Background:Triple-negative breast cancer(TNBC)is the most aggressive subtype and occurs in approximately 15%–20%of diagnosed breast cancers.TNBC is characterized by its highly metastatic and recurrent features,as wel...Background:Triple-negative breast cancer(TNBC)is the most aggressive subtype and occurs in approximately 15%–20%of diagnosed breast cancers.TNBC is characterized by its highly metastatic and recurrent features,as well as a lack of specific targets and targeted therapeutics.Epidermal growth factor receptor(EGFR)is highly expressed in a variety of tumors,especially in TNBC.LR004-VC-MMAE is a new EGFR-targeting antibody–drug conjugate produced by our laboratory.This study aimed to evaluate its antitumor activities against EGFR-positive TNBC and further studied its possible mechanism of antitumor action.Methods:LR004-VC-MMAE was prepared by coupling a cytotoxic payload(MMAE)to an anti-EGFR antibody(LR004)via a linker,and the drug-to-antibody ratio(DAR)was analyzed by HIC-HPLC.The gene expression of EGFR in a series of breast cancer cell lines was assessed using a publicly available microarray dataset(GSE41313)and Western blotting.MDA-MB-468 and MDA-MB-231 cells were treated with LR004-VC-MMAE(0,0.0066,0.066,0.66,6.6 nmol/L),and the inhibitory effects of LR004-VC-MMAE on cell proliferation were examined by CCK-8 and colony formation.The migration and invasion capacity of MDA-MB-468 and MDA-MB-231 cells were tested at different LR004-VCMMAE concentrations(2.5 and 5 nmol/L)with wound healing and Transwell invasion assays.Flow cytometric analysis and tumorsphere-forming assays were used to detect the killing effects of LR004-VC-MMAE on cancer stem cells(MDA-MB-468 and MDA-MB-231 cells).The mouse xenograft models were also used to evaluate the antitumor efficacy of LR004-VC-MMAE in vivo.Briefly,BALB/c nude mice were subcutaneously inoculated with MDA-MB-468 or MDAMB-231 cells.Then they were randomly divided into 4 groups(n=6 per group)and treated with PBS,naked LR004(10 mg/kg),LR004-VC-MMAE(10 mg/kg),or doxorubicin,respectively.Tumor sizes and the body weights of mice were measured every 4 d.The effects of LR004-VC-MMAE on apoptosis and cell cycle distribution were analyzed by flow cytometry.Western blotting was used to detect the effects of LR004-VC-MMAE on EGFR,ERK,MEK phosphorylation and tumor stemness marker gene expression.Results:LR004-VC-MMAE with a DAR of 4.02 were obtained.The expression of EGFR was found to be significantly higher in TNBC cells compared with non-TNBC cells(P<0.01).LR004-VC-MMAE inhibited the proliferation of EGFRpositive TNBC cells,and the ICvalues of MDA-MB-468 and MDA-MB-231 cells treated with LR004-VC-MMAE for 72 h were(0.13±0.02)nmol/L and(0.66±0.06)nmol/L,respectively,which were significantly lower than that of cells treated with MMAE[(3.20±0.60)nmol/L,P<0.01,and(6.60±0.50)nmol/L,P<0.001].LR004-VC-MMAE effectively inhibited migration and invasion of MDA-MB-468 and MDA-MB-231 cells.Moreover,LR004-VC-MMAE also killed tumor stem cells in EGFR-positive TNBC cells and impaired their tumorsphere-forming ability.In TNBC xenograft models,LR004-VC-MMAE at 10 mg/kg significantly suppressed tumor growth and achieved complete tumor regression on day 36.Surprisingly,tumor recurrence was not observed until the end of the experiment on day 52.In a mechanistic study,we found that LR004-VC-MMAE significantly induced cell apoptosis and cell cycle arrest at G/M phase in MDAMB-468[(34±5)%vs.(12±2)%,P<0.001]and MDA-MB-231[(27±4)%vs.(18±3)%,P<0.01]cells.LR004-VC-MMAE also inhibited the activation of EGFR signaling and the expression of cancer stemness marker genes such as Oct4,Sox2,KLF4 and EpCAM.Conclusions:LR004-VC-MMAE showed effective antitumor activity by inhibiting the activation of EGFR signaling and the expression of cancer stemness marker genes.It might be a promising therapeutic candidate and provides a potential therapeutic avenue for the treatment of EGFR-positive TNBC.展开更多
Purpose:To investigate nursing students’knowledge of and attitudes about problem-based learning(PBL).Methods:A total of 1200 students were surveyed at eight nursing colleges in Hunan Province.Results:In all,1037 vali...Purpose:To investigate nursing students’knowledge of and attitudes about problem-based learning(PBL).Methods:A total of 1200 students were surveyed at eight nursing colleges in Hunan Province.Results:In all,1037 valid questionnaires were returned,for an effective return rate of 86.4%.Some 54.4%of the students learned that PBL was a pedagogical method from teachers,and 27.8%of the students had participated in PBL courses.Almost all of students(97.6%)were interested in PBL,and 66.7%of survey participants believed that students who were not good at solving problems would have difficulty in PBL courses.Conclusion:Nursing educators should guide students to adapt to new learning approaches,and encourage students to participate in the teaching reform to promote students’autonomous learning ability,innovation ability,and comprehensive ability.展开更多
