Phytochemical profile and biological activities of Bromelia antiacantha extracts

Abstract Reports from popular medicine usually act as a basis for the development of new drugs from natural compounds with therapeutic actions for serious diseases and prevalence such as cancer. Bromelia antiacantha Bertol. is a species of the Bromeliaceae family, considered an unconventional food plant, found in the south and midwest regions of Brazil. Despite the high nutritional content and pharmacological potential of its fruits, few scientific studies report its biological actions. Thus, this study evaluates the phytochemical profile of aqueous and ethanol extracts obtained from B. antiacantha fruits, as well as their possible antioxidant, antitumor, and cytotoxic activities. The aqueous extract exhibited phenolic compounds and flavonoids, while ethanol extracts indicated the presence of flavonoids and coumarin in their composition, regardless of the region of collection. The ethanolic extract demonstrated a more promising antioxidant effect than the aqueous extract and also induced a significant inhibition in the viability of human cervical cancer cells of the SiHa strain. In addition, treatment with both extracts did not alter the viability of non-tumor cells of the immortalized human keratinocyte lineage (HaCaT). These results bring new data about extracts obtained from a native plant, edible and traditionally used in popular medicine, opening new perspectives for its possible therapeutic application.

Resumo Relatos da medicina popular costumam atuar como referencial para o desenvolvimento de novos fármacos a partir de moléculas naturais com ações terapêuticas para doenças de alta gravidade e prevalência como o câncer. Bromelia antiacantha Bertol. é uma espécie da família Bromeliaceae, considerada uma planta alimentícia não convencional (PANC), encontrada nas regiões sul e centro-oeste do Brasil. Apesar do alto teor nutritivo e potencial farmacológico de seus frutos, poucos estudos científicos relatam suas ações biológicas. Desta forma, este estudo avalia o perfil fitoquímico de extratos aquoso e etanólico obtidos de frutos de B. antiacantha, bem como a sua possível ação antioxidante, antitumoral e citotóxica. O extrato aquoso apresentou compostos fenólicos e flavonoides, enquanto os extratos etanólicos apontam a presença de flavonóides e cumarina em sua composição, independente da região de coleta. O extrato etanólico demonstrou efeito antioxidante mais promissor do que o extrato aquoso e também induziu uma inibição significativa na viabilidade de células humanas de câncer cervical da linhagem SiHa. Além disso, o tratamento com ambos extratos não alterou a viabilidade de células não tumorais da linhagem de queratinócitos humanos imortalizados (HaCaT). Estes dados trazem novas informações sobre extratos obtidos de uma espécie vegetal nativa, comestível e já utilizada tradicionalmente, mas abrindo novas perspectivas quanto a possíveis aplicações terapêuticas.

Phytochemical profile and biological activities of Bromelia antiacantha extracts / Perfil fitoquímico e atividades biológicas de extratos de Bromelia antiacantha

Reports from popular medicine usually act as a basis for the development of new drugs from natural compounds with therapeutic actions for serious diseases and prevalence such as cancer. Bromelia antiacantha Bertol. is a species of the Bromeliaceae family, considered an unconventional food plant, found in the south and midwest regions of Brazil. Despite the high nutritional content and pharmacological potential of its fruits, few scientific studies report its biological actions. Thus, this study evaluates the phytochemical profile of aqueous and ethanol extracts obtained from B. antiacantha fruits, as well as their possible antioxidant, antitumor, and cytotoxic activities. The aqueous extract exhibited phenolic compounds and flavonoids, while ethanol extracts indicated the presence of flavonoids and coumarin in their composition, regardless of the region of collection. The ethanolic extract demonstrated a more promising antioxidant effect than the aqueous extract and also induced a significant inhibition in the viability of human cervical cancer cells of the SiHa strain. In addition, treatment with both extracts did not alter the viability of non-tumor cells of the immortalized human keratinocyte lineage (HaCaT). These results bring new data about extracts obtained from a native plant, edible and traditionally used in popular medicine, opening new perspectives for its possible therapeutic application.

