Objective: To develop a RHCE genotyping assay based on kompetitive allele-specific PCR (KASP) and assess its clinical accuracy for RhCE blood group determination. Methods: KASP primers were designed to interrogate three RHCE loci: the 109 bp insertion/deletion in intron 2, c. 307T>C, and c. 676C>G. A total of 1 194 RhD-positive inpatients from Chengdu were typed by both KASP genotyping and manual tube serology. Discordant samples (n=10) were retested by both methods and further resolved by Sanger sequencing. An additional 377 cases were tested for the c. 48C>G locus to evaluate the predictive accuracy of individual loci and combined locus testing for RhC antigen. Results: Genotyping concordance with serology was 100.0% for both the c. 676C>G locus (RhE/Rhe) and the c. 307T>C locus (Rhc). For RhC prediction using the 109 bp insertion, overall accuracy was 99.7% (1 191/1 194); the 3 discordant cases were confirmed by Sanger sequencing to be false negatives attributable to 109 bp deletion in intron 2. Testing the c. 48C>G allele for RhC prediction yielded 7 false positives, with an accuracy of 98.1% (370/377). RhC antigen status was determined by combining the 109 bp insertion and the c. 48C allele. After excluding 10 samples with inconsistent results between the two loci, the accuracy reached 100% in the remaining 367 samples. When both loci were applied in combination, accuracy reached 100% in the 367 cases with concordant results. Among the 1 194 patients, CCee (45.8%) and CcEe (31.7%) were the most common RhCE phenotypes. The e antigen had the highest positivity rate (92.2%), and the Ce haplotype was the most frequent (66.9%). Conclusion: The KASP-based RHCE genotyping method achieves high accuracy for clinical RhCE typing. Combining the 109 bp insertion/deletion with the c. 48C allele significantly improves RhC antigen prediction compared with either locus alone. This method was applied to RhCE genotyping of 1 194 RhD-positive inpatients in Chengdu, providing local RhCE phenotype and haplotype distribution data to support RhCE-matched transfusion practice.

Background/Aims: The pathophysiology of refractory gastroesophageal reflux disease (RGERD), which differs from proton pump inhibitor dependent gastroesophageal reflux disease (DGERD), remains incompletely elucidated. This study aims to compare esophageal motility patterns, transdiaphragmatic pressure gradients (TPG), and reflux profiles between RGERD and DGERD patients, and to delineate dynamic pressure gradient-esophagogastric junction (EGJ) interactions in these patients. Methods: In this retrospective study, 274 patients who underwent 24-hour impedance-pH monitoring and high-resolution manometry, along with an assessment of proton pump inhibitor responsiveness, were classified as RGERD (32.5%), DGERD (54.4%), or non-GERD (13.1%). Clinical characteristics, TPG, esophageal motility, and reflux metrics were compared between RGERD and DGERD patients. Subgroup analysis excluding hiatal hernia (HH) was conducted to investigate the pathophysiology of RGERD. Results: The RGERD group exhibited a significantly higher proportion of chest pain compared to the DGERD group. Regarding reflux profiles, RGERD patients without HH (RGERDHH- group) experienced increased weakly acidic reflux (P < 0.001) and prolonged bolus exposure (P = 0.006) compared to their counterparts (DGERDHH- group). Mechanistically, the RGERDHH- group showed reduced lower esophageal sphincter basal pressure (P = 0.010) and EGJ contractile integral (P = 0.005). Notably, following a wet-swallow, the RGERDHH- group experienced the significant elevation in gastric pressure and TPG. Correlation analyses revealed weakly acidic reflux and bolus exposure were positively correlated with gastric pressure variation, and inversely correlated with lower esophageal sphincter basal pressure. Conclusions Transient gastric pressure elevation and compromised EGJ barrier function drive weakly acidic reflux and esophageal bolus exposure. This pressure gradient-barrier mismatch underpins the refractoriness of RGERD.

