ObjectiveTo explore the effects of Jingui Shenqiwan （JSW） and Liuwei Dihuangwan （LDW） on the reproductive ability and brain function of normal mice and compare the actions of the two medications. MethodsSeven groups of female and male mice were divided at a ratio of 2∶1. Except for the control group， the other six groups were as follows： a group of both males and females receiving JSW （3.0 g·kg^-1）， a group of both males and females receiving LDW （4.5 g·kg^-1）， a group of males receiving water and females receiving JSW， a group of males receiving water while females receiving LDW， a group of females receiving water while males receiving JSW， and a group of females receiving water while males receiving LDW. Each group was administered the drug for 14 days and then caged together at a 2∶1 （female∶male） ratio to detect the number of pregnant mice and calculate the pregnancy rate. Pregnant mice continued receiving the drug until they naturally gave birth， which was followed by the observation of newborn mice， calculation of their average number， and the measurement of the offspring's preference for sugar water and neonatal recognition index. At the end of the experiment， the weights of the thymus and spleen were measured to calculate the organ coefficients， and mRNA or protein expression was analyzed in the brain and testes or ovaries. A 1% sucrose solution was used to examine the euphoria of their brain reward systems， while novel object recognition test （NOR） was applied to assess their memory capabilities. mRNA expression was detected using real-time quantitative polymerase chain reaction （Real-time PCR） assay， and protein expression was analyzed with Western blot. ResultsCompared with the control group， oral administration of JSW to both male and female mice for 14 days significantly increased the pregnancy rate of female mice on day 2 after being caged together （P<0.05）， while LDW showed a trend but no statistical significance. Additionally， compared with the control group， JSW could upregulate the gene expression of gonadotropin-releasing hormone （GnRH） in the thalamus， as well as reproductive stem cell factor （SCF） and tyrosine kinase receptor （c-Kit） in the testes and reproductive stem cell marker mouse vasa homologue （MVH） in the ovaries， upregulate the expression of proteins influencing neuronal functional activity， such as brain-derived neurotrophic factor （BDNF）， in hippocampal neurons （P<0.05）， and enhance sucrose preference in male mice （P<0.05）. Compared with the control group， JSW significantly increased sucrose preference and novel object recognition index in offspring mice （P<0.05）， which was related to the upregulation of hippocampal dopamine D1 receptor （D1R） and N-methyl-D-aspartate receptor （Nmdar） gene expression. Compared with the control group， both JSW and LDW could upregulate the protein expression of glucocorticoid receptor （GR）， BDNF， and tyrosine kinase receptor B （TrkB） in the hippocampus of offspring mice （P<0.05）. ConclusionJSW significantly enhances the reproductive ability of normal mice， which is not only related to the release of gonadotropin but also associated with its regulation of brain function. Additionally， JSW has a certain regulatory effect on the brain function of the offspring mice.

ObjectivePancreatic ductal adenocarcinoma (PDAC) exhibits a limited response to current treatments due to its dense fibrotic stroma and highly immunosuppressive tumor microenvironment. In recent years, advancements in cellular immunotherapy, particularly chimeric antigen receptor macrophage (CAR-M) therapy, have offered new hope for pancreatic cancer treatment. Although CAR-M therapy demonstrates dual potential in directly killing tumor cells and remodeling the immune microenvironment, it still faces challenges such as complex in vitro preparation processes and low in vivo targeting and delivery efficiency. Therefore, developing strategies for efficient and targeted in vivo delivery of CAR genes has become crucial for overcoming current therapeutic limitations. This study aims to develop an orally administrable nano-gene delivery system for the targeted delivery of CAR genes to pancreatic tumor sites. MethodsCore nano-gene particles (PNP/pCAR) were constructed by loading plasmid DNA encoding CAR (pCAR) with cationic polypeptides (PNP). Subsequently, PNP/pCAR was surface-modified with β-glucan to prepare the targeted nanoparticles (βGlus-PNP/pCAR). The loading efficiency of PNP for pCAR was quantitatively assessed by gel retardation assay. The particle size, Zeta potential, morphology, and storage stability of PNP/pCAR were characterized using a Malvern particle size analyzer and transmission electron microscopy. At the cellular level, RAW 264.7 macrophages were selected. The cytotoxicity of PNP/pCAR was evaluated using the CCK-8 assay. The