Relatos da medicina popular costumam atuar como referencial para o desenvolvimento de novos fármacos a partir de moléculas naturais com ações terapêuticas para doenças de alta gravidade e prevalência como o câncer. Bromelia antiacantha Bertol. é uma espécie da família Bromeliaceae, considerada uma planta alimentícia não convencional (PANC), encontrada nas regiões sul e centro-oeste do Brasil. Apesar do alto teor nutritivo e potencial farmacológico de seus frutos, poucos estudos científicos relatam suas ações biológicas. Desta forma, este estudo avalia o perfil fitoquímico de extratos aquoso e etanólico obtidos de frutos de B. antiacantha, bem como a sua possível ação antioxidante, antitumoral e citotóxica. O extrato aquoso apresentou compostos fenólicos e flavonoides, enquanto os extratos etanólicos apontam a presença de flavonóides e cumarina em sua composição, independente da região de coleta. O extrato etanólico demonstrou efeito antioxidante mais promissor do que o extrato aquoso e também induziu uma inibição significativa na viabilidade de células humanas de câncer cervical da linhagem SiHa. Além disso, o tratamento com ambos extratos não alterou a viabilidade de células não tumorais da linhagem de queratinócitos humanos imortalizados (HaCaT). Estes dados trazem novas informações sobre extratos obtidos de uma espécie vegetal nativa, comestível e já utilizada tradicionalmente, mas abrindo novas perspectivas quanto a possíveis aplicações terapêuticas.

Monotropein Induced Apoptosis and Suppressed Cell Cycle Progression in Colorectal Cancer Cells / 中国结合医学杂志

OBJECTIVE@#To determine whether monotropein has an anticancer effect and explore its potential mechanisms against colorectal cancer (CRC) through network pharmacology and molecular docking combined with experimental verification.@*METHODS@#Network pharmacology and molecular docking were used to predict potential targets of monotropein against CRC. Cell counting kit assay, plate monoclonal assay and microscopic observation were used to investigate the antiproliferative effects of monotropein on CRC cells HCT116, HT29 and LoVo. Flow cytometry and scratch assay were used to analyze apoptosis and cell cycle, as well as cell migration, respectively in HCT116, HT29, and LoVo cells. Western blotting was used to detect the expression of proteins related to apoptosis, cell cycle, and cell migration, and the expression of proteins key to the Akt pathway.@*RESULTS@#The Gene Ontology and Reactome enrichment analyses indicated that the anticancer potential of monotropein against CRC might be involved in multiple cancer-related signaling pathways. Among these pathways, RAC-beta serine/threonine-protein kinase (Akt1, Akt2), cyclin-dependent kinase 6 (CDK6), matrix metalloproteinase-9 (MMP9), epidermal growth factor receptor (EGFR), cell division control protein 42 homolog (CDC42) were shown as the potential anticancer targets of monotropein against CRC. Molecular docking suggested that monotropein may interact with the 6 targets (Akt1, Akt2, CDK6, MMP9, EGFR, CDC42). Subsequently, cell activity of HCT116, HT29 and LoVo cell lines were significantly suppressed by monotropein (P<0.05). Furthermore, our research revealed that monotropein induced cell apoptosis by inhibiting Bcl-2 and increasing Bax, induced G1-S cycle arrest in colorectal cancer by decreasing the expressions of CyclinD1, CDK4 and CDK6, inhibited cell migration by suppressing the expressions of CDC42 and MMP9 (P<0.05), and might play an anticancer role through Akt signaling pathway.@*CONCLUSION@#Monotropein exerts its antitumor effects primarily by arresting the cell cycle, causing cell apoptosis, and inhibiting cell migration. This indicates a high potential for developing novel medication for treating CRC.

