Objective: To investigate the therapeutic effects of human umbilical cord mesenchymal stem cell derived exosomes (hucMSC-Exo) on acute myocardial infarction (AMI) and to elucidate the underlying mechanisms. Methods: Human umbilical cord mesenchymal stem cells (hucMSCs) were isolated and cultured. Exosomes were extracted from the culture media. AMI model was constructed by ligating the anterior descending branch of the coronary artery in mice. The mice were randomly divided into Sham, AMI, Exo and Fer-1 groups, and PBS, PBS, hucMSC-Exo and Ferrostatin-1 were injected into the marginal zone of the ischemic myocardium of each group, respectively. The levels of the antioxidants glutathione (GSH) content and malondialdehyde (MDA) content in mouse myocardial tissues measured by corresponding assay kits, and reactive oxygen species (ROS) levels detected by DHE probe were used to assess the severity of ferroptosis in mouse myocardial tissue; Cardiac ultrasound and Masson staining were used to evaluate the cardiac function and fibrosis level in mice; HL-1 cells were treated with DMSO, Erastin and hucMSC-Exo and subjected to RNA sequencing, and the potential mechanism of the therapeutic effect of hucMSC-Exo was evaluated by KEGG enrichment analysis and clustering heat map analysis. The viability of HL-1 cells after hucMSC-Exo or hucMSC-Exo+BI-6C9 co-treatment was detected by CCK-8 assay. BH3 interacting domain death agonist (Bid) expression levels in infarcted myocardial tissues of mice were detected by qRT-PCR. Results: Compared with the AMI group, the Exo and Fer-1 groups of mice showed a significant decrease in MDA and ROS levels in the infarcted region of the myocardium, an increase in GSH content, a reduction in myocardial fibrosis, and an improvement in cardiac function; among the mice in the Exo group, the treatment effect was more pronounced compared to the Fer-1 group on the 28th day after AMI. KEGG enrichment analysis showed that the reversed genes by hucMSC-Exo were mainly enriched in the p53 signaling pathway; clustering heatmap analysis of p53-related genes revealed that Bid gene expression was significantly higher in HL-1 cells in the Erastin group compared with the control group, and the level of Bid expression in HL-1 cells in the Exo group was significantly lower than that in the Erastin group. The therapeutic effect of hucMSC-Exo in HL-1 cells was not significantly increased after treatment with the Bid inhibitor BI-6C9. Bid mRNA level in the heart tissues of mice in the AMI group was substantially increased compared with that in the control group, and was significantly decreased in the Exo group compared with that in the AMI group. Conclusion: hucMSC-Exo inhibits Bid expression to mitigate ferroptosis after AMI, thereby alleviate myocardial injury.

Objective: To investigate the therapeutic effect and mechanism of Clitoria ternatea flower (CF) on nonalcoholic fatty liver disease (NAFLD) in mice induced by a highfat diet (HFD). Methods: Twentyfour healthy ICR mice were randomly divided into four groups: normal control group (NC group), NAFLD model group (NAFLD group), highdose CF group (CFH group, 200 mg/kg), and lowdose CF group (CFL group, 100 mg/kg), with 6 mice in each group. The NC group was fed a normal diet, while the NAFLD, CFH, and CFL groups were fed a 60% highfat diet to induce NAFLD. Additionally, the CFH and CFL groups received corresponding doses of CF by gavage, and the NC and NAFLD groups received an equal volume of normal saline (06 mL) by gavage daily for 12 weeks. Serum levels of hepatic enzymes, blood lipids, and oxidative stress factors were detected, combined with untargeted metabolomics analysis. Fecal microbiota composition was analyzed via 16S rRNA gene sequencing, and the contents of three major shortchain fatty acids (acetate, propionate, and butyrate) were determined. Meanwhile, hepatic inflammationrelated indicators were examined by quantitative realtime polymerase chain reaction (qRTPCR). Results: Compared with the NAFLD group, the CFL and CFH groups showed significant reductions in serum levels of alanine aminotransferase (ALT), triglyceride (TG), total cholesterol (TC), and lowdensity lipoprotein cholesterol (LDLC) (all P<005). In the liver, the mRNA expressions of proinflammatory cytokines IL1β and TNFα were decreased, while those of antiinflammatory cytokines IL4 and IL10 were increased (all P<001). Additionally, serum superoxide dismutase (SOD) activity was markedly enhanced, whereas malondialdehyde (MDA) content was significantly decreased (all P<001). 16S rRNA sequencing results indicated that compared with the NAFLD group, the CF groups had increased abundances of multiple beneficial gut bacteria including Prevotella and Lactobacillus, along with elevated concentrations of acetate, propionate, and butyrate in feces. Serum untargeted metabolomics analysis revealed that compared with the NAFLD group, the CF groups had higher levels of various antiinflammatory and antioxidative stress metabolites, which were mainly involved in glycerophospholipid metabolism, pyrimidine metabolism, and other pathways. Conclusion: CF can alleviate liver damage in NAFLD mice by improving lipid levels and hepatic inflammation, regulating gut microbiota balance and serum metabolic pathways, showing potential therapeutic effects for NAFLD.

