ObjectiveTo observe the clinical efficacy and safety of Bushen Ruyan Formulation （补肾乳岩方， BRF） in treating premenopausal breast cancer patients of disharmony of the chong （冲） and ren （任） meridian type under endocrine intensive therapy. MethodsA total of 88 premenopausal breast cancer patients who received endocrine intensive therapy and were diagnosed with disharmony of the chong and ren meridian by traditional Chinese medicine （TCM） were included and randomly divided into a control group and a treatment group， with 44 cases in each group. The control group received ovarian function suppression （OFS） combined with endocrine therapy， while the treatment group was given oral BRF additionally. Both groups were treated for three months. The clinical efficacy was evaluated by comparing the pre- and post-treatment results of the Functional Assessment of Cancer Therapy-Breast （FACT-B）， modified Kupperman score， T cell subsets （CD3⁺， CD4⁺， CD8⁺， CD4⁺/CD8⁺）， sex hormone levels， including estradiol （E₂）， luteinizing hormone （LH）， follicle-stimulating hormone （FSH）， progesterone （P）， testosterone （T）， and prolactin （PRL）， tumor markers， such as carcinoembryonic antigen （CEA）， carbohydrate antigen 125 （CA125）， and carbohydrate antigen 153 （CA153）， TCM syndrome score， and TCM syndrome efficacy. Blood routine tests， liver function and kidney function were monitored to assess safety. ResultsThe FACT-B scores of each domains and the total scores of the treatment group increased， while the social/family status score of the control group decreased. The treatment group had significantly higher scores in all domains and total score compared to the control group （P＜0.05 or P＜0.01）. The modified Kupperman score and TCM syndrome score of the treatment group decreased， and were lower than those of the control group （P＜0.01）. The levels of CD3⁺ and CD4⁺ in the treatment group increased， and were higher than those of the control group （P＜0.05）. Serum testosterone levels in the treatment group were lower than those in the control group （P＜0.05）. The total effective rate of TCM syndrome efficacy in the treatment group was 67.44%， significantly higher than the 23.26% in the control group （P＜0.05）. No significant abnormalities were observed in the blood routine tests or liver/kidney function indicators in either group before or after treatment. ConclusionBRF can effectively improve quality of life， alleviate symptoms， increase serum CD3⁺ and CD4⁺ levels， and enhance clinical efficacy in premenopausal breast cancer patients undergoing endocrine intensive endocrine therapy. It is also safe with no significant adverse effects.

Objective:This study aims to develop a rapid identification technique for various genotypes of extended-spectrum β-lactamase (ESBL) producing Escherichia coli using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) in conjunction with machine learning algorithms. Methods:A total of 158 Escherichia coli strains were isolated from the clinical laboratory of the First Hospital of Jilin University from August 2018 to December 2022. Polymerase chain reaction (PCR) was employed to detect the CTX-M-1, CTX-M-8, CTX-M-9, and SHV genes. Mass spectral data of the bacterial strains were acquired by MALDI-TOF MS with a cooperative matrix of (E)-propyl α-cyano-4-hydroxycinnamate (CHCA-C3). Models based on random forest (RF), logistic regression (LR), and support vector machine (SVM) algorithms were constructed. The performance of the constructed models was evaluated using metrics including accuracy, sensitivity, specificity, and the area under the receiver operating characteristic curve (AUC). Mass spectral peaks exhibiting sensitivity and specificity exceeding 80% in the models were designated as characteristic peaks. To validate the efficacy of the cooperative matrix of CHCA-C3, clinical isolates of ESBL-producing Escherichia coli were analyzed by MALDI-TOF MS using the conventional CHCA matrix for comparative purposes. Results:Among the 158 strains of Escherichia coli, 91 strains produced ESBL, all of which were CTX-M genotype. The AUC values for the respective models were as follows: CTX-M-1 genotype exhibited AUC values of 0.98 for LR, 1.00 for RF, and 0.73 for SVM; CTX-M-9 genotype exhibited AUC values of 0.93 for LR, 0.99 for RF, and 0.76 for SVM; for CTX-M-8, all models achieved an AUC of 1.00, indicating excellent classification performance with respect to accuracy, specificity, and sensitivity. The characteristic mass spectral peaks associated with each genotype included: CTX-M-1 genotype at m/z 6 390; CTX-M-8 genotype at m/z 5 224, m/z 5 393, and m/z 9 021; CTX-M-9 genotype at m/z 5 161 and m/z 5 273. In the MALDI-TOF MS analysis conducted with the conventional CHCA matrix, the characteristic peak at m/z 9 021 for CTX-M-8 was the only one detected, with the characteristic peaks for CTX-M-1 and CTX-M-9 remaining undetected. Conclusion:The application of cooperative matrix of CHCA-C3 in conjunction with MALDI-TOF MS and machine learning algorithms facilitates the rapid and precise identification of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli. This approach offers a feasible solution for evidence-based clinical therapy and the control of healthcare-associated infections.

