P < 0.05). Significant differences in immune cell densities were observed among the 4 molecular subtypes (all P < 0.01); compared to NSMP subtype, POLE-mutated, MMRd and p53 abnormal subtypes had higher densities of CD3 + and CD8 + cells, MMRd subtype had higher CD4 + cell density, and POLE-mutated and MMRd subtypes had higher CD68 + and CD20 + cell densities (all P < 0.05). The median follow-up was 43 months (range: 7-75 months). Among the molecular subtypes, p53 abnormal patients had the worst DFS, and POLE-mutated patients had the best DFS, and the difference in DFS among the 4 subtypes was statistically significant ( P = 0.046). Grouping according to the median density of immune cells in the entire group, patients with high CD8 + cell density (45 cases) had better DFS than those with low density (45 cases) ( P = 0.010), PD-L1 ICS-positive patients had worse DFS than negative patients ( P = 0.019), and NSMP subtype patients with high CD4 + cell density (24 cases) had better DFS than those with low density (23 cases) ( P < 0.001). There was no statistically significant difference in DFS among patients grouping with other PD-L1 scoring modes and other immune cell infiltration density (all P > 0.05). Cox regression analysis indicated that high CD8 + cell density ( HR = 0.335, 95% CI: 0.113-0.990, P = 0.048) was an independent protective factor for poor DFS in endometrial cancer patients, and high CD4 + cell density was an independent protective factor for poor DFS in NSMP subtype patients ( HR = 0.035, 95% CI: 0.003-0.345, P = 0.004). Conclusions:There are significant differences in PD-L1 expression and immune cell infiltration density among the different molecular subtypes of endometrial cancer, which are correlated with the prognosis of patients, and may provide reference for the selection of immunotherapy strategies and prognosis judgment.

Objective:To investigate the immunomodulatory effect of mixed konjac glucomannan(MKGM)on cyclophosphamide-induced immunosuppressed mice.Methods:The immunosuppressed mice model was established by cyclophosphamide.After treatment with MKGM for 25 d,organ index,lymphocyte proliferation,macrophage function,NK cell killing,and cytokine secretion of mice were observed.Results:Immunomodulatory effect of MKGM was firstly enhanced and then declined.Compared to model group,there were significant differences in organ index,lymphocyte proliferation,macrophage function,NK cell killing,and hemolysin in the medium-dose MKGM group(P＜0.01).HE staining showed that the low-dose MKGM had the best effect on repairing spleen injury caused by cyclophosphamide.However,medium-and high-dose MKGM had relatively weak immunomodulatory effects.Conclusion:The appropriate dose of MKGM can play an immunomodulatory role in the cyclophosphamide-induced immunosuppressed mice.