Objective To study the related molecular mechanism of andrographolide(AGP)in inhibi-ting the proliferation of human colon cancer LoVo cells and promoting their apoptosis.Methods The LoVo cells were set up as the control group and experimental group.The cells were treated with different concentra-tions of AGP(0,20,40,80 μmol/L).The cell viability was detected by CCK-8,the cellular cycle and apoptosis were detected by flow cytometery,and Western blot was used to detect protein level of PCNA,Bad and Bcl-2,and the effect of AGP of BMP9 overexpression or silencing AGP on PCNA,Bad and Bcl-2 protein levels;West-ern blot was used to analyze the effect of AGP on PTEN,Akt1/2/3 and p-Akt1/2/3 levels.Results AGP could inhibit the proliferation of LoVo cells,promoted their apoptosis and increased the BMP9 expression lev-el.The BMP9 overexpression could enhance the effect of AGP for inhibiting the proliferation and promoting the apoptosis,silencing BMP9 could weaken the above effects of AGP(P＜0.05).AGP decreased the expres-sion level of p-Akt1/2/3 in the LoVo cells and increased the PTEN expression level(P＜0.05).BMP9 over-expression enhanced AGP and decreased the p-Akt 1/2/3 expression level in the LoVo cells,and silencing BMP9 could weaken the above effects of AGP(P＜0.05).Conclusion AGP could inhibit the proliferation of human colon cancer LoVo cells,its mechanism may be related with up-regulating BMP9 expression,thus in-crease PTEN protein level and inhibit P13K/Akt signal.

Objective:The purpose of this study was to investigate the characteristics of viral pathogen spectrum and the epidemiological characteristics of each viral pathogen in hospitalized cases associated with severe acute respiratory infection (SARI) in Luohe City, Henan Province from 2017 to 2019.Methods:Based the SARI Case Surveillance Platform, SARI cases were collected in Central Hospital of Luohe City, Henan Province from November 2017 to February 2019. In the end, 783 SARI cases were included, whose throat swabs were taken within 24 h of admission, as well as their demographic characteristics, onset time, clinical characteristics and other information recorded. At the same time, viral identification was performed, and the age and time distribution of each virus were analyzed.Results:The age of 783 SARI cases shown as M ( P 25, P 75) was 3 (1, 5) years old, ranging from 1 month to 95 years old. Children under 5 years old were the majority (71.01%). The males (61.81%) were more than females (38.18%). Among the 783 SARI cases, a total of 9 kind of viruses were identified with 64.88% (508/783) of the throat swabs tested positive for at least one virus. The positive rate of influenza virus and human respiratory syncytial virus were both 20.18% (158 cases), which was the highest among all the detected respiratory virus. The co-infection rate was 15.84% (124/783), among which double infection was the most common, accounting for 85.48% (106/124) of the co-infected cases. And human respiratory syncytial virus, human rhinovirus and influenza virus were the most common pathogen in co-infection cases. Moreover, the viral positive rate was 68.71% in children aged 5 years and 63.27% in people aged 60-95 years. Influenza and human respiratory syncytial virus dominated in winter and spring, while human parainfluenza virus was the main infection in summer. Conclusion:Influenza virus and human respiratory syncytial virus were the main viruses in throat swabs of SARI cases from 2017 to 2019 in Luohe City, Henan Province. There were differences in the age and seasonal epidemiological characteristics of each virus.

Objective:The purpose of this study was to investigate the characteristics of viral pathogen spectrum and the epidemiological characteristics of each viral pathogen in hospitalized cases associated with severe acute respiratory infection (SARI) in Luohe City, Henan Province from 2017 to 2019.Methods:Based the SARI Case Surveillance Platform, SARI cases were collected in Central Hospital of Luohe City, Henan Province from November 2017 to February 2019. In the end, 783 SARI cases were included, whose throat swabs were taken within 24 h of admission, as well as their demographic characteristics, onset time, clinical characteristics and other information recorded. At the same time, viral identification was performed, and the age and time distribution of each virus were analyzed.Results:The age of 783 SARI cases shown as M ( P 25, P 75) was 3 (1, 5) years old, ranging from 1 month to 95 years old. Children under 5 years old were the majority (71.01%). The males (61.81%) were more than females (38.18%). Among the 783 SARI cases, a total of 9 kind of viruses were identified with 64.88% (508/783) of the throat swabs tested positive for at least one virus. The positive rate of influenza virus and human respiratory syncytial virus were both 20.18% (158 cases), which was the highest among all the detected respiratory virus. The co-infection rate was 15.84% (124/783), among which double infection was the most common, accounting for 85.48% (106/124) of the co-infected cases. And human respiratory syncytial virus, human rhinovirus and influenza virus were the most common pathogen in co-infection cases. Moreover, the viral positive rate was 68.71% in children aged 5 years and 63.27% in people aged 60-95 years. Influenza and human respiratory syncytial virus dominated in winter and spring, while human parainfluenza virus was the main infection in summer. Conclusion:Influenza virus and human respiratory syncytial virus were the main viruses in throat swabs of SARI cases from 2017 to 2019 in Luohe City, Henan Province. There were differences in the age and seasonal epidemiological characteristics of each virus.