cellular uptake efficiency and lysosomal escape ability of the nanoparticles were assessed via flow cytometry and confocal microscopy. Transfection efficiency was quantitatively evaluated by detecting the expression of the reporter gene GFP using flow cytometry. At the in vivo level, an orthotopic pancreatic cancer mouse model was established. Cy7-labeled βGlus-PNP/pCAR nanoparticles were administered orally, and the fluorescence distribution in mice was dynamically monitored at 1, 2, 4, 8, and 16 h post-administration using a small animal in vivo imaging system. Forty-eight hours after oral gavage, the mice were euthanized, and pancreatic tumor tissues were collected for further analysis of intratumoral fluorescence signals using the imaging system. Additionally, βGlus-PNP/pCAR-GFP nanoparticles loaded with the reporter gene (GFP) were administered orally. Forty-eight hours post-administration, pancreatic tumor tissues were harvested to prepare frozen sections, and GFP expression was observed and analyzed under a fluorescence microscope. ResultsThe PNP carrier exhibited a high loading capacity for pCAR. The successfully prepared PNP/pCAR nanoparticles were regular spheres with a hydrodynamic diameter of approximately (120±10) nm and a Zeta potential of about +(6±1) mV. They maintained good structural stability after incubation in PBS buffer for 7 d. Cell experiments demonstrated that PNP/pCAR exhibited no significant cytotoxicity in RAW 264.7 cells while being efficiently internalized and effectively escaping lysosomal degradation. The transfection positive rate of PNP/pCAR-GFP in RAW 264.7 cells reached (25±3)%, surpassing that of Lipofectamine 2000-loaded pCAR-GFP (Lipo/pCAR-GFP), which was (20±1)%.In vivo experiments revealed that, compared to unmodified PNP/pCAR, βGlus-PNP/pCAR exhibited strongerin situ pancreatic tumor targeting ability after oral administration. Furthermore, oral administration of βGlus-PNP/pCAR-GFP resulted in significant GFP protein expression detectable within pancreatic tumor tissues. ConclusionThis study successfully constructed and validated an orally administrable, pancreatic cancer-targeting polypeptide-based nano-gene delivery system. It provides an important technological foundation in delivery systems and experimental basis for the subsequent development of in situ CAR-M-based therapeutic strategies for pancreatic cancer.

ObjectivePancreatic ductal adenocarcinoma (PDAC) exhibits a limited response to current treatments due to its dense fibrotic stroma and highly immunosuppressive tumor microenvironment. In recent years, advancements in cellular immunotherapy, particularly chimeric antigen receptor macrophage (CAR-M) therapy, have offered new hope for pancreatic cancer treatment. Although CAR-M therapy demonstrates dual potential in directly killing tumor cells and remodeling the immune microenvironment, it still faces challenges such as complex in vitro preparation processes and low in vivo targeting and delivery efficiency. Therefore, developing strategies for efficient and targeted in vivo delivery of CAR genes has become crucial for overcoming current therapeutic limitations. This study aims to develop an orally administrable nano-gene delivery system for the targeted delivery of CAR genes to pancreatic tumor sites. MethodsCore nano-gene particles (PNP/pCAR) were constructed by loading plasmid DNA encoding CAR (pCAR) with cationic polypeptides (PNP). Subsequently, PNP/pCAR was surface-modified with β-glucan to prepare the targeted nanoparticles (βGlus-PNP/pCAR). The loading efficiency of PNP for pCAR was quantitatively assessed by gel retardation assay. The particle size, Zeta potential, morphology, and storage stability of PNP/pCAR were characterized using a Malvern particle size analyzer and transmission electron microscopy. At the cellular level, RAW 264.7 macrophages were selected. The cytotoxicity of PNP/pCAR was evaluated using the CCK-8 assay. The cellular uptake efficiency and lysosomal escape ability of the nanoparticles were assessed via flow cytometry and confocal microscopy. Transfection efficiency was quantitatively evaluated by detecting the expression of the reporter gene GFP using flow cytometry. At the in vivo level, an orthotopic pancreatic cancer mouse model was established. Cy7-labeled βGlus-PNP/pCAR nanoparticles were administered orally, and the fluorescence distribution in mice was dynamically monitored at 1, 2, 4, 8, and 16 h post-administration using a small animal in vivo imaging system. Forty-eight hours after oral gavage, the mice were euthanized, and pancreatic tumor tissues were collected for further analysis of intratumoral fluorescence signals using the imaging system. Additionally, βGlus-PNP/pCAR-GFP nanoparticles loaded with the reporter gene (GFP) were administered orally. Forty-eight