The current study was designed to investigate the mechanisms by which ropivacaine may act within the central nervous system （CNS） to produce cardiotoxicity. Eighty New Zealand rabbits were divided into four groups r...The current study was designed to investigate the mechanisms by which ropivacaine may act within the central nervous system （CNS） to produce cardiotoxicity. Eighty New Zealand rabbits were divided into four groups randomly. In Group 1,20 rabbits received intracerebroventricular （icv） saline, and then received icv ropivacaine 30 min later. In Group 2, 20 rabbits received icv ropivacaine. Whenever dysrhythmias continued for more than 5 min, 0.1 ml saline was administered into the left cerebral ventricle. Ten minutes later, 0.1 ml midazolam was given into the left lateral ventricle. In Group 3, 20 rabbits received icv ropivacaine, and once the dysrhythmias developed, the inspired isoflurane concentration was increased from 0.75% to 1.50%. In Group 4, 20 animals received an intravenous （iv） phenylephrine infusion until dysrhythmias occurred. In Group 1, the rabbits did not develop dysrhythmias in response to icv saline, whereas dysrhythmias did develop in these animals after icv ropivacaine. In Group 2, icv saline had no effect on the dysrhythmias; however, icv midazolam terminated cardiac dysrhythmias. In Group 3, an increase in the concentration of the inspired isoflurane had no effect on dysrhythmias. In Group 4, icv midazolam had no effect on dysrhythmias in response to iv phenylephdne. Ropivacaine administered directly into the CNS is capable of producing cardiac dysrhythmias; midazolam terminated dysrhythmias presumably by potentiation of y-aminobutyric acid （GABA） receptor activity. Our results suggest that ropivacaine produces some of its cardiotoxicity not only by the direct cardiotoxicity of the drug, but also by the CNS effects of ropivacaine.展开更多
基金supported by the CAMS Innovation Fund for Medical Sciences(CIFMS)(2021-1-I2M-026)the Beijing Natural Science Foundation(7202133)the Non-profit Central Research Institute Fund of Chinese Academy of Medical Sciences(2021-RW350-002)。
文摘Background:Triple-negative breast cancer(TNBC)is the most aggressive subtype and occurs in approximately 15%–20%of diagnosed breast cancers.TNBC is characterized by its highly metastatic and recurrent features,as well as a lack of specific targets and targeted therapeutics.Epidermal growth factor receptor(EGFR)is highly expressed in a variety of tumors,especially in TNBC.LR004-VC-MMAE is a new EGFR-targeting antibody–drug conjugate produced by our laboratory.This study aimed to evaluate its antitumor activities against EGFR-positive TNBC and further studied its possible mechanism of antitumor action.Methods:LR004-VC-MMAE was prepared by coupling a cytotoxic payload(MMAE)to an anti-EGFR antibody(LR004)via a linker,and the drug-to-antibody ratio(DAR)was analyzed by HIC-HPLC.The gene expression of EGFR in a series of breast cancer cell lines was assessed using a publicly available microarray dataset(GSE41313)and Western blotting.MDA-MB-468 and MDA-MB-231 cells were treated with LR004-VC-MMAE(0,0.0066,0.066,0.66,6.6 nmol/L),and the inhibitory effects of LR004-VC-MMAE on cell proliferation were examined by CCK-8 and colony formation.The migration and invasion capacity of MDA-MB-468 and MDA-MB-231 cells were tested at different LR004-VCMMAE concentrations(2.5 and 5 nmol/L)with wound healing and Transwell invasion assays.Flow cytometric analysis and tumorsphere-forming assays were used to detect the killing effects of LR004-VC-MMAE on cancer stem cells(MDA-MB-468 and MDA-MB-231 cells).The mouse xenograft models were also used to evaluate the antitumor efficacy of LR004-VC-MMAE in vivo.Briefly,BALB/c nude mice were subcutaneously inoculated with MDA-MB-468 or MDAMB-231 cells.Then they were randomly divided into 4 groups(n=6 per group)and treated with PBS,naked LR004(10 mg/kg),LR004-VC-MMAE(10 mg/kg),or doxorubicin,respectively.Tumor sizes