Effect and mechanism of gracillin-induced autophagy in lung cancer A549 cells / 中国药房

OBJECTIVE To investigate the effect and mechanism of gracillin from Reineckia carnea on autophagy in non- small cell lung cancer A549 cells. METHODS Using A549 cells as subjects， the effects of different concentrations of gracillin （0.25， 0.5， 1， 2， 4 μmol/L） on the proliferation of cells were detected by CCK-8 after being treated for different time （12， 24， 48 h）. Compared with the control group without medication， the effect of gracillin （2 μmol/L） on the formation of autophagosomes in cells was observed by transmission electron microscope after 24 h of exposure. The aggregation of GFP-LC3 on autophagosome membrane was detected by GFP-LC3 plasmid transfection after being treated with gracillin （0.25， 0.5， 1， 2 μmol/L） for 24 h. Quantitative real-time PCR and Western blot assay were used to detect the mRNA and protein expressions of family with sequence similarity 102 member A（FAM102A）， the expressions of autophagy-related proteins [p62， Beclin-1， microtubule-associated protein 1 light chain 3B （LC3B）]， and the expressions of phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway-related proteins in A549 cells after being treated with gracillin （0.25， 0.5， 1 and 2 μmol/L） for 24 h. RESULTS Gracillin significantly inhibited the proliferation of A549 cells in a concentration- and time-dependent manner. The IC50 was 2.55 μmol/L at 24 h. After 24 h of gracillin treatment， autophagosomes with bilayer membrane structure were found in the cell cytoplasm， and GFP-LC3 green fluorescent spots on autophagosome membrane were obvious， representing an increasing trend as drug concentration. Compared with the control group， mRNA and protein expressions of FAM102A （0.5， 1， 2 μmol/L groups）， protein expression of Beclin-1 （1， 2 μmol/L groups） and LC3B-Ⅱ/LC3B-Ⅰ ratio （2 μmol/L group） were significantly increased in different concentrations of gracillin groups， while the protein expression of p62 （1， 2 μmol/L groups）， and the protein phosphorylations of Akt （1， 2 μmol/L groups） and PI3K （2 μmol/L group） were all decreased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS Gracillin can promote excessive autophagy in A549 cells by up-regulating mRNA and protein expressions of FAM102A and inhibiting PI3K/Akt signaling pathway， thus inhibiting cell proliferation.

Diallyl disulfide augments sensitivity of DJ-1 overexpressed human gastric cells to 5-FU / 中国药理学通报

Aim To investigate whether diallyl disul-fide (DADS) augments the sensitivity of DJ-1 (protein/ nucleic acid deglycase) overexpressed human gastric SGC7901 cells to 5-FU (5-fluorouracil). Methods The experimental groups include control group, DADS group, VCR (vincristine) group, VCR + DADS group, DJ-1 group, DJ-1 + DADS group. MTT was used to analyze the effect of DADS on 5 -FU (5 -fluorou- racil) induced proliferation inhibition. Flow cytometry was performed to examine the effect of DADS on cell apoptosis. RT-PCR, Western blot, and immunofluo-rescence were used for determine the effect of DADS on the drug resistance associated gene expression. Results DADS enhanced the proliferation inhibitory effect of 5-FU on DJ-1 overexpressed cells and VCR resistant cells. DADS could induce apoptosis in VCR-resistant cells. DADS downregulated the expression of DJ-1 while inducing apoptosis in DJ-1 overexpressed cells. DJ-1 overexpression upregulated the expression of P-gp (P-glycoprotein), Bcl-2, and XIAP (X-linked inhibitor of apoptosis protein), downregulated the expression of caspase-3. DADS decreased the expression of P-gp, Bcl-2, and XIAP, while increased the expression of caspase-3 in DJ-1 overexpressed cells and VCR-resistant cells. Conclusions DADS can augment the sensitivity of DJ-1 overexpressed cells to 5-FU, which is related to its antagonism against DJ-1 mediated upregula- tion of P-gp, Bcl-2, XIAP, and downregulation of caspase-3.

