OBJECTIVE To investigate the activity and mechanism of tetrandrine(TET)against influenza A virus in vitro and in vivo.METHODS(1)Cell experiments.① Human non-small cell lung cancer cells(A549)were divided into TET 0(cell control),1.25,2.5,5,10,20 and 25 μmol·L-1 groups,and H1N1+TET 0,1.25,2.5,5,10,20 and 25 μmol·L-1 groups.The TET groups were treated with the corresponding concentrations of TET while the H1N1+TET groups were infected with H1N1 for 1 h before the corresponding concentrations of TET were added.After 48 h,cell viability was detected using the CCK-8 method.② The cells were divided into cell control,H1N1+TET 0,2.5,5,and 10 μmol·L-1 groups and treated as in ①.After 24 h of incubation,the mRNA expressions of matrix protein 1(M1),hemagglutinin(HA),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),interferon-β(IFN-β)were tested by the real-time quantitative PCR(RT-qPCR).The expression levels of M1,HA,neuraminidase(NA),nucleoprotein(NP),and phosphorylation of signal transducer and activator of transcription 3(STAT3)protein were detected by Western blotting.(2)Animal experiments.① Male BALB/c mice were randomly divided into the solvent control group,H1N1 group,H1N1+oseltamivir phosphate(Ose)20 mg·kg-1 group,and H1N1+TET 25,50 and 100 mg·kg-1 groups.The solvent control group and the H1N1 group were ig administered with 0.5％carboxymethyl cellulose sodium(CMC-Na),while the H1N1+Ose group and the H1N1+TET 25,50 and 100 mg·kg-1 groups were ig given suspensions of the respective concentrations of drugs in 0.5％CMC-Na.After three consecutive days of pretreatment,all these groups except the solvent control group were intranasally inoculated with H1N1 to establish an influenza-infected mouse model.The survival rate and body mass of mice were monitored and recorded for 15 consecutive days post-H1N1 infection.② The grouping and treatment were the same as ①.After infection,mice were sacrificed on day 3 and 5.The expression levels of M1,HA,TNF-α,IL-1βand IL-6in lung tissues were detected by RT-qPCR,and those of M1,HA,NA,NP,and phosphoryla-tion of STAT3 protein in mice lung tissues by Western blotting.Hematoxylin-Eosin(HE)staining was performed to observe the pathological changes of lung tissues in mice.The levels of IL-6,TNF-α and IFN-β in bronchoalveolar lavage fluid(BALF)were determined by enzyme-linked immunosorbent assays(ELISA).RESULTS(1)① The half maximal inhibitory concentration study showed a value of 18.06 μmol·L-1 for A549 effected by TET.Compared with the H1N1 group,TET 2.5,5 and 10 μmol·L-1 significantly increased cell viability.② The expression levels of M1,HA mRNA and M1,HA,NA protein in the TET 2.5,5 and 10 μmol·L-1 groups were significantly lowered compared with the H1N1 group.TET 5 μmol·L-1 significantly decreased H1N1-induced IL-6,TNF-α and IFN-β mRNA expression levels in A549 cells.TET 5 and 10 μmol·L-1 could significantly mitigate the phosphorylation of STAT3.(2)① Com-pared with the H1N1 group,TET 50 mg·kg-1 significantly improved the survival rate of H1N1-infected mice while TET 25 mg·kg-1 significantly elevated the body-weight of H1N1-infected mice.In the TET 50 mg·kg-1 group,expressions of HA and M1 mRNA,and HA,M1,NA and NP protein in the lung tissues of H1 N1-infected mice were significantly reduced compared with the H1N1 group.Compared with the H1N1 group,TET 50 mg·kg-1 significantly decreased the lung index,improved inflammatory lesions in lung tissues,inhibited the mRNA expressions of TNF-α,IL-6 and IFN-β in lung tissues,and down regu-lated the expressions of TNF-α,IL-6 and IFN-β proinflammatory cytokines in the BALF of the H1N1-infected mice.In addition,TET 50 mg·kg-1 also significantly inhibited STAT3 phosphorylation in lung tissues of mice infected with H1N1.CONCLUSION TET can inhibit H1N1 infection both in vivo and in vitro.The potential mechanism may be related to the inhibition of the IL-6/STAT3 pathway,which subse-quently suppresses the inflammatory response induced by H1N1.