and the body weights of mice were measured every 4 d.The effects of LR004-VC-MMAE on apoptosis and cell cycle distribution were analyzed by flow cytometry.Western blotting was used to detect the effects of LR004-VC-MMAE on EGFR,ERK,MEK phosphorylation and tumor stemness marker gene expression.Results:LR004-VC-MMAE with a DAR of 4.02 were obtained.The expression of EGFR was found to be significantly higher in TNBC cells compared with non-TNBC cells(P<0.01).LR004-VC-MMAE inhibited the proliferation of EGFRpositive TNBC cells,and the ICvalues of MDA-MB-468 and MDA-MB-231 cells treated with LR004-VC-MMAE for 72 h were(0.13±0.02)nmol/L and(0.66±0.06)nmol/L,respectively,which were significantly lower than that of cells treated with MMAE[(3.20±0.60)nmol/L,P<0.01,and(6.60±0.50)nmol/L,P<0.001].LR004-VC-MMAE effectively inhibited migration and invasion of MDA-MB-468 and MDA-MB-231 cells.Moreover,LR004-VC-MMAE also killed tumor stem cells in EGFR-positive TNBC cells and impaired their tumorsphere-forming ability.In TNBC xenograft models,LR004-VC-MMAE at 10 mg/kg significantly suppressed tumor growth and achieved complete tumor regression on day 36.Surprisingly,tumor recurrence was not observed until the end of the experiment on day 52.In a mechanistic study,we found that LR004-VC-MMAE significantly induced cell apoptosis and cell cycle arrest at G/M phase in MDAMB-468[(34±5)%vs.(12±2)%,P<0.001]and MDA-MB-231[(27±4)%vs.(18±3)%,P<0.01]cells.LR004-VC-MMAE also inhibited the activation of EGFR signaling and the expression of cancer stemness marker genes such as Oct4,Sox2,KLF4 and EpCAM.Conclusions:LR004-VC-MMAE showed effective antitumor activity by inhibiting the activation of EGFR signaling and the expression of cancer stemness marker genes.It might be a promising therapeutic candidate and provides a potential therapeutic avenue for the treatment of EGFR-positive TNBC.
文摘Purpose:To investigate nursing students’knowledge of and attitudes about problem-based learning(PBL).Methods:A total of 1200 students were surveyed at eight nursing colleges in Hunan Province.Results:In all,1037 valid questionnaires were returned,for an effective return rate of 86.4%.Some 54.4%of the students learned that PBL was a pedagogical method from teachers,and 27.8%of the students had participated in PBL courses.Almost all of students(97.6%)were interested in PBL,and 66.7%of survey participants believed that students who were not good at solving problems would have difficulty in PBL courses.Conclusion:Nursing educators should guide students to adapt to new learning approaches,and encourage students to participate in the teaching reform to promote students’autonomous learning ability,innovation ability,and comprehensive ability.
基金Project (No. 2006K13-G7-4) supported by the Key Sci-Tech Research Project of Shaanxi Province,China
文摘The current study was designed to investigate the mechanisms by which ropivacaine may act within the central nervous system （CNS） to produce cardiotoxicity. Eighty New Zealand rabbits were divided into four groups randomly. In Group 1,20 rabbits received intracerebroventricular （icv） saline, and then received icv ropivacaine 30 min later. In Group 2, 20 rabbits received icv ropivacaine. Whenever dysrhythmias continued for more than 5 min, 0.1 ml saline was administered into the left cerebral ventricle. Ten minutes later, 0.1 ml midazolam was given into the left lateral ventricle. In Group 3, 20 rabbits received icv ropivacaine, and once the dysrhythmias developed, the inspired isoflurane concentration was increased from 0.75% to 1.50%. In Group 4, 20 animals received an intravenous （iv） phenylephrine infusion until dysrhythmias occurred. In Group 1, the rabbits did not develop dysrhythmias in response to icv saline, whereas dysrhythmias did develop in these animals after icv ropivacaine. In Group 2, icv saline had no effect on the dysrhythmias; however, icv midazolam terminated cardiac dysrhythmias. In Group 3, an increase in the concentration of the inspired isoflurane had no effect on dysrhythmias. In Group 4, icv midazolam had no effect on dysrhythmias in response to iv phenylephdne. Ropivacaine administered directly into the CNS is capable of producing cardiac dysrhythmias; midazolam terminated dysrhythmias presumably by potentiation of y-aminobutyric acid （GABA） receptor activity. Our results suggest that ropivacaine produces some of its cardiotoxicity not only by the direct cardiotoxicity of the drug, but also by the CNS effects of ropivacaine.
