Objective: To establish an accurate and rapid typing method for Rh typing of samples from patients who have received recent blood transfusions by utilizing the difference in osmotic fragility between fresh and old red blood cells. Methods: A lysing solution suitable for destroying old RBCs was prepared. Sixty-one samples collected in our hospital in 2024 with Rh typing of double groups were treated with the lysing solution to remove the old allogeneic red blood cells while preserving the patient's own fresh red blood cells, followed by repeat Rh typing tests. Results: For 61 samples with Rh typing in double groups, 41 were accurately detected identified through the red blood cell lysis method, yielding an identification rate of 67.21%. No significant difference was observed compared to the detection rate of the commonly used capillary centrifugation modified method (χ =0.103, P>0.05). Conclusion: The red blood cell lysis method provides a novel and rapid experimental approach for clinical use in processing Rh-typed samples that are of double groups, thereby offering a basis for Rh compatibility blood transfusion.

BACKGROUND:The Guilu Erxian Glue consists of Testudinis Plastrum,Cornu Cervi,Lycii Fructus,and Ginseng Radix.In earlier clinical observations,it is discovered that using Guilu Erxian Glue to treat patients with liver-kidney deficiency type knee osteoarthritis effectively alleviated knee pain,increased the range of motion,and improved walking ability.However,the exact mechanism by which oral administration of Guilu Erxian Glue can produce local therapeutic effects on the knee joint is still unclear.OBJECTIVE:To investigate the effects of Guilu Erxian Glue on gut microbiota in rats with knee osteoarthritis and to evaluate its mechanism using 16S rDNA sequencing and machine learning analysis.METHODS:Totally 18 female SD rats were randomly divided into three groups:blank group,model group,and Guilu Erxian Glue group,with 6 rats in each group.A knee osteoarthritis model was prepared using the destabilization of the medial meniscus surgical method.After successful modeling,the Guilu Erxian Glue group was given a decoction of Guilu Erxian Glue by gavage,while the blank and model groups were given an equal amount of distilled water.After 28 days of continuous intervention,high performance liquid chromatography was used to detect the active ingredients of Guilu Erxian Glue.MRI imaging was used to observe the condition of rat knee articular cartilage.Fecal samples were collected;DNA was extracted using a kit,amplified and purified by PCR,and an Illumina sequencing library was constructed.The Illumina MiSeq platform was used for high-throughput sequencing to generate raw sequence data.After obtaining the raw data,QIIME2 software was used to process the data.Linear Discriminant Analysis Effect Size analysis and random forest algorithm were used to screen for differential species in microbial data.KEGG and MetaCyc functional pathway analyses were used to explore the association between key microbial communities and experimental groups.Linear discriminant analysis effect values and random forest algorithm were used to screen for differential species.Association networks were used to analyze the interactions between microbial communities,and machine learning methods were used to analyze the composition and changes of gut microbiota.RESULTS AND CONCLUSION:(1)LC-MS component identification was conducted on the traditional Chinese medicine formula of Guilu Erxian Glue,and a total of 7 effective ingredients were identified.(2)MRI imaging showed that synovitis scope of high-density shadows in rats of the Guilu Erxian Glue group was reduced,and the degeneration of medial femoral condyle cartilage was less than that in the model group.(3)16S rDNA sequencing showed that the model group rats exhibited significant microbial imbalance,with a significant decrease in the abundance of Firmicutes and Bacteroidetes at the phylum level,while the proportion of Proteobacteria increased significantly(P＜0.05).The gut microbiota structure of rats in the Guilu Erxian Glue group was significantly improved,and the proportion of Firmicutes and Bacteroidetes increased,restoring a more diverse microbiota composition,approaching that of the blank group(P＜0.05).(4)KEGG and MetaCyc functional pathway analysis showed that the Guilu Erxian Glue group significantly activated multiple metabolic pathways,including amino acid metabolism,lipid metabolism,and biotin synthesis pathways(P＜0.05).(5)The results indicate that Guilu Erxian Glue contains seven active ingredients,and the changes in gut microbiota of knee osteoarthritis rats were analyzed using 16S rDNA sequencing.Guilu Erxian Glue can significantly improve the imbalance of gut microbiota,restore the abundance of beneficial bacteria,and have a significant impact on the composition of gut microbiota,providing scientific basis for the efficacy and mechanism of Guilu Erxian Glue.