Objective To explore the influence of combination of recombinant human collagen dressing and promestriene ointment on symptoms and vaginal microecology in patients with atrophic vaginitis.Methods The data of patients with atrophic vaginitis admitted to the General Hospital of the People's Liberation Army were retrospectively collected from April 2017 to April 2024.According to treatment methods,the enrolled patients were divided into a study group(recombinant human collagen dressing combined with promestriene ointment for 7 days)and a control group(promestriene ointment for 7 days).The efficacy,symptom disappearance time,vaginal microecology and adverse reactions were compared between groups,and recurrence rate of atrophic vaginitis within 1 month was observed.Results A total of 150 patients were screened and included,77 in the study group and 73 in the control group.After treatment,the total therapeutic efficacy in the study group was higher than that in the control group(89.61％vs.76.71％,P＜0.05).The disappearance durations of abnormal leucorrhea,vulva pruritus and vulva burning pain in the study group were significantly shorter compared with those in the control group(all P＜0.05).The vaginal pH value in the study group was lower,while the positive rate of Lactobacillus and proportions of vaginal flora density grade Ⅱ-Ⅲ and diversity grade Ⅱ-Ⅲ were higher compared to the control group(all P＜0.05).During treatment,no significant difference was exhibited in the total incidence rate of adverse reactions between the two groups(P＞0.05).The recurrence rate was lower in the study group than that in the control group within 1 month of follow-up(P＜0.05).Conclusion Recombinant human collagen dressing combined with promestriene ointment is more effective than promestriene ointment alone in improving the efficacy of patients with atrophic vaginitis,and can better shorten the disappearance durations of symptoms such as abnormal leucorrhea,vulva pruritus and vulva burning pain,correct the disorder of vaginal microecology,and reduce the short-term recurrence rate of vaginitis,and offer good safety.

OBJECTIVE To investigate the activity and mechanism of tetrandrine(TET)against influenza A virus in vitro and in vivo.METHODS(1)Cell experiments.① Human non-small cell lung cancer cells(A549)were divided into TET 0(cell control),1.25,2.5,5,10,20 and 25 μmol·L-1 groups,and H1N1+TET 0,1.25,2.5,5,10,20 and 25 μmol·L-1 groups.The TET groups were treated with the corresponding concentrations of TET while the H1N1+TET groups were infected with H1N1 for 1 h before the corresponding concentrations of TET were added.After 48 h,cell viability was detected using the CCK-8 method.② The cells were divided into cell control,H1N1+TET 0,2.5,5,and 10 μmol·L-1 groups and treated as in ①.After 24 h of incubation,the mRNA expressions of matrix protein 1(M1),hemagglutinin(HA),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),interferon-β(IFN-β)were tested by the real-time quantitative PCR(RT-qPCR).The expression levels of M1,HA,neuraminidase(NA),nucleoprotein(NP),and phosphorylation of signal transducer and activator of transcription 3(STAT3)protein were detected by Western blotting.(2)Animal experiments.① Male BALB/c mice were randomly divided into the solvent control group,H1N1 group,H1N1+oseltamivir phosphate(Ose)20 mg·kg-1 group,and H1N1+TET 25,50 and 100 mg·kg-1 groups.The solvent control group and the H1N1 group were ig administered with 0.5％carboxymethyl cellulose sodium(CMC-Na),while the H1N1+Ose group and the H1N1+TET 25,50 and 100 mg·kg-1 groups were ig given suspensions of the respective concentrations of drugs in 0.5％CMC-Na.After three consecutive days of pretreatment,all these groups except the solvent control group were intranasally inoculated with H1N1 to establish an influenza-infected mouse model.The survival rate and body mass of mice were monitored and recorded for 15 consecutive days post-H1N1 infection.