3CL protease (3CLpro) of SARS-CoV-2 is a pivotal enzyme required in coronavirus replication and transcription. Its highly conserved structure and the absence of homologous proteins in the host make it an ideal target for broad-spectrum anti-coronavirus drug development. In this work, we systematically investigated and compared the binding modes and dynamic properties of the four stereoisomers of a pyridyl-urea diyne ester (PyDU) molecule with two chiral centers within the 3CLpro active site. Through molecular docking, MD simulations, MM/GBSA binding free-energy calculations, and DCCM analysis, all four stereoisomers were stabilized primarily by hydrophobic packing. Among them, the (R,S) isomer exhibited the best overall performance, including docking score, binding free-energy components, and key residue interactions. The (R,S) and (S,R) isomers also enhanced the cooperative motions around the binding pocket, while the (R,S) isomer further modulated the flexibility of domain III, which may influence 3CLpro dimerization. Conversely, the (S,S) isomer exhibited the weakest affinity due to insufficient hydrophobic contact. By innovatively integrating chirality, binding energy and protein dynamical features, we revealed the dual role of chirality in modulating affinity and dynamic responses, which provides a theoretical basis for the chiral-guided design of coronavirus inhibitors.

Objective To gain insights into the biological characteristics of different populations of Culex tritaeniorhynchus within and around the Yellow Sea wetland from the perspective of the occurrence of resistance, we investigated the levels of resistance to deltamethrin and kdr gene mutation in the wetland and its peripheral areas. Methods Specimens were collected from Cx. tritaeniorhynchus populations at two monitoring sites in the Rare Bird National Nature Reserve and Tiaozi Ni Wetland Scenic Area, and also from two populations in Yancheng City and the Liuhe District of Nanjing, and the resistance of these mosquitoes to deltamethrin was determined using the CDC biotest bottle method. For each concentration of deltamethrin assessed, a random subset of exposed specimens was selected for amplification of the kdr gene fragment, followed by Sanger sequencing to identify and analyze resistance-associated mutations. Results The LC50 levels of deltamethrin among mosquitoes from the four populations in Luhe, Yancheng, the Rare Bird National Nature Reserve and the Tiaozi Ni Wetland Scenic Area were 2.048 5, 7.798 2, 3.473 3, and 17.695 5 mg/mL, respectively, with corresponding concentrations of deltamethrin ranging from 0.005 to 5.000,0.050 to 50.000,0.050 to 25.000 and 0.050 to 50.000 mg/mL, respectively. Furthermore, the ranges of the KT50 values were 11.76-107.43, 67.05-216.30,29.77-107.43 and 28.40-329.51 min; the 1-h knockdown rates were 34.58%-99.15%, 9.52%-43.80%, 55.09%-73.01%, and 10.09%-68.07%; and the 24-h mortality rates were 12.15%-67.52%,9.52%-79.56%,13.17%-82.21%, and 11.01%-78.99%, respectively. With respect to kdr gene mutation, we assayed a total of 63,70,59, and 57 mosquitoes for the four populations, for which we detected L1014F mutation frequencies of 14.29%, 35.00%, 20.34%, and 31.58%, respectively, with a majority of these mutations being heterozygous for resistance. In addition, five adult mosquitoes were identified has having synonymous mutations at site 1011[i. e. , AAT(asparagine)mutation to AAC(asparagine)]. Conclusions Our findings revealed the clear resistance of Cx. tritaeniorhynchus to deltamethrin in the Yancheng region of the Yellow Sea wetland, and the resistance phenotype and kdr frequency of Cx. tritaeniorhynchus in the wetland environment were comparable to those of Cx. tritaeniorhynchus in the wetland environment, thereby indicating that the resistance of different populations of Cx. tritaeniorhynchus was homogeneous under the pressure of different insecticide selection within and around the wetland. However, the underlying mechanisms need to be further studied.

