OBJECTIVE To explore the mechanism of action of active components of Nostoc commune in anti-triple-negative breast cancer(TNBC) by the network pharmacology method and molecular biology experiment. METHODS The active components of Nostoc commune were collected by consulting the literature and combined with the preliminary research in the laboratory, the Swiss Target Prediction database was used for target prediction, and the disease targets were obtained in the TTD, Genecards and OMIM databases. The STRING online platform was used for protein-protein interaction, and the KEGG signaling pathway and GO gene function enrichment analysis were performed using the Metascape database. Molecular docking of N-acetyltryptamine, a component of Nostoc commune, and target AKT1 by AutoDock software. Annexin V-FITC/PI double staining method was performed to analyze the apoptotic rate of cells. RT-qPCR and Western blotting were used to detect the mechanism of action of the active components of Nostoc commune on anti-TNBC. RESULTS The results of network pharmacology showed that there were 8 effective components, such as N-acetyltryptamine, Scytonemin and Nostocionone, involved 75 key targets such as signal transduction and AKT1, STAT3 and CCND1. The KEGG signaling pathway and GO gene function enrichment analysis results involved cancer-related signaling pathways, PI3K-Akt signaling pathways and MAPK signaling pathways. Molecular docking showed that N-acetyltryptamine had better affinity with AKT1. N-acetyltryptamine could not significantly promote apoptosis of breast cancer cells. Western blotting showed that N-acetyltryptamine could down-regulate the protein expressions of AKT1. The results of RT-qPCR showed that N-acetyltryptamine could effectively reduce the mRNA expression of AKT1 in cells. CONCLUSION N-acetyltryptamine may inhibit the proliferation of TNBC cells by inhibiting the AKT1 signaling pathway, thereby exerting anti-TNBC effects.

OBJECTIVE: To investigate the effect of licochalcone A (LCA) on the proliferation and cell cycle of human lung squamous carcinoma cells and explore its possible molecular mechanism. METHODS: MTT assay was used to detect the changes in proliferation of H226 cells after treatment with different concentrations of LCA for 48 h, and the IC₅₀ of LCA was calculated. Flow cytometry was used to analyze cell cycle changes in H226 cells treated with 10, 20, and 40 μmol/L LCA, and the expressions of cyclin D1, cyclin-dependent kinase CDK2 and CDK4, and p-PI3K, PI3K, p-Akt, and Akt in the treated cells were detected using Western blotting. The effect of intraperitoneal injection of LCA for 24 days on tumor volume and weight was assessed in a BALB/c-nu mouse model bearing lung squamous carcinoma xenografts. RESULTS: MTT assay showed that LCA significantly decreased the viability of H226 cells with an IC₅₀ of 28.3 μmol/L at 48 h. Flow cytometry suggested that LCA treatment induced obvious cell cycle arrest at the G1 phase. LCA treatment also significantly decreased the expressions of cyclin D1, CDK2, and CDK4, and inhibited the phosphorylation of PI3K and Akt in H226 cells. In the tumor-bearing mice, LCA treatment for 24 days significantly reduced the tumor volume and weight. CONCLUSION LCA is capable of inhibiting the proliferation and inducing cell cycle arrest in lung squamous carcinoma cells possibility by regulating the PI3K/Akt singling pathway.
Humans ; Animals ; Mice ; Cyclin D1 ; Phosphatidylinositol 3-Kinases ; Proto-Oncogene Proteins c-akt ; Carcinoma, Non-Small-Cell Lung ; Carcinoma, Squamous Cell ; Cell Cycle Checkpoints ; Lung Neoplasms ; Signal Transduction ; Lung