BACKGROUND:The Guilu Erxian Glue consists of Testudinis Plastrum,Cornu Cervi,Lycii Fructus,and Ginseng Radix.In earlier clinical observations,it is discovered that using Guilu Erxian Glue to treat patients with liver-kidney deficiency type knee osteoarthritis effectively alleviated knee pain,increased the range of motion,and improved walking ability.However,the exact mechanism by which oral administration of Guilu Erxian Glue can produce local therapeutic effects on the knee joint is still unclear.OBJECTIVE:To investigate the effects of Guilu Erxian Glue on gut microbiota in rats with knee osteoarthritis and to evaluate its mechanism using 16S rDNA sequencing and machine learning analysis.METHODS:Totally 18 female SD rats were randomly divided into three groups:blank group,model group,and Guilu Erxian Glue group,with 6 rats in each group.A knee osteoarthritis model was prepared using the destabilization of the medial meniscus surgical method.After successful modeling,the Guilu Erxian Glue group was given a decoction of Guilu Erxian Glue by gavage,while the blank and model groups were given an equal amount of distilled water.After 28 days of continuous intervention,high performance liquid chromatography was used to detect the active ingredients of Guilu Erxian Glue.MRI imaging was used to observe the condition of rat knee articular cartilage.Fecal samples were collected;DNA was extracted using a kit,amplified and purified by PCR,and an Illumina sequencing library was constructed.The Illumina MiSeq platform was used for high-throughput sequencing to generate raw sequence data.After obtaining the raw data,QIIME2 software was used to process the data.Linear Discriminant Analysis Effect Size analysis and random forest algorithm were used to screen for differential species in microbial data.KEGG and MetaCyc functional pathway analyses were used to explore the association between key microbial communities and experimental groups.Linear discriminant analysis effect values and random forest algorithm were used to screen for differential species.Association networks were used to analyze the interactions between microbial communities,and machine learning methods were used to analyze the composition and changes of gut microbiota.RESULTS AND CONCLUSION:(1)LC-MS component identification was conducted on the traditional Chinese medicine formula of Guilu Erxian Glue,and a total of 7 effective ingredients were identified.(2)MRI imaging showed that synovitis scope of high-density shadows in rats of the Guilu Erxian Glue group was reduced,and the degeneration of medial femoral condyle cartilage was less than that in the model group.(3)16S rDNA sequencing showed that the model group rats exhibited significant microbial imbalance,with a significant decrease in the abundance of Firmicutes and Bacteroidetes at the phylum level,while the proportion of Proteobacteria increased significantly(P＜0.05).The gut microbiota structure of rats in the Guilu Erxian Glue group was significantly improved,and the proportion of Firmicutes and Bacteroidetes increased,restoring a more diverse microbiota composition,approaching that of the blank group(P＜0.05).(4)KEGG and MetaCyc functional pathway analysis showed that the Guilu Erxian Glue group significantly activated multiple metabolic pathways,including amino acid metabolism,lipid metabolism,and biotin synthesis pathways(P＜0.05).(5)The results indicate that Guilu Erxian Glue contains seven active ingredients,and the changes in gut microbiota of knee osteoarthritis rats were analyzed using 16S rDNA sequencing.Guilu Erxian Glue can significantly improve the imbalance of gut microbiota,restore the abundance of beneficial bacteria,and have a significant impact on the composition of gut microbiota,providing scientific basis for the efficacy and mechanism of Guilu Erxian Glue.