Objective:This study aims to develop a rapid identification technique for various genotypes of extended-spectrum β-lactamase (ESBL) producing Escherichia coli using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) in conjunction with machine learning algorithms. Methods:A total of 158 Escherichia coli strains were isolated from the clinical laboratory of the First Hospital of Jilin University from August 2018 to December 2022. Polymerase chain reaction (PCR) was employed to detect the CTX-M-1, CTX-M-8, CTX-M-9, and SHV genes. Mass spectral data of the bacterial strains were acquired by MALDI-TOF MS with a cooperative matrix of (E)-propyl α-cyano-4-hydroxycinnamate (CHCA-C3). Models based on random forest (RF), logistic regression (LR), and support vector machine (SVM) algorithms were constructed. The performance of the constructed models was evaluated using metrics including accuracy, sensitivity, specificity, and the area under the receiver operating characteristic curve (AUC). Mass spectral peaks exhibiting sensitivity and specificity exceeding 80% in the models were designated as characteristic peaks. To validate the efficacy of the cooperative matrix of CHCA-C3, clinical isolates of ESBL-producing Escherichia coli were analyzed by MALDI-TOF MS using the conventional CHCA matrix for comparative purposes. Results:Among the 158 strains of Escherichia coli, 91 strains produced ESBL, all of which were CTX-M genotype. The AUC values for the respective models were as follows: CTX-M-1 genotype exhibited AUC values of 0.98 for LR, 1.00 for RF, and 0.73 for SVM; CTX-M-9 genotype exhibited AUC values of 0.93 for LR, 0.99 for RF, and 0.76 for SVM; for CTX-M-8, all models achieved an AUC of 1.00, indicating excellent classification performance with respect to accuracy, specificity, and sensitivity. The characteristic mass spectral peaks associated with each genotype included: CTX-M-1 genotype at m/z 6 390; CTX-M-8 genotype at m/z 5 224, m/z 5 393, and m/z 9 021; CTX-M-9 genotype at m/z 5 161 and m/z 5 273. In the MALDI-TOF MS analysis conducted with the conventional CHCA matrix, the characteristic peak at m/z 9 021 for CTX-M-8 was the only one detected, with the characteristic peaks for CTX-M-1 and CTX-M-9 remaining undetected. Conclusion:The application of cooperative matrix of CHCA-C3 in conjunction with MALDI-TOF MS and machine learning algorithms facilitates the rapid and precise identification of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli. This approach offers a feasible solution for evidence-based clinical therapy and the control of healthcare-associated infections.

