ObjectiveTo investigate the role of DEAD-box helicase 3 X-linked （DDX3X） in immune-mediated liver injury （ILI）， and to clarify its mechanism by regulating endoplasmic reticulum stress （ERS）-dependent apoptotic pathway and its association with the clinical progression of hepatitis B. MethodsMice were given injection of concanavalin A （ConA） via the caudal vein to establish a model of ILI， PBS （control group） and different concentrations of ConA were injected into the tail vein of hepatocyte-specific DDX3X-knockout mice （DDX3X^ΔHep and DDX3X-flox mice （DDX3X^fl/fl）， respectively.. The log-rank survival analysis， measurement of the serum levels of aspartate aminotransferase （AST） and alanine aminotransferase （ALT）， and HE staining of liver tissue were performed to assess liver injury， and qRT-PCR and Western Blot were used to measure the mRNA and protein expression levels of glucose-regulated protein 78 （GRP78）， CCAAT/enhancer-binding protein homologous protein （CHOP）， and DDX3X in liver tissue. Intraperitoneal injection of 4-phenylbutyric acid （4-PBA， 100 mg/kg） was performed to inhibit ERS. Serum samples （n=30） and liver tissue samples （n=6） were collected from healthy controls， chronic hepatitis B （CHB） patients， and hepatitis B virus-associated liver failure （HBV-LF） patients； ELISA was used to measure the serum level of DDX3X， and qRT-PCR/Western Blot was used to analyze the expression of targets in liver tissue. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsCompared with the control group of mice， the expression of DDX3X in the liver of mice induced by ConA was significantly increased after liver injury （P<0.05）， and hepatocyte-specific DDX3X knockout increased the 72-hour survival rate of mice by 55% （compared with 20% in the DDX3X^fl/fl group）， with significant reductions in the serum levels of ALT and AST （P<0.000 1） and the expression levels of the ERS markers GRP78 and CHOP （P<0.05）. After ERS was inhibited by 4-PBA， there was alleviation of liver injury （with reductions in ALT and AST， P <0.001） and a reduction in DDX3X expression （P<0.01）. The analysis of clinical samples showed that the mRNA and protein expression levels of liver DDX3X in CHB patients and HBV-LF patients were significantly higher than those in healthy controls （all P<0.01）， and there was a significant increase in the serum level of DDX3X in HBV-LF patients （P<0.000 1）. ConclusionDDX3X exacerbates ILI by regulating the ERS-dependent apoptotic pathway （GRP78/CHOP）， and its expression is associated with the progression of hepatitis B. Therefore， it can be used as a potential therapeutic target.

ObjectiveTo investigate the mechanism of Zuogui Wan （ZGW） in improving ovarian inflammatory microenvironment and stemness of ovarian germline stem cells （OSCs） for treating ovarian aging via the cyclic guanosine monophosphate/adenosine monophosphate synthase （cGAS）/stimulator of interferon genes （STING） signaling pathway. MethodsForty C57BL/6 female mice were randomly divided into a blank group， a model group， a low-dose ZGW group （2.7 g·kg^-1）， a high-dose ZGW group （5.4 g·kg^-1）， and an estradiol valerate group （0.15 mg·kg^-1）， with 8 mice in each group. Except the blank group， all other groups received a single intraperitoneal injection of cyclophosphamide at 120 mg·kg^-1 to establish an ovarian aging mouse model. After successful modeling， each group was continuously administered for 4 weeks， once daily. The physiological status of the mice was observed， and the ovarian index was calculated. The estrus cycle of the mice was monitored. Hematoxylin-eosin （HE） staining was used to observe pathological changes in ovarian