Background: and Purpose Live cell-based assays (LCBA) are increasingly used for serological antibody detection due to their ability to preserve antigen conformation, offering moderately higher sensitivity than fixed cell-based assays. However, the clinical necessity of prioritizing LCBA for the detection of aquaporin-4 immunoglobulin G (AQP4-IgG) in neuromyelitis optica spectrum disorder (NMOSD) remains unclear, especially when compared to its established role in diagnosing myelin oligodendrocyte glycoprotein antibody-associated disease. Methods: We compared the performance of live cell-based assays using immunofluorescence (LCBA-IF) and fixed cell-based assays using immunofluorescence (FCBA-IF) in detecting AQP4-IgG in 90 cases of NMOSD meeting 2015 International Panel for Neuromyelitis Optica Diagnosis criteria, alongside 40 controls. Additionally, we further investigated the relationship between AQP4-IgG titers as measured by LCBA-IF and FCBA-IF and clinical parameters in NMOSD patients. Results: Results showed 96.9% agreement between LCBA-IF and FCBA-IF (Cohen’s κ=0.935), with a strong Spearman correlation (0.977, p<0.001). Both methods demonstrated 100% specificity, with LCBA-IF showing slightly higher sensitivity compared to FCBA-IF. Within LCBAIF-tested groups, statistically significant differences in annualized relapse rates were observed across all pairwise comparisons (low-titer vs. moderate-titer, low-titer vs. high-titer, and moderate-titer vs. high-titer; all p<0.050). However, this association reached statistical significance in some FCBA-IF-tested groups. Conclusions Overall, there is a strong concordance between LCBA-IF and FCBA-IF in detecting AQP4-IgG, where LCBA-IF shows slightly higher sensitivity. Furthermore, there is a potential link between elevated AQP4-IgG titers and an increased risk of relapse, and this correlation may appear more pronounced when using LCBA-IF.

Glaucoma is a group of optic nerve disorders characterized by progressive optic nerve atrophy and visual field defects, which can lead to irreversible blindness. Early diagnosis of glaucoma is essential for preventing visual loss. However, due to the absence of obvious early symptoms, the diagnosis of glaucoma remains challenging. Visual electrophysiological examinations, an objective approach for evaluating visual function, have the potential to be used in the early diagnosis of glaucoma. This review integrates the latest publications to introduce visual electrophysiological examination techniques, including electroretinography(ERG)and visual evoked potential(VEP). It also explores the mechanisms underlying these techniques and their application value in the early diagnosis of glaucoma. In addition, this review summarizes the advantages, limitations, and applicable scenarios of different visual electrophysiological techniques. Finally, the review provides an outlook on the development prospects of visual electrophysiological techniques in the early diagnosis of glaucoma. The findings of this review can assist clinicians in selecting appropriate diagnostic methods, promote the innovation and development of early visual electrophysiological diagnostic techniques for glaucoma, and contribute to reducing the risk of blindness caused by glaucoma.

Objective To observe the value of non-parallel transmission(non-PTX)reduced field-of-view(rFOV)echo planar imaging(EPI)sequence applied in diffusion weighted imaging(DWI)for displaying prostate lesions.Methods Conventional EPI-DWI and non-PTX rFOV-EPI-DWI were prospectively acquired in 30 patients with prostate lesions,including 22 cases of prostatic hyperplasia and 8 cases of prostate cancer.Subjective scoring of imaging quality,as well as objective evaluation on indexes including signal-to-noise ratio(SNR),contrast ratio(CR),contrast-to-noise ratio(CNR)and lesions'apparent diffusion coefficient(ADC)values were performed and compared between two kinds of DWI.Results The subjective score of non-PTX rFOV-EPI-DWI was higher than that of conventional EPI-DWI(P＜0.001).SNR,CR,CNR of non-PTX rFOV-EPI-DWI and lesions'ADC values measured on non-PTX rFOV-EPI-DWI were all higher than those of conventional EPI-DWI(all P＜0.05).Conclusion non-PTX rFOV-EPI DWI could display prostate lesions better than conventional EPI-DWI.