② The grouping and treatment were the same as ①.After infection,mice were sacrificed on day 3 and 5.The expression levels of M1,HA,TNF-α,IL-1βand IL-6in lung tissues were detected by RT-qPCR,and those of M1,HA,NA,NP,and phosphoryla-tion of STAT3 protein in mice lung tissues by Western blotting.Hematoxylin-Eosin(HE)staining was performed to observe the pathological changes of lung tissues in mice.The levels of IL-6,TNF-α and IFN-β in bronchoalveolar lavage fluid(BALF)were determined by enzyme-linked immunosorbent assays(ELISA).RESULTS(1)① The half maximal inhibitory concentration study showed a value of 18.06 μmol·L-1 for A549 effected by TET.Compared with the H1N1 group,TET 2.5,5 and 10 μmol·L-1 significantly increased cell viability.② The expression levels of M1,HA mRNA and M1,HA,NA protein in the TET 2.5,5 and 10 μmol·L-1 groups were significantly lowered compared with the H1N1 group.TET 5 μmol·L-1 significantly decreased H1N1-induced IL-6,TNF-α and IFN-β mRNA expression levels in A549 cells.TET 5 and 10 μmol·L-1 could significantly mitigate the phosphorylation of STAT3.(2)① Com-pared with the H1N1 group,TET 50 mg·kg-1 significantly improved the survival rate of H1N1-infected mice while TET 25 mg·kg-1 significantly elevated the body-weight of H1N1-infected mice.In the TET 50 mg·kg-1 group,expressions of HA and M1 mRNA,and HA,M1,NA and NP protein in the lung tissues of H1 N1-infected mice were significantly reduced compared with the H1N1 group.Compared with the H1N1 group,TET 50 mg·kg-1 significantly decreased the lung index,improved inflammatory lesions in lung tissues,inhibited the mRNA expressions of TNF-α,IL-6 and IFN-β in lung tissues,and down regu-lated the expressions of TNF-α,IL-6 and IFN-β proinflammatory cytokines in the BALF of the H1N1-infected mice.In addition,TET 50 mg·kg-1 also significantly inhibited STAT3 phosphorylation in lung tissues of mice infected with H1N1.CONCLUSION TET can inhibit H1N1 infection both in vivo and in vitro.The potential mechanism may be related to the inhibition of the IL-6/STAT3 pathway,which subse-quently suppresses the inflammatory response induced by H1N1.

Objective To explore the influence of combination of recombinant human collagen dressing and promestriene ointment on symptoms and vaginal microecology in patients with atrophic vaginitis.Methods The data of patients with atrophic vaginitis admitted to the General Hospital of the People's Liberation Army were retrospectively collected from April 2017 to April 2024.According to treatment methods,the enrolled patients were divided into a study group(recombinant human collagen dressing combined with promestriene ointment for 7 days)and a control group(promestriene ointment for 7 days).The efficacy,symptom disappearance time,vaginal microecology and adverse reactions were compared between groups,and recurrence rate of atrophic vaginitis within 1 month was observed.Results A total of 150 patients were screened and included,77 in the study group and 73 in the control group.After treatment,the total therapeutic efficacy in the study group was higher than that in the control group(89.61％vs.76.71％,P＜0.05).The disappearance durations of abnormal leucorrhea,vulva pruritus and vulva burning pain in the study group were significantly shorter compared with those in the control group(all P＜0.05).The vaginal pH value in the study group was lower,while the positive rate of Lactobacillus and proportions of vaginal flora density grade Ⅱ-Ⅲ and diversity grade Ⅱ-Ⅲ were higher compared to the control group(all P＜0.05).During treatment,no significant difference was exhibited in the total incidence rate of adverse reactions between the two groups(P＞0.05).The recurrence rate was lower in the study group than that in the control group within 1 month of follow-up(P＜0.05).Conclusion Recombinant human collagen dressing combined with promestriene ointment is more effective than promestriene ointment alone in improving the efficacy of patients with atrophic vaginitis,and can better shorten the disappearance durations of symptoms such as abnormal leucorrhea,vulva pruritus and vulva burning pain,correct the disorder of vaginal microecology,and reduce the short-term recurrence rate of vaginitis,and offer good safety.