Objective: To explore the regulatory role and mechanism of high mobility group box 1 (HMGB1) in the radiosensitivity of triple-negative breast cancer (TNBC) cells. Methods: The expression level of HMGB1 in breast cancer subtypes and its relationship with the survival prognosis of breast cancer patients were analyzed by using The Cancer Genome Atlas (TCGA). Breast epithelial cells MCF-10A, non-TNBC cells MCF-7, SK-BR-3, and TNBC cells MDA-MB-231, MDA-MB-436, MDA-MB-468 were cultured in vitro. The mRNA and protein expression levels of HMGB1 were detected by qRT-PCR and Western blotting. Cell proliferation was assessed at different time points (0, 24, 48, 72 and 96 h) after 4 Gy X-ray irradiation, and the radiosensitivity of the cells was evaluated after different doses (0, 2, 4, 6, 8 Gy) of irradiation. HMGB1 expression was knocked down or overexpressed in MDA-MB-231 cells using plasmids or lentivirus, respectively, and cell proliferation, clone formation ability and apoptosis level after 4 Gy X-ray irradiation were detected by CCK-8 assay, colony formation assay, and flow cytometry, respectively. Changes in nuclear and intracellular HMGB1 in MDA-MB-231 cells after 0, 4, and 8 Gy irradiation were detected by immunofluorescence assay. MDA-MB-231 cells were divided into shControl, shHMGB1-2, Vector, and Flag-HMGB1 groups, irradiated with 4 Gy for 72 h, and changes in nuclear, HMGB1, and γ-H2AX were detected by immunofluorescence assay. Protein expression of p-AKT, AKT, p-GSK-3β, GSK-3β, Caspase 9, p-H2AX, and HMGB1 were detected by Western blotting in MDA-MB-231 cells after 0, 4, 8 Gy irradiated, and in shControl, shHMGB1-2, Vector, Flag-HMGB1 groups cells after 4 Gy irradiation for 72 h. The proliferation inhibition rate was assessed by CCK-8 assay in control, radiotherapy, AKT inhibitor, and AKT inhibitor + radiotherapy groups. Cell proliferation activity was examined by CCK-8 assay, and protein expression of p-AKT/AKT, Caspase 9, and p-H2AX were detected by Western blotting in Vector, Flag-HMGB1, Vector+AKT inhibitor, and Flag-HMGB1+AKT inhibitor groups after 4 Gy irradiation. Results: HMGB1 was highly expressed in TNBC, and its high expression was associated with poor prognosis. The mRNA and protein expression levels of HMGB1 was higher in MDA-MB-231/436/468 cells than in MCF-7 and SK-BR-3 cells, with the former having lower radiosensitivity. Knockdown of HMGB1 significantly increased the radiosensitivity of MDA-MB-231 cells, while overexpression of HMGB1 significantly decreased it. Knockdown of HMGB1 resulted in a significant decrease in p-AKT expression levels and a significant increase in p-GSK-3β and p-H2AX expression levels post 4 Gy irradiation. Overexpression of HMGB1 resulted in a significant increase in p-AKT expression levels and a significant decrease in p-GSK-3β and pH2AX expression levels post 4 Gy irradiation. The resistance-promoting effect of HMGB1 on radiotherapy was abolished by AKT inhibitor AKTi-1/2 treatment. Conclusion: HMGB1 is highly expressed in TNBC cells and regulates the radio-sensitivity of cells through the PI3K/AKT pathway. Thus, it may be implicated that HMGB1 may potentially serve as an important molecular target for radio-resistance in TNBC.

Objective: To investigate the therapeutic effects and underlying mechanisms of human umbilical cord mesenchymal stem cell-derived small extracellular vesicles (HucMSC-sEVs) on renal injury in diabetic kidney disease (DKD) rats. Methods: HucMSCs were isolated, cultured, and used to extract HucMSC-sEVs from their culture supernatant. A DKD rat model was induced by a high-fat diet combined with STZ injection. After successful modeling validation, rats were randomized into the DKD group and the HucMSC-sEVs group, with normal rats as controls. At 8 weeks, HucMSC-sEVs were injected via the tail vein. After 24 weeks, kidney tissues were collected. Renal histopathology was assessed by HE staining, while Masson staining was employed to evaluate collagen deposition. The expression levels of necroptosis markers (p-RIPK1/RIPK1, p-RIPK3/RIPK3, and p-MLKL/MLKL) were quantified using qRT-PCR, Western blotting, and immunohistochemical staining. NRK-52E cells were treated with high glucose and HucMSC-sEVs. Transmission electron microscopy was used to observe necrotic phenomena in NRK-52E cells, and the expression levels of necroptosis marker proteins such as p-RIPK1/RIPK1, p-RIPK3/RIPK3, and p-MLKL/MLKL were detected by Western blotting and qRT-PCR. Results: Compared with the control group, the glomerular basement membrane in the DKD group rats was significantly thickened, and the interstitium exhibited fibrotic characterization, with a marked increase in the expression of necroptosis marker proteins. After treatment with HucMSC-sEVs, the pathological damage and fibrosis degree of the rat kidney tissues were alleviated, and the expression of necroptosis execution proteins was reduced. In vitro experiments revealed that high glucose-treated NRK-52E cells exhibited necrotic features including mitochondrial cristae fragmentation, endoplasmic reticulum dilation, and elevated expression of necroptosis markers (p-RIPK1/RIPK1, p-RIPK3/RIPK3, and p-MLKL/MLKL). Treatment with HucMSC-sEVs significantly attenuated these cellular alterations and reduced the expression levels of necroptosis-related proteins. Conclusion: HucMSC-sEVs can improve renal function and delay the progression of DKD in rats, and the mechanism may be attributed to the inhibition of necroptosis of renal tubular epithelial cells.