Objective PINK1 and Parkin are directly invoveled in the regulation and maintenance of mitochondrial functional morphology. We aim to explore the effect of Wnt2 overexpression on PINK1^B9 Mutant Drosophila and its mechanism in this study. Methods The GAL4-UAS system was used to construct the normal control flies（W1118/ + ；MHC-GAL4/+）, PINK1^B9 transgenic Drosophila model flies（UAS-PINK1^B9 /y；MHC-GAL4 / +；Parkinson's disease model of Drosophila melanogaster), the Wnt2 overexpression flies（UAS-PINK1^B9 /y；MHC-GAL4 / Wnt2 OE） and the Wnt2 RNAi flies（UAS-PINK1^B9 /y；MHC-GAL4 /Wnt2）. On the 5th day, the abnormal wings phenotype rate and flying rate of flies were observed. The contents of Ndufs3 proteins were detected by Western blot. The mRNA expression levels of PGC-1α, Nrf1 and TFAM related to mitochondrial metabolism and synthesis were detected by real-time fluorescence quantitative PCR. The morphology of mitochondria was observed by electron microscopy. Complex I and Complex II function was detected by high-resolution mitochondrial respiratory system. Results Compared with the normal control flies, PINK1^B9 transgenic Drosophila model flies showed increased abnormal wings phenotype rate([1.87±0.06]% vs [68.79±0.70]%), decreased flying rate([ 97.51±0.52)% vs(3.95±0.53)%], and the differences were statistically significant(P<0.05); Compared with PINK1^B9 transgenic Drosophila model flies, the Wnt2 RNAi flies showed decreased abnormal wings phenotype rate[(10.14±1.72)%], increased flying rate([41.83±2.57]%)(P<0.05). Compared with the normal control flies, PINK1^B9 transgenic Drosophila model flies showed decreased expression levels of PGC-1α and Nrf1,Ndufs3 proteins, Complex I and Complex II(P<0.05);On the contrary, the Wnt2 RNAi flies showed increased trends compared with PINK1^B9 transgenic Drosophila model flies(P<0.05). Conclusion Overexpression of Wnt2 protects PINK1^B9 transgenic Drosophila models, which is related to the improvement of mitochondrial function.

[Abstract] Objective To evaluate the effect of Mycobacterium tuberculosis Direct Assay (MTD) for rapid detecting Mycobacterium tuberculosis rRNA and Multi-locus PCR for M.bovis BCG strain typing in patients with suspected extra-pulmonary tuberculosis.Methods From June 2010 to December 2011,47 children and 75 adult patients with suspected extra-pulmonary tuberculosis in Shanghai public health clinical center were recruited.Also 48 non-tuberculosis patients were taken as a negative control.Clinical specimens from these patients were collected.Acid fast stain,solid culture,liquid culture,and MTD were used to detect all clinical specimens simultaneously.Screen tuberculosis strains of the culture isolates by MPT64 antigen assay and use Multi-locus PCR for the BCG strain genotyping of the isolates without MPT64 antigen.SPSS16.0 was used to analyse the results.Results The sensitivity for acid fast stain,solid culture,liquid culture and MTD test was 10.7％ (13/122),11.5％ (14/122),16.4％ (20/122) and 37.7％ (46/122),respectively.And the specificity of MTD was 100.0％.Six clinical isolates from children were identified as BCG by Multi-locus PCR typing,the same with chemical tests.Conclusions The MTD assay and the MGIT960 liquid culture are effective and reliable method for diagnosing extra-pulmonary tuberculosis.And Multi-locus PCR can be assisted for the early diagnosis of extra-pulmonary tuberculosis patients with suspected BCG infection.

Objective Toinvestigatetheroleofcontrolledanticoagulationinocclusionofvertebral arteriesforthetreatmentofgiantfusiformaneurysmsofthebasilartrunkinchildren.Methods The clinical data of 3 children with giant fusiform aneurysms of the basilar trunk were analyzed retrospectively. Three children underwent bilateral vertebral artery occlusion with endovascular intervention,and conducted controlled anticoagulation by intravenous infusion of heparin sodium (10-30 U/[kg·h])after procedure. Results Intheprocessofanticoagulation,2patientshadsomeseverebleedingcomplications,including epistaxis,gastrointestinal bleeding,and after transient cessation of anticoagulation,they had severe brainstem ischemic symptoms,including vomiting,hemiplegia,loss of consciousness and respiratory dysfunctions (one recovered after using low-molecular weight heparin and one died),another patient did not have any serious complicationsandwascuredafteradjustingtheanticoagulationstrategy.Conclusion Inthetreatmentof basilar artery trunk giant fusiform aneurysms in children through the bilateral vertebral artery occlusion,the controlled anticoagulation after procedure may reduce the occurrence of brain stem infarction and severe bleeding complications. It may be an important measure for improving the safety and effectiveness of bilateral vertebral artery occlusion.

