Objective To develop a composite nutritional preparation that can effectively improve exercise performance under calorie restriction(CR)condition.Methods A total of 24 male C57BL/6J mice(weighing 23～26 g,8 weeks old)were randomly divided into control group(CON),CR,CR+basal nutrient group(CRN1),and CR+compound nutrient group(CRN2).All groups underwent moderate-intensity running training 5 d per week,for totally 3 weeks.The grip strength of the forelimbs were measured weekly,and in 3 weeks after training,exhaustion and post-exhaustion distance tests were conducted to evaluate exercise performance.Blood biochemical indicators,levels of skeletal muscle and liver redox biomarkers,and histopathological conditions were measured and observed.Results After 21 d of intervention,the CR group and CRN1 group had the post-exhaustion running distance prolonged by 278%and 289%,respectively,reduced blood glucose level,and decreased muscle mass,subcutaneous fat and epididymal fat mass when compared with the CON group(P<0.05).Compared with the CON group,the CRN1 and CRN2 groups demonstrated significantly higher gastrocnemius glycogen content.The CRN2 group obtained even longer post-exhaustion distance(increased by 52%and 36%respectively,compared with the CON group and CRN1 group,P<0.05),enhanced grip strength of the forelimbs(raised by 9%,17%and 15%,respectively than the CON,CR and CRN1 groups,P<0.05),elevated brown fat mass(compared to the CON group and CRN1 group,P<0.05),increased blood glucose level(compared to the CRN1 group,P<0.05),decreased blood low-density lipoprotein cholesterol level(compared to the CON and CR groups,P<0.05),and increased glutathione peroxidase content in the gastrocnemius muscle(compared to the CON group,P<0.05).Conclusion Supplementing with compound nutritional supplements in mice under CR can promote exercise performance,including improving fatigue recovery after exhaustive exercise and enhancing forelimb grip strength.

SARS-CoV-2 and its emerging variants continue to pose a significant global public health threat. The SARS-CoV-2 main protease (M^pro) is a critical target for the development of antiviral agents that can inhibit viral replication and transcription. In this study, we identified chebulagic acid (CHLA), isolated from Terminalia chebula Retz., as a potent non-peptidomimetic and non-covalent M^pro inhibitor. CHLA exhibited intermolecular interactions and provided significant protection to Vero E6 cells against a range of SARS-CoV-2 variants, including the wild-type, Delta, Omicron BA.1.1, BA.2.3, BA.4, and BA.5, with EC₅₀ values below 2 μmol/L. Moreover, in vivo studies confirmed the antiviral efficacy of CHLA in K18-hACE2 mice. Notably, CHLA bound to a unique groove at the interface between M^pro domains I and II, which was revealed by the high-resolution crystal structure (1.4 Å) of the M^pro-CHLA complex, shrinking the substrate binding pocket of M^pro and inducing M^pro aggregation. CHLA was proposed to act as an allosteric inhibitor. Pharmacokinetic profiling and safety assessments underscore CHLA's potential as a promising broad-spectrum antiviral candidate. These findings report a novel binding site on M^pro and identify antiviral activity of CHLA, providing a robust framework for lead compounds discovery and elucidating the underlying molecular mechanisms of inhibition.

Objective To detect diatoms in tissues and water samples from drowning cases by gene array,and explore the application value of DNA microarray.Methods Diatoms in the lung,liver,kidney samples,and on-site water samples from 19 drowned corpses that appeared in the rivers of Zhongshan City between July 2021 and April 2022,were detected by gene arrays.Moreover,diatom detection was also performed on 28 samples from 7 corpses by microscope.Then the testing results were compared and analyzed.Results For those 28 samples,diatom types in both tissue and water samples by gene detection were more than those detected by microscopy,the detection was statistically significant(P<0.05).However,there was no statistically significant difference(P>0.05)in the detection of diatom types between tissue samples and water samples both using gene arrays.In terms of tissue samples,diatoms were easier to be detected in lung than liver or kidney,and there were also more types of diatoms detected in lung.Diatom detection by gene arrays showed that the diatom types showed a decreasing trend as the tissue was further away from the entrance of body,and the difference of diatom types increased among tissues.Conclusion The gene array can effectively detect the diatom types and distribution characteristics in drowned corpses,and has great potential in the research of the traceability analysis of corpses found in water and the causes of death.