Objective: To explore the disrupting effects of low-dose methyl-2-benzimidazole carbamate (CBZ) on glycolipid metabolism in mice during pubertal development period and its mechanism. Methods: A developmental (4-week-old) and adult (8-week-old) ICR mouse model with low-dose CBZ exposure was established. The mice were randomly divided into a blank control group (administered with corn oil) and different dose CBZ exposure groups (0.3, 3, 30 and 60 mg/kg ), and were given intragastric administration for 28 days. Commercial kits were used to detect the serum levels of total cholesterol (TC), total triglyceride (TG), low density lipoprotein cholesterol (LDLC), high density lipoprotein cholesterol (HDLC), creatinine (CRE), blood urea nitrogen (BUN), aspartate aminotransferase (AST), alanine aminotransferase (ALT), as well as the activity of glucose-6-phosphate dehydrogenase (G6PD) in the liver. Fasting blood glucose (FBG) was measured by a blood glucose meter. Body weight was monitored by weighing. The expression levels of genes related to glycolipid metabolism and endoplasmic reticulum stress pathway were detected by quantitative realtime PCR. Results: Compared with the blank control group, CBZ exposure significantly increased the serum lipid levels and various blood biochemical indexes in developing and adult mice (P<0.05 or P<0.01), and only significantly increased the FBG level and decreased the body weightof adult mice (P<0.05). CBZ exposure upregulated the expression of lipid synthesis and transport genes such as CD36 and SREBP-1, down-regulated the expression of lipid metabolism genes such as PPARα and HNF4α, as well as gluconeogenesis and glycogen synthesis genes such as G6PC, PEPCK and GSK3β in developing and adult mice(P<0.05). Meanwhile, CBZ up-regulated the expression of GRP78, PERK, CHOP, Bax, Caspase3 and other genes (P<0.05), down-regulated the expression of ATF4 and Bcl-2 genes (P<0.05), and activated the endoplasmic reticulum stress and apoptotic pathways. Conclusion: CBZ exerts endocrine and metabolic toxicity to developing mice, which can interfere with glycolipid metabolism and cause metabolic dysfunction by affecting the expression of glycolipid metabolism-related genes and activating the endoplasmic reticulum stress signaling pathway.

Objective: To explore the effects of recombinant avirulent Newcastle disease virus La Sota strain expressing rabies virus glycoprotein (rL-RVG) on the polarization of murine macrophage RAW264.7 cells and to analyze the effect of polarized RAW264.7 cells on the viability of Lewis lung cancer cells. Methods: Murine macrophage RAW264.7 cells were divided into four groups: blank control group and rL-RVG groups at different multiplicities of infection(0.5, 1, 5 MOI); and the viability of RAW264.7 cells was assessed by using the CCK-8 assay. RAW264.7 cells were divided into three groups: RAW264.7-M0 type (complete media), RAW264.7-M1 type (induced by 1 μg/mL lipopolysaccharide +50 ng/mL IFN-γ), and RAW264.7-M2 type (induced by 20 ng/mL IL-4); and the morphological characteristics of these phenotypes were observed under phase-contrast microscopy. The protein expression levels of CD86 and Arg-1 were measured by immunofluorescence in four groups: Raw264.7-M1, Raw264.7-M2, rL-RVG+Raw264.7-M0, and rL-RVG+Raw264.7-M2. The mRNA expression levels of cytokines IL-4, IL-6, IL-10 and tumor necrosis factor-α (TNF-α) in the blank control group and rL-RVG groups were also assessed by qRT-PCR. Lewis lung cancer cells were co-cultured with conditioned media from the above groups and divided into four groups: blank control group and rL-RVG groups at different MOIs (0.5, 1, 5 MOI). The viability of Lewis lung cancer cells was evaluated by using the CCK-8 assay. Results: Compared with the blank control group, the cell proliferation rates in the 1, 5 MOI rL-RVG groups were significantly increased (both P<0.01). The morphological characteristics of Raw264.7-M0, M1 and M2 macrophages were round or oval, flat or irregular, and fibrous, elongated or spindle-shaped, respectively. Compared with RAW264.7-M0 type macrophages, the relative expression level of CD86 mRNA in RAW264.7-M1 type was significantly increased (P<0.01), and the relative expression level of Arg-1 mRNA in RAW264.7-M2 type was greatly increased (P<0.01). Compared with Raw264.7-M1 type, the relative fluorescence expression level of CD86 in rL-RVG+Raw264.7-M0 type was significantly increased (P<0.05); compared with Raw264.7-M2 type, the relative fluorescence expression level of CD86 in rL-RVG+Raw264.7-M2 type was significantly increased (P<0.05), and the relative fluorescence expression level of Arg-1 was significantly decreased (P<0.05). rL-RVG treatment led to an increase in the mRNA expressions of TNF-α and IL-6, while the mRNA expressions of IL-4 and IL-10 decreased. Compared with the blank control group, the viability of Lewis lung cancer cells in the 0.5, 1, and 5 MOI rL-RVG groups were significantly decreased (all P<0.01). Conclusion: rL-RVG could induce the polarization of RAW264.7 macrophages towards the M1 phenotype and subsequently inhibit the viability of Lewis lung cancer cells at MOIs ranging from 0.5 to 5.

Objective: To explore the effect of crocin on the migration and invasion ability of anaplastic thyroid carcinoma (ATC) cells and its molecular mechanism of regulating epithelial-mesenchymal transition (EMT) via the Smad-dependent signaling pathway. Methods: ATC cells were treated with different concentrations of crocin. Cell proliferation and apoptosis were evaluated by CCK-8 assay and flow cytometry, respectively. The effects of crocin on ATC cell migration and invasion were detected by Transwell assays. Western blotting analysis was used to examine the expression of EMT markers (E-cadherin, N-cadherin, vimentin, fibronectin) and Smad signaling pathway-related proteins. A subcutaneous tumor model was established using BHT-101 cells to evaluate the inhibitory effect of crocin on tumor invasiveness. Immunohistochemistry and Western blotting were performed to detect the expression changes of MMP-2, MMP-9, N-cadherin and vimentin in tumor tissues. Results: At concentrations below 40 μmol/L, crocin treatment significantly reduced cell invasion and migration compared with the untreated group (P<0.01), accompanied by increased E-cadherin expression and decreased N-cadherin, vimentin and fibronectin expression. The inhibitory effect of crocin on Smad2/3 phosphorylation was alleviated after exogenous TGF-β intervention. In vivo experiments showed that crocin notably downregulated the expression of MMP-2, MMP-9, N-cadherin and vimentin in BHT-101 subcutaneous xenograft tumors. Conclusion: Crocin can inhibit the invasion and EMT of ATC cells by modulating the Smad-dependent signaling pathway.

