Objective: To identify an optimal back-flush solution for leukocyte-depleted filters that maximizes peripheral blood mononuclear cell (PBMC) recovery with high viability, long-term storage stability, and sterility of the harvested residues, thereby providing a clinically translatable strategy. Methods: Three sterile bag-packaged solutions—Saline, Solvent, and Hanks' balanced salt solution (HBSS)—were used to back-flush randomly assigned leukocyte-depleted filters. Nucleated cell recovery rate and viability of the harvested residues were compared. The optimal solution identified was applied to an expanded sample set. PBMC viability and yield were evaluated after 1h vs 48h storage of the residues. PBMCs isolated from the residues were cryopreserved in liquid nitrogen for 1 month, followed by post-thaw comparisons of viability and T-cell expansion capacity. Results: The Solvent group achieved the highest and most consistent nucleated cell recovery rate. Post-flush recovery rate from filters after 400 mL whole blood processing was (21.3±1.6)% for the Solvent group, significantly higher than Saline group (19.2±6.3)% and HBSS group (11.2±5.0)%, with residues from all groups maintaining viability >90%. No biologically significant difference in residue viability was observed between 48h vs 1h storage groups (93.3±2.3)% vs (95.7±1.8)%). PBMC recovery rates from residues showed no statistical difference between 48h vs 1h storage groups [(48.2%±9.5%)vs (40.41%±8.35%), P>0.05], with (17.7±2.6)×10 cells. After 1-month cryopreservation and 10-day expansion, PBMCs isolated from 48-hour-stored residues retained (91.2±3.2)% viability and achieved a (61.9±15.9)-fold expansion. Conclusion: The bag-packaged Solvent, as a back-flush solution, enables sterile acquisition of leukocyte-depleted filter residues through closed-system tubing connections. These residues maintained PBMC viability and recovery rates after 48h storage at 2℃-8℃, with post-cryopreservation (1-month liquid nitrogen) viability and expansion capacity remaining stable. This protocol complies with blood bank regulatory criteria, addresses the concerns about the infectious window period in cell therapy raw materials, and provides a clinically translatable strategy for PBMC-based applications.

ObjectiveTo investigate the distribution of traditional Chinese medicine （TCM） syndromes in intrahepatic cholestasis of pregnancy （ICP） and its association with perinatal outcomes， and to provide a basis for precise treatment based on TCM syndrome differentiation. MethodsA cross-sectional study was conducted among 275 patients with ICP who were admitted to The Affiliated Hospital of Chengdu University of Traditional Chinese Medicine from April 2023 to April 2025. A hierarchical cluster analysis was used to summarize TCM syndromes. The Kruskal-Wallis H test was used for comparison of non-normally distributed continuous data between groups， and the chi-square test was used for comparison of categorical data between groups. A multivariate Logistic regression analysis was used to identify the clinical features significantly associated with TCM syndrome. ResultsThe cluster analysis identified three core TCM syndromes among the 275 patients with ICP， i.e.， liver-gallbladder damp-heat syndrome （45.8%）， syndrome of blood deficiency generating wind （30.9%）， and liver depression and spleen deficiency syndrome （23.3%）. There was a significant difference in the distribution of TCM syndromes between different groups stratified by maternal age at delivery， parity， history of ICP recurrence， gestational weeks at disease onset， total bile acid （TBA）， alanine aminotransferase （ALT）， and comorbidity with gestational diabetes mellitus （GDM） （all P<0.05）. The multivariate Logistic regression analysis showed that<34 gestational weeks at disease onset was significantly associated with all three syndromes （damp-heat： odds ratio ［OR］=3.769， P<0.001； blood deficiency： OR=4.031， P<0.001； liver stagnation： OR=3.552， P<0.001）. Liver-gallbladder damp-heat syndrome was associated with maternal age ≥35 years at disease onset （OR=2.048， P=0.014）， parity ≥2 times （OR=1.921， P=0.034）， history of ICP recurrence （OR=2.404， P=0.030）， ALT ≥200 U/L （OR=2.051， P=0.018）， comorbidity with GDM （OR=1.944， P=0.029）， and TBA ≥40 μmol/L （OR=2.542， P=0.024）. The syndrome of blood deficiency generating wind syndrome was associated with maternal age ≥35 years （OR=2.939， P=0.003）， parity ≥2 time （OR=3.222， P=0.003）， history of ICP recurrence （OR=3.809， P=0.010）， ALT ≥200 U/L （OR=2.889， P=0.006）， comorbidity with GDM （OR=3.711， P=0.001）， and comorbidity with hypertensive disorders of pregnancy （OR=4.472， P=0.011）. Liver depression and spleen deficiency syndrome was associated with TBA ≥40 μmol/L （OR=2.995， P=0.044）. The analysis of perinatal outcomes showed that there were significant differences in mode of delivery， gestational weeks at the time of delivery， postpartum blood loss， and neonatal birth weight between the three groups with different TCM syndromes （all P<0.05）. ConclusionLiver-gallbladder damp-heat syndrome， syndrome of blood deficiency generating wind， and liver depression and spleen deficiency syndrome are the main TCM syndrome types in ICP， and the distribution of TCM syndromes is closely associated with clinical factors and perinatal outcomes， which provides a basis for precise TCM syndrome differentiation and individualized treatment.