Silver nanoparticles-resistance of HeLa cell associated with its unusually high concentration of α-ketoglutarate and glutathione / 生物工程学报

Silver nanoparticles (AgNPs) is known as one of the most valuable metal nanoparticles in antibacterial and anticancer application. AgNPs-resistant bacteria has been documented, but it is unclear whether cancer cells can also escape the anti-cancer effect of AgNPs. In this study, we aimed to investigate this phenomenon and its underlying mechanism. The antibacterial activity and cytotoxicity of AgNPs were measured in the presence of HeLa cell metabolites. The status of AgNPs in the system associated with metabolites were characterized by UV-Vis, Zetasizer Nano ZS, and transmission electron microscopy. Non-targeted metabolomics was used to reveal the metabolites components that bind with AgNPs. HeLa cells were injected intraperitoneally to establish the tumor-bearing mice model, and the stability of AgNPs in mice serum was analyzed. The results manifested that HeLa cell metabolites inhibited the anticancer and antibacterial effects of AgNPs in a dose-dependent manner by causing AgNPs aggregation. Effective metabolites that inhibited the biological activity of AgNPs were stable in 100 ℃, insoluble in chloroform, containing sulfur elements, and had a molecular weight less than 1 kDa in molecular weight. There were 115 compounds bound with AgNPs. In vitro experiments showed that AgNPs aggregation occurred only when the concentration of α-ketoglutarate (AKG) and glutathione (GSH) together reached a certain threshold. Interestingly, the concentration of AKG and GSH in HeLa cellular metabolites was 10 and 6 times higher than that in normal cervical epithelial cells, respectively, which explained why the threshold was reached. Furthermore, the stability of AgNPs in the serum of tumor-bearing mice decreased by 20% (P < 0.05) compared with the healthy mice. In conclusion, our study demonstrates that HeLa cells escaped the anti-cancer effect of AgNPs through the synergistic effect of AKG and GSH, suggesting the need to develop strategies to overcome this limitation.

Effects of live attenuated measles vaccine Hu191 strain on epithelial mesenchymal transition,proliferation and migration of 4T1 breast cancer cells / 中国生物制品学杂志

@#Objective To investigate the effects of live attenuated measles vaccine Hu191 strain(MV-Hu191)on epithelial mesenchymal transition(EMT),proliferation and migration of 4T1 breast cancer cells.MethodsCCK-8 and clone formation assay were used to analyze the effect of MV-Hu191 on the proliferation of 4T1 cells;The effect of MV-Hu191 on 4T1cell migration was analyzed by cell scratch test;The expression of EMT pathway proteins(MMP-2,MMP-9,E-cadherin)in4T1 cells was detected by Western blot;4T1 tumor-bearing mouse model was established in female BALB/c mice. The model mice were divided into control group(PBS),MV-Hu191(1 × 106TCID50)group and paclitaxel group(15 mg/kg),with 10 mice in each group,and injected into tumor at the dosage of 100 μL every 2 d for 5 times. At 28 d after administration,the effects of MV-Hu191 on survival time,tumorigenicity and metastasis in vivo were observed;The pathological characteristics of lung tissue and tumor tissue were observed by HE staining under microscope;The expression of EMT pathway proteins(MMP-2,MMP-9 and E-cadherin)in tumor tissue was detected by immunohistochemical staining.Results The results of in vitro experiment showed that,compared with the control group,MV-Hu191 inhibited the proliferation and migration of 4T1 cells(F = 2. 811 and 13. 535,P = 0. 001 and 0. 002,respectively),down regulated the expression of MMP-2 and MMP-9(F = 45. 433 and 9. 744,P = 0. 011 and 0. 038,respectively),and up regulated the expression of Ecadherin(F = 7. 001,P = 0. 032);The results of in vivo experiment showed that MV-Hu191 significantly prolonged the survival time of tumor-bearing mice,and decreased the tumor quality(F = 8. 301,P = 0. 003)and the number of pulmonary nodules metastasis compared with the control group(F = 33. 792,P = 0. 000);MV-Hu191 treated tumor tissue gap was small,the cells were round,and the alveolar contour was clearly visible;The expression of MMP-2 and MMP-9 in MVHu191 treated tumor tissue decreased significantly(F = 6. 705 and 9. 047,P = 0. 028 and 0. 023,respectively),while the expression of E-cadherin increased significantly(F = 3. 468,P = 0. 039).ConclusionMV-Hu191 signi-ficantly inhibits the proliferation and migration of 4T1 breast cancer cells,antagonizes the tumorigenicity and lung meta-stasis of 4T1 tumorbearing mice,and prolongs the survival time of mice. The possible mechanism of MV-Hu191 against breast cancer is closely related to the regulation of EMT pathway protein expression.