Objective To detect diatoms in tissues and water samples from drowning cases by gene array,and explore the application value of DNA microarray.Methods Diatoms in the lung,liver,kidney samples,and on-site water samples from 19 drowned corpses that appeared in the rivers of Zhongshan City between July 2021 and April 2022,were detected by gene arrays.Moreover,diatom detection was also performed on 28 samples from 7 corpses by microscope.Then the testing results were compared and analyzed.Results For those 28 samples,diatom types in both tissue and water samples by gene detection were more than those detected by microscopy,the detection was statistically significant(P<0.05).However,there was no statistically significant difference(P>0.05)in the detection of diatom types between tissue samples and water samples both using gene arrays.In terms of tissue samples,diatoms were easier to be detected in lung than liver or kidney,and there were also more types of diatoms detected in lung.Diatom detection by gene arrays showed that the diatom types showed a decreasing trend as the tissue was further away from the entrance of body,and the difference of diatom types increased among tissues.Conclusion The gene array can effectively detect the diatom types and distribution characteristics in drowned corpses,and has great potential in the research of the traceability analysis of corpses found in water and the causes of death.

Objective To evaluate the detection performance of common strains and the antimicrobial binding capacity of four blood culture systems made by BMX-FA/N Plus,Zhengzhou Anto,Zhuhai DIER and Chongqing Zhongyuan.Methods According to the common pathogens of clinical bloodstream infections in Wuhan No.1 Hospital,ATCC standard isolates of Staphylococcus aureus,Streptococcus pneumoniae,Enterococcus faecalis,Escherichia coli,Pseudomonas aeruginosa,Haemophilus influenzae,Stenotrophomonas Maltophil,Bacteroides tenuis and Candida glabla were chosen to explore.Prefabricated bacterial suspension and sterile horse blood were injected into different brands of blood culture systems to simulate the blood samples of patients with non-antibiotic treatment and antibiotic treatment.Six commonly used clinical antibiotics,Imipenem,Piperacillin/Tazobactam,Cefoperazone/Sulbactam,Levofloxacin,Vancomycin and Micafungine,were added to the blood samples after simulated antibiotic treatment.The performance was evaluated by recording the positive bottles and the detection time of each brand culture systems within five days with and without antibiotic.Results In the absence of antibiotic,four blood culture systems showed 100％recovery on all of the pathogens.Staphylococcus aureus,Pseudomonas aeruginosa and Haemophilus influenzae were recovered earlier in DIER aerobic bottles than BMX-FA Plus aerobic bottles,and the differences were statistically significant(t=2.608,5.547,12.247,all P＜0.05).The time to detection of Staphylococcus aureus,Streptococcus pneumoniae,Enterococcus faecalis and Haemophilus influenzae in Zhongyuan anaerobic bottles was significantly faster than that of BMX-FA Plus anaerobic bottles,with an average shortening of 1.92～10.80 h,and the differences were statistically significant(t=30.187,5.367,33.068,24.855,all P＜0.05).When antibiotics added,BMX-FA Plus culture bottle showed 100％recovery to all the detecting pathogens with the peak concentration antibiotics except Cefoperazone/Sulbactam,while the recovery in Zhongyuan blood culture bottle also was 100％with peak concentration antibiotics of Piperacillin/Tazobactam,Levofloxacin and Micafunzin.The peak concentration of imipenem antibiotics in domestic bottles was only detected in Anto anaerobic bottles,with lower positive detection rate(66.7％)lower than that of BMX-FA Plus(100％)and a later detection,and the difference was statistically significant(t=-21.000,P=0.030).Conclusion In the absence of antibiotic interference,the positive detection rate of the above pathogens are the same for four blood culture systems,and the time to detection of DIER and Zhongyuan systems is shorter than that of BMX-FA/N Plus.In the presence of antibiotic interference,the detection ability to pathogens in BMX-FA/N Plus system is the best,followed by domestic Zhongyuan system.