BACKGROUND:Although the clinical application of Masquelet technology has achieved extensive success,the research on optimizing all aspects of Masquelet technology is still being carried out.The focus of doctors is to speed up bone healing and shorten bone healing time after bone grafting. OBJECTIVE:To observe the effect of calcium phosphate combined with recombinant human bone morphogenetic protein-2 in repairing tibial infectious bone defects. METHODS:Thirty-one patients with tibial infectious bone defects were selected from The People's Hospital of Jianyang City from June 2017 to June 2022.They were treated with the Masquelet membrane induction technique.During the second stage of operation,they were divided into a control group(n=15)and a study group(n=16)according to different bone graft materials.Patients in the control group were implanted with autologous bone/allogeneic bone particles,and those in the study group were implanted with calcium phosphate combined with recombinant human bone morphogenetic protein-2/autologous bone particles.Six months after the second stage operation,peripheral blood inflammatory indexes such as white blood cell count,C-reactive protein,and erythrocyte sedimentation rate were detected.Imaging bone healing time,bone healing X-ray score,bone defect healing classification,and adjacent joint function were recorded.The presence of nail track infection,implant absorption,pain,and infection in the bone extraction area were observed. RESULTS AND CONCLUSION:(1)White blood cell count,erythrocyte sedimentation rate,and C-reactive protein levels of the two groups 6 months after the second stage operation were significantly lower than those before the first stage operation(P<0.05).There was no significant difference in each index between the two groups(P>0.05).(2)Bone healing time in the study group was shorter than that in the control group(P<0.05).(3)The Samantha X-ray score of the study group 6 months after the second stage operation was higher than that of the control group(P<0.05).The excellent and good rate of bone defect healing and adjacent joint function of the study group was higher than that of the control group(P<0.05).There was no significant difference in the recurrence rate and complication rate between the two groups(P>0.05).(4)These findings indicate that the effect of calcium phosphate combined with recombinant human bone morphogenetic protein-2 during the second stage operation of the Masquelet membrane induction technique in the treatment of tibial infectious bone defect is good and safe.

OBJECTIVE To analyze the differences in gait features between patients with cardiovascular and cerebrovascular dis-eases and normal people,and to explore new objective features of traditional Chinese medicine(TCM)whole-body inspection.METHODS A monocular camera was used to collect frontal walking videos of subjects,and the diagnosis results of TCM practitioners were used as disease annotation data;a deep learning model was used to estimate the three-dimensional coordinates of key points;the gait features were defined and calculated based on the three-dimensional coordinates of key points of the lower limbs;differences in gait features among people with cardiovascular and cerebrovascular diseases were collected and verified.RESULTS The three-di-mensional coordinates of key points of the lower limbs were automatically extracted and 8 types of TCM gait features were calculated:step width,stride length,foot lift height,limb angle,left and right hip joint angles,and left and right knee joint angles.It was found that there were significant differences in the features between people with cardiovascular and cerebrovascular diseases and healthy peo-ple(P<0.05).CONCLUSION The TCM inspection gait extracted by this study can effectively distinguish patients with cardiovas-cular and cerebrovascular diseases from healthy people,expands the research scope of TCM whole-body inspection,and provides new ideas for the early detection and prevention of cardiovascular and cerebrovascular diseases.

OBJECTIVE To analyze the differences in gait features between patients with cardiovascular and cerebrovascular dis-eases and normal people,and to explore new objective features of traditional Chinese medicine(TCM)whole-body inspection.METHODS A monocular camera was used to collect frontal walking videos of subjects,and the diagnosis results of TCM practitioners were used as disease annotation data;a deep learning model was used to estimate the three-dimensional coordinates of key points;the gait features were defined and calculated based on the three-dimensional coordinates of key points of the lower limbs;differences in gait features among people with cardiovascular and cerebrovascular diseases were collected and verified.RESULTS The three-di-mensional coordinates of key points of the lower limbs were automatically extracted and 8 types of TCM gait features were calculated:step width,stride length,foot lift height,limb angle,left and right hip joint angles,and left and right knee joint angles.It was found that there were significant differences in the features between people with cardiovascular and cerebrovascular diseases and healthy peo-ple(P<0.05).CONCLUSION The TCM inspection gait extracted by this study can effectively distinguish patients with cardiovas-cular and cerebrovascular diseases from healthy people,expands the research scope of TCM whole-body inspection,and provides new ideas for the early detection and prevention of cardiovascular and cerebrovascular diseases.