Objective: To investigate the correlation between the characteristic parameters of brown adipose tissue (BAT) and coronary artery calcification in 18F-FDG PET/CT images. Methods: A total of 2 399 patients who underwent 18F-FDG PET/CT examination at the Affiliated Hospital of Jiangsu University from December 2013 to April 2024 were collected. After 1∶1 propensity score matching (PSM), the patients were divided into BAT positive group (n=171) and BAT negative group (n=171) according to whether BAT was positive. Differences in general clinical data and the incidence of coronary artery calcification between the two groups were analyzed. Differences in BAT imaging characteristic parameters among six anatomical sites (the neck, supraclavicular fossa, axilla, mediastinum, paravertebral region, and abdomen) in patients of the BAT-positive group were analyzed, and the site with the highest BAT activity (the paravertebral region) was screened out. Furthermore, differences in paravertebral BAT imaging characteristic parameters among groups with different numbers of coronary artery calcification-involved branches and different severities of coronary artery calcification were analyzed. Results: After PSM, the incidence of coronary artery calcification and the coronary artery calcium score in the BAT positive group were significantly lower than those in the BAT negative group. In the BAT positive group, the SUVmax of paravertebral BAT was the highest among the six anatomical sites, and the difference was statistically significant compared with the SUVmax of abdominal and axillary BAT (all P<0.001). The SUVmean of paravertebral BAT significantly differed from that of axillary BAT (P<0.001), but showed no significant differences with other anatomical sites (all P>0.05). Both the volume and surface area of paravertebral BAT demonstrated statistically significant differences compared to abdominal and axillary BAT (all P<0.001). As both the number of coronary artery branches involved and the severity of calcification increased, all characteristic parameters of paravertebral BAT progressively decreased (all P<0.01). Conclusion: The presence of BAT is associated with a lower risk of coronary artery calcification. Paravertebral BAT may have a potential inhibitory effect on the occurrence and development of coronary artery calcification.

Objective: To investigate the median effective dose (ED50) and 95% effective dose (ED95) of ciprofol for combined procedural sedation during spinal anesthesia in patients with different ages. Methods: A total of 67 patients scheduled for lower limb orthopedic surgery under subarachnoid block from December 2023 to March 2024 were enrolled and divided into three groups by age: the youth group (18-40 years old, 22 cases), the middleaged group (41-64 years old, 24 cases), and the elderly group (65-80 years old, 21 cases). After completion of spinal anesthesia, the Dixon up-and-down sequential method was used to administer ciprofol for procedural sedation, with an initial dose of 0.2 mg/kg and a dose gradient of 0.05 mg/kg between consecutive patients. Two minutes after administration, a modified observer′s assessment of alertness/sedation scale (MOAA/S) score ≤3 and a bispectral index (BIS) <85 were defined as satisfactory sedation. The dose for subsequent patients was adjusted, and the study was terminated after MOAA/S score ≥7 cross inflection points appeared. The Probit regression analysis was used to calculate ED50 and ED95 in each group. Vital signs, MOAA/S scores, BIS values, and the incidence of adverse reactions were recorded. Results: The ED50 and ED95 of the youth group were 0.263 mg/kg (95%CI: 0.232-0.300 mg/kg) and 0.318 mg/kg (95%CI: 0.288-0.509 mg/kg), respectively; those of the middle-aged group were 0.208 mg/kg (95%CI: 0.178-0.238 mg/kg) and 0.264 mg/kg (95%CI: 0.235-0.417 mg/kg); those of the elderly group were 0.178 mg/kg (95%CI: 0.130-0.217 mg/kg) and 0244 mg/kg (95%CI: 0.209-0.526 mg/kg). There were no statistically significant differences in vital signs, MOAA/S scores, BIS values, or the incidence of adverse reactions among the three groups during procedural sedation (all P>0.05). Conclusion: The effective doses of ciprofol for combined procedural sedation during spinal anesthesia depend on patients, age groups, and ED50 and ED95 decrease with increasing age.

Objective: To investigate the risk factors for diuretic resistance in patients with acute exacerbation of chronic heart failure and to construct a Nomogram model, aiming to provide a reference for the early identification of patients with diuretic resistance. Methods: A retrospective study was designed by including 215 patients with acute exacerbation of chronic heart failure who were admitted to the Affiliated People′s Hospital of Jiangsu University from January 2021 to August 2024. Patients were grouped into two groups, one is resistance group, the other non-resistance based on the criteria for diagnosing diuretic resistance, and baseline data were collected for both groups. Using the least absolute shrinkage and selection operator (LASSO) regression for univariate screening, followed by multifactorial Logistic regression for further screening, a nomogram model was constructed and subjected to internal validation. Results: LASSO regression selected 6 variables, and further analysis using multiple Logistic regression revealed that an elevated Nterminal pro B-type natriuretic peptide (NT-proBNP) level was an independent risk factor for diuretic resistance ［OR (95%CI): 2.342 (1.087-5.043), P<0.05］, while higher levels of estimated glomerular filtration rate (eGFR), hemoglobin, serum sodium, and left ventricular ejection fraction (LVEF) were identified as protective factors ［OR (95%CI): 0.978 (0.965-0.990), 0.973 (0.958-0.989), 0.893 (0.822-0.971), 0.944 (0.918-0.971), all P<0.05］. A nomogram model was constructed based on the above risk factors. The area under the receiver operating characteristic (ROC) curve was 0.846 (95%CI: 0.795-0.897). The calibration curve and Hosmer-Lemeshow test indicated that the model fit well (P=0.159). The clinical decision curve analysis demonstrated that the model provided a favorable net clinical benefit when the threshold probability ranged from 0% to 87.0%. Conclusion: The Nomogram model, incorporating NT-proBNP, hemoglobin, blood sodium, eGFR, and LVEF, effectively assesses the risk of diuretic resistance in patients with acute exacerbation of chronic heart failure.