This paper summarizes Professor ZHANG Wei's experience in treating post-stroke dysphagia by using"unblocking pharyngeal orifice"grouping acupoints based on the concept of"harmonization and balance".Professor ZHANG Wei based on the theoretical of"the treatments should focus on where the meridians and collaterals pass through""the treatments should focus on where the acupoints are",target treatment,yin and yang balance,etc.,aiming at the basic pathogenesis of post-stroke dysphagia,that is,the obstruction of meridians and pharyngeal orifices,the obstruction of qi movement,as well as the fundamental pathogenesis of yin and yang imbalance and qi and blood disorder in stroke.She puts forward the concept of"harmonization and balance",followed the treatment principle of"unblocking the collaterals by opening and closing,relieving sore throat and opening the orifices,balancing yin and yang".The treatment characteristics of"selecting local acupoint and make direct action""taking its own advantages by selecting multi-channel acupoint""taking tongue,neck and body as a trinity acupuncture"and"combining supplementation and drainage to make a proper acupuncature therapy"have formed a unique"unblocking pharyngeal orifice"grouping acupoints.Professor ZHANG initially constructed a treatment plan combining tongue group acupoints,neck group acupoints and body group acupoints,and achieved unique effects in the clinical treatment of post-stroke dysphagia.

Objective To study the value of hemodynamics parameters,homocysteine(Hcy)and D-dimer levels in uterine artery during early pregnancy in patients with recurrent spontaneous abortion(RSA).Methods A total of 72 patients with RSA in Xuancheng People's Hospital from November 2020 to November 2022 were enrolled,and another 50 healthy women with normal results on prenatal ultrasonography were set as control group.Both groups underwent color Doppler ultrasonography and serum detection during early pregnancy,then parameters including pulsatile index(PI),ratio of peak flow velocity during systole to diastole(S/D),resistance index(RI),Hcy and D-dimer were compared between two groups.Logistic regression model was used to analyze the influencing factors of RSA occurrence,and receiver operating characteristic(ROC)curve was used to analyze the diagnostic value of each index to RSA.Results The values of PI,S/D,RI,Hcy and D-dimer in RSA group were higher than those in control group(P<0.05).The values of PI,S/D,RI,Hcy and D-dimer increased as risk factors for RSA(P<0.05).ROC curve denoted that the area under the curve(AUC)of PI,S/D,RI,Hcy and D-dimer dimerin the diagnosis of RSA were 0.778,0.826,0.819,0.778 and 0.756,respectively,and the AUC of combined detection of the above five parameters was 0.858,with a sensitivity of 75.0%and a specificity of 80.0%,indicating that the diagnostic value of joint detection was better than that of single detection.Conclusion The ascending uterine artery hemodynamic parameters,elevated Hcy and D-dimer level during early pregnancy are the risk factors for RSA patients,and the combined detection of all indicators has certain predictive value for RSA diagnosis.

Objective:To construct a novel dual-specific antibody targeting human CD123 (CD123 DuAb) and study its effects in acute myeloid leukemia (AML) .Methods:Based on the variable region of the CD123 monoclonal antibody independently developed at our institution, the CD123 DuAb expression plasmid was constructed by molecular cloning and transfected into ExpiCHO-S cells to prepare the antibody protein. Through a series of in vitro experiments, its activation and proliferation effect on T cells, as well as the effect of promoting T-cell killing of AML cells, were verified.Results:① A novel CD123 DuAb plasmid targeting CD123 was successfully constructed and expressed in the Expi-CHO eukaryotic system. ②The CD123 DuAb could bind both CD3 on T cells and CD123 on CD123 + tumor cells. ③When T cells were co-cultured with MV4-11 cells with addition of the CD123 DuAb at a concentration of 1 nmol/L, the positive expression rates of CD69 and CD25 on T cells were 68.0% and 44.3%, respectively, which were significantly higher than those of the control group ( P<0.05). ④Co-culture with CD123 DuAb at 1 nmol/L promoted T-cell proliferation, and the absolute T-cell count increased from 5×10 5/ml to 3.2×10 6/ml on day 9, and CFSE fluorescence intensity decreased significantly. ⑤ With the increase in CD123 DuAb concentration in the culture system, T-cell exhaustion and apoptosis increased. When the CD123 DuAb was added at a concentration of 1 nmol/L to the culture system, the proportion of CD8 + PD-1 + LAG-3 + T cells was 10.90%, and the proportion of propidium iodide (PI) - Annexin Ⅴ + T cells and PI + Annexin Ⅴ + T cells was 18.27% and 11.43%, respectively, which were significantly higher than those in the control group ( P<0.05). ⑥ The CD123 DuAb significantly activated T cells, and the activation intensity was positively correlated with its concentration. The expression rate of CD107a on T cells reached 16.05% with 1 nmol/L CD123 DuAb, which was significantly higher than that of the control group ( P<0.05). ⑦The CD123 DuAb promoted cytokine secretion by T cells at a concentration of 1 nmol/L, and the concentration of IFN-γ and TNF-α in the supernatant of the co-culture system reached 193.8 pg/ml and 169.8 pg/ml, respectively, which was significantly higher than that of the control group ( P<0.05). ⑧When CD123 DuAb was added at a concentration of 1 nmol/L to the co-culture system of T cells and CD123 + tumor cells, the killing intensity of T cells significantly increased, and the residual rates of CD123 + MV4-11 cells, CD123 + Molm13 cells, and CD123 + THP-1 cells were 7.4%, 6.7%, and 14.6% on day 3, respectively, which were significantly lower than those in the control group ( P<0.05) . Conclusion:In this study, a novel CD123 DuAb was constructed and expressed. In vitro experiments verified that the DuAb binds to CD123 + tumor cells and T cells simultaneously, promotes T-cell activation and proliferation, and facilitates their anti-leukemia effect, which provides a basis for further clinical research.