Background/Aims: Natural killer (NK) cell function is generally considered dampened in chronic hepatitis B virus (HBV) infection; however, the NK cell pool exhibits phenotypic and functional heterogeneity, and the antibody--mediated effect of NK cells remains less characterized. This study evaluated the dynamic changes in antibody-mediated NK cell responses and the involvement of distinct NK subsets across disease stages and during antiviral treatment. Methods: A T-cell receptor-like antibody specific for the HBV core 18–27 peptide (cTCRL-Ab) was used to determine the antibody-mediated effect of NK cells, and an array of NK cell surface markers were analyzed in cross-sectional and longitudinal cohorts of patients with chronic HBV infection. Single-cell RNA sequencing (scRNA-seq) was performed to identify the heterogeneity of NK subsets. Results: The cTCRL-Ab enabled the detection of NK cell cytolytic activity and IFNγ production. Notably, cTCRL-Ab-mediated NK cell responses were compromised in chronically HBV-infected patients, particularly in those receiving pegylated interferon-α (Peg-IFNα), which was associated with the downregulation of CD16 expression. Correspondingly, Peg-IFNα inhibited cTCRL-Ab-mediated NK cell function by reducing CD16 expression in vitro. scRNA-seq revealed that CD16 downregulation occurred mainly within a dysfunctional CD16hi NK subset exhibiting exhaustion properties. In contrast, an activated CD16hiNK subpopulation (CX3CR1⁺KLRC2–CD16hi) with high cytotoxicity was enriched in patients who experienced favorable treatment responses. Furthermore, the intrahepatic CX3CR1+KLRC2–CD16hi subset tended to exhibit functional restoration in HBsAg-loss individuals. Conclusions Our data contribute to the understanding of antibody-mediated responses of NK cells in chronic HBV infection, and highlight a previously unappreciated functional CX3CR1+KLRC2–CD16hiNK subset as a potential therapeutic target.

Objective To investigate clinical features and severity influencing factors of misophonia in psychiatric outpatients. Methods A total of 130 misophonia patients from Weifang People's Hospital (Oct 2023-Jun 2025) were assessed with scales and psychiatric interviews. Linear regression was used to analyze influencing factors. Results Among the 52.3% reported gradually worsening symptoms. Primary triggers were human oronasopharyngeal sounds (65.3%). Core emotional responses were disgust (86.2%) and anger (61.5%), with 75.4% experiencing loss of control. Common coping strategies included covering ears/wearing earplugs (66.1%) and avoidance (64.6%). Mean Amsterdam Misophonia Scale-Revised score was 26.4±5.2, indicating severe social dysfunction. Common comorbidities were obsessive-compulsive disorder (11.5%), depressive disorder (8.5%), and social anxiety disorder (5.4%). Multivariate analysis identified depressive disorder, social anxiety disorder, obsessive-compulsive disorder and post-traumatic stress disorder as independent factors affecting severity (all P<0.05). Conclusion Misophonia follows a chronic course with severe impairment in social functioning and quality of life. Obsessive-compulsive disorder, depressive disorder, social anxiety disorder, and post-traumatic stress disorder are key factors influencing symptom severity.

ObjectiveTo investigate the clinical efficacy and safety of Yanling Yishou prescription combined with aerobic exercise in the intervention of heart and spleen deficiency syndrome in middle-aged white-collar workers with "stress-induced aging". MethodsA randomized， controlled， and open-label trial was conducted involving 156 patients with stress-induced aging and heart and spleen deficiency syndrome who were recruited from the Affiliated Hospital of Nanjing University of Chinese Medicine. Participants were randomly assigned via a central randomization system into an intervention group and a control group， with 78 cases in each group. The intervention group received Yanling Yishou prescription combined with a standard aerobic exercise protocol， while the control group received the standard aerobic exercise protocol only. The composition of Yanling Yishou prescription was derived from Qinggong Medical Records： 10 g Ziziphi Spinosae Semen， 10 g Codonopsis Radix， 10 g Astragali Radix， 10 g Paeoniae Radix Alba， 10 g Angelicae Sinensis Radix， 10 g Rehmanniae Radix， 10 g Atractylodis Macrocephalae Rhizoma， 10 g Poria， and 3 g Glycyrrhizae Radix et Rhizoma， decocted in water and administered 2 hours after breakfast and dinner， respectively. The aerobic exercise protocol was prescriptionted according to the 2017 guidelines of the American College of Sports Medicine， consisting of a 5-minute warm-up， a progressively increasing exercise stage （starting at 5 minutes and increasing by 1-2 minutes daily up to a maximum of 20 minutes）， and a 10-minute cool-down. Both groups were treated for 28 days and followed up for 35 days. Outcome measures included Checklist Individual Strength （CIS）， sleep quality indicators （ total sleep time， light sleep， and deep sleep）， traditional Chinese medicine （TCM） symptom scores for heart and spleen deficiency syndrome， Hospital Anxiety and Depression Scale （HADS）， physical function score from the SF-36 scale （SF-36 PF）， Pain Visual Analog Scale （Pain VAS）， and vital signs. ResultsCompared with baseline， the intervention group showed a significantly greater reduction in CIS scores and improvement in sleep quality （P<0.05， P<0.01）， while the control group showed no significant changes. Post-treatment comparisons between groups revealed that the intervention group achieved significantly better outcomes in fatigue reduction and sleep improvement （P<0.05， P<0.01）. In terms of TCM syndrome scores， the intervention group showed significant improvements in fatigue， frequent awakening， vivid dreaming， poor appetite， palpitations， and loose stools （P<0.05， P<0.01）， while the control group showed no significant improvement in poor appetite， palpitations， or loose stools. After treatment， the intervention group showed significantly better improvements in fatigue， frequent awakening， vivid dreaming， and palpitations compared to the control group （P<0.05， P<0.01）. No significant differences were observed between the two groups in SF-36 PF， HADS， and Pain VAS scores before and after treatments. No serious adverse events occurred in either group during the study period. ConclusionYanling Yishou prescription combined with aerobic exercise can alleviate symptoms such as fatigue， frequent awakening， vivid dreaming， and palpitations， improve sleep quality， and alleviate heart and spleen deficiency symptoms， thereby improving the quality of life in middle-aged white-collar workers with stress-induced aging.