hours post-administration, pancreatic tumor tissues were harvested to prepare frozen sections, and GFP expression was observed and analyzed under a fluorescence microscope. ResultsThe PNP carrier exhibited a high loading capacity for pCAR. The successfully prepared PNP/pCAR nanoparticles were regular spheres with a hydrodynamic diameter of approximately (120±10) nm and a Zeta potential of about +(6±1) mV. They maintained good structural stability after incubation in PBS buffer for 7 d. Cell experiments demonstrated that PNP/pCAR exhibited no significant cytotoxicity in RAW 264.7 cells while being efficiently internalized and effectively escaping lysosomal degradation. The transfection positive rate of PNP/pCAR-GFP in RAW 264.7 cells reached (25±3)%, surpassing that of Lipofectamine 2000-loaded pCAR-GFP (Lipo/pCAR-GFP), which was (20±1)%.In vivo experiments revealed that, compared to unmodified PNP/pCAR, βGlus-PNP/pCAR exhibited strongerin situ pancreatic tumor targeting ability after oral administration. Furthermore, oral administration of βGlus-PNP/pCAR-GFP resulted in significant GFP protein expression detectable within pancreatic tumor tissues. ConclusionThis study successfully constructed and validated an orally administrable, pancreatic cancer-targeting polypeptide-based nano-gene delivery system. It provides an important technological foundation in delivery systems and experimental basis for the subsequent development of in situ CAR-M-based therapeutic strategies for pancreatic cancer.

ObjectiveHepatocellular carcinoma (HCC) represents 90% of all primary liver cancers. The main risk factors associated with HCC include viral hepatitis (B and/or C), alcohol abuse, and metabolic dysfunction-associated steatotic liver disease (MASLD), which progressively advance to liver fibrosis, cirrhosis, and ultimately evolve into HCC. Surgical resection represents the most effective treatment for HCC, while recent advances in immunotherapy, including immune checkpoint inhibitors and adoptive cell therapies, have provided improved treatment prospects for patients with unresectable HCC. However, the complex metabolic heterogeneity of HCC limits the therapeutic efficacy. Metabolic intermediates acyl-CoA not only provide energy and substrates for numerous biochemical reactions but also serve as donors for protein lysine acylation, a major class of post-translational modification (PTM). Therefore, a deeper understanding of the molecular mechanisms underlying protein lysine acylation and hepatocarcinogenesis is urgently needed. MethodsThe levels of protein lysine acylation and silence information regulator 5 (SIRT5) expression levels in clinical HCC samples were analyzed by Western blot. Quantitative malonylome and succinylome of HCC samples were analyzed by antibody-based affinity enrichment coupled with tandem mass spectrometry. The proliferation of HCC cells was analyzed with Cell Counting Kit-8 (CCK-8) assays, the apoptosis was quantified by Annexin V-FITC/propidium iodide (PI) staining coupled with flow cytometry, and the ability of cells to migrate was assayed by Transwell assays. The enzymatic activity of glutathione S-transferase Mu 1 (GSTM1) was quantified. Transgenic mice with hepatic overexpression of SIRT5 were constructed using CRISPR-Cas9, and primary hepatocarcinogenesis was induced by administration of diethylnitrosamine. ResultsWestern blot analysis indicated that the expression level of SIRT5 was elevated in clinical samples from HCC patients, and the levels of lysine malonylation, glutarylation, and succinylation were significantly reduced in HCC tissues. Knockout of SIRT5 in MHCC-97H and MHCC-97L hepatoma cells suppressed cell proliferation, and increased the percentage of apoptotic cells significantly. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of the differentially malonylome and succinylome of HCC samples revealed significant enrichment in two major classes of biological processes: core energy metabolism (e.g., glycolysis/gluconeogenesis, tricarboxylic acid metabolic process, fatty acid beta oxidation) and detoxification and oxidative stress response (e.g., response to toxic substance, chemical carcinogenesis, reactive oxygen species (ROS)). SIRT5 removes malonylation from lysine residues in GSTM1 and restores its detoxification activity, which is crucial for the survival of hepatocytes under stressed conditions. More importantly, in vivo experiment indicated that hepatic-specific overexpression of SIRT5 in mice accelerated diethylnitrosamine-induced liver fibrosis and hepatocarcinogenesis, indicating the critical role of SIRT5 in HCC progression. ConclusionThis study highlights the previously unrecognized SIRT5-GSTM1 axis as a key regulator in hepatocarcinogenesis, and suggests a potential target for the treatment of patients with HCC.