Objective:To construct a novel chimeric antigen receptor T (CAR-T) cell targeting CD138 and to investigate its cytotoxicity against myeloma cells.Methods:The hybridoma strain that can stably secrete the CD138 monoclonal antibody (mAb) was prepared and obtained through monoclonal antibody screening technology. The hybridoma strain cells were intraperitoneally injected into mice to produce ascites containing monoclonal antibodies, which were then collected and purified to obtain pure CD138 mAb. Further examinations were performed to assess the biological characteristics of CD138 mAb. The variable region sequence of this antibody was amplified through reverse transcription polymerase chain reaction and was used as the antigen recognition domain of CD138 CAR, which was subsequently expressed on the surface of T cells by lentiviral infection. Flow cytometry was employed to assess the phenotype of CD138 CAR-T cells. In vitro cytotoxicity and degranulation assays were performed to evaluate their antitumor effects.Results:① We successfully prepared anti-human CD138 antibody hybridoma cell lines and screened a hybridoma cell strain, 5G2, which could persistently and stably secrete the anti-CD138 antibody. ② The purified CD138 (5G2) mAb can especially recognize CD138 + cells with a binding affinity constant (K D) of 6.011×10 -9 mol/L and showed no significant binding activity with CD138 - cells. ③The variable region sequence of the CD138 (5G2) antibody was obtained using molecular cloning technology, and CD138 (5G2) CAR was successfully constructed and expressed on T cells through lentivirus infection and, concurrently, demonstrated effective binding to recombinant human CD138 protein.④ The proliferation of T cells transduced with the CD138 (5G2) CAR was highly efficient. The phenotype analysis revealed that CD138 (5G2) CAR-T cells exhibited a greater tendency to differentiate into central memory T cells and memory stem T cells, with a reduced proportion of terminally differentiated effector memory subsets. ⑤CD138 (5G2) CAR-T cells demonstrated specific cytotoxicity against CD138 + myeloma cell line H929, whereas CD138 - cell line K562 remained unaffected. The percentage of residual H929 cells was (12.92±8.02) % after co-culturing with CD138 (5G2) CAR-T cells, while (54.25±15.79) % was left in the Vector-T group (E∶T=1∶2; P<0.001). ⑥Results of degranulation assays demonstrated a significant activation of CD138 (5G2) CAR-T cells after co-culture with the H929 cell line, whereas no significant activation was observed in Vector-T cells [ (25.78±3.35) % vs (6.13±1.30) %, P<0.001]. ⑦After co-culturing with CD138 + cells, CD138 (5G2) CAR-T cells exhibited a significant increase in cytokine secretion compared to the Vector-T group [interleukin-2: (1 697.52±599.05) pg/ml vs (5.07±1.17) pg/ml, P<0.001; interferon-γ: (3 312.20±486.38) pg/ml vs (9.28±1.46) pg/ml, P<0.001; and tumor necrosis factor-α: (1 837.43±640.49) pg/ml vs (8.75±1.65) pg/ml, P<0.001]. However, no significant difference was observed in cytokine secretion levels between the two groups after co-culturing with CD138 - cells. Conclusion:This study successfully prepared a novel monoclonal antibody against CD138, and CAR-T cells constructed with the antigen recognition domain derived from this 5G2 mAb demonstrated effective antitumor activity against myeloma cells. This can be used as a new option for the detection of the CD138 antigen and proposes a novel strategy for multiple myeloma immunotherapy.

Animals ; SARS-CoV-2 ; Antibodies, Neutralizing ; Macaca mulatta ; COVID-19/prevention & control* ; Antibodies, Viral ; Vaccines