The mechanism of Tibetan medicine Zuotangkaca pills (ZTKCW) for the treatment of hypertension was explored by network pharmacology and in vivo experiments. 68 active ingredients of ZTKCW and 518 drug-disease targets were screened by network pharmacology. Eight core components of ZTKCW (vasicolinone, luteolin, (–)-isocorypalmine, esculetin, liquiritigenin, etc.) and eight key targets (AKT1, TNF, IL6, and STAT3, etc.) were screened by network topology analysis. KEGG enrichment analysis showed that the core targets were mainly enriched in lipids and atherosclerosis, JAK/STAT, and inflammation-related pathways. An in vivo experiment was conducted using spontaneously hypertensive rats (SHR), which were gavaged with ZTKCW at doses of 0.41, 0.82, and 1.64 g/kg for 12 weeks, respectively. The results showed that ZTKCW at a dose of 1.64 g/kg significantly reduced both systolic and diastolic pressure in SHR rats and decreased the phosphorylation levels of AKT1, PI3K, STAT3, and JAK2 in the thoracic aorta and heart tissues. This study demonstrates that ZTKCW may exert its antihypertensive effects through PI3K/AKT and JAK2/STAT3 pathways, providing some insights and a theoretical basis for the use of ZTKCW in hypertension.

OBJECTIVES: To characterize the biological properties of double-negative T (DNT) cells isolated from leukoreduction filter residues. METHODS: Leukoreduction filters containing residues from 400 mL whole blood units (n=6) were collected from a blood center. Filters were back-flushed with normal saline, and the eluate was concentrated to obtain leukoreduction filter residues. Leukocytes in the residues were counted by dual-fluorescence staining. DNT cells were then isolated from the residues using antibody-mediated adsorption and density gradient centrifugation. Both cryopreserved and fresh unstimulated DNT cells derived from the residues were subjected to in vitro culture. Following culture, cells were assessed for expansion fold, viability, immunophenotype, differentiation status, and cytotoxicity against target cells using dual-fluorescence staining and flow cytometry, with comparisons made to DNT cells derived from whole blood. RESULTS: The leukocyte recovery rate achieved through reverse flushing of the leukocyte reduction filter was (41.9±14.7)%. Compared to whole blood, the DNT cell starting material obtained from filter residues showed no significant difference in total T-cell content (P>0.05). However, the viability and purity of the resulting DNT cell starting materials were significantly lower (both P<0.05). After 17 days of culture, DNT cells from filter residues and whole blood showed no significant differences in expansion fold, immunophenotype, differentiation status, or cytotoxicity toward target cells (all P>0.05). However, the viability of DNT cells from residues was significantly lower than that of whole blood-derived DNT cells [(86.0±4.2)% vs. (92.2±1.2)%, P<0.05]. After thawing (post 3 or 15 days of cryopreservation) and 17 days of culture, DNT cell starting materials from residues showed comparable immunophenotype, expansion fold, and differentiation status to their non-cryopreserved counterparts from the same source (all P>0.05). However, the viability of DNT cells cryopreserved for 3 days [92.4% (91.8%, 92.8%)] and the cytotoxicity against target cells of those cryopreserved for 15 days [91.3% (89.4%, 95.1%)] were significantly higher than those of non-cryopreserved DNT cells [87.8% (82.0%, 89.0%) and 70.9% (67.3%, 80.2%), respectively] (P<0.05). CONCLUSIONS DNT cells derived from leukoreduction filter residues exhibited highly comparable characteristics to those from whole blood in terms of expansion, purity, differentiation, and biological potency. Furthermore, their biological activity post-cryopreservation and revival remained largely similar to non-cryopreserved cells. These findings suggest that leukoreduction filter residues represent a promising alternative source of starting material for manufacturing off-the-shelf, allogeneic DNT cell therapeutics.