Aim To investigated the possible effect of COX-2 on the BMP9-induced activation of PI3K/Akt signal in progenitor cells.Methods The activity of alkaline phosphatase(ALP) was measured using histochemical staining or chemiluminescence.The mRNA level of ALP was determined using real-time PCR assay.The protein levels of osteopontin(OPN), osteocalcin(OCN), COX-2, Akt1/2 and phosphorylated Akt1/2 were detected by Western blot.The mRNA level of COX-2 was assayed with RT-PCR, and the mineralization was measured with Alizarin Red staining.Results The ALP activity was apparently increased by BMP9 in C2C12 cells, as well as the protein level of OPN and OCN.The mineralization was also markedly induced by BMP9 in C2C12 cells.BMP9 increased the level of phosphorylated Akt1/2 greatly, although no substantial effect was observed on total protein level of Akt1/2.The BMP9-induced ALP activity was dramatically decreased by the inhibitor of PI3K.The mRNA and protein level of COX-2 were both increased by BMP9 in C2C12cells, and the BMP9-induced ALP activity and mineralization were greatly attenuated by the inhibitor of COX-2.The BMP9-induced phosphorylation level of Akt1/2 was increased by the exogenous expression of COX-2, but decreased by the inhibitor of COX-2.Conclusion Activation of PI3K/Akt signaling may be a critical event in BMP9-induced osteogenic differentiation, and this process may be mediated by the BMP9-upregulated COX-2 in stem cells at least.

Aim To investigate the role of TGF-β3 in the anti-proliferation effect of ursolic acid(UA)in co-lon cancer cells and the possible molecular mechanism underlying this effect.Methods We introduced crys-tal violet staining,flow cytometry and Western blot as-say to determine the effect of UA on proliferation and apoptosis in HCT1 1 6 cells.The levels of TGF-β3, Smad2 /3 and β-catenin in HCT1 1 6 cell were evaluated by RT-PCR and Western blot.Finally,TGF-β3 inhibi-tor and recombinant adenovirus,and luciferase reporter assay were used to analyze the possible mechanism through which TGF-β3 mediated the anti-cancer effect of UA in HCT1 1 6 cells.Results UA inhibited the proliferation and induced apoptosis apparently in HCT1 1 6 cells.UA down-regulated TGF-β3 both in mRNA and in protein level.Meanwhile,UA decreased the phosphorylation of Smad2 /3 concentration depend-ently,although no significant effect was found on the total protein level of Smad2 /3 in HCT1 1 6 cells.Over-expression of TGF-β3 attenuated the inhibitory effect of UA on the proliferation of HCT1 1 6 cells,while the TGF-β3 inhibitor potentiated this effect. UA sup-pressed the transconduction of Wnt/β-catenin signaling in HCT1 1 6 cells through decreasing the level of β-catenin.Exogenous expression of TGF-β3 increased the level of β-catenin and partly reversed the UA-in-duced decrease of β-catenin.However,TGF-β3 inhib-itor potentiated the inhibitory effect of UA on β-catenin in HCT1 1 6 cells.Conclusion The anti-proliferation activity of UA in colon cancer may be partly mediated through down-regulating TGF-β3 to suppress Wnt/β-catenin signaling at least.

At present,pharmacy experiment is mainly in accordance with the traditional teaching mode,and students mechanically complete the experiments according to the demonstration of teachers.There is lack of interaction between teachers and students,and students lack independent thinking and experimental content innovation,which lead to the inefficient teaching effects.By flipping the classroom in the pharmacy experimental teaching,teachers prepare teaching courseware,teaching video and learning tasks before class,in class emphasize the experiment key and the difficult point,and carry on the preliminary experiment,and after class,summarize and analyze the practical problems that students have in the experiment,which have effectively aroused students' professional interest in learning and promoted their practical operation and independent thinking ability,and achieved good teaching results.

Aim To investigate the relationship be-tween the anti-proliferation effect of resveratrol ( Res ) and p38 MAPK in colon cancer cells .Methods Crys-tal violet staining , Western blot and flow cytometry were employed to analyze the effect of Res on the pro-liferation in LoVo cells.Western blot assay was used to detect the effect of Res on the apoptosis of LoVo cells and the phosphorylation of p 38 MAPK.Crystal violet staining and Western blot assay were used to analyze whether p38 MAPK was involved in the Res-induced proliferation inhibition and apoptosis in LoVo cells .Re-sults Res inhibited the proliferation , arrested cell cy-cle at S phase , and increased the protein level of PC-NA in LoVo cells apparently .Res increased the level of Bad in LoVo cells, but decreased the level of Bcl-2. Although Res exerted no substantial effects on total lev-el of p38 MAPK, it markedly increased the phospho-rylation level of p38 MAPK in LoVo cells.p38 MAPK inhibitor promoted the proliferation , and decreased the anti-proliferation effect of Res on LoVo cells .Moreo-ver , the effects of Res on the level of Bcl-2 and Bad were both reduced by the p 38 MAPK inhibitor .Con-clusions Res can inhibit the proliferation of LoVo cells, which may be partly mediated by promoting the phosphorylation of p38 MAPK.