Objective: To investigate the effects of extracellular matrix (ECM) protein SVEP1 on lipolysis and thermogenesis in adipocytes. Methods: The RNA-seq database of abdominal subcutaneous adipose tissue (SAT) of 236 normal and obese individuals constructed previously was used to analyze the correlation between SVEP1 expression level and body mass index (BMI), fat mass, fat percentage and waist circumference. 3T3-L1 preadipocytes and human stromal vascular fraction (SVF) cells were differentiated into mature adipocytes respectively. The expression of Svep1 in adipocytes stimulated by rosiglitazone and Forskolin was detected by realtime quantitative PCR (qRT-PCR). Based on the Pearson correlation coefficient (r>0.4, P<0.05) between RPKM values of SVEP1 and other genes in the RNA-seq database of human SAT, co-expressed gene set was screened and KEGG enrichment analysis was performed. Svep1 was knocked down in mouse primary adipocytes by lentivirus-mediated shRNA. qRT-PCR and Western blotting were used to assess the expression of thermogenic genes and proteins in adipocytes. Glycerol detection kit was used to detect the level of lipolysis stimulated by isoproterenol (ISO), and Western blotting was used to detect the phosphorylation level of lipolysis-related proteins. Results: The expression level of SVEP1 in human abdominal SAT was positively correlated with obesity related indicators (BMI, fat mass, fat percentage and waist circumference). Cold stimulation could down-regulate Svep1 transcription level in brown and beige adipocytes in mice. The transcription level of Svep1 was down-regulated in mature 3T3-L1 adipocytes and human adipocytes when stimulated by Rosi or Forskolin. KEGG enrichment analysis showed that Svep1 co-expressed gene set was enriched in the regulation of lipolysis and insulin resistance related pathways. Svep1 knockdown could significantly promote ISO-stimulated lipolysis in mouse white adipocytes and increase the expression of thermogenic genes (Ucp1, Dio2, Pgc1α, Elovl3 and Cidea) in mouse beige adipocytes. Conclusion: This study demonstrated an association between Svep1 expression and adipocyte function, and it may be implicated that Svep1 might be involved in obesity-related adipose tissue dysfunction and contribute to the development and progression of obesity.

Objective: To prepare shikonin liposomes (SHK-L) and preliminarily evaluate their anti-aging effects. Methods: The SHK-L prescription was optimized through single factor experiments to investigate its in vitro release and oral bioavailability in rats. The in vivo safety was evaluated through zebrafish toxicity experiments, and the anti-aging effect was examined by staining the β-galactosidase in aging zebrafish. Results: The optimal prescription was SHK 6 mg, lecithin 70 mg, cholesterol 10 mg, vitamin E polyethylene glycol succinate (TPGS) 30 mg, and sodium dodecyl sulphate (SDS) 1.5 mg, with a particle size of (206.96±0.75) nm, polydispersity coefficient (0.186±0.030), encapsulation rate of (96.88±2.19)% and drug loading was (5.36±0.01)%. SHK-L improved the drug release rate compared to SHK. According to in vivo pharmacokinetic data, SHK-L had a relative bioavailability of 413.37% of SHK. At 96 hours post-administration, LC₅₀ of shikonin and SHK-L in zebrafish were (0.112±0.007) μg/mL and (0.141±0.012) μg/mL, respectively. Conclusion: SHK-L can enhance the solubility and oral bioavailability of insoluble drugs, as well as their anti-aging effect and safety.

Objective: To investigate the association between the polymorphisms of meteorinlike protein (METRNL) gene rs4986080 and type 2 diabetes mellitus (T2DM). Methods: A total of 467 subjects were selected, including 315 T2DM patients and 152 individuals with normal glucose tolerance (NGT). The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was used to detect the polymorphisms of METRNL gene rs4986080 in all subjects, then analyzed the association between these polymorphisms and T2DM. Results: The constituent ratio of GG genotype and the frequency of G allele at rs4986080 locus of METRNL gene was higher in the T2DM group compared to the NGT group (P<0.05). Compared to individuals with GG genotype at the METRNL gene rs4986080 locus, those with AA+AG genotypes showed significant lower waisttohip ratio, fasting plasma glucose (FPG), 2hour postprandial glucose (2hPG), glycated hemoglobin (HbA1c), and homeostasis model assessment of insulin resistance (HOMA-IR) (P<0.05). Logistic regression analysis showed that A allele at rs4986080 locus of METRNL gene was probably a protective factor for T2DM. Conclusion: The polymorphisms of METRNL gene rs4986080 may be associated with the occurrence of T2DM, and A allele may be a protective factor for T2DM.