Alzheimer's disease (AD) is a common neurodegenerative disorder among the elderly, and BuChE has emerged as a potential therapeutic target. In this study, we reported the development of compound 8e, a selective reversible BuChE inhibitor (eqBuChE IC₅₀ = 0.049 μmol/L, huBuChE IC₅₀ = 0.066 μmol/L), identified through extensive virtual screening and lead optimization. Compound 8e demonstrated favorable blood-brain barrier permeability, good drug-likeness property and pronounced neuroprotective efficacy. Additionally, 8e exhibited significant therapeutic effects in zebrafish AD models and scopolamine-induced cognitive impairments in mice. Further, 8e significantly improved cognitive function in APP/PS1 transgenic mice. Proteomics analysis demonstrated that 8e markedly elevated the expression levels of very low-density lipoprotein receptor (VLDLR), offering valuable insights into its potential modulation of the Reelin-mediated signaling pathway. Thus, compound 8e emerges as a novel and potent BuChE inhibitor for the treatment of AD, with significant implications for further exploration into its mechanisms of action and therapeutic applications.

The PA-PB1 interface of the influenza polymerase is an attractive site for antiviral drug design. In this study, we designed and synthesized a mini-library of indazole-containing compounds based on rational structure-based design to target the PB1-binding interface on PA. Biological evaluation of these compounds through a viral yield reduction assay revealed that compounds 27 and 31 both had a low micromolar range of the half maximal effective concentration (EC₅₀) values against A/WSN/33 (H1N1) (8.03 μmol/L for 27; 14.6 μmol/L for 31), while the most potent candidate 24 had an EC₅₀ value of 690 nM. Compound 24 was effective against different influenza strains including a pandemic H1N1 strain and an influenza B strain. Mechanistic studies confirmed that compound 24 bound PA with a K _d which equals to 1.88 μmol/L and disrupted the binding of PB1 to PA. The compound also decreased the lung viral titre in mice. In summary, we have identified a potent anti-influenza candidate with potency comparable to existing drugs and is effective against different viral strains. The therapeutic options for influenza infection have been limited by the occurrence of antiviral resistance, owing to the high mutation rate of viral proteins targeted by available drugs. To alleviate the public health burden of this issue, novel anti-influenza drugs are desired. In this study, we present our discovery of a novel class of indazole-containing compounds which exhibited favourable potency against both influenza A and B viruses. The EC₅₀ of the most potent compounds were within low micromolar to nanomolar concentrations. Furthermore, we show that the mouse lung viral titre decreased due to treatment with compound 24. Thus our findings identify promising candidates for further development of anti-influenza drugs suitable for clinical use.

Human carboxylesterase 2A (hCES2A) plays pivotal roles in prodrug activation and hydrolytic metabolism of ester-bearing chemicals. Targeted inhibition of intestinal hCES2A represents a feasible strategy to mitigate irinotecan-triggered gut toxicity (ITGT), but the orally active, selective, and efficacious hCES2A inhibitors are rarely reported. Here, a novel drug-like hCES2A inhibitor was developed via three rounds of structure-based drug design (SBDD) and structural optimization. Initially, donepezil was identified as a moderate hCES2A inhibitor from 2000 US Food and Drug Administration (FDA)-approved drugs. Following two rounds of SBDD and structural optimization, a donepezil derivative (B7) was identified as a strong reversible hCES2A inhibitor. Subsequently, nine B7 carbamates were rationally designed, synthesized and biologically assayed. Among all synthesized carbamates, C3 showed the most potent time-dependent inhibition on hCES2A (IC₅₀ = 0.56 nmol/L), excellent specificity and favorable drug-like properties. C3 could covalently modify the catalytic serine of hCES2A with high selectivity, while this agent also showed favorable safety profiles, high intestinal exposure, and impressive effects for ameliorating ITGT in both human intestinal organoids and tumor-bearing mice. Collectively, this study showcases a rational strategy for developing drug-like and serine-targeting covalent inhibitors against target serine hydrolase(s), while C3 emerges as a promising orally active drug candidate for ameliorating ITGT.

Objective To design a positive and negative pressure sequential irrigation suction device and explore its efficacy when applied to unblocking nasoenteric tubes.Methods The device consisted of a positive-pressure irrigation component,a negative-pressure suction component,a three-way adapter and a precise unblocker.The positive-pressure irrigation compo-nent was composed of a positive-pressure irrigation unit,a positive pressure regulator,a pressure sensor,a pressure bag and etc;the negative-pressure suction component was made up of a negative-pressure suction unit,a negative pressure regulator,a negative-pressure gauge,a negative-pressure bottle and etc.The pressure bag was hollow cylinder-shaped,and made of ordinary polyvinyl chloride for its inner layer and acrylonitrile-styrene-butadiene copolymer for its outer layer.Sixty patients with nasoenteric tube blockage were selected and randomly divided into a control group and an observation group,with 30 cases in each group.Irrigation suction was carried out with a 10 mL syringe for the control group and with the device for the observation group,and then unblocking was performed with lukewarm water between 38 and 40 degrees.The two groups were compared in terms of the unblocking efficacy,time required for unblocking and the days and cost of nutritional support.SPSS 22.0 software was used for statistical analysis.Results The observation group gained advantages over the control group in terms of the total effective rate(93.3％vs 73.4％),unblocking efficacy,time required for unblocking((7.63±4.97)min vs(10.73±4.57)min),days of nutritional support((10.07±3.07)d vs(13.03±4.05)d)and cost of nutritional support((3 180±260)yuan vs(4 550±660)yuan).Conclusion The device developed effectively enhances the efficacy for unblocking nasoenteric tubes,shortens the time required and decreases the days and cost of nutritional support,and thus is worthy promoting practi-cally.[Chinese Medical Equipment Journal,2025,46(1):114-117]