tissue. Enzyme-linked immunosorbent assay （ELISA） was used to detect serum sex hormone levels. Serum inflammatory factors interleukin-1β （IL-1β）， tumor necrosis factor-α （TNF-α）， and mouse interleukin-6 （IL-6） levels were detected using kits. Western blot was used to detect the protein expression of ovarian cGAS， STING， p-STING， TANK-binding kinase 1 （TBK1）， p-TBK1， interferon-induced transmembrane protein 3 （Fragilis）， and Vasa homolog protein （MVH）. Quantitative real-time polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of inflammatory factors in ovarian tissue. Immunofluorescence double labeling was performed to locate OSCs in ovarian tissues， and fluorescence intensities of OSCs markers MVH and octamer binding transcription factor 4 （Oct4） were calculated. ResultsCompared with the blank group， the model group showed reduced body weight， ovarian wet weight， and ovarian index （P<0.01） and a disordered estrus cycle （P<0.01）. In addition， the levels of serum follicle-stimulating hormone （FSH）， TNF-α， IL-6， and IL-1β were increased （P<0.01）， while anti-Müllerian hormone （AMH） and estradiol （E₂） levels were decreased （P<0.01）. The protein expression of cGAS， p-STING/STING， and p-TBK1/TBK1 in ovarian tissue was increased （P<0.05， P<0.01）， while that of OSCs stemness factors MVH and Fragilis was reduced （P<0.01）. Immunofluorescence indicated a reduction in MVH and Oct4 expression in OSCs （P<0.01）. The mRNA expression of inflammatory factors TNF-α， IL-6， and IL-1β in ovarian tissue was increased （P<0.05， P<0.01）. Compared with the model group， the treatment groups exhibited improved body weight， ovarian wet weight， and ovarian index （P<0.05） and a reduced rate of estrus cycle disorder （P<0.05， P<0.01）. The levels of serum FSH， TNF-α， IL-6， and IL-1β were decreased （P<0.05， P<0.01）， while AMH and E₂ levels were increased （P<0.01）. The protein expression levels of cGAS， p-STING/STING， and p-TBK1/TBK1 in ovarian tissue were decreased （P<0.05）， while the protein expression of MVH and Fragilis was increased （P<0.05）， and the fluorescence intensities of MVH and Oct4 were increased （P<0.05， P<0.01）. The mRNA expression of inflammatory factors in ovarian tissue was decreased （P<0.05）. ConclusionZGW alleviate ovarian inflammatory response， regulate ovarian microenvironment homeostasis， and maintain stemness of OSCs in ovarian aging mice probably by modulating the cGAS-STING signaling pathway， thereby improving ovarian function and delaying ovarian aging.

Despite breakthroughs in immunotherapy for solid tumors, significant variations in treatment efficacy persist. Up to 80% of cancer patients suffer from malnutrition, which leads to: lymphoid atrophy and reduced T-cell reserves; deficiency of substrates required for T-cell activation and expansion; concurrent inflammation hindering T-cell infiltration into tumors; and cachexia accelerating PD-1 antibody clearance. Clinical studies confirm that severe malnutrition significantly impairs immune responses and increases the risk of treatment toxicity. Therefore, implementing standardized nutritional therapy is crucial for optimizing the reserve, activation, expansion, and infiltration capacity of immune cells, thereby providing a sound immune system foundation for immunotherapy. Immunonutrition therapy, by enhancing immunonutrients such as arginine, omega-3 polyunsaturated fatty acids, and nucleotides, reduces the secretion of pro-inflammatory mediators and promotes T-cell activation and proliferation. This enhances anti-tumor immune responses, prolongs survival, and advances cancer treatment towards multimodal combination and precision approaches.