Objective: To explore the feasibility of preoperative prediction of perineural invasion (PNI) and lymphovascular invasion (LVI) in advanced gastric cancer using dual-energy computed tomography (DECT) venous phase imaging features and spectral parameters of a hypotonic water-filled stomach, along with clinical laboratory information. Methods: A retrospective analysis was conducted on 161 cases of advanced gastric cancer that underwent DECT imaging within one week before surgery, and the cases were randomly divided into training sets and test sets in a 7∶3 ratio. Based on postoperative pathological assessment, 115 cases demonstrated LVI and/or PNI positivity, whereas 46 cases were negative for both. A predictive model for LVI/PNI was developed using venous phase imaging of a hypotonic water-filled stomach, DECT parameters ［including the slope of the spectral Hounsfeld unit curve (between 40 keV and 100 keV), normalized iodine concentration (NIC), and effective atomic number］, and clinical laboratory data (inflammatory and tumor markers). The predictive performance of the model was evaluated using the area under the ROC curve (AUC), and its clinical utility was assessed using decision curve analysis. Results: The AUC values of the radiomics model (Rad-score) in the training sets and test sets were 0.776 (95%CI: 0.653-0.821) and 0.781（95%CI：0.582~0.847）, respectively. The independent predictors for the DECT parametric model was NIC, with AUC values of 0.729（95%CI：0.615~0.790 in the training sets and 0.771（95%CI：0.604~0.864）in the test sets. For the clinical information predictive model, the independent predictor was lymphocyte percentage, with AUC values of 0.693 (95%CI: 0.638-0.805) in the training sets and 0.502 (95%CI: 0.352-0.648) in the test sets. The combined model integrating the Rad-score, DECT parameters, and clinical information had independent predictors including Rad-score, NIC, and lymphocyte percentage. The AUC values for this combined model were 0.880 (95%CI: 0.701-0.859) in the training sets and 0830 (95%CI: 0.602-0.857) in the test sets, demonstrating superior performance compared to the radiomics model, DECT parametric model, and clinical model. The DeLong test showed that the AUC of the combined model was significantly higher than that of the radiomics model, DECT parametric model, and clinical information model in the training sets (Z=1.979, P=0.048; Z=3.199, P=0.001; Z=3.053, P=0.001). In the test sets, the AUC of the combined model was also significantly higher than that of the clinical information model (Z=2.417, P=0.015). Decision curve analysis revealed that when the risk threshold ranges from 0.15 to 0.96, adopting the combined model for treatment guidance yielded a higher clinical net benefit rate. Conclusion: The integrated model, incorporating radiomics, NIC, and lymphocyte percentage, serves as a comprehensive predictive model for assessing lymphovascular invasion and perineural invasion status in advanced gastric cancer.

Objective: To investigate the role of the long noncoding RNA (lncRNA) NONRATT021203.2 in the progression of cancer-induced bone pain (CIBP) and its potential molecular mechanisms. Methods: A CIBP model was constructed in Sprague-Dawley (SD) rats by intratibial inoculation of Walker 256 breast cancer cells. Sixteen adult female SD mice weighing 180-200 g were randomly divided into control group and CIBP group, with 8 rats in each group, and injected into tibiae with 10 μL normal saline and 10 μL Walker 256 breast cancer cell suspension (1×10⁸ cells/mL), respectively. Paw withdrawal threshold (PWT) and paw withdrawal latency (PWL) of the modeled lower limb of rats were detected by behavioral analysis. The expression levels of lncRNA NONRATT021203.2 and miRNA-138-5p in DRG were measured by using quantitative real-time PCR (qRT-PCR), and the localization of miRNA-138-5p in DRG tissue was examined by fluorescence in situ hybridization (FISH). Seven days after modeling, 10 rats with CIBP were selected and divided into CIBP+siRNA group (n=5) and CIBP+siNC group (n=5), lncRNA NONRATT021203.2-siRNA and siNC were injected intrathecally, respectively. The relative expression levels of lncRNA NONRATT021203.2 and miRNA-138-5p in the two groups were detected by qRT-PCR. Additionally, seven days after modeling, 16 rats with CIBP were divided into CIBP+mimics (n=8) and CIBP+NC group (n=8), miRNA-138-5p mimics and negative control oligonucleotide were injected intrathecally, respectively. PWT and PWL were detected by behavioral test. Finally, the binding sites of lncRNA NONRATT021203.2 and miRNA-138-5p were analysed by using miRand and miRwalk3.0 software. Results: Compared with control group, CIBP group showed significantly reduced PWT and PWL (P<0.05), markedly increased expression of lncRNA NONRATT021203.2 (P<0.01), and significantly decreased expression of miRNA-138-5p (P<0.001). FISH results showed that miRNA-138-5p was localized in neuronal cytoplasm. Compared with CIBP+siNC group, CIBP+siRNA group exhibited significantly reduced lncRNA NONRATT021203.2 expression (P<0.05) and increased miRNA-138-5p expression (P<0.05). Compared with CIBP+NC group, PWT was significantly increased in CIBP+miRNA-138-5p mimics group (P<0.01). Bioinformatics analysis revealed the presence of a potential binding site between lncRNA NONRATT021203.2 and miRNA-138-5p. Conclusion: LncRNA NONRATT021203.2 could reduce the intracellular free miRNA-138-5p level by competitively binding to miRNA-138-5p, thereby promoting the occurrence and development of bone cancer pain.

Objective: To develop a wound dressing incorporating Clostridium butyricum (C. butyricum, anaerobic probiotic), which is capable of producing antibacterial substances and exerting hydrogen-producing properties under normoxic conditions, and investigate the effects of its selective bacteria inhibitionand reactive oxygen species (ROS) scavenging. Methods: C. butyricum and Bacillus subtilis (B. subtilis, aerobic probiotic) were simultaneously encapsulated in hydrogel to prepare a dual-probiotic hydrogel (BC-gel). B. subtilis was used to consume oxygen within the hydrogel. This creates an anaerobic microenvironment for C. butyricum to maintain its viability. The selective bacteria inhibition of BC-gel was evaluated by plate counting method, while the selective scavenging of ROS was evaluated through flow cytometry. Results: BC-gel can effectively inhibit the proliferation of Escherichia coli and methicillin-resistant Staphylococcus aureus. While maintaining the normal growth of Bifidobacterium and Lactobacillus, BC-gel has the ability to scavenge ·OH and ONOO- free radicals, but has no significant decreasing effect on O-2· and NO·. Conclusion: BC-gel exhibits favorable selective bacteria inhibition and ROS scavenging properties.