Baicalin Induces Ferroptosis in Gastric Cancer Cells via p53-mediated SLC7A11 Down-regulation / 中国实验方剂学杂志

ObjectiveTo explore the inhibitory effect of different concentration of baicalin （0， 100， 200， 400 μmol·L-1） on the proliferation of human gastric cancer SGC-7901 cells and the underlying mechanism. MethodSGC-7901 cells were treated with baicalin. Then methyl thiazolyl tetrazolium （MTT） assay was employed to examine the inhibitory effect of baicalin on the cells. At the same time， ferrostatin-1 （Fer-1） was added to observe the viability of cells after baicalin treatment. The expression of ferroptosis-related genes was detected by Real-time polymerase chain reaction （Real-time PCR） and Western blot. The content of malondialdehyde （MDA） and the level of glutathione （GSH） were detected respectively by MTT assay and enzyme-linked immunosorbent assay. The role of tumor protein 53 （p53）/solute carrier family 7 member 11 （SLC7A11） pathway in the regulation of ferroptosis was investigated respectively via overexpression and small interfering RNA （siRNA） methods. ResultCompared with the blank group， baicalin decreased the viability of SGC-7901 （P<0.05， P<0.01） in a dose- and time-dependent manner. The intervention of Fer-1 significantly alleviated the decrease of SGC-7901 cell viability caused by baicalin （P<0.01）. In addition， compared with the baicalin group， Fer-1+baicalin group showed decrease in MDA content and the mRNA and protein levels of prostaglandin-endoperoxide synthase 2 （PTGS2） in the cells （P<0.01）， and increase in GSH activity and mRNA and protein levels of glutathione peroxidase 4 （GPX4） （P<0.01）. The protein level of SLC7A11 in the baicalin group was decreased compared with that in the blank group （P<0.05， P<0.01） in a dose-dependent manner. Compared with the baicalin group， the reactive oxygen species （ROS） level and MDA content in SLC7A11-overexpressing cells were significantly decreased after baicalin treatment （P<0.01）， and the GSH activity was significantly increased （P<0.01）. The fluorescence intensity of p53 in the cells of the baicalin group was increased compared with that of the blank group （P<0.01）. Compared with the baicalin group， the expression level of p53 protein in the cells transfected with p53 siRNA was significantly decreased after baicalin treatment （P<0.01）， and the expression level of SLC7A11 was significantly increased （P<0.01）. ConclusionBaicalin can effectively inhibit the proliferation of SGC-7901 cells by regulating p53/SLC7A11-mediated ferroptosis.

Oenothein B inhibits proliferation and migration of breast cancer cells by regulating P53 / 中国中药杂志

The effects of oenothein B(OEB) on the proliferation, apoptosis, invasion, and migration of breast cancer MCF-7 and MDA-MB-231 cells were investigated by cell culture in vitro, network pharmacology, and molecular docking. In vitro cell experiments revealed that OEB inhibited the proliferation and colony formation ability, and promoted the apoptosis and formation of apoptotic bodies in breast cancer cells, as well as inhibited the invasion and migration of breast cancer cells. The targets of OEB were obtained using SwissTargetPrediction database and breast cancer targets were obtained from GeneCards. The targets of OEB and breast cancer were entered separately in Venny 2.1 software to obtain the Venn diagram of common targets of OEB and breast cancer. The common targets of OEB and breast cancer were input into STRING database to construct a protein-protein interaction(PPI) network, which was entered into Cytoscape 3.7.2 software for network topology analysis. Key targets were screened according to protein association strength, and analyzed for KEGG pathway enrichment. Molecular docking of OEB to key targets using AutoDock software revealed that OEB stably bound to the active pocket of P53, while OEB promoted the expression of P53 protein. MCF-7 and MDA-MB-231 cell viability and migration ability increased after silencing P53, and this change was reversed after treatment with OEB. Therefore, this study showed that OEB inhibited the proliferation, migration, and invasion of breast cancer MCF-7 and MDA-MB-231 cells, and promoted the apoptosis of breast cancer MCF-7 and MDA-MB-231 cells, which may be related to the targeted regulation of P53.