ObjectiveAs a common lifestyle habit, alcohol consumption has a controversial association with the onset of Parkinson’s disease (PD). To demonstrate the correlation between alcohol consumption and PD and to identify associated genes, we integrated findings from clinical surveys, genomics, transcriptomics, and animal experiments. MethodsWe investigated the alcohol consumption rates (including both before and after disease onset) among 244 PD patients in China and 177 PD patients from the U.S. NHANES database. Mendelian randomization (MR) analysis was performed using genome-wide association study (GWAS) data for three alcohol-related traits and seven PD-related datasets from the MRC IEU OpenGWAS database. Transcriptomic data from the substantia nigra of PD patients were obtained from three GEO datasets (GSE7621, GSE20141, and GSE49036) to analyze RIT2 gene transcription. Finally, three groups of animal experiments (water/20% ethanol/20% liquor, with 4 C57BL/6J mice per group) were conducted to examine changes in brain RIT2 gene expression and transcriptomic profiles following alcohol consumption. ResultsThe alcohol consumption rates among PD patients in China and the U.S. (9%-18.87%) were significantly lower than the general population rates of 15%-45% in their respective regions (P<0.001), suggesting a possible negative association between alcohol consumption and PD. Subsequently, in 21 bidirectional MR analyses using 3 alcohol-related GWAS datasets and 7 PD-related GWAS datasets, the forward MR analyses (alcohol intake as exposure, PD as outcome) yielded 12 negative associations (OR_IVW<1) and 9 positive associations (OR_IVW>1). Among these, only two negative associations reached statistical significance: alcohol intake frequency (OR_IVW=0.75, 95% CI: 0.60-0.93, P=0.010) and alcohol consumption (OR_IVW=0.20, 95% CI: 0.05-0.83, P=0.026). The forward MR analysis (alcohol intake→PD) identified 235 SNPs, annotated to 316 genes, while the reverse MR analyses (PD→alcohol intake) identified 37 SNPs, annotated to 53 genes. Notably, only the RIT2 gene appeared in both the forward and reverse MR analyses (alcohol intake→PD: rs28597806, rs8083110; PD→alcohol intake: rs4588066). RIT2 is selectively expressed in the human brain (FPKM: 5.259±2.103), with low or no expression in peripheral tissues (FPKM: <1). Analysis of three human substantia nigra transcriptomic datasets revealed a decreasing trend in RIT2 gene expression in PD patients (GSE20141 array signal: 3.49±1.23 vs. 2.33±0.87, P=0.044). Animal experiments demonstrated that administration of 20% ethanol or 20% liquor (approximately 8% ethanol) stimulated a >2-fold upregulation of RIT2 gene expression in the mouse brain. Furthermore, transcriptomic sequencing revealed that the two alcohol-treated groups exhibited 96 (20% ethanol vs. water control) and 4 (20% liquor vs. water control) differentially expressed genes, respectively, indicating that low-dose alcohol consumption can achieve RIT2 upregulation while minimizing impact on other brain genes. In addition to its anti-infective effects, low-dose alcohol consumption primarily influences signaling pathways related to neurodegenerative diseases such as PD and Prion diseases. ConclusionAlcohol consumption is generally considered as a harmful lifestyle habit. However, some studies have also shown a lower risk of mortality among individuals who consume low doses of alcohol (100 g/week of ethanol) or drink occasionally. Currently, one of the research focuses on alcohol consumption is whether the human body can benefit from low-dose alcohol intake. This study provides new evidence supporting a negative association between alcohol consumption and PD, and for the first time, through MR analysis, identifies the RIT2 gene as a potential mediator of the effect of alcohol consumption on PD. RIT2 is selectively expressed in the human brain. Building upon existing evidence indicating downregulated RIT2 gene expression in PD pathogenesis, our experiments confirm that low-dose alcohol consumption can upregulate RIT2 expression in the brain. In brief, alcohol consumption may suppress the pathogenesis of PD by upregulating RIT2 expression in the substantia nigra. China is facing a serious problem of population aging. This study offers important insights for long-term PD prevention and treatment strategies, with the aim of benefiting more potential PD patients through lifestyle modifications, thereby improving the quality of life of the aging population and reducing the economic burden on healthcare.