AIM: To investigate the effects of insulin-like growth factor 1(IGF-1)on the secretion of transforming growth factor β2(TGF-β2), matrix metalloproteinase 2(MMP-2)and hypoxia-inducible factor 1α(HIF-1α)in human scleral fibroblasts(HSF)and their mechanism.METHODS: The cells were cultured with IGF-1 and PI3K/AKT pathway inhibitor LY294002, respectively. CCK-8 method was used to detect cell viability and determine the optimal concentration and time of drug action. Cell migration activity was observed by cell scratch method. To determine the effects of IGF-1 on HSF cells and the regulatory role of PI3K/AKT pathway, HSF cells were divided into control group(without drugs), IGF-1(80 μg/L)group, IGF-1+LY294002(80 μg/L+5 mmol/L)group, and LY294002(5 mmol/L)group, and were cultured for 24 h; the protein expression levels of TGF-β2, MMP-2, HIF-1α, PI3K and AKT were detected by Western blot; the fluorescence expression of TGF-β2, MMP-2 and HIF-1α was detected by cellular immunofluorescence.RESULTS: The results of CCK-8 showed that the cell viability of the 80 μg/L IGF-1 group cultured with different concentrations of IGF-1 was the highest(all P<0.05), and the cell viability of the 80 μg/L IGF-1 group at 24 h was the highest under different culture times. Therefore, the concentration of IGF-1 was selected as 80 μg/L for 24 h. The viability of cells cultured with different concentrations of LY294002 gradually decreased from 6 h(all P<0.05). According to the IC50 value, therefore, the concentration of LY294002 was selected as 5 mmol/L for 24 h. The cell scratch results showed that compared with the control group, the cell mobility of 40 μg/L and 80 μg/L IGF-1 groups was increased(all P<0.05). Compared with the control group, cell mobility in the 2.5 and 5 mmol/L LY294002 groups was decreased(all P<0.05). Western blot results showed that compared with the control group, the protein expressions of TGF-β2, MMP-2, HIF-1α, PI3K and AKT in the IGF-1 group were increased, while those in the LY294002 group were decreased(all P<0.05). Compared with the IGF-1 group, the expression levels of TGF-β2, MMP-2, HIF-1α, PI3K and AKT in the IGF-1+LY294002 group were decreased(all P<0.05). The results of cell immunofluorescence showed that compared with the control group, the fluorescence expressions of TGF-β2, MMP-2 and HIF-1α in the IGF-1 group were increased, while those in the LY294002 group were decreased(all P<0.05). Compared with the IGF-1 group, the fluorescence expressions of TGF-β2, MMP-2 and HIF-1α in the IGF-1+LY294002 group were significantly decreased(all P<0.05).CONCLUSION: IGF-1 promoted the proliferation and migration of human HSF. IGF-1 may up-regulate the expression of TGF-β2, MMP-2 and HIF-1α in HSF through the PI3K/AKT signaling pathway, and participate in the occurrence and development of myopia.

OBJECTIVE To observe the effects of sodium p-hydroxybenzoate on the central nervous system and cardiovascular system of experimental animals. METHODS Kunming mice were given a single dose of sodium p-hydroxybenzoate of 20, 50 and 100 mg·kg-1 by oral gavage, the effects of sodium p-hydroxybenzoate on the central nervous system were observed by mice tail-flick experiment, mice autonomic activity experiment, pole-climbing experiment, coordinating hypnosis test and Morris water maze experiment. SD rats were given a single dose of sodium p-hydroxybenzoate of 14, 35 and 70 mg·kg-1 and Beagle dogs were given a single dose of sodium p-hydroxybenzoate of 4.2, 10.5 and 21 mg·kg-1 by oral gavage, the effects of sodium p-hydroxybenzoate on cardiovascular system and body temperature were observed by measuring blood pressure and body temperature in Beagle dogs, and measuring electrocardiogram in SD rats. RESULTS There was no significant influence of sodium p-hydroxybenzoate on sensory-motor reflex, autonomic activity, coordinated movements, sleep rate of mice with the sub-threshold sleep dose of pentobarbital sodium and learning-memory ability. Similarly, there were no significant effects on electrocardiogram of SD rats and there were no significant effects on blood pressure and body temperature of Beagle dogs. CONCLUSION Single oral gavage of sodium p-hydroxybenzoate has no significant effects on the cardiovascular system and the central nervous system of experimental animal under the condition.