Misophonia is a psychophysiological and behavioral disorder characterized by an individual's low tolerance to specific sounds， leading to intense negative emotional experiences and physiological responses. Currently， there is no standardized and universally effective treatment for misophonia in clinical practice worldwide. This article reports the case of an 18-year-old male patient with misophonia who showed poor response to sertraline combined with exposure and response prevention therapy. Subsequently， the patient received 8 weeks of Morita therapy （once a week， 50 minutes per session）， with symptomatic improvement. By presenting this case， we explore the potential efficacy of Morita therapy in treating misophonia， aiming to provide a reference for its clinical management. ［Funded by Scientific Research Development Fund Project of Shandong Second Medical University （number， 2024FYM034）］

Abstract: To explore the relationship between the replication-associated bromodomain protein 4 ( BRD4)and human papillomavirus (HPV) 16(HPV16) viral load in cervical squamous cell carcinoma and CIN Ⅰ tissues , confirm the effects of BRD4 degradation agent MZ1 on viral load . Methods : Thirty HPV16-positive cervi- cal cancer specimens and 30 non-cervical cancer specimens were collected , and the viral load of the samples was detected by real-time fluorescence quantitative PCR , and the expression of BRD4 was analyzed by immunohisto- chemistry and Western blot. Results : The viral load was higher in the samples of cervical cancer group than in the samples of non-cancer group , the difference is statistically significant ( P < 0. 01) . Immunohistochemistry results showed that the expression of BRD4 were significantly higher in cervical cancer specimens than in noncancerous specimens , the difference is statistically significant (P < 0. 05) . BRD4 expression was significantly and positively correlated with high viral loads , the difference is statistically significant (P < 0. 001) . the BRD4 degradation agent MZ1 significantly reduced the viral load , the difference is statistically significant (P < 0. 01) . Conclusion BRD4 may be involved in the replication of HPV16 virus , and BRD4 degradation agent MZ1 can inhibit the replication of HPV16 virus .

