ObjectiveGastrodia elata has evolved ecological types with shortened rhizome internodes and diversified flower and fruit coloration in response to different altitudes. Studying the genetic mechanisms of different ecotype germplasm is significant for guiding variety breeding in different cultivation areas. MethodsThe bHLH gene family was identified based on the whole-genome datasets of G. elata f. elata and G. elata f. glauca. Subsequently， the gene family members were subject to analysis， including gene structure， chromosomal localization， cis-acting elements， gene synteny， and phylogeny. Combined with transcriptome data and quantitative Real-time PCR， the expression patterns of bHLH genes in the stems of the different G. elata ecotype germplasm were analyzed. Finally， correlation analysis was conducted between gene expression patterns and color to obtain the key bHLH genes regulating the color formation of stem. ResultsA total of 63 bHLH genes were identified in both G elata f. elata and G. elata f. glauca， unevenly distributed across 17 chromosomes and clustered into 16 subfamilies， with significant expansion in some family members. Obvious inversions of bHLH genes on the same chromosome and interchromosomal translocations were detected in the two ecotype germplasm. Among these genes， 12 bHLH genes （such as bHLH62-3 and bHLH74） were associated with the bright yellow color of G elata f. elata stem， while 9 bHLH genes （such as PIL13， UNE12， and bHLH130） were correlated with the red color of G. elata f. glauca stem. Compared to G. elata f. glauca， the bHLH48 expression level was significantly higher in flowers and scale leaves of G elata f. elata， and the bHLH62-3 expression level was significantly higher in all organs of G elata f. elata. ConclusionsFunctional pathway divergence of the bHLH family members has occurred across different chromosomes in G elata f. elata and G. elata f. glauca. Through synergism or antagonism with other genes， 21 bHLH genes participate in the coloration metabolic pathway regulation of stems， flowers， and fruits. Specifically， bHLH62-3 is involved in regulating stem color differentiation in the anthocyanin biosynthesis pathway of G. elata， thus relevant to the color formation of stem. Additionally， GebHLH48 positively regulates flowering-related pathways to promote the early-flowering phenotype of G. elata f. elata. These findings have laid the foundation for analyzing the genetic regulatory mechanisms underlying the color formation of the G. elata stem.

ObjectiveGastrodia elata has evolved ecological types with shortened rhizome internodes and diversified flower and fruit coloration in response to different altitudes. Studying the genetic mechanisms of different ecotype germplasm is significant for guiding variety breeding in different cultivation areas. MethodsThe bHLH gene family was identified based on the whole-genome datasets of G. elata f. elata and G. elata f. glauca. Subsequently， the gene family members were subject to analysis， including gene structure， chromosomal localization， cis-acting elements， gene synteny， and phylogeny. Combined with transcriptome data and quantitative Real-time PCR， the expression patterns of bHLH genes in the stems of the different G. elata ecotype germplasm were analyzed. Finally， correlation analysis was conducted between gene expression patterns and color to obtain the key bHLH genes regulating the color formation of stem. ResultsA total of 63 bHLH genes were identified in both G elata f. elata and G. elata f. glauca， unevenly distributed across 17 chromosomes and clustered into 16 subfamilies， with significant expansion in some family members. Obvious inversions of bHLH genes on the same chromosome and interchromosomal translocations were detected in the two ecotype germplasm. Among these genes， 12 bHLH genes （such as bHLH62-3 and bHLH74） were associated with the bright yellow color of G elata f. elata stem， while 9 bHLH genes （such as PIL13， UNE12， and bHLH130） were correlated with the red color of G. elata f. glauca stem. Compared to G. elata f. glauca， the bHLH48 expression level was significantly higher in flowers and scale leaves of G elata f. elata， and the bHLH62-3 expression level was significantly higher in all organs of G elata f. elata. ConclusionsFunctional pathway divergence of the bHLH family members has occurred across different chromosomes in G elata f. elata and G. elata f. glauca. Through synergism or antagonism with other genes， 21 bHLH genes participate in the coloration metabolic pathway regulation of stems， flowers， and fruits. Specifically， bHLH62-3 is involved in regulating stem color differentiation in the anthocyanin biosynthesis pathway of G. elata， thus relevant to the color formation of stem. Additionally， GebHLH48 positively regulates flowering-related pathways to promote the early-flowering phenotype of G. elata f. elata. These findings have laid the foundation for analyzing the genetic regulatory mechanisms underlying the color formation of the G. elata stem.