BACKGROUND:Autoimmune regulator gene(Aire)and Wnt signaling pathway play an important role in the maintenance and differentiation of mouse embryonic stem cell pluripotency.However,whether the Wnt signal and Aire are involved in the differentiation of embryonic stem cells to thymic epithelial progenitor cells remains poorly understood. OBJECTIVE:To investigate the relationship of the Wnt signaling pathway and Aire with the differentiation of embryonic stem cells. METHODS:A two-step differentiation method was used to induce mouse embryonic stem cells to differentiate into endoderm and then into thymic epithelial progenitor cells.Mouse embryonic stem cells were infected with Aire shRNA lentivirus,and monoclonal stable strains were screened by puromycin.Mouse embryonic stem cells were collected on days 0,3 and 10 of the directed induction of differentiation after the induced differentiation by the two-step differentiation method.Cellular immunofluorescence,flow cytometry,western blot assay,and real-time qPCR were used to detect the expression changes of related genes and proteins. RESULTS AND CONCLUSION:(1)Immunofluorescence staining showed positive expression of SSEA1 and OCT4 on day 0 of targeted induction of differentiation.(2)Immunofluorescence staining showed double-positive expression of SOX17 and FOXA2 on day 3 of targeted induction of differentiation.(3)Flow cytometry results showed positive expression of EPCAM1,K5 and K8 on day 10 of targeted induction of differentiation.(4)Compared with undifferentiated mouse embryonic stem cells,the expressions of Wnt7a,β-catenin,and Gsk-3β proteins were elevated,and the expression level of Aire protein was decreased in induced differentiated thymic epithelial progenitor cells.(5)Compared with undifferentiated mouse embryonic stem cells,the expressions of Wnt7a,β-catenin,Gsk-3β and Aire mRNA were elevated in thymic epithelial progenitor cells.(6)Compared with normal cultured mouse embryonic stem cells and their ultimately differentiated thymic epithelial progenitor cells,the expression levels of Wnt7a,β-catenin and Gsk-3β proteins were reduced in mouse embryonic stem cells with knockdown of Aire genes and their final differentiated thymic epithelial progenitor cells.In conclusion,the Wnt signaling pathway and Aire are jointly involved in the process of targeted induction of differentiation of mouse embryonic stem cells into mouse thymic epithelial progenitor cells.

Objective To investigate the role of mitochondrial ATP-sensitive potassium(mito-KATP)channels in attenuation of ischemia-reperfusion(I/R)injury by lidocaine pretreatment in the isolated rat heart.Methods Adult female Wistar rats weighing 220-250 g were anesthetized with intraperitoneal 3% pentobarbital 35 mg/kg.Their hearts were excised and perfused in a Langendorff apparatus with K-H solution saturated with 95%O2-5%CO2 at 37 ℃.Twenty-four isolated rat hearts with I/R injury were randomly divided into 3 groups(n = 8 each):group I/R,lidocaine group(group L)and lidocaine + glibenclamide group(group LG).After 10 min of equilibration,group C,L and LG received 20 min of perfusion with K-H solution,K-H solution containing lidocaine 2.5 mg/L and K-H solution containing lidocaine 2.5 mg/L + glibenclamide(a blocker of mito-KATP channels)10 μmol/L,respectively,then subjected to 30 min of ischemia followed by 60 min of reperfusion.HR,left ventricular developed pressure(LVDP),+ dp/dtmax and - dp/dtmax were recorded at the end of equilibration(T0)and at 15,30,45 and 60 min of reperfusion(T1-4).Coronary effluent was collected at T0 and T4 for determination of lactate dehydrogenase(LDH)and creatine kinase(CK)activities.Myocardial tissues were obtained from cardiac apex at T4 for determination of Na+ -K+ -ATPase and SOD activities and MDA and Ca2+ contents.Results Compared with group I/R,HR,LVDP,+ dp/dtmax and - dp/dtmax were significantly increased,CK and LHD activities were decreased,Na+ -K+-ATPase and SOD activities were increased,and MDA and Ca2+ contents were decreased in group L(P ＜0.05).Compared with group L,HR,LVDP,+ dp/dtmax and -dp/dtmax were significantly decreased,CK and LHD activities were increased,Na+ -K+ -ATPase and SOD activities were decreased,and MDA and Ca2+ contents were increased in group LG(P＜0.05).Conclusion The mechanism by which lidocaine pretreatment attenuates I/R injury to the isolated rat heart is related to mito-KATP channel opening.

CTL responses by autologous colorectal cancer cells were observed in five cases. The specific CTL was induced from two cases, in which tumor associated antigen, CA-Hb_3, was strongly positive. The CTL could not induced from other three cases in which CA-Hb_3 antigen was weakly positive. Weakly positive tumor cells in the three cases were loaded CA-Hb_3 antigen into cytoplasmas by the cationic liposomes and tested by immunohistochemical assay. The results showed that CA-Hb_3 antigen of tumor cells in two cases changed to strongly positive, and could induce CTL responses successfully. With blocking agents BFA and anti-MHC-I monoclonal antibody, CTL responses could be blocked. Our results suggested that soluble CA-Hb_3 antigen could enhance the immunogenity of colorectal cancer cells and induce effective anti-tumor cellular immune responses.