Aim To investigate the effect of licochalcone A (LCA) on proliferation and apoptosis of osteosarcoma HOS and U2OS cells and to explore its possible molecular mechanism. Methods The HOS and U2OS cells were cultured in vitro. MTT assay was used to detect the proliferation of the cells after being treated with different concentrations of LCA at different intervention time. Then HOS and U2OS cells were treated with 0. 1% DMSO, or different concentrations of LCA (5, 10, 20 μmol/L), and flow cytometry was used to assess the cell apoptosis. The expression of apoptosis-related protein cleaved PARP1, Bcl-2, Bax, and Akt, ERK were detected by Western blot. The antitumor effect of LCA was detected on U2OS xenograft mice in vivo. Results LCA could inhibit the proliferation of HOS and U2OS cells in a time-and dose-dependent manner. Flow cytometry showed that LCA treatment could induce cell apoptosis. Western blot results showed that LCA could inhibit the phosphorylation of Akt and ERK, increase the expression of cleaved PARP1 and Bax, and decrease the expression of Bcl-2. In the tumor-bearing mouse models, LCA significantly decreased the tumor volume (P < 0. 05) and weight (P < 0. 01). Conclusions LCA could inhibit the proliferation of HOS and U2OS and induce apoptosis possibly by inhibiting the Akt/ERK signaling pathway.

OBJECTIVETo investigate the effect of Luteolin on proliferation and cell death of human multiple myeloma cell line RPMI-8226 and its mechanisms.

METHODSThe effect of Luteolin on the growth of human multiple myeloma cell line RPMI-8226 was detected by CCK-8, and then the minimal effective concentration was determined and was used to treat RPMI-8226 cells. The effects of luteolin and chloroquine on expression of cleaved-caspase 3 and LC3 in RPMI8826 cells was detected by Western blot.

RESULTSThe luteolin significant inhibited RPMI-8226 cell proliferation in a dose-dependent manner. Treatment with lutedin 40-80µmol/L for 24 hours and luteolin 20-80µmol/L for 48 hours inhibited RPMI-8226 cell proliferation in a dose-dependent manner (24 h, r= -0.983; 48 h, r= -0.985). After treatment with lutelin 20µmol/L for 48 h, the expression of cleaved caspase3 and LC3 Ⅱ/Ⅰ in RPMI 8826 cells significantly increased; after treatment with chloroquine at the same time, the expression of cleaved-caspase 3 and LC3 Ⅱ/Ⅰ significantly decreased.

CONCLUSIONLuteolin inhibits the proliferation of RPMI-8226 cells by inducing the pathways of both apoptosis and autophagy, moreover the actions of apoptosis and autophagy are interactive or/and promotive each other.

OBJECTIVE: To study the effects of epigallocatechin-3-gallate (EGCG) on proliferation and apoptosis of nasopharyngeal carcinoma CNE-2 cell line and analyze the expression of Bcl-2, Bax and Caspase-3 in the cell line which treated with EGCG. METHOD: MTT assay and flow cytometry were used to analyze cell proliferation and cell cycle. Hoechst33258 fluorescence staining was adopted to study cell apoptosis. RT-PCR was used to detect the expression of Bcl-2, Bax, Caspase-3. RESULT: EGCG could significantly inhibit proliferation of CNE-2 cell line and induce its apoptosis with dose-independent relationship. EGCG could suppress the expression of Bcl-2 and induce expression of Bax, Caspase-3. CONCLUSION EGCG in vitro has efficacy of anti-nasopharyngeal carcinoma cells, which may be through regulating the expression of cell proliferation and apoptosis genes involved.
Apoptosis ; drug effects ; Carcinoma ; Caspase 3 ; metabolism ; Catechin ; analogs & derivatives ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Humans ; Nasopharyngeal Carcinoma ; Nasopharyngeal Neoplasms ; metabolism ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; bcl-2-Associated X Protein ; metabolism