目的： 探讨临床诊断活动性肺结核的影响因素，建立并验证活动性肺结核临床诊断模型。方法： 选择2020年1月至2023年12月于柳州市人民医院感染性疾病科住院患者958例，其中活动性肺结核658例，根据病原学结果将402例病原学阳性肺结核纳入建模组，256例病原学阴性肺结核纳入验模组；社区获得性肺炎300例，根据2∶1随机原则将200例纳入建模组，100例纳入验模组。对可能与活动性肺结核诊断相关的自变量，包括性别、年龄、体重指数(BMI)、个人史评分、临床表现评分、影像学指标评分、免疫学指标评分、血清腺苷脱氨酶和红细胞沉降率(ESR)等进行单因素分析；将有意义的变量(P＜005)作为自变量，以活动性肺结核作为结局指标进行多因素Logistic回归分析；建立活动性肺结核的临床诊断模型，用ROC曲线评价模型的诊断预测效能；最后利用验模组数据进一步验证该模型的诊断效能。结果： 单因素分析结果显示，性别、年龄、BMI、临床表现评分、影像学指标评分、免疫学指标评分、腺苷脱氨酶和ESR等8个变量差异有统计学意义(P＜0.05)，个人史评分差异无统计学意义(P＞0.05)；多因素分析结果显示，BMI、临床表现评分、影像学指标评分和免疫学指标评分是诊断活动性肺结核的独立影响因素(P<0.05)；活动性肺结核临床诊断模型=BMI×10＋临床表现评分×14＋免疫学指标评分×27＋影像学指标评分×10。ROC曲线结果显示，建模组和验模组AUC分别为0.922(95％CI：0.868～0.976)和0.766(95%CI 0.723～0.830)。结论： BMI、临床表现评分、影像学指标评分和免疫学指标评分是临床诊断活动性肺结核的独立影响因素，基于此构建的临床诊断模型对病原学阴性肺结核活动性的判断有一定的预测价值。

Objective: To construct an end-to-end deep learning model for the multi-class classification task of T staging in advanced gastric cancer using CT images. Methods: This retrospective study included enhanced venous phase images from 423 gastric cancer patients, which were randomly divided into the training set and the test set at an 8∶2 ratio. We employed a deep learning automatic segmentation model based on the 3D-nnUNet for segmenting tumors. Simultaneously, a multi-class classification model based on the SmallFocusNet was developed for classification of T staging of advanced gastric cancer. Finally, these models were integrated to construct an end-to-end deep learning model for CT-T staging diagnosis of advanced gastric cancer. The performance of the segmentation model was evaluated using the Dice similarity coefficient (DSC), Intersection over Union (IoU), and 95% Hausdorff Distance (HD_95). The prediction efficacy of the deep learning model was assessed using area under the ROC curve (AUC) values, sensitivity, and specificity. Additionally, the diagnostic performance of the deep learning model for classification of T staging of advanced gastric cancer was compared with that of radiologists. Results: In the test set, the DSC and IoU of the automatic segmentation model were 0.869±0.095 and 0.779±0.137, respectively. The macro-average AUC value of the deep learning model was 0.882 (95%CI: 0.812-0.926). The AUC values for distinguishing T2, T3 and T4a stage tumors were 0.960 (95%CI: 0.915-0.990), 0.739 (95%CI: 0.616-0.849) and 0.917 (95%CI: 0.812-0.926), respectively. The average sensitivity was 0.769 (95%CI: 0.676-0.853), with sensitivities for distinguishing T2, T3 and T4a stage tumors of 0.808 (95%CI: 0.654-0.923), 0.750 (95%CI: 0.571-0.893) and 0.750 (95%CI: 0.594-0.906), respectively. Furthermore, the deep learning model outperformed radiologists in the diagnostic performance of T staging for advanced gastric cancer. Conclusion: The end-to-end deep learning model, which integrates multi-channel and attention mechanisms based on enhanced CT images, demonstrates high accuracy and consistency in preoperative T staging diagnosis of advanced gastric cancer.

Objective: To investigate the expression levels of Rho GTPase-activating protein 12 (ARHGAP12) within tertiary lymphoid structures (TLS) in gastric cancer and its impact on tumor immune response. Methods: ARHGAP12 mRNA expression differences between gastric cancer (n=414) and normal tissues (n=211) were analyzed using TCGA databases. Western blotting was employed to validate the protein expression levels of ARHGAP12 in 12 pairs of gastric cancer and adjacent tissue samples. Immunofluorescence was used to detect ARHGAP12 protein expression in TLS and tumor-draining lymph nodes (TDLN) in gastric tissue pathological sections. Hallmark and KEGG pathway enrichment analyses were conducted to assess the correlation between ARHGAP12 mRNA expression and the activation of immune-related pathways. Immunohistochemical staining was performed to examine the protein expression levels of ARHGAP12 in pathological sections of tumor tissues from 28 gastric cancer patients, and clinical data were analyzed to explore the correlation between cytokine expression levels and ARHGAP12 immunohistochemical staining scores. KM plotter database was used to analyze the relationship between ARHGAP12 mRNA expression and prognosis of patients with gastric cancer. Results: TCGA database analysis showed that ARHGAP12 mRNA expression in gastric cancer tissues was significantly higher than that in adjacent tissues (P<0.001). Western blotting results indicated that there was no significant difference in ARHGAP12 protein expression levels between the gastric adenocarcinoma and adjacent tissue samples (P>0.05). Multiplex immunofluorescence results demonstrated higher expression of ARHGAP12 protein in gastric cancer cells, TLS, and TDLN regions (P<0.05). Hallmark and KEGG pathway enrichment analysis suggested that ARHGAP12 mRNA expression is involved in the activation of interferon response pathways and antigen presentation processes. Furthermore, high ARHGAP12 immunohistochemical staining score was positively correlated with preoperative serum IFN-γ levels in patients with gastric cancer (P<0.05). KM plotter database analysis showed that high expression of ARHGAP12 mRNA in gastric cancer tissues is positively correlated with longer overall survival period (P<0.05). Conclusion: ARHGAP12 is highly expressed in gastric cancer TLS, and it is positively correlated with the activation of interferon-γ-related anti-tumor immune pathways.