Objective To screen hub genes and signaling pathways associated with acute ischemic stroke(AIS)using bioinformatics methods,identify potential biomarkers,and provide new evidence for the mechanism research of AIS.Methods The gene expression dataset GSE37587 of AIS patients and healthy controls was obtained from the public database gene expression omnibus(GEO).The differentially expressed genes(DEGs,|log2 FC|≥1.2,FDR＜0.05)were screened using the limma package.The enrichment analysis of GO/KEGG was performed with the DAVID database.The weighted correlation network analysis(WGCNA)was used to construct a gene co-expression network for screening key modules.Then,a protein-protein interaction(PPI)network was constructed based on the STRING database and Cytoscape software to identify hub genes.The dataset GSE16561 was used to validate.Meanwhile,the clinical samples from 30 AIS patients and 30 healthy controls visited Zibo First Hospital from January to May 2025 were validated by the real time fluorescence quantitative PCR(qRT-PCR).The diagnostic efficacy was evaluated using the receiver operating characteristics(ROC)curve.Results A total of 653 DEGs were identified,including 252 up-regulated and 401 down-regulated genes.They were mainly enriched in biological processes such as ribosome biogenesis,endoplasmic reticulum protein processing,and oxidative phospho-rylation,as well as signaling pathways such as viral infection-related pathways and PD-L1/PD-1 checkpoint pathways in cancer.The core genes in the light green module identified by the WGCNA analysis were significantly enriched in the pathways such as mitophagy,ribosome,and endocytosis.The hub genes such as RPL34 and DDIT3 were screened from the PPI network,and their expression levels were significantly correlated with AIS.The analysis of the ROC curve showed that the areas under the ROC curve(AUCROC)of the hub genes for the diagnosis of AIS were 0.78-0.82,which had high clinical application value.Conclusion Ribosomal proteins,endoplas-mic reticulum stress-related genes,and viral infection response pathways are key molecular events in the occurrence of AIS.The genes such as RPL34 and DDIT3 are expected to be potential biomarkers for AIS,providing experimental evidence for the development of di-agnostic markers.

Objective:To establish an oxidative stress injury model by using hydrogen peroxide (H 2O 2) to induce human ovarian granulosa cells COV434. Methods:Human ovarian granulosa cells line COV434 were randomly divided into 6 groups, control group was not treated, H 2O 2 groups were treated with H 2O 2 of 200 μmol/L, 400 μmol/L, 600 μmol/L, 800 μmol/L and 1 000 μmol/L for 0.5 h, 1 h, 2 h, 4 h and 6 h, respectively, and the cell viability was determined by CCK-8 method. The follow-up experiments were treated with different concentrations of H 2O 2 for 1 h. β-galactosidase staining was used to determine the degree of cell senescence. DCFH-DA fluorescence staining was determined by flow cytometry, and the level of reactive oxygen species (ROS) in cells was determined. JC-1 staining was used to determine the mitochondrial membrane potential of cells. Western blotting was used to determine the expression levels of apoptosis-related proteins Caspase-3 and Caspase-9. After the successful establishment of the model, in order to verify the usability of the cell model, the cells were pretreated with the antioxidant vitamin E for 12 h, followed by the addition of H 2O 2 for intervention, and the ROS level and mitochondrial membrane potential were measured. Results:The cell viability of the 200 μmol/L and 400 μmol/L groups decreased first and then increased compared with the control, and tended to be stable after 1 h of intervention, and there was no significant difference in cell viability at each time point (all P>0.05). When the concentration of H 2O 2 increased to 600 μmol/L, the cell viability gradually decreased with the treatment time and tended to stabilize after 1 h, and decreased significantly to nearly 50% ( P<0.001). When the concentration of H 2O 2 continued to increase to 800 μmol/L and 1 000 μmol/L, the cell viability gradually decreased with the treatment time and stabilized after 1 h, and decreased to less than 10% (all P<0.001). When the concentration of H 2O 2 was 200 μmol/L and 400 μmol/L, there was no significant difference in the ratio of β-galactosidase-positive cells and the relative ROS intensity after 1 h compared with the control (all P>0.05). When the concentration of H 2O 2 increased to 600 μmol/L, 800 μmol/L and 1 000 μmol/L, the ratio of β-galactosidase-positive cells and the relative ROS intensity increased significantly (β-galactosidase staining: P=0.011 at 600 μmol/L, P=0.003 at 800 μmol/L, P=0.005 at 1 000 μmol/L; the relative ROS intensity: P=0.002 at 600 μmol/L, P<0.001 at 800 μmol/L and 1 000 μmol/L). Compared with the control, the mitochondrial membrane potential of cells decreased gradually after intervention with different concentrations of H 2O 2, and was negatively correlated with H 2O 2 concentration (all P<0.001). There was no difference in the expression of Cleaved-Caspase-3 in the H 2O 2 group at 200 μmol/L compared with the control, and the expression was significantly increased at 400 μmol/L, 600 μmol/L, 800 μmol/L and 1 000 μmol/L (all P<0.001). The expressions of Caspase-3 and Caspase-9 were significantly increased in all H 2O 2 treated groups (all P<0.001). Compared with the model group, the relative ROS intensity of the vitamin E group was significantly reduced ( P=0.009), and the mitochondrial membrane potential was significantly increased ( P<0.001), but it could not be restored to the level of the control. Conclusion:Using 600 μmol/L H 2O 2 to continuously treat COV434 cells for 1 h can quickly establish a stable and effective oxidative stress injury model of human ovarian granulosa cells.