Tumor cell lysate with low content of HMGB1 enhances immune response of dendritic cells against lung cancer in mice / 南方医科大学学报

OBJECTIVE@#To assess the effect of tumor cell lysate (TCL) with low high-mobility group B1 (HMGB1) content for enhancing immune responses of dendritic cells (DCs) against lung cancer.@*METHODS@#TCLs with low HMGB1 content (LH-TCL) and normal HMGB1 content (NH-TCL) were prepared using Lewis lung cancer (LLC) cells in which HMGB1 was inhibited with 30 nmol/L glycyrrhizic acid (GA) and using LLC cells without GA treatment, respectively. Cultured mouse DCs were exposed to different doses of NH-TCL and LH-TCL, using PBS as the control. Flow cytometry was used to detect the expressions of CD11b, CD11c and CD86 and apoptosis of the stimulated DCs, and IL-12 levels in the cell cultures were detected by ELISA. Mouse spleen cells were co-cultured with the stimulated DCs, and the activation of the spleen cells was assessed by detecting CD69 expression using flow cytometry; TNF-β production in the spleen cells was detected with ELISA. The spleen cells were then co-cultured with LLC cells at the effector: target ratios of 5:1, 10:1 and 20:1 to observe the tumor cell killing. In the animal experiment, C57/BL6 mouse models bearing subcutaneous LLC xenograft received multiple injections with the stimulated DCs, and the tumor growth was observed.@*RESULTS@#The content of HMGB1 in the TCL prepared using GA-treated LLC cells was significantly reduced (P < 0.01). Compared with NH-TCL, LH-TCL showed a stronger ability to reduce apoptosis (P < 0.001) and promote activation and IL- 12 production in the DCs. Compared with those with NH-TCL stimulation, the DCs stimulated with LH-TCL more effectively induced activation of splenic lymphocytes and enhanced their anti-tumor immunity (P < 0.05). In the cell co-cultures, the spleen lymphocytes activated by LH-TCL-stimulated DCs showed significantly enhanced LLC cell killing activity (P < 0.01). In the tumor-bearing mice, injections of LH-TCL-stimulated DCs effectively activated host anti-tumor immunity and inhibited the growth of the tumor xenografts (P < 0.05).@*CONCLUSION@#Stimulation of the DCs with LH-TCL enhances the anti-tumor immune activity of the DCs and improve the efficacy of DCbased immunotherapy for LLC in mice.

Porphyromonas gingivalis enhances the proliferation of colorectal cancer Caco-2 cells via the JAK2-STAT3 pathway / 口腔疾病防治