ObjectiveTo develop and validate a diagnostic prediction model for differentiating pulmonary tuberculosis from community-acquired pneumonia based on routine blood parameters. MethodsA total of 642 patients with pulmonary tuberculosis and 503 patients with community-acquired pneumonia were retrospectively enrolled from the Seventh Hospital of Tangshan. They were randomly divided into a training set and an internal validation set at a 7∶3 ratio. Additionally， 218 patients from the 981st Hospital were independently included as an external validation set. The Boruta algorithm and recursive feature elimination were employed to select predictors from 82 blood parameters， sex， and age. A multivariate Logistic regression model was established and evaluated using receiver operating characteristic （ROC） curves， calibration curves， and decision curve analysis. ResultsThe final model incorporated eight predictors， namely uric acid， urea nitrogen， alkaline phosphatase， monoamine oxidase， alanine aminotransferase， glutathione， neutrophil percentage， and age. The model achieved areas under the curve （AUCs） of 0.800 （95%CI： 0.769-0.830）， 0.787 （95%CI： 0.738-0.836）， and 0.736 （95%CI： 0.667-0.835） in the training， internal validation， and external validation sets， respectively. The model demonstrated good calibration， and decision curve analysis showed clinical net benefit within the threshold probability range of 10%-80%. ConclusionThe developed model exhibits good discriminative ability and clinical utility， serving as an effective early screening tool for primary healthcare institutions.

The protein kinase A/protein kinase G/protein kinase C-family (AGC kinase family) of eukaryotes is involved in regulating numerous biological processes. The 3-phosphoinositide- dependent protein kinase 1 (PDK1), is a conserved serine/threonine kinase in eukaryotes. To understand the roles of PDK1 homologous genes in cell death and immunity in tetraploid Nicotiana tabacum, the previuosly generated transgenic CRISPR/Cas9 lines, in which 5-7 alleles of the 4 homologous PDK1 genes (NtPDK1a/1b/1c/1d homologs) simultaneously knocked out, were used in this study. Our results showed that the hypersensitive response (HR) triggered by transient overexpression of active Pto (Pto^Y207D) or soybean GmMEKK1 was significantly delayed, whereas the resistance to Pseudomonas syrangae pv. tomato DC3000 (Pst DC3000) and tobacco mosaic virus (TMV) was significantly elevated in these partial knockout lines. The elevated resistance to Pst DC3000 and TMV was correlated with the elevated activation of NtMPK6, NtMPK3, and NtMPK4. Taken together, our results indicated that NtPDK1s play a positive role in cell death but a positive role in disease resistance, likely through negative regulation of the MAPK signaling cascade.