Objective To explore the impact of number of positive regional lymph nodes (nPRLN) in N1 stage on the prognosis of non-small cell lung cancer (NSCLC) patients. Methods Patients with TxN1M0 stage NSCLC who underwent lobectomy and mediastinal lymph node dissection from 2010 to 2015 were screened from SEER database (17 Regs, 2022nov sub). The optimal cutoff value of nPRLN was determined using X-tile software, and patients were divided into 2 groups according to the cutoff value: a nPRLN≤optimal cutoff group and a nPRLN>optimal cutoff group. The influence of confounding factors was minimized by propensity score matching (PSM) at a ratio of 1 : 1. Kaplan-Meier curves and Cox proportional hazards models were used to evaluate overall survival (OS) and lung cancer-specific survival (LCSS) of patients. Results A total of 1316 patients with TxN1M0 stage NSCLC were included, including 662 males and 654 females, with a median age of 67 (60, 73) years. The optimal cutoff value of nPRLN was 3, with 1165 patients in the nPRLN≤3 group and 151 patients in the nPRLN>3 group. After PSM, there were 138 patients in each group. Regardless of before or after PSM, OS and LCSS of patients in the nPRLN≤3 group were superior to those in the nPRLN>3 group (P<0.001). N1 stage nPRLN>3 was an independent prognostic risk factor for OS [HR=1.52, 95%CI (1.22, 1.89), P<0.001] and LCSS [HR=1.72, 95%CI (1.36, 2.18), P<0.001]. Conclusion N1 stage nPRLN>3 is an independent prognostic risk factor for NSCLC patients in TxN1M0 stage, which may provide new evidence for future revision of TNM staging N1 stage subclassification.

Objective To examine the impact of extreme temperature and drought stress on the survival of Bulinus globosus, so as to provide the theoretical evidence for the genomic research of Bulinus in absence of reference genes. Methods B. globosus snail samples were collected from Kiwani Shehia in Pemba Island, Zanzibar, Tanzania, and offspring snails were obtained through laboratory breeding and reproduction. A total of 120 10-week-old B. globosus snails from the same generation were selected and randomly assigned into four groups, including the high-temperature drought (HD) group, normal temperature drought (D) group, low-temperature drought (LD) group, and the control (C) group, of 30 snails in each group. Snails in HD, D, and LD groups were placed in beakers containing dry soil at the bottom and subsequently housed in climate chambers at 35, 26 ℃, and 10 ℃, respectively, while snails in Group C were maintained in 500 mL petri dishes containing dechlorinated tap water at 26 ℃. Following 3 days of breeding, living snails in each group were collected, and soft tissues were dissected and isolated. Total RNA was extracted from snail soft tissues for library construction, followed by high-throughput sequencing on the Illumina HiSeq 4000 sequencing system. De novo transcriptome assembly was performed using the Trinity software, and the longest transcripts were selected as unigenes. Gene functional annotations of unigenes were conducted using the Diamond software against Gene Ontology (GO) knowledgebase, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database, NCBI non-redundant (NR) protein sequences database, Protein Family (Pfam) database, and UniProtKB/Swiss-Prot (Swiss-Prot) knowledgebase. GO and KEGG enrichment analyses of differentially expressed genes (DEGs) were performed using the topGO and clusterProfiler software, respectively. In addition, four relevant genes were selected for validation using a real-time quantitative PCR (qRT-PCR) assay to verify the reliability of transcriptome sequencing results. Results Following 3 days of breeding, there were 7, 20, 28, and 30 survival B. globosus snails in HD, LD, D, and C groups, with corresponding survival rates of 23.33% (7/30), 66.67% (20/30), 93.33% (28/30), and 100.00% (30/30), respectively (χ² = 52.72, P < 0.001). De novo transcriptome assembly generated 176 942 unigenes, with annotation rates of 0.98%, 13.49%, 26.46%, 12.48%, and 14.39% against GO knowledgebase, KEGG pathway database, NR protein sequences database, Pfam database, and Swiss-Prot knowledgebase, respectively. There were 33 up-regulated and 72 down-regulated genes in Group D, 483 up-regulated and 815 down-regulated genes in Group HD, and 245 up-regulated and 172 down-regulated genes in Group LD relative to in Group C. Following removal of overlapping genes across groups and unmatched genes, 11 candidate genes were identified. GO and KEGG analyses revealed 3 heat shock protein (HSP)-related DEGs in these 11 candidate genes, which were annotated as HSP12.2, HSP70, and HSP20 genes and were all significantly up-regulated in each treatment group. Three immune and nervous system-related DEGs were identified, and were all significantly down-regulated in each treatment group, which were involved in the neural cell adhesion molecule L1-like protein pathway, fibrinogen binding protein pathway, and leukocyte elastase inhibitor-like protein pathway. qRT-PCR assay quantified that the expression trends of four genes related to temperature and drought stress across different treatment groups were highly consistent with transcriptome sequencing data. Conclusion The survival rate of B. globosus significantly reduces under combined stresses of extreme temperature and drought, possibly due to an imbalance in its cellular homeostasis regulatory system.