ObjectiveTo determine the effects of tuina at "Weizhong （BL 40）" on mechanical pain hypersensitivity in a rat model of chronic dorsal root ganglion （CCD）， and to preliminarily explore the anatomical distribution characteristics of neural afferents associated with "Weizhong （BL 40）" acupoint and their central projections. MethodsThirty SD rats were randomly assigned to a sham operation group， a model group， and a tuina group， with 10 rats in each group. In the model group and the tuina group， a chronic compression of CCD model was established by implanting stainless-steel rods into the right L4/5 and L3/4 intervertebral foramina. Rats in the sham operation group underwent surgical exposure only. From day 4 to day 28 after modeling， rats in the tuina group received daily manipulation at the right "Weizhong （BL 40）" （pressure： 5 N； frequency： 2 Hz； duration： 10 min/session）. Rats in the sham operation group and the model group were subjected only to fixation. Mechanical withdrawal threshold （MWT） was assessed before modeling and on day 3， 7， 14， 21， and 28 after modeling. On day 29 after modeling， pseudorabies virus （PRV）， a retrograde trans-multisynaptic tracer， was injected into the right "Weizhong （BL 40）" acupoint in rats from the model group. After 120 h， rats were perfusion-fixed through the heart， and local tissues surrounding the "Weizhong （BL 40）" acupoint， corresponding lumbar dorsal root ganglia， lumbar spinal cord， and brain tissues were collected for fluorescence imaging of PRV-positive signals. ResultsCompared to the sham operation group， rats in the model group and the tuina group showed significantly reduced MWT on day 3， 7， 14， 21， and 28 after modeling （P＜0.05）. Compared to the model group， rats in the tuina group exhibited significantly increased MWT on day 21 and 28 after modeling （P＜0.05）. Fluorescence imaging revealed PRV-positive signals in the deep fascial layer of local muscle tissue at "Weizhong （BL 40）"， the L4 and L5 dorsal root ganglia， the dorsal horn and central canal region of the spinal cord， as well as the primary somatosensory cortex （S1）， secondary motor cortex （M2）， and anterior cingulate cortex （ACC）. Among the examined cortical regions， M2 contained the largest number of PRV-positive neurons. ConclusionTuina at "Weizhong （BL 40）" alleviates mechanical hyperalgesia in rats with the chronic compression of CCD model. A traceable neuroanatomical connection exists between the linking deep tissues within "Weizhong （BL 40）" region and the corresponding lumbar dorsal root ganglia， the spinal dorsal horn， and specific cortical areas.

Objective:To explore the value and related mechanism of preoperative serum sialic acid (SA) on evaluating microvascular invasion (MVI) in patients with intrahepatic cholangiocarcinoma (ICC).Methods:A total of 91 patients who underwent surgical resection and were pathologically diagnosed with ICC from December 2020 to September 2024 at the Oriental Hepatobiliary Surgery Hospital affiliated to the Naval Medical University were included in this retrospective analysis. The patients were divided into non-MVI (41 cases) and MVI groups (50 cases). The general data and laboratory examination indexes were collected and analyzed. Univariate and multivariate logistic regression analyses were performed to identify independent risk factors for predicting MVI. The predictive value of serum indicators for MVI was evaluated by receiver operating characteristic curves. The correlation between MVI and SA was analyzed by point-biserial correlation. ICC cells stably overexpressing β-galactoside α2, 6-sialyltransferase 1 (ST6GAL1) were generated through lentiviral transfection. ST6GAL1 protein expression and mRNA expression were detected by Western blot and quantitative real-time polymerase chain reaction, respectively. Sambucus nigra (SNA) lectin fluorescence staining was used to detect α2, 6-sialylation levels on cells. Cell migration ability was assessed by wound healing and Transwell assays, and cell proliferation was evaluated by colony formation assays.Results:Compared with the non-MVI group, patients in the MVI group exhibited significantly higher levels of fibrinogen, aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, SA and 5′-nucleotidase (5′-NT) (all P<0.05). Multivariate logistic regression analysis revealed that SA ( OR=1.01,95% CI 1.01-1.02, P=0.023) was the only independent predictor for MVI. The area under curve of SA in predicting MVI was 0.757 (95% CI 0.640-0.870), sensitivity 67.65%, specificity 77.78%. SA was positively correlated with MVI ( r=0.443, P<0.001). ICC cells overexpressing ST6GAL1 were featured with increased mean fluorescence intensity of SNA lectin, and increased level of α2, 6-sialylation on the cell surface (both P<0.05). The number of colonies formed by hypersialylated ICC cells was also increased ( P<0.05), and both the migration rate and the number of migrating cells were significantly higher ( P<0.05). Conclusions:Serum SA is an independent predictor for MVI in ICC patients. Hypersialylation in ICC cells is associated with higher malignancy.