Objective To analyze the trends in disease burden and risk factors of depression among the elderly population in China from 1990 to 2021,and to provide a theoretical basis for the prevention,treatment,and policy-making of geriatric depression in China.Methods Data on the disease burden of geriatric depres-sion in China from 1990 to 2021,including the number of incident cases,disability-adjusted life years(DALYs),incidence rate,and DALY rate,were extracted from the 2021 Global Burden of Disease(GBD)database.The Joinpoint regression model was used to analyze the trends by calculating the annual percentage change(APC)and average annual percentage change(AAPC).The autoregressive integrated moving average(ARIMA)model was employed to predict the disease burden of geriatric depression over the next five years.Population attributable fractions(PAFs)were used to describe the risk factors for geriatric depression in China in 1990 and 2021.Results From 1990 to 2021,the number of incident cases and the incidence rate of geri-atric depression in China showed an overall upward trend.The most significant increase in incidence was ob-served in the 60-64 age group,while the prevalence rate increased notably in the ≥ 95 age group.The DALY rate showed the most pronounced upward trend in the 65-69 age group.The incidence,prevalence,and DALY rates of geriatric depression were higher in women than in men.Major risk factors included child-hood sexual abuse and intimate partner violence,with the impact of intimate partner violence being particular-ly significant among women.The ARIMA model predicted that the incidence,prevalence,and DALY rates of geriatric depression in China would decline over the next five years,with a greater decline observed in women than in men.Conclusions From 1990 to 2021,the incidence,prevalence,and DALY rates of geriatric de-pression in China showed an overall upward trend,with higher rates observed in women than in men.Greater attention should be paid to the elderly female population,with a focus on early prevention to reduce the dis-ease burden of geriatric depression.

Objective:To construct curved microgrooves with gradient surface roughness on polycaprolactone (PCL) members by regulating alkali etching time and to investigate the synergistic effect of surface roughness and curved microgrooves on the in vitro osteogenic differentiation of mouse pre-osteoblasts (MC3T3-E1), aiming to determine the optimal PCL surface modification strategy. Methods:Soft lithography and melt-casting techniques were used to fabricate PCL membranes with regularly arranged curved microgrooves (CMP). Alkali etching was performed for 24, 48, and 72 h. Groups: smooth PCL (control), CMP (curved microgrooves only), CMP-24 h, CMP-48 h, CMP-72 h (CMP etched for 24, 48, 72 h, respectively). Surface physicochemical properties were characterized: surface morphology was observed by scanning electron microscopy (SEM), surface roughness was measured by atomic force microscopy (AFM), and surface hydrophilicity was evaluated by contact angle measurement. MC3T3-E1 cells were cultured in vitro. Cell adhesion, proliferation, and osteogenic differentiation were assessed using cell counting (CCK-8), immunofluorescence staining, alkaline phosphatase (ALP) and Alizarin red staining with quantification. The mRNA expression levels of osteogenesis-related genes [ALP, collagen type Ⅰ (COL-1), Runt-related transcription factor 2 (RUNX-2), osteocalcin (OCN), osteopontin (OPN)] were detected by real-time fluorescence quantitative PCR (RT-qPCR). Results:Curved microgrooves were successfully fabricated on PCL membranes. Alkali treatment improved surface hydrophilicity and increased roughness. The CMP-72 h group exhibited the best hydrophilicity (contact angle: 33.2°±5.5°), with significant differences compared to all other groups (all P<0.05). The CMP-72 h group showed the highest roughness [(59.966±4.729) nm], which was significantly different from all other groups (all P<0.05). CCK-8 results on day 5 showed that both curved microgrooves and roughness promoted cell proliferation: CMP-24 h (0.292±0.003) and CMP-72 h (0.383±0.004) were significantly higher than those in the smooth group (0.270±0.005) (all P<0.05). Immunofluorescence staining revealed that curved microgrooves induced significant contact guidance of cells; this effect weakened with increasing etching time. ALP and Alizarin red staining indicated the deepest osteogenic staining in the CMP-48 h group. Both ALP activity (0.013 021±0.000 032) and Alizarin red quantification (0.290±0.003) were highest in the CMP-48 h group, significantly different from all other groups (all P<0.05). RUNX-2 expression in CMP-24 h and CMP-48 h groups (1.845±0.087 and 1.837±0.027, respectively) was significantly higher than in other groups (all P<0.05), with no significant difference between these two groups ( P>0.05). CMP-48 h group exhibited the highest mRNA expression of all osteogenic genes tested, specifically ALP (2.194±0.028), COL-1 (1.983±0.024), OCN (7.644±0.156), and OPN (2.648±0.031), all significantly greater than other groups (all P<0.05). Conclusions:Both curved microgrooves and surface roughness modification enhance the in vitro osteogenic differentiation of cells on PCL membranes. Among the tested strategies, alkali etching of curved microgrooves for 48 hours (CMP-48h) provided the optimal enhancement of osteogenic capability for MC3T3-E1 cells and represented a promising surface modification strategy for future PCL membranes.