Objective:To investigate the relationship between nerve growth factor (NGF) concentration in follicular fluid and abnormal menstrual cycle in infertile patients with polycystic ovary syndrome (PCOS).Methods:A retrospective cohort study was conducted on 100 infertile patients with PCOS who underwent in vitro fertilization and embryo transfer (IVF-ET) at Department of Obstetrics and Gynecology, Peking University Third Hospital from March 2017 to June 2019. For comparison, the 100 patients with PCOS were divided into low NGF group ( n=50) and high NGF group ( n=50) based on the median NGF concentration (1 644.03 ng/L) in follicular fluid. Baseline characteristics, menstrual status and clinical outcomes of assisted reproductive technology were compared. We performed multiple linear regression analysis to examine the effect of NGF in follicular fluid on menstrual cycle length for multivariate analysis. Results:1) PCOS patients in the low NGF group had significantly higher body mass index [(27.24±5.17) kg/m 2] and white blood cell count [7.31(5.99, 8.43)×10 9/L ] than those in the high NGF group [(25.03±4.46) kg/m 2, P=0.024; 5.95(5.08,7.01)×10 9/L, P=0.001], while high-density lipoprotein cholesterol [1.15 (0.98, 1.36) mmol/L] and basic follicle-stimulating hormone level [6.51 (5.10,7.95) U/L] in the low NGF group were significantly lower than those in the high NGF group [1.36 (1.09,1.52) mmol/L, P=0.039;6.51 (5.10,7.95)U/L, P=0.040]. 2) PCOS patients in the low NGF group had significantly higher menstrual cycle length [60.00 (35.00, 180.00) d] than the high NGF group [32.50 (27.00,67.50) d, P=0.001]. 3) Multiple linear regression analysis revealed that after adjustment for body mass index, age, infertility duration, infertility type, and glucose and lipid metabolic parameters, the NGF concentration in the follicular fluid independently and negatively correlated with menstrual cycle length ( P<0.05). 4) The NGF concentration in follicular fluid was not correlated with assisted reproductive outcomes. Conclusion:NGF concentration in follicular fluid is closely related to the degree of menstrual cycle abnormalities in patients with PCOS.