Objective To construct a graded nursing program for exercise rehabilitation of children with hematopoietic stem cell transplantation(HSCT)and carry out a preliminary application,aiming to provide a basis for healthcare personnel to guide the exercise rehabilitation of children with HSCT.Methods Literature analysis and Delphi expert correspondence were used to construct a graded nursing program of exercise rehabilitation for children with HSCT.Through the convenience sampling method,children with HSCT in the hematology department of a tertiary-level children's hospital in Suzhou City were selected as the study subjects from June-December 2023,of which 26 children with HSCT from September-December 2023 served as an experimental group,and 23 children with HSCT from June-August 2023 as a control group.The experimental group was applied with the graded nursing program of exercise rehabilitation,and the control group was applied with the conventional nursing care.The comparison of cardiorespiratory fitness and adverse events between the 2 groups after 8 weeks of exercise intervention was conducted,and the adherence to the exercise intervention programme in the experimental group was recorded.Results The final constructed graded care program of exercise rehabilitation for children with HSCT included 6 primary indicators,20 secondary indicators,and 44 tertiary indicators.Finally,21 children in the experimental group and 23 children in the control group were included in the study.After the intervention,the 6 min walking distance of the experimental group increased compared with that of the control group,and the difference was statistically significant(P＜0.001).85.71％of children in the experimental group had good exercise compliance,and there was no adverse events in both groups.Conclusion The constructed graded nursing program of exercise rehabilitation for children with HSCT is scientific,reasonable,safe and feasible,which can improve the cardiorespiratory capacity of children with HSCT and provide the basis for their exercise rehabilitation.

Nitrogen oxides(NOx=NO+NO2)are important precursors of ozone(O3),and NOx and its oxides together constitute reactive nitrogen oxides(NOy)in the atmosphere.A comprehensive understanding of the total NOy level in the atmosphere is of great significance for a deeper understanding of the atmospheric nitrogen cycle and oxidation,as well as for formulating strategies for air pollution prevention and control.In this work,a thermal decomposition-broadband cavity enhanced absorption spectroscopy(TD-BBCEAS)technique for online measurement of total NOy in the atmosphere was developed.With this method,the NOy was efficiently converted into NO2,and the total NOy concentration in the atmosphere was indirectly obtained by measuring NO2.Focusing on the key factors affecting the measurement of total NOy,the influence of NO titration efficiency and other NOy component TD efficiency on measurement accuracy was emphasized.By changing the oxygen(O2)flow rate through the mercury lamp to alter the O3 concentration for titrating NO,the conversion efficiency of NO was evaluated.At O2 flow rate of 6 mL/min,the conversion efficiency of NO was greater than 99%.TD efficiency testing and analysis on NO2,peroxyacetyl nitrate(PAN),nitric acid(HNO3),and nitrous acid(HONO),which account for a large proportion of atmospheric NOy components,was carried out using 680℃as the optimal TD temperature for efficient conversion of NOy.With NO and HONO sample gases as typical verification gases,the conversion efficiency of NOy and the accuracy of NOy measurement by TD-BBCEAS system were verified by switching the on and off modes of mercury lamp and TD device.At integration time of 60 s,the detection limit of the system for NOy was 2.83×1010 molecules/cm3(60 s,2σ).A comparative measurement of actual atmospheric NOy was conducted between the TD-BBCEAS system and the NOy analyzer.The observation results showed a correlation coefficient(R2)of 0.98 and a slope of 0.93,further verifying the feasibility and accuracy of applying the TD-BBCEAS system to measurement of total NOy.