Objective: To investigate the changes in gene expression in pancreatic cancer during radiotherapy combined with immunotherapy and to screen and validate key genes involved in mice. Methods: Twenty C57BL/6J male mice aged 6-8 weeks were randomly divided into 4 groups: control group, radiotherapy group, immunotherapy group and radiotherapy combined with immunotherapy group, with 5 mice in each group. The tumor-bearing mouse model was established by subcutaneous injection of pancreatic cancer KPC cells, and the radiotherapy group was irradiated with 8 Gy radiation, the immunotherapy group received a single intraperitoneal injection of PD-1 monoclonal antibody (200 μg/mouse), the radiotherapy combined with immunotherapy group received a single dose of 8 Gy radiation followed by intraperitoneal injection of PD-1 monoclonal antibody (200 μg/mouse), and the control group did not receive any treatment. The tumor tissues were harvested 7 days later, and the transcriptome sequencing was performed. The differentially expressed genes (DEGs) with significant expression were analyzed by GO, KEGG and Reactome functional annotation to explore the related signaling pathways. Venn analysis was performed to identify the intersection of DEGs, and survival analysis was used to identify key genes. The expression levels of these key genes were analyzed, and their correlation with immune cell infiltration and immune checkpoint expression was examined. A proteinprotein interaction (PPI) network was constructed to analyze the interactions of these key genes. Two subcutaneous tumorbearing models of male mice with pancreatic cancer KPC and PANC02 cell lines C57BL/6J were constructed. The combined radiotherapy and immunotherapy treatment was administered on the following day and maintained for 3 days. Quantitative reverse transcription PCR (qRT-PCR) was used to assess the relative mRNA expression of key genes in tumor tissues. Results: The results of transcriptome sequencing analysis showed that compared with the radiotherapy group and the immunotherapy group, the radiotherapy combined with immunotherapy group had the richest DEGs and mainly participated in the signal pathways related to metabolism, the immune system and signal transduction. Venn analysis of upregulated genes in the radiotherapy, immunotherapy, and combined radiotherapy and immunotherapy groups identified 11 common genes. Survival analysis revealed that myxovirus resistance 1 (MX1) was a key gene, which was highly expressed in pancreatic cancer and significantly negatively correlated with the prognosis of pancreatic cancer patients (P<0.05). The MX1 mRNA expression in pancreatic cancer was positively correlated with the infiltration levels of neutrophils and dendritic cells (r=0.413, 0.333, both P<0.05), and was also positively correlated with the expression of immune checkpoints CD80, CD274, and CD86 (all P<0.05). The gene cluster interacting with MX1 in pancreatic cancer included OASL1, RSAD2, IFIT1, IFIT2 and IFIT3. qRT-PCR results showed that in the tumor-bearing mouse models of pancreatic cancer KPC and PANC02 cells, compared with the control group, the relative expression level of MX1 mRNA in the radiotherapy combined with immunotherapy group increased significantly (P<0.05). Conclusion: The relative expression of MX1 mRNA significantly increased in radiotherapy combined with immunotherapy for pancreatic cancer, and it may be one of the key genes.

Objective: To construct a bioaffinity chromatography of TrkA-lipid rafts@CNBr-Sepharose 4B gels and apply it to the assessment of the efficacy of the affinity adsorption of TrkA targeted drugs. Methods: TrkA-lipid rafts@CNBr-Sepharose 4B gels stationary phase was prepared and characterized through the coupling reaction of cyanogen bromide immobilized ligand. The stability was assessed by repeating the up-sampling experiment over a period of 30 days. Affinity adsorption experiments were conducted to investigate the affinity adsorption behavior of TrkA-targeted drugs on both blank agarose gels columns and TrkA-lipid rafts@CNBr-Sepharose 4B gels columns. Results: The Western blotting results revealed that the lipid rafts contained Flotillin-1, Caveolin-1, and TrkA proteins. Following the coupling of U251 cell lipid rafts coupled Sepharose 4B gels, it was observed that the lipid rafts maintained a stable solid loading on the column bed even after rinsing with multiple column volumes of buffer. Adsorption experiments confirmed the specific affinity adsorption of the TrkA-lipid rafts@CNBr-Sepharose 4B gels stationary phase. Conclusion: The bioaffinity chromatography of TrkA-lipid rafts@CNBr-Sepharose 4B gels was successfully constructed, thus preliminarily validating its feasibility and stability.

Objective: Screening and identifying of core genes in clear cell renal cell carcinoma (ccRCC) based on bioinformatics analysis techniques to search for novel biomarkers for the diagnosis of ccRCC and therapeutic targets. Methods: Differentially expressed genes (DEGs) between ccRCC samples and normal kidney samples were obtained from three datasets (including GSE100666, GSE168845, and GSE96574) using the GEO2R tool; DAVID online tool was used to annotate the intersection DEGs of these three datasets with GO and KEGG functions; A protein interaction network of these intersecting DEGs was constructed using the STRING website, and then the core genes through the CytoHubba plugin in Cytoscape software were determined; The expression of core genes in ccRCC were verified using data from Oncomine and TIMER 2.0 databases, the relationship between core genes and the survival of ccRCC patients were further analysed using GEPIA database, and also the relationship between core genes and immune cell infiltration were further analysed using TIMER 2.0 database. Results: A total of 189 DEGs were screened, which involve various biological functions and pathways such as tumor growth regulation. Ten DEGs were identified as core genes, includingPTPRC, CTLA4, CCR5, CXCL9, GZMB, CD27, CXCL13, IDO1, LCP2, and CASR . The expression analysis results of GEO dataset, Oncomine, and TIMER database all showed that compared with normal samples, the expression of CASR in ccRCC was significantly reduced, while the expression of the other 9 core genes was significantly increased. Survival analysis showed that CTLA4, CXCL13, and CASR were associated with overall survival in ccRCC patients, while CXCL13 and CASR were associated with disease-free survival in ccRCC patients. The TIMER 2.0 results indicated that upregulation of core gene expression was associated with immune cell infiltration in ccRCC. Conclusion: Ten core genes were identified that may be closely related to the pathogenesis of ccRCC, which may become new diagnostic markers and therapeutic targets for ccRCC.