OBJECTIVE: To analyze the phenotype and genotype of a neonate with Osteo-oto-hepato-enteric syndrome (O2HE) and review the literature. METHODS: A female neonate diagnosed with O2HE syndrome on December 13, 2024 at the First Affiliated Hospital of Zhengzhou University was selected as the study subject, and her clinical characteristics were analyzed, and pathogenic variants were explored by whole exome sequencing (WES). This study was approved by the Medical Ethics Committee of the Hospital (Ethics No.: 2025-KY-1038). RESULTS: The proband, a female infant, was delivered by Cesarean section at 36⁺¹ weeks of gestation. Five days after birth, she had developed severe diarrhea, mild cholestasis, sensorineural hearing loss, and growth retardation. WES revealed that she has harbored novel compound heterozygous variants c.512delA (p.Lys171Serfs*64) and c.698C>A (p.Thr233Asn) of the UNC45A gene, which were inherited from her mother and father, respectively. A total of 8 English papers were retrieved, which involved 16 patients from 14 families. Combined with our case, the 17 patients included 13 (76.5%) females and 4 (23.5%) males. Four patients (23.5%) had consanguineous parents. One case was excluded from further genetic analysis due to co-morbidity with other genetic variants. The primary clinical features included diarrhea (87.5%), cholestasis (81.3%), sensorineural hearing loss (31.3%), bone fragility (37.5%), and developmental delay (50.0%). Bi-allelic compound heterozygous mutations were identified in 12 patients (75.0%), and homozygous variants in 4 (25.0%). These included missense, nonsense, frameshift and deletional variants. The c.710T>C (p.Leu237Pro) variant was identified for 5 times, 3 of which were in homozygote forms. CONCLUSION O2HE syndrome should be suspected in cases with diarrhea, cholestasis, and hearing abnormalities during early postnatal period. Genetic testing facilitate early identification, genetic diagnosis and treatment.
Humans ; Female ; Infant, Newborn ; Male ; Mutation ; Hearing Loss, Sensorineural/genetics* ; Diarrhea, Infantile/genetics* ; Exome Sequencing ; Phenotype ; Fetal Growth Retardation ; Hair Diseases ; Facies

Background/Aims: Liver transplantation (LT) following total hepatectomy is a life-saving treatment for hepatocellular carcinoma (HCC). The HCC recurrence after LT hinders the effectiveness of the procedure. The objective of this study is to develop a pre-operative risk stratification model based on a liquid biopsy. Methods: We conducted a comprehensive multi-omics study of 260 HCC patients from three centers, including clinical data, low-coverage whole-genome sequencing of cell-free DNA (cfDNA) from plasma, as well as whole-exome, single-nucleus RNA, and spatial transcriptomics from matched tumor and non-tumor tissues. Results: We identified cfDNA-derived copy number alteration (CNA) signatures associated with post-transplant recurrence. By integrating cfDNA-derived CNA profiles with single-cell transcriptomic data, we traced recurrence-associated cfDNA to a distinct subpopulation of malignant cells within the primary tumor. These cells were embedded in a pro-metastatic microenvironment of specialized endothelial subtypes and cancer-associated fibroblasts. Notably, most recurrence-associated lesions were detectable in cfDNA prior to liver transplantation (LT). Building on these insights, we developed the ZJU Criteria based on CNA fragments and tumor markers, a pre-LT risk prediction tool that integrates conventional clinical factors with cfDNA-derived CNA signatures, and validated it using internal and independent external cohorts. Conclusion Our findings suggest that post-transplant recurrence commonly originates from advanced subclones that emerge late during tumor evolution. The ZJU Criteria provides an accurate, non-invasive strategy that significantly improves pre-LT risk stratification and clinical decision-making for patients with HCC.