Objective:To analyze the clinical application of protocol biopsy (PB) during postoperative follow-up in recipients of allogeneic kidney transplantation.Methods:A prospective observational cohort study was conducted. Recipients who underwent allogeneic kidney transplantation at the Second Affiliated Hospital of Nanjing Medical University between January 2022 and September 2024 and received PB at 3 months (±1 week) and/or 12 months (±4 weeks) post-transplantation in the Department of Nephrology were enrolled. The implementation, complications, and pathological results of PB were summarized. The safety and diagnostic efficacy of PB were analyzed.Results:A total of 143 allogeneic kidney transplant recipients were included, and 200 PB procedures were performed. The overall implementation rate of protocol biopsy (PB) was 84.1% (143/170). Among them, 170 recipients completed 3-month follow-up, and 136 PBs were performed at 3 months (±1 week) post-transplantation, with an implementation rate of 80.0%. Seventy-nine recipients completed 12-month follow-up, and 64 PBs were performed at 12 months (±4 weeks), with an implementation rate of 81.0%. One major PB-related complication occurred (0.5%), presenting as gross hematuria and diagnosed as a transplant renal arteriovenous fistula. At 3 months post-transplantation, 58 biopsies (42.6%) showed pathological abnormalities, including rejection in 12 cases (8.8%), borderline changes in 18 cases (13.2%), BK virus nephropathy (BKVN) in 10 cases (7.4%), calcineurin inhibitor (CNI) nephrotoxicity in 13 cases (9.6%), and recurrent kidney disease in 5 cases (3.7%). At 12 months post-transplantation, 22 biopsies (34.4%) revealed pathological abnormalities, including rejection in 13 cases (20.3%), borderline changes in 4 cases (6.3%), BKVN in 3 cases (4.7%), CNI nephrotoxicity in 1 case (1.6%), and recurrent disease in 1 case (1.6%).Conclusions:Protocol kidney allograft biopsy after allogeneic kidney transplantation is highly safe and feasible in clinical practice. PB provides significant diagnostic value for the early detection of subclinical rejection and BKVN, thereby supporting its clinical utility in postoperative monitoring and management.

Objective To systematically investigate biomechanical characteristics of one-finger Zen push method applied at the Fengchi acupoint under different force conditions,in order to provide precise quantitative data and enhance the treatment efficacy.Methods Ten senior Tuina practitioners were recruited.The German Novel Pliance-X 32 Expert dynamic pressure distribution system was used to record the mechanical parameters during the application of one-finger Zen push at the Fengchi acupoint under three force intensities:light,medium,and heavy,for 3 minutes.Data from the stable 1-minute segment of the mechanical output was selected for data analysis,and key biomechanical parameters such as the maximum force,average force,peak pressure,mean pressure,force-time integral(FTI),pressure-time integral(PTI),and operational frequency were evaluated.Results Under light,medium,and heavy force conditions,the mean maximum force applied by senior Tuina practitioners at the Fengchi acupoint were 6.31,9.45,and 18.27 N,respectively,while the mean force were 3.31,5.64,and 9.05 N,respectively.The mean peak pressures were 26.10,34.80,and 70.00 kPa,while the mean pressures were 11.95,21.00,and 26.15 kPa,respectively.The mean FTIs were 55.65,182.10,and 225.21 N·s,and the mean PTIs were 167.10,489.59,and 795.83 kPa·s,respectively.The mean operational frequencies were 156.00,150.60,and 154.80 times/min,respectively.Conclusions Ten senior Tuina practitioners showed a high degree of consistency between their subjective definitions of light,medium,and heavy force and the objectively measured mechanical parameters.This reflected their precise control over the applied force under different force conditions,verifying the practicality and reproducibility of the one-finger Zen push method in clinical applications.This study provides a reliable basis for quantitative research and development of standardized clinical operation guidelines.