Objective To investigate the effect of nerve growth factor (NGF) on osteogenesis induced by bone morphogenetic protein-9 (BMP-9) in mouse embryonic fibroblasts (MEFs).Methods MEFs were respectively transfected with adenovirus-mediated NGF (NGF group), BMP-9 (BMP-9 group) and NGF + BMP-9 (combined group) and green fluorescence protein (GFP) (control group).Cytochemical staining was used to test the activity of alkaline phosphatase (ALP) 3 d and 5 d after treatment.Level of osteopontin (OPN) mRNA was detected by RT-PCR 9 d after treatment.Level of OPN protein was assayed by Western blot and immunocytochemistry 9 d after treatment.Mineralization was detected by Alizarin red staining 14 d after treatment.Results ALP activity in MEFs was elevated in BMP-9 group rather than in NGF group, but a significant increase in ALP activity was noted in combined group.In control group, BMP-9 group, NGF group and combined group, level of OPN mRNA was 0.92 ± 0.03, 1.28 ± 0.04, 0.94 ± 0.03 and 1.62 ± 0.04 respectively (F =214.60, P ＜ 0.01);level of OPN protein was 0.60 ± 0.05, 0.84 ± 0.03, 0.53 ± 0.05 and 1.27 ± 0.05 respectively (F =162.5, P ＜ 0.01).In comparison, OPN mRNA and protein were significantly up-regulated in combined group than in BMP-9 group (t =10.569 and 11.778,P ＜ 0.05).In control group, BMP-9 group, NGF group and combined group, relative density of OPN protein was 3.63 ±0.17, 6.27 ±0.30, 3.86 ±0.18 and 10.16 ±0.18respectively (F =602.6, P ＜ 0.01), with a significant higher level in combined group than in BMP-9 group (t =22.280, P ＜ 0.05).Level of mineralization was significantly higher in combined group than in BMP-9 or NGF group.Conclusion NGF can potentiate the osteogenesis induced by BMP-9 in MEFs.

Aim To study the anti-proliferation effect of resveratrol (Res)and the role of Res-induced bone morphorgenetic protein 9 (BMP9 )in this process in colon cancer cells.Methods Crystal violet staining and flow cytomtry were introduced to assay the anti-proliferation effect of Res in LoVo cells.The effect of Res on apoptosis in LoVo cells was also detected with flow cytometry.Then,RT-PCR and Western blot assay were employed to unveil the effect of Res on the ex-pression of BMP9 .The effect of BMP9 on the anti-pro-liferation of Res in LoVo cells was analyzed with crystal violet staining and flow cytometry too.Finally,the effect of Res on the expression of ALK2 and ALK3 was assayed with RT-PCR,and the inhibitor of ALK2 and ALK3 was used to figure out the possible mechanism of BMP9 on Res-induced proliferation inhibition in LoVo cells.Results Res apparently inhibited the prolifera-tion,arrested the cell cycle at S phase in LoVo and in-creased the percentage of apopotic cells in LoVo cells. Res increased the expression of mRNA and protein of BMP9 concentration dependently. Exogenous ex-pressed-BMP9 enhanced the anti-proliferation and ap-optosis inducing effects of Res in LoVo cells, but BMP9 knockdown decreased these effects of Res.Al-though Res had no apparent effect on increasing the phosphorylation of Smad1/5/8,it increased the ex-pression of ALK2 and ALK3 .Inhibition of ALK2 and ALK3 decreased the anti-proliferation effect of Res partly in LoVo cells.Conclusion Res is potent to in-hibit the proliferation of LoVo cells,Which may be mediated by up-regulating the expression of BMP9 and its receptor at least.

Aim To investigate the anti-proliferation effect of resveratrol (Res)on human colon cancer cells and dissect the possible mechanism underlaying this effect.Methods We introduced crystal violet staining and Western blot to analyse the anti-proliferation effect of Res on HCT1 1 6 cells.Then,we used flow cytome-try and Western blot assay to detect the Res induced apoptosis in HCT1 1 6 cells.Next,we employed the well established TCF4 /LEF luciferase reporter to meas-ure the effect of Res on Wnt/β-catenin signaling trans-duction.Finally,we took Western blot and PCR assay to analyse the effect of Res on the expression of β-cate-nin in HCT1 1 6 cells.Results Crystal violet staining and Western blot analysis showed that Res could inhib-it the proliferation of HCT1 1 6 cells in a concentration-and time dependent fashion.What’s more,Res could promote apoptosis in HCT1 1 6 cells.The transcriptional activities of TCF4 /LEF reporter were reduced by Res in a concentration-dependent fashion (P <0.05 when the concentration of Res was 20 μmol·L -1 ,and P <0.01 when the concentration of Res was 40 μmol·L -1 or 80 μmol·L -1 ).Res could decrease not only the protein level of β-catenin in HCT1 1 6 cells,but also the mRNA expression of β-catenin.Conclusion Res can inhibit the proliferation of HCT1 1 6 cells,which may be mediated at least by down-regulating the ex-pression of β-catenin to inhibit the Wnt/β-catenin sig-naling transduction.