Objective:To investigate the intervention effect of exercise rehabilitation based on constructivism teaching mode on pelvic girdle pain (PGP) in pregnant women in late pregnancy, and to provide guidance for improving the pregnancy experience of PGP pregnant women.Methods:A quasi-experimental study was conduced, from November 2023 to February 2024, using convenience sampling method to select late pregnant women with PGP who had been filed and had regular obstetric checkups in the Second Hospital of Shandong University as the study subjects, they were randomly divided into intervention group (granted exercise rehabilitation based on the constructivism teaching model) and control group (granted exercise rehabilitation based on the traditional teaching model) according to the random number table method. Compare the pain scores and teaching effectiveness of pregnant women before and after the intervention in the two groups.Results:The final number of the two groups were 50 in each group, aged (32.38 ± 3.93) years in the intervention group; aged (31.34 ± 3.19) years in the control group. After the intervention, the pain scores of pregnant women in the intervention group (1.48 ± 1.26) scores were significantly lower than those in the control group (2.96 ± 1.42) scores, and the difference was statistically significant ( t = -5.45, P<0.05). The scores of the evaluation of the teaching effect memory problem scores, movement mastery scores, and classroom evaluations were (7.96 ± 1.30) scores, (13. 96 ± 0.88) scores, (27.54 ± 3.08) scores in the intervention group, which were significantly higher than those in the control group (5.80 ± 1.29) scores, (8.48 ± 1.82) scores, (16.24 ± 3.80) scores, the differences were statistically significant ( t = 8.30, 19.16, 16.31, all P<0.05). Conclusions:Based on the constructivism teaching model, exercise rehabilitation for pregnant women with PGP in late pregnancy can effectively alleviate their pelvic girdle pain, and the teaching effect is better than the traditional teaching model, which has the potential to be promoted and applied in the teaching of obstetric maternity schools.

BACKGROUND:The utilization of miniscrews for assisting in molar distalization with clear aligners is a commonly adopted clinical approach.However,treatment outcomes may be influenced by the implantation position of miniscrew and the hooks in various tooth location.OBJECTIVE:To analyze the biomechanical effects of hooks at different tooth positions and the assistance of buccal and palatal miniscrews in distalizing molars with clear aligners by using the finite element method.METHODS:By integrating volunteer cone beam CT and Itero intraoral scan data,high-precision finite element models were constructed to simulate the combined use of miniscrews and hooks to assist in the distalization of the maxillary first and second molars with clear aligners,aiming for a designed distalization magnitude of 0.2 mm.Four conditions were established based on the placement of buccal or palatal miniscrews,as well as the located of hooks at the canine or first premolar.The buccal anchorage screw was used in combination with hooks on the buccal side of the canine;the buccal anchorage screw was used in combination with hooks on the buccal side of the first premolar;the palatal anchorage screw was used in combination with hooks on the lingual side of the canine,and the palatal anchorage screw was used in combination with hooks on the lingual side of the first premolar.Analysis using ANSYS software was conducted to evaluate the three-dimensional displacement trends of maxillary teeth and the maximum equivalent stress on the periodontal ligament under each condition.RESULTS AND CONCLUSION:(1)Regardless of whether hooks were placed on canines or first premolars,palatal miniscrews had been shown to enhance the distal movement of maxillary molars and reduce the mesial movement of premolars as well as the labial tipping tendency of anterior teeth,compared to buccal miniscrews.The maximum equivalent stress in the periodontal ligament of molars was increased,while that of anterior teeth was decreased.(2)Regardless of whether the miniscrews were located on the buccal or palatal side,compared with hooks at the first premolar,when hooks were done at the canine,the labial inclination trend of the anterior teeth was reduced,the distal movement of the molars was reduced,the mesial movement of the premolars was increased,and the maximum equivalent stress of the molar and the periodontal membrane of the anterior teeth was reduced.(3)Regardless of whether miniscrews were located on the buccal or palatal side,and the hooks were located on the canine or the first premolar,the molar could not achieve overall movement.(4)The results show that clear aligner combined with palatal miniscrews and hooks at the first premolar are more conducive to improving the efficiency of maxillary molar distalization,but attention should be paid to protecting the anterior tooth miniscrews and molar torque control;clear aligner combined with palatal miniscrews and hooks at the canine is more conducive to reducing the labial tipping tendency of anterior teeth,but the efficiency of molar distalization will be relatively reduced.