Objective: To evaluate the cost utility of different influenza vaccination strategies for primary and secondary school students in Anshun City, so as to provide healtheconomic evidence for influenza prevention and control in this population. Methods: Based on the demographic data of 380 700 students aged 5-14 in Anshun City in 2024, a decision tree model for different influenza vaccination strategies was constructed. The incremental cost utility ratio (ICUR) and net monetary benefit (NMB) were calculated, and sensitivity analysis was performed to assess the robustness of the model results. Results: For influenza vaccination among primary and secondary school students in Anshun City, compared with the selfpaid strategy (2% coverage), the free strategy (45% coverage) and the target strategy (70% coverage) both yielded an ICUR of -7 874.29 yuan per quality adjusted life year (QALY). When using 1 and 3 times the per capita Gross Domestic Product (GDP) as the willingness to pay thresholds, the per capita Net Monetary Benefit (NMB) was 1 000, 1 600 and 2 900, 4 500 RMB, respectively. The number of influenza infections or school absenteeism decreased by 25% and 40%, while saving a total cost of (46.779 5 and 73.976 8) million yuan, and increasing QALY by 5 940.79 and 9 394.73, respectively. Sensitivity analysis showed that within the reasonable fluctuation range of parameters, the ICUR remained consistently lower than the one and three times willingness to pay. Furthermore, 99% of the simulation results for both the free and target strategies fell below those thresholds, indicating high robustness of the model results. Conclusion Implementing free and target influenza vaccination strategies for primary and secondary school students in Anshun City offer distinct cost utility advantages, and the advantage enhances as the vaccination coverage rate increases.

ObjectiveTo investigate the therapeutic effect of Bazhen Lihe Kangxian prescription on rats with liver fibrosis induced by carbon tetrachloride （CCl₄）， and to explore its mechanism of action against liver fibrosis. MethodsA total of 50 male Sprague-Dawley rats were randomly divided into blank group， model group， silybin group （43.19 mg/kg）， and low-， middle-， and high-dose Bazhen Lihe Kangxian prescription groups （7.96， 15.93， and 31.86 g/kg， respectively）. Subcutaneous injection of 40% CCl₄-olive oil solution （twice a week for 8 consecutive weeks） was performed to establish a model of liver fibrosis. After successful modeling， the rats in the blank group and the model group were given normal saline by gavage， while those in the treatment groups were given the corresponding drug by gavage， once a day for 4 consecutive weeks. HE staining and Masson staining were used to observe liver histopathological changes； an automatic biochemical analyzer was used to measure the serum levels of albumin （Alb）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT）； ELISA was used to measure the serum levels of hyaluronic acid （HA）， collagen Ⅳ （Col-Ⅳ）， procollagen Ⅲ N-terminal peptide （PⅢNP）， and laminin （LN）； Western blotting was used to measure the protein expression levels of phosphoinositide 3-kinase （PI3K）， phosphorylated PI3K （p-PI3K）， protein kinase B （Akt）， phosphorylated Akt （p-Akt）， mammalian target of rapamycin （mTOR）， and phosphorylated mTOR （p-mTOR） in liver tissue； RT-qPCR was used to measure the mRNA expression levels of collagen Ⅰ （Col-Ⅰ） and α-smooth muscle actin （α-SMA） 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 blank group， the model group had disrupted hepatic lobular structure， disordered arrangement of hepatic cell cords， hepatocyte fatty degeneration， severe inflammatory cell infiltration， and deposition of massive blue collagen fibers， as well as significant increases in the levels of AST， ALT， HA， LN， Col‑Ⅳ， and PⅢNP and a significant reduction in the level of Alb （all P<0.01）. Compared with the model group， the silybin group and the three Bazhen Lihe Kangxian prescription groups had varying degrees of improvement in inflammatory cell infiltration， hepatocyte fatty degeneration， and collagen deposition in liver tissue， as well as significant reductions in the levels of AST， ALT， HA， LN， Col‑Ⅳ， and PⅢNP and a significant increase in the level of Alb （all P<0.01）. Compared with the blank group， the model group had significant increases in the protein expression levels of p‑PI3K/PI3K， p-Akt/Akt， and p‑mTOR/mTOR and the mRNA expression levels of Col-I and α-SMA in liver tissue （all P<0.01）； compared with the model group， the silybin group and the high- and middle-dose Bazhen Lihe Kangxian prescription groups had significant reductions in the protein expression levels of p-PI3K/PI3K （all P<0.01）， and the silybin group and the three Bazhen Lihe Kangxian prescription groups had significant reductions in the protein expression levels of p-Akt/Akt and p-mTOR/mTOR and the mRNA expression levels of Col-Ⅰ and α-SMA in liver tissue （all P<0.01）. ConclusionBazhen Lihe Kangxian prescription can improve CCl₄-induced liver fibrosis in rats and alleviate liver injury， possibly by inhibiting the PI3K/Akt/mTOR pathway.