Objective:To analyze the clinical application of protocol biopsy (PB) during postoperative follow-up in recipients of allogeneic kidney transplantation.Methods:A prospective observational cohort study was conducted. Recipients who underwent allogeneic kidney transplantation at the Second Affiliated Hospital of Nanjing Medical University between January 2022 and September 2024 and received PB at 3 months (±1 week) and/or 12 months (±4 weeks) post-transplantation in the Department of Nephrology were enrolled. The implementation, complications, and pathological results of PB were summarized. The safety and diagnostic efficacy of PB were analyzed.Results:A total of 143 allogeneic kidney transplant recipients were included, and 200 PB procedures were performed. The overall implementation rate of protocol biopsy (PB) was 84.1% (143/170). Among them, 170 recipients completed 3-month follow-up, and 136 PBs were performed at 3 months (±1 week) post-transplantation, with an implementation rate of 80.0%. Seventy-nine recipients completed 12-month follow-up, and 64 PBs were performed at 12 months (±4 weeks), with an implementation rate of 81.0%. One major PB-related complication occurred (0.5%), presenting as gross hematuria and diagnosed as a transplant renal arteriovenous fistula. At 3 months post-transplantation, 58 biopsies (42.6%) showed pathological abnormalities, including rejection in 12 cases (8.8%), borderline changes in 18 cases (13.2%), BK virus nephropathy (BKVN) in 10 cases (7.4%), calcineurin inhibitor (CNI) nephrotoxicity in 13 cases (9.6%), and recurrent kidney disease in 5 cases (3.7%). At 12 months post-transplantation, 22 biopsies (34.4%) revealed pathological abnormalities, including rejection in 13 cases (20.3%), borderline changes in 4 cases (6.3%), BKVN in 3 cases (4.7%), CNI nephrotoxicity in 1 case (1.6%), and recurrent disease in 1 case (1.6%).Conclusions:Protocol kidney allograft biopsy after allogeneic kidney transplantation is highly safe and feasible in clinical practice. PB provides significant diagnostic value for the early detection of subclinical rejection and BKVN, thereby supporting its clinical utility in postoperative monitoring and management.

Objective:To investigate the use of non-vitamin K antagonist oral anticoagulants(NOACs)and their associated comorbidities in patients aged 80 years and older with non-valvular atrial fibrillation(NVAF), as well as to understand the challenges faced by elderly patients receiving NOAC therapy.Methods:We retrospectively enrolled elderly patients(≥80 years old)with NVAF who were treated with NOACs at a hospital in Beijing from January 2018 to August 2023.Patients were categorized into two age groups: 80-89 years and ≥90 years.We collected baseline data, including demographic characteristics, details of atrial fibrillation, comorbidities, laboratory test results, and medication combinations, for descriptive statistical analysis and intergroup comparisons.Results:A total of 695 elderly patients with NVAF receiving NOACs were included in the study, with a median age of 84 years.Among these patients, there were 328 males(47.19%, 328/695)and 422 cases of paroxysmal atrial fibrillation(60.72%, 422/695).The age group of 80-89 years comprised 640 cases(92.09%, 640/695), while the group aged 90 years and above included 55 cases(7.91%, 55/695).The use of NOACs in patients aged 90 and older exhibited an increasing trend over the years.Inter-group comparisons indicated that the ≥90 years group had lower body mass index, longer hospital stays, increased bedridden time, poorer renal function, lower levels of albumin and hemoglobin, and higher D-dimer levels.Inappropriate dosing of DOACs occurred in 49.64%(345/695)of cases, with 90.72%(313/345)receiving doses lower than recommended.Lower-than-recommended doses were more prevalent in the ≥90 years group, while higher-than-recommended doses were more common in the 80-89 years group.Polypharmacy was noted in 61.29%(426/695)of patients.The concurrent use of antiplatelet drugs, rhythm control medications, and ventricular rate control drugs was observed in 12.52%(87/695), 19.57%(136/695), and 54.53%(379/695)of patients, respectively, with no significant differences between groups.Conclusions:Inappropriate dosing and polypharmacy are prevalent issues among elderly NVAF patients.Therefore, it is essential to enhance multidisciplinary collaboration to optimize anticoagulation treatment strategies.