Objective:To explore the effects of fluoride exposure from early life to adulthood on learning and memory abilities of offspring mice and the expression of genes related to the NOTCH signaling pathway in hippocampal tissue.Methods:Thirty 8-week-old (body weight 18 - 24 g) clean-grade C57BL/6J mice, half male and half female, were randomly divided into 3 groups by body weight and then paired 1 ∶ 1 (10 mice per group). Successfully pregnant female mice were given tap water containing 0 (control), 20, or 40 mg/L sodium fluoride (NaF) during gestation and lactation until weaning. Offspring mice were exposed to fluoride until 12 weeks of age after weaning at 21 days, with fluoride exposure doses identical to those of the maternal mice. The 24-hour urine of the offspring mice was collected to determine urine fluoride level using fluoride ion selective electrode method, and Morris water maze was used to evaluate their spatial learning and memory ability. The offspring mice were euthanized under anesthesia, and protein and RNA were extracted from hippocampal tissue. Western blot was used to detect the protein expression levels of postsynaptic density protein 95 (PSD95) and synapsin-1 (SYN-1) in hippocampal tissues. Quantitative real-time PCR and Western blot were used to detect the mRNA and protein expression levels of NOTCH signaling pathway related genes (including NOTCH1, DLL3, HES5) in hippocampal tissue.Results:Compared with the control group [(0.86 ± 0.25) mg/L], the offspring mice of the 20 and 40 mg/L NaF groups had higher urinary fluoride levels [(3.77 ± 0.51), (6.04 ± 1.63) mg/L, P < 0.05]. The Morris water maze results showed that compared with the control group, the offspring mice in the 20 and 40 mg/L NaF groups had longer escape latency (day 5), fewer platform crossings, and shorter target platform dwell time ( P < 0.05). Compared with the control group, the protein expression level of synaptic related protein PSD95 in the hippocampal tissues of the offspring mice in the 40 mg/L NaF group was lower, and the protein expression level of SYN-1 in the 20 and 40 mg/L NaF groups was lower ( P < 0.05). The protein expression level of NOTCH1, a gene related to the NOTCH signaling pathway, was lower in the hippocampal tissue of offspring mice in the 40 mg/L NaF group. The mRNA and protein expression levels of DLL3 and HES5 were also lower in the 20 and 40 mg/L NaF groups, and the mRNA expression level of HES5 in the 40 mg/L NaF group was lower than that in the 20 mg/L NaF group ( P < 0.05). Conclusions:Exposure to fluoride from early life to adulthood can lead to a decline in the learning and memory abilities of offspring mice, as well as a decrease in the expression of synaptic related proteins. The mechanism of action may be related to the inhibition of the NOTCH signaling pathway in hippocampal tissue.

OBJECTIVE: To explore the clinical significance of circulating tumor DNA (ctDNA) in response evaluation and relapse monitoring for patients with primary mediastinal large B-cell lymphoma (PMBCL). METHODS: The clinical characteristics, efficacy and survival of 38 PMBCL patients in our hospital from January 2010 to April 2020 were retrospectively analyzed. The ctDNA monitoring was conducted by targeted next-generation sequencing (NGS). RESULTS: Among the 38 patients, 26 cases were female, and 32 cases were diagnosed with Ann Arbor stage I-II. The 5-year overall survival (OS) rate and progression-free survival (PFS) rate were 74.7% and 61.7%, respectively. Males and those with high aaIPI scores (3 points) had a relatively poor prognosis. The NGS results of 23 patients showed that STAT6 (65.2%), SOCS1 (56.5%), and TNFAIP3 (56.5%) were the most common mutated genes. Patients with stable disease (SD)/progressive disease (PD) exhibited enrichment in cell cycle, FoxO, and TNF signaling pathways. A total of 29 patients underwent end-of-treatment PET/CT (EOT PET/CT), and 16 of them received ctDNA monitoring with 12 negative. Among 6 patients with EOT PET/CT positive (Deauville 4), 4 underwent ctDNA monitoring, and 3 of them were negative, being still in continuous remission without any subsequent anti-tumor therapy. CONCLUSION CtDNA may be combined with PET/CT to assess efficacy, monitor relapse, and guide treatment of PMBCL.