Nicotiana/virology* ; Disease Resistance/genetics* ; Plant Diseases/immunology* ; Plants, Genetically Modified/genetics* ; Gene Knockout Techniques ; Plant Proteins/genetics* ; CRISPR-Cas Systems ; Protein Serine-Threonine Kinases/genetics* ; 3-Phosphoinositide-Dependent Protein Kinases/genetics* ; Pyruvate Dehydrogenase Acetyl-Transferring Kinase ; Tobacco Mosaic Virus/pathogenicity*

Objective:To investigate the association between homocysteine (Hcy) and recurrent pregnancy loss (RPL), as well as its impact on clinical pregnancy outcomes in patients undergoing in vitro fertilization/intracytoplasmic sperm injection and embryo transfer (IVF/ICSI-ET). Methods:This retrospective cohort study collected clinical data from patients undergoing IVF/ICSI-ET at the Reproductive Health Hospital of the Third Affiliated Hospital of Zhengzhou University between December 2020 and March 2024. Patients were divided into no history of pregnancy loss group (named control group, n=1 027) and RPL group ( n=743) based on history of pregnancy loss. Peripheral blood Hcy levels were compared between the two groups. Multivariate logistic regression was performed to adjust for confounding factors and determine whether Hcy is an independent risk factor for RPL. RPL patients were divided into four subgroups based on Hcy quartiles, named Q1 subgroup (Hcy<7.03 μmol/L), Q2 subgroup (7.03 μmol/L≤Hcy<8.63 μmol/L), Q3 subgroup (8.63 μmol/L≤Hcy<10.44 μmol/L), and Q4 subgroup (Hcy≥10.44 μmol/L), to further analyze the impact of Hcy level on pregnancy outcomes after IVF/ICSI-ET in these patients. Results:1) Baseline characteristics between control and RPL groups: statistically significant differences were observed in female age, male age, female body mass index (BMI), duration of infertility, cause of infertility, and peripheral blood Hcy levels (all P<0.05). 2) After adjusting for female age, male age, female BMI, duration of infertility, and cause of infertility via multivariate logistic regression, elevated Hcy levels was identified as an independent risk factor for RPL (a OR=1.366, 95% CI: 1.298-1.438, P<0.001). 3) Baseline characteristics of the four RPL subgroups: antral follicle count (AFC) differed significantly among Q1, Q2, Q3 and Q4 subgroups [17.00 (11.00, 24.00), 15.00 (10.00, 24.00), 14.00 (7.00, 22.25), 15.50 (8.00, 22.00), P=0.043]. No statistically significant differences were observed in other baseline characteristics (all P>0.05). 4) Pregnancy outcomes across the four RPL subgroups: miscarriage rates in the Q1, Q2, Q3 and Q4 subgroups were 18.18% (18/99), 30.61% (30/98), 33.70% (31/92), and 35.96% (32/89), respectively, live birth rates were 44.26% (81/183), 36.17% (68/188), 32.80% (61/186), and 30.65% (57/186), respectively. Intergroup differences in miscarriage rate and live birth rate were statistically significant ( P=0.033, P=0.036). Specifically, miscarriage rate in the Q3 and Q4 subgroups, and live birth rate in the Q4 subgroup were significantly higher than those in the Q1 subgroup (all q<0.05). However, no significant differences were observed in clinical pregnancy rate or early miscarriage rate among the four groups (all P>0.05). After adjusting for confounding factors using multivariate logistic regression, taking the Q1 subgroup as the control, there were no statistically significant differences in the clinical pregnancy rate between the remaining groups and the Q1 subgroup (all P>0.05). The early miscarriage rate in the Q3 subgroup (a OR=2.184, 95% CI: 1.077-4.426, P=0.030) and the early miscarriage rate in the Q4 subgroup (a OR=2.290, 95% CI: 1.116-4.697, P=0.024) were significantly higher than those in the Q1 subgroup; the miscarriage rate in the Q3 subgroup (a OR=2.207, 95% CI: 1.125-4.330, P=0.021) and the miscarriage rate in the Q4 subgroup (a OR=2.377, 95% CI: 1.209-4.674, P=0.012) were significantly higher than those in the Q1 subgroup; the live birth rate in the Q3 subgroup (a OR=0.615, 95% CI: 0.401-0.944, P=0.026) and the live birth rate in the Q4 subgroup (a OR=0.560, 95% CI: 0.364-0.863, P=0.009) were significantly lower than those in the Q1 subgroup. Conclusion:Elevated Hcy is a high-risk factor for RPL in IVF/ICSI-ET patients and may adversely affect pregnancy outcomes.