ObjectiveTo investigate the protective effect and mechanism of Artemisia capillaris Thunb. decoction (YCHT), a classic heat-clearing and cholagogic traditional Chinese medicine (TCM) prescription, on rats with severe acute pancreatitis (SAP) induced by sodium taurocholate. MethodsA total of 30 Sprague-Dawley rats were randomly divided into sham-operation (SO) group, SAP model group, and YCHT (4.0 g/kg) treatment group, with 10 rats in each group. At 24 hours after successful modeling, pancreatic tissue and plasma samples were collected for analysis. HE staining was used to observe pathological injury of the pancreas; ELISA was used to measure the plasma levels of amylase, tumor necrosis factor-α (TNFα), and interleukin-1β (IL-1β); immunofluorescent staining was used to measure the fluorescence intensity of LC-3 protein, and TUNEL was used to measure cell apoptosis. Western blot was used to measure the protein expression of LC-3, Beclin-1, X-linked inhibitor of apoptosis protein (XIAP), caspase-3, and nuclear factor-kappa B (NF-κB) in the pancreas, and quantitative real-time PCR was used to measure the expression levels of lncRNA PVT1 and miRNA-30a-5p. A one-way analysis of variance and the Tukey’s test were used to analyze the differences between multiple independent samples. ResultsYCHT significantly alleviated the pathological injury of the pancreas of SAP rats, such as edema, necrosis, hemorrhage, and inflammatory cell infiltration. Compared with the SO group, the SAP group had significant increases in the plasma levels of amylase and the inflammatory factors TNFα and IL-1β, and there were significant reductions in the plasma levels of amylase, TNFα, and IL-1β after YCHT treatment (all P＜0.05). Compared with the SO group, the SAP group had significant increases in LC-3II/LC-3I ratio and the protein expression of Beclin-1, XIAP, caspase-3, and NF-κB, and compared with the SAP group, the YCHT group had significant reductions in LC-3II/LC-3I ratio and the protein expression of Beclin-1, XIAP, and NF-κB (all P＜0.05). Compared with the SO group, the SAP group had a significant increase in the expression of lncRNA PVT1 and a significant reduction in the expression of miRNA-30a-5p in the pancreas (both P＜0.05), and compared with the SAP group, the YCHT group had a significant reduction in the expression of lncRNA PVT1 and a significant increase in the expression of miRNA-30a-5p (both P＜0.05). ConclusionCell autophagy and apoptosis mediated by lncRNA PVT1/miRNA-30a-5p may be a drug target for YCHT treatment of SAP, which provides experimental and theoretical bases for further development of the TCM prescription YCHT for the treatment of SAP.

OBJECTIVE： To provide reference for the establishment and improvement of the post-marketing re-evaluation of drugs （shorted for re-evaluation） legal system in China. METHODS： Through sorting out American re-evaluation system， this paper focused on the current situation and procedures of the implementation of American re-evaluation system and put forward the suggestions for improving drug re-evaluation system in China. RESULTS & CONCLUSIONS： American re-evaluation system takes enterprises as the main body of execution and the government as the main body of supervision. It has the characteristics of highly informatized and transparent process. The work includes the report of ADR implementation of monitoring systems， the periodic reporting system and the post-listing clinical trials and research systems. The implementation process is to find clues， FDA preliminary review and notification， enterprise further self-examination and review， corporate actions and accept FDA supervision. It is suggested that when establishing the legal system of re-evaluation system in China， the main role of patients should be highlighted， and risk communication should be guided by the public. For example， the Medwatch voluntary reporting system of FDA can be imitated. The unified data collection， storage system and scientific data processing methods can be established. Continuously strengthen the main responsibility consciousness of pharmaceutical enterprises in testing and reporting， and constantly reduce the risk of drug use of patients.