Purpose To explore the ultrasound characteristics of female breast cancer patients with different molecular types,and improve the diagnostic value of ultrasound.Materials and Methods The ultrasound diagnostic data of breast cancer patients in Xijing Hospital,the Air Force Medical University from January 2021 to July 2023 were retrospectively analyzed,and the ultrasound diagnosis of breast cancer patients from the aspects of molecular typing and related human epidermal growth factor receptor-2(HER-2)expression and Ki-67 expression were analyzed.Results A total of 1 405 patients were included in this study,including 240 patients with Luminal A type,631 patients with Luminal B type,162 patients with HER-2+hormone receptor(HR)-type,207 patients with HER-2+HR+type and 165 patients with triple-negative breast cancer.Among the ultrasound characteristics of female breast cancer with different molecular classifications,there were statistical differences in breast imaging reporting and data system classification,edge angulation,spiculation,posterior echo and calcification(χ2=13.652-143.178,all P<0.05).According to the analysis of different degrees of invasion,there were statistical differences in breast imaging reporting and data system classification,edge angulation,spiculation,posterior echo and calcification from the Lumina group(Luminal A,Luminal B)and the aggressive group(HER-2+HR-,HER-2+HR+and triple-negative)(χ2=4.702-17.587,all P<0.05).The HER-2-negative and HER-2-positive groups of patients had significant differences in shape,edge angulation,spiculation and calcification,with statistical differences(χ2=3.947-7.944,all P<0.05).According to the Ki-67 level analysis,it was found that edge angulation,spiculation and posterior echo were influencing factors,with statistical differences(χ2=9.309-56.380,all P<0.05).Conclusion Breast cancer with different molecular expression has its own ultrasound characteristics.Preoperative ultrasound can provide a certain reference basis for the diagnosis,treatment and prognosis assessment of breast cancer.

Objective To observe the value of automated fiber quantification(AFQ)for analyzing relationships of abnormal white matter microstructures and executive dysfunction in children with attention deficit hyperactivity disorder(ADHD).Methods Brain diffusion tensor imaging(DTI)and white matter structure AFQ analysis were prospectively performed on 83 children with ADHD(ADHD group)and 52 healthy children(healthy control[HC]group).The axial diffusivity(AD),radial diffusivity(RD),mean diffusivity(MD),and fractional anisotropy(FA)of complete fiber bundles and respective segments were obtained and compared between groups.Partial correlation analysis was conducted of fiber bundle diffusion parameters and the scores of Swanson Nolan and Pelham-Ⅳ(SNAP-Ⅳ)and behavior rating inventory of executive function(BRIEF)in ADHD group.Receiver operating characteristic curves were applied to assess the efficacy of each diffusion parameter for evaluating ADHD.Results AD and MD of right anterior thalamic radiation(ATR),and AD of right cingulum bundle(CB)in ADHD group were all higher than those in HC group(FWE corrected,all P＜0.000 7).Significant differences of AD of the 54-78 segments of right ATR,MD of the 13-24 and 53-77 segments of right ATR,AD of the 24-26,53-56 and 93-97 segments of right CB,as well as MD of the 72-85 segments of left inferior longitudinal fasciculus(ILF)were found between groups(FWE corrected,all P＜0.001).In ADHD group,AD of 54-78 segments of right ATR were negatively correlated with the attention deficit scores,while of 24-26 segments of right CB were negatively correlated with the emotional control scores.MD of right ATR and some segments,AD of right CB and some segments in ADHD group were positively correlated with the scores of task initiation,planning,organization,monitoring and metacognitive index(all P＜0.05).The area under the curve(AUC)of segmental fiber bundle diffusion parameter and intact fiber bundle diffusion parameter for evaluating ADHD was 0.881 and 0.808,respectively,the former was higher than the latter(Z=2.435,P＜0.05).Conclusion AFQ could be used to analyze relationships of abnormal white matter microstructures and executive dysfunction in ADHD,which could be effectively evaluated based on specific fiber bundle segments.