Objective:To evaluate the application effect of spiral cyclic intervention with information support based on knowledge, attitude/belief, practice (KABP) Model on the quality of life among hospitalized people living with HIV/AIDS (PLWHA), in order to provide a scientific basis for improving quality of life intervention strategies.Methods:This was a random-controlled trial. A total of 50 hospitalized PLWHA admitted to the Infectious Diseases Department of Xi'an Eighth Hospital from October 2021 to November 2023 were randomly divided into an intervention group and a control group by random number table method. The intervention group received a 12-week spiral cyclic intervention with information support, while the control group received routine nursing care. The quality of life data were collected at baseline, week 4, and week 12. Data analysis was conducted using Chi-square test, t-test and generalized estimating equation(GEE). Results:A total of 50 PLWHA were enrolled with 25 cases in each group. The control group comprised 22 males and 3 females aged 17-60 years, while the intervention group included 23 males and 2 females aged 21-60 years. GEE results showed that after 12 weeks of intervention, the intervention group scored significantly higher than the control group in physical health summary and eight dimensions: mental health, health distress, quality of life, cognitive functioning, pain, role functioning, general health perceptions, and health transition. Specifically, the intervention group′s scores were (49.45 ± 1.45), (64.99 ± 3.25), (69.29 ± 4.13), (61.71 ± 2.63), (76.46 ± 3.85), (81.54 ± 3.80), (82.05 ± 5.68), (55.80 ± 3.52), and (64.15 ± 3.28) points, compared to the control group′s (45.39 ± 1.17), (52.75 ± 2.19), (54.08 ± 4.15), (51.03 ± 3.07), (65.14 ± 3.36), (71.41 ± 3.39), (60.84 ± 6.15), (44.57 ± 2.40), and (51.43 ± 3.08) points. All differences were statistically significant (Wald χ2 values were 3.97-9.53, all P<0.05). Conclusions:The spiral cyclic intervention with information support based on the KABP Model effectively improves the quality of life of hospitalized PLWHA, particularly in physical health summary.

Objective:To analyze the current situation of health-related quality of life (HRQOL) and its influencing factors among community-dwelling elderly with mild cognitive impairment (MCI) based on the health ecology model (HEM).Methods:From December 2023 to September 2024, a cross-sectional survey was conducted among 997 community-dwelling elderly in Beijing, Shandong, Chongqing and other regions. The EuroQol five-dimensional questionnaire 5 level (EQ-5D-5L), the Montreal cognitive assessment (MoCA), and the Pittsburgh sleep quality index (PSQI) were used to assess the subjects.Descriptive analysis, Mann-Whitney U test, Kruskal-Wallis test, and Tobit regression analysis were performed using Stata 18.0 software. Results:A total of 324 valid samples were finally obtained, and the health utility value of MCI elderly was 0.842(0.815, 0.951). Tobit regression analysis showed that ≥80 years old( β=-0.121, 95% CI=-0.193--0.050) in individual traits level, don't exercise( β=-0.196, 95% CI=-0.255--0.137) and poor quality of sleep ( β=-0.064, 95% CI=-0.116--0.013) in behavior characteristics level, and poor subjective economic status( β=-0.153, 95% CI=-0.261--0.045) in living and working conditions level were risk factors for the health utility value of the elderly with MCI. Drinking( β=0.096, 95% CI=0.022-0.171) in behavior characteristics level and participating in social activities( β=0.126, 95% CI=0.062-0.190) in interpersonal network level were protective factors for the health utility value of the elderly with MCI. Conclusions:The HRQOL of the community-dwelling elderly with MCI in China is low, and its influencing factors are multi-level. HEM should be combined to strengthen the intervention and management of the elderly with MCI from personal constitution to policy environment to improve their HRQOL.

This article explores the construction and practice of the"Party Building+Health Science Popularization"model,using the"Yixian Health Science Popularization Guangdong Tour"campaign conducted by Sun Yat-sen Memorial Hospital as a case study.The initiative has achieved remarkable results.Additionally,it summarizes innovative measures,as well as uni-versal and exemplary experiences,providing new insights and pathway recommendations for public hospitals to develop the"Party Building+Health Science Popularization"model.