Objective:To explore the mediating effect of job embeddedness and job satisfaction on psycholog-ical capital and organizational silence in clinical nurses.Methods:Totally 412 clinical nurses(145 males and 267 females)were selected and assessed with the Nurse Organizational Silence Assessment Questionnaire(NOSAQ),Psychological Capital Questionnaire-24(PCQ-24),Job Embedding Scale(JES)and Nurses Job Satisfaction Scale(NJSS).SPSS macro program PROCESS and Bootstrap method were used to explore the role of mediation.Results:The NOSAQ scores were negatively correlated with the scores of PCQ-24,JES and NJSS(r=-0.55,-0.59,-0.51,Ps＜0.01).Job embeddedness and job satisfaction played a chain mediating role between psychological capital and organizational silence of clinical nurses(95％CI:-0.26--0.04),and the mediating effect accounted for 14.7％of the total effect.Conclusion:It suggests that organizational silence is closely related to psychological capital,job embeddedness and job satisfaction in clinical nurses.

Objective To study the predictive value of aspartate aminotransferase to platelet ratio index(APRI)combined with AIMS65score for death and rebleeding within 6 weeks in patients with cirrhosis with upper gastrointestinal bleeding(UGIB).Methods The clinical data of 302 patients with cirrhosis with UGIB who attended the Affiliated Hospital of Xuzhou Medical University from June 2019 to April 2024 were retrospectively collected.Binary Logistic regression was used to analyse the independent risk factors for death within 6 weeks and rebleeding.The receiver operating characteristic(ROC)curve was plotted and the area under the curve(AUC)was calculated to compare its efficacy in predicting death and rebleeding within 6 weeks.Results Univariate and multivariate Logistic regres-sion analyses showed that APRI and AIMS65score were independent risk factors for death within 6 weeks in cirrhosis with UGIB;ROC curves showed that the AUC values of APRI combined with AIMS65score were higher than those of APRI,AIMS65score,Child-Pugh score,and MELD score(P＜0.05),and their AUC values were 0.889,0.775,0.859,0.832 and 0.735.History of previous bleed-ing,APRI,and AIMS65score were independent risk factors for rebleeding within 6 weeks in cirrhosis with UGIB;ROC curves showed that the AUC values of APRI,AIMS65score,Child-Pugh score,MELD score,and APRI combined with AIMS65score had AUC values of 0.709,0.729,0.735,0.688 and 0.774,respectively.And the predictive efficacy of APRI combined with AIMS65score was better than that of APRI,AIMS65score,and MELD score(P＜0.05).Conclusion APRI and AIMS65score are independent risk factors for death and rebleeding within 6 weeks in cirrhosis with UGIB,and the combination of the two has better predictive value for the short-term prog-nosis of patients with cirrhosis with UGIB.