Humans ; Circulating Tumor DNA/blood* ; Female ; Mediastinal Neoplasms ; Male ; Retrospective Studies ; High-Throughput Nucleotide Sequencing ; Prognosis ; Lymphoma, Large B-Cell, Diffuse/genetics* ; Middle Aged ; Adult ; Aged ; Neoplasm Recurrence, Local ; Mutation

BACKGROUND:Kaempferol has anti-osteoporotic effects,but the mechanisms by which kaempferol regulates gut microbiota and metabolites to prevent and treat osteoporosis remain unclear.OBJECTIVE:To exploring the potential mechanisms by which kaempferol inhibit osteoporosis based on gut microbiota and comprehensive targeted metabolomics.METHODS:Eighteen female Sprague-Dawley rats were randomly divided into three groups:sham operation group,model group,and kaempferol group,with 6 rats in each group.Animal models of osteoporosis were made in the latter two groups through removal of bilateral ovaries.Eight weeks after modeling,the sham operation and model groups were gavaged with distilled water,and the kaempferol group was gavaged with 40 mg/kg kaempferol.Continuous administration in each group was carried out for 12 weeks.Rat fecal samples were collected for 16S rDNA amplicon sequencing to observe changes in the gut microbiota structure.Serum samples were subjected to comprehensive targeted metabolomics analysis using ultra-high performance liquid chromatography-tandem mass spectrometry technology,along with a proprietary database and multivariate statistical analysis.RESULTS AND CONCLUSION:After 12 weeks of continuous intervention,the results of 16S rDNA amplicon sequencing showed that compared with the sham operation group,the abundance of gut microbiota increased in the model group.Compared with the model group,kaempferol group exhibited a statistically significant increase in the abundance of the genus Latilactobacillus(P=0.021),while the abundances of Pantoea(P=0.034),Enterorhabdus(P=0.000),Monoglobus(P=0.024),Butyricimonas(P=0.034),Rothia(P=0.043),and Clostridia(P=0.004)were significantly downregulated.After 12 weeks of continuous intervention,the results of the serum samples analyzed by broad-targeted metabolomics revealed that 120 and 79 metabolites were identified between the sham operation and model groups and between the model and kaempferol groups,respectively.Among the three groups,there were 17 overlapping differentially expressed metabolites,including Cis-aconitic acid,barbituric acid,L-homocitrulline,3,4,5-trimethoxycinnamic acid,L-3-phenyllactic acid,cyclo(pro-pro),L-phenylalanine-L-serine,proline-isoleucine,L-donoraminoacetic acid-L-phenylalanineacetic acid,and phenylalanine-aspartic acid.Most of them belong to amino acids and their metabolites,glycerophospholipids and fatty acyls.The Kyoto Encyclopedia of Genes and Genomes pathways involved in the differential metabolites were mainly enriched in D-amino acid metabolism,histidine metabolism,propionate metabolism,lysine degradation,fatty acid metabolism and sphingolipid metabolism.After 12 weeks of continuous intervention,combined analysis revealed that genera such as Enterorhabdus,Latilactobacillus,Rothia,and Ruminococcus were closely associated with differential serum metabolites.To conclude,kaempferol may exert its anti-osteoporotic effects by modulating the abundance,diversity,and structure of gut microbiota,thereby regulating the metabolism of amino acids,their metabolites,and fatty acids.

In recent years,viral vector-mediated chimeric antigen receptor(CAR)cell therapy has emerged as a prominent topic in the field of cancer treatment and has demonstrated significant clinical efficacy in patients with hematological malignancies.However,the permanent expression of CAR induced by this therapy may give rise to severe adverse reactions.In contrast,messenger RNA(mRNA)chimeric antigen receptor therapy is anticipated to become a new generation of safer and more effective treatments due to its lack of insertional mutagenesis and minimal risk of off-tar-get toxicity.This therapeutic approach involves transfecting effector immune cells with CAR-en-coding mRNA to elicit an immune response within the body.It has been employed for treating B-cell malignancies,lymphoma,acute myelogenous leukemia(AML),and other cancers.The continuous optimization of mRNA purification,modification,and transfection technology enhances CAR ex-pression durability,while increasing killing efficiency and expanding the range of action.This pa-per reviews the research advances on CAR-encoding mRNA delivery,its application and advan-tages in cancer therapies.