Background Trans-corneally subretinal injection in rodent model is a useful method for genetic therapy,stem cell transplantation and the study on the ophthalmic research.Standarized operation process is critical for the successful treatment.However,there is no literature to report the detailed procedure and the influence of this technique on morphology and function of retina.Objective This sudy was to introduce a method of trans-corneally subretinal injection and evaluate its influence on the morphology and function of retina.Methods Trans-corneallly subretinal injection was performed on the left eyes of 2-month-old SPF C57BL/6J mice after dilation of pupils.A 301/2G disposable needle was used to puncture the cornea within the pupil area near limbus and avoid touching the lens and irises under eye surgery microscope.Then,a 33G blunt needle was used to insert into the vitreous and toward subretinal space via corneal puncture.Normal saline with 0.1％ fluorescein sodium of 1 μl was slowly injected into the space,and 2.5％ hydroxypropyl methylcellulose was dropped on ocular surface for the observation of the fundus clearly.According to the percentage of the retina filled with subretinally injected solution,the experimental eyes were divided into 80％-100％ area group,50％-70％ area group after injection,and the mice in the pseudo-injected group,in which injection procedure stopped just before the solution was pushed in to the subretinal space did not inject any solution after punctured.The right uninjected eyes of the mice served as normal control group.Four eyes were selected for each group.The structural changes were evaluated by optical coherance tomograpby (OCT) 1 day,2 days,3 days and 5 weeks after injection,and retinal function was assessed by the recored of electroretinography (ERG) 5 weeks after injection.The retinal sepcimens were prepared to examin the morphological changes by hematoxylin and esosin staning.The use of care followd the Regulations for the Administration of Affair Concerning Experimental Animals of Zhejiang Province.Results About 70％ of the injected eyes showed that retinal blebs filled with injected green fluorescein solution occupied 50％ or more retinal area with minimal damages.The focal detachment between neurosensory retinal layer and retinal pigment epithelium (RPE) was exhibited 1 day postinjection,and almost all the retinas retached 2 days after injection.In the fifth week after injection,the amplitudes of ERG b wave were (386.25±37.88),(357.50±41.03),(324.25±53.45) and (410.50±14.88) μV in the sham operation group,50％-70％ area group,80％-100％ area group and normal control group,respectively,showing a significant difference among the 4 groups (F=3.574,P=0.047),and the amplitudes of b wave in the normal control group were higher than those in the 80％-100％ area group (all at P ＜ 0.05).The detachment between retinal neuroepithelium layer and RPE layer,cell proliferation and transposition in the outer nuclear layer were dispalyed under the light microscope in the sham operation group,50％-70％ area group and 80％-100％ area group,and the disordered outer segment of photoreceptors at the injecting area was seen in the 50％-70％ area and 80％-100％ area groups at five weeks after injection.However,retinal sructure and morphology were normal at the non-injection area.Conclusions Trans-corneally subretinal injection is an effective and safe way for subretinal injection.

OBJECTIVETo analyze the effect of small interfering RNA (siRNA) targeting mouse epididymis-specific colipase-like (meClps) gene on mouse sperm mobility.

METHODSThe eukaryotic expression vector pDsRed2.0-C1-meClps was constructed and transfected into NIH-3T3 cells, and the protein expression was detected with anti-meClps serum. Three interfering sequences targeting meClps (RNAi-251, 224 and 286) were inserted into lentiviral vectors pRNAT-U6.2/lenti, which were co-transfected with pDsRed2.0-C1-meClps into NIH-3T3 cells. The RNA interfering efficiency was confirmed by semi-quantitative PCR and Western blotting. The lentivirus, packed with the lentiviral vector with the highest interfering efficiency, was injected into the caput tissues of mouse epididymis, and its effect on sperm mobility of the cauda epididymis was evaluated.

RESULTSAll the 3 lentiviral RNAi vectors targeting meClps could inhibit the mRNA and protein expressions of meClps, among which pRNAT-U6.2/lenti-RNAi-251 had the highest interfering efficiency. The lentivirus packed with pRNAT-U6.2/lenti-RNAi-251 significantly reduced the path velocity of cauda sperm after injection into the caput epididymis of the mice (P<0.05).

CONCLUSIONKnock-down meClps expression by lentiviral-mediated RNA interference can lower sperm mobility of mice.

Animals ; Epididymis ; Gene Targeting ; Genetic Vectors ; Lentivirus ; Male ; Mice ; NIH 3T3 Cells ; RNA Interference ; RNA, Messenger ; RNA, Small Interfering ; Sperm Motility ; Spermatozoa ; Transfection