Objective To establish an inductively coupled plasma-mass spectrometry(ICP-MS)method for the analysis of 29 mineral elements in Artemisiae Argyi Folium(AAF),and to develop a model for judging the origin of AAF based on the elemental fingerprint combined with chemometrics.Methods The variance method was used to compare the contents of 29 mineral elements in AAF samples from Qichun of Hubei province,Hebei province and Henan province,respectively.The discriminant models of AAF from different habitats were established by discriminant analysis,PLS-DA and PCA-Logistic regression algorithms.Results The differences of the 22 elements(V,Cr,Ni,As,Se,Rb,Mo,Cd,Sb,Ba,Hg,Pb,K,Li,B,Mg,Al,P,Ca,Fe,Zn,and Ga)in samples from Qichun of Hubei province,Hebei province and Henan province are extremely significant(P＜0.01).The discriminant analysis model showed that eight kinds of element variables(P,Cr,K,Li,Hg,Ba,Mg,and Mo),which have a significant effect on the origin discrimination are gradually introduced into the discriminant model,and the correct rate of back test is 100％.The correct rate of"leave one method"cross-validation is 96.7％.The PLS-DA model showed good prediction ability,and the variable importance projection values of K,P and Mg elements were greater than one,which could be used as the difference marker of AAF from different habitats.The accuracy of the origin discrimination model constructed by PC A-Logistic regression analysis was 100％.The elements with high first principal component loadings were Li,B,Mg,Al,P,K,Ca,V,Cr,Fe,Ni,Zn,As,Rb,Cd,Sb,and Ba.Conclusion This study shows that elemental fingerprint technology combined with chemometric analysis can identify AAF from Qichun of Hubei province,Hebei province,or Henan,providing technical support for the traceability of the origin of Artemisia argyi.

Objective:This study aims to explore the characteristics of body temperature(BT)of the elderly population in China and the relationship with health status.Methods:This is a cross-sectional study.The data were derived from the China Health and Nutrition Study(CHNS)in 2015.A total of 4 176 elderly people over 60 years old were enrolled for analysis(1968 males, and 2208 females). The participants were divided into three groups according to their BT values: hypothermia group(BT<36.5℃), moderate BT group(36.5℃≤BT<37.0℃), and hyperthermia group(37.0℃≤BT<37.5℃). The differences in health status among each group were compared based on the disease conditions(four-week prevalence rate)in the past four weeks.Multivariate logistic regression analysis was performed to explore the relationships between BT and the four-week prevalence rate.Results:Among the elderly population included in the analysis, the average age was 68.4 ± 6.9 years(ranging from 60 to 99 years). The average BT of the elderly was 36.40 ± 0.37℃(males: 36.42 ± 0.37℃; females: 36.38 ± 0.37℃).3 348 study subjects were aged between 60-74 years old, while 828 study subjects were aged between75-99 years old.The older the age, the lower the body temperature.Of the participants, 2 212(53.0%)elderly people belonged to the hypothermia group, 1 656(39.6%)belonged to the moderate BT group, and 308(7.4%)belonged to the hyperthemia group.Among 4 176 elderly people, a total of 1368 had suffered from disease in the past four weeks(31.3%)The four-week prevalence rates of each group were as follows: the hypothermia group(33.1%), the moderate BT group(29.8%); hyperthemia group(26.6%)( χ2=8.403, P=0.015). Multivariable logistic regression analysis indicated that for every 1℃ increases in BT among the elderly, the four-week prevalence rate decreased by 22%( OR=0.78, 95% CI=0.65-0.94, P=0.01). Conclusions:The BT of the elderly is generally lower than the current standard, and the lower the BT, the higher the four-week prevalence rate.Raising the BT of the elderly may help them improve their physical condition.