Objective @# To investigate the effect of pathogenic bacterium-Porphyromonas gingivalis (P.g) on the proliferation and inflammatory factor expression of human colorectal cancer Caco-2 cells, to determine whether the Janus kinase 2-signal transducers and activators of transcription 3 (JAK2-STAT3) pathway is involved in the regulation of Caco-2 cell proliferation by P.g and to provide an experimental basis for further exploring the relationship between P.g and colorectal cancer. @*Methods @# Caco-2 cells were cultured in vitro, and P.g at different multiplicities of infection (MOIs) (0, 1, 10, 25) was selected to stimulate for 12, 24 and 48 h. The effect of P.g on the proliferation of Caco-2 cells was detected by CCK8. The stimulation time was set as 12, 24 and 48 h. MOI=0 was the control group, and MOI=1, 10 and 25 comprised the experimental group. qRT-PCR and Western blot were used to detect the changes in interleukin-6 (IL-6), interleukin-10(IL-10), JAK2 and STAT3 gene and protein (phosphorylated protein) levels in each group. @*Results @# After P.g infection of Caco-2 cells, P.g had a sustained stimulatory effect on the cells for 12, 24 and 48 h at MOI=1 and MOI=10 compared with the control group. Compared with that in the control group, the expression of pro-inflammatory factor IL-6 and related proliferative pathway protein JAK2 and STAT3 in Caco-2 cells with P.g infection increased in a concentration- and time-dependent manner (P<0.05). Additionally, the expression of IL-10, an anti-inflammatory factor, in Caco-2 cells infected with P.g decreased (P<0.05). After the addition of the JAK2 inhibitor AZ960, the proliferation of Caco-2 cells infected with P.g decreased, and the mRNA expression of STAT3 and JAK2 and the protein expression of p-STAT3 and p-JAK2 decreased (P<0.05). @*Conclusion @#P.g can promote the proliferation of the colorectal cancer cell line Caco-2, and the effect of P.g on Caco-2 cells may promote cell proliferation through the JAK2-STAT3 pathway while promoting the expression of the proinflammatory factor IL-6 and inhibiting the expression of the anti-inflammatory factor IL-10, creating an inflammatory environment conducive to cell proliferation, which may be the mechanism by which P.g affects the proliferation of Caco-2 cells.

Jianpi Yangzheng Xiaozheng Decoction Affect Proliferation and Stemness of Gastric Cancer HGC-27 Cells by Inhibiting Aerobic Glycolysis / 中国实验方剂学杂志

ObjectiveTo observe the effect of Jianpi Yangzheng Xiaozheng decoction （JYXD） on the proliferation and stemness of the human gastric cancer （GC） cell line HGC-27 by inhibiting aerobic glycolysis， and explore the underlying mechanism. MethodMethyl thiazolyl tetrazolium （MTT） assay was employed to determine the survival rate and chemotherapy sensitivity of HGC-27 cells treated with JYXD （0.25， 0.5， 1， 2， 4， 8， 16， 32 g·L-1）. Colony formation assay was employed to detect the effect of JYXD （2， 4， 8 g·L-1） on the colony formation of the cells. The aerobic glycolysis level of HGC-27 cells after treatment with JYXD was measured by glucose assay kit and lactic acid assay kit. The proportion of stem cell subsets in HGC-27 cells was detected by flow cytometry. Western blot was employed to determine the expression of glycolysis-associated proteins such as lactate dehydrogenase （LDH）， hexokinase 2 （HK2）， glucose transporter 1 （GLUT1）， and pyruvate kinase isozyme M2 （PKM2）， and the expression of stemness-associated proteins such as octamer-binding transcription factor 4 （OCT4）， SRY-box transcription factor 2 （SOX2）， and Nanog. ResultJYXD （0.5， 1， 2， 4， 8， 16， 32 g·L-1） inhibited the activity of HGC-27 cells （P<0.05， P<0.01）， with the inhibitory concentration 50（IC50） of 4.83 g·L-1， and it improved the sensitivity of HGC-27 cells to cisplatin chemotherapy. Compared with the control group， JYXD （2， 4， 8 g·L-1） reduced the colony formation number of HGC-27 cells （P<0.01） in a concentration-dependent manner. Flow cytometry showed that compared with that in the control group， the proportion of CD44+CD24+ALDH+ population in the cells treated with JYXD （2， 4， 8 g·L-1） decreased （P<0.05）. In addition， JYXD （2， 4， 8 g·L-1） inhibited the glucose uptake and lactic acid production of HGC-27 cells. Western blot showed that compared with the control group， JYXD （2， 4， 8 g·L-1） down-regulated the expression levels of SOX2， Nanog， OCT4， PKM2， LDH， GLUT1， and HK2 （P<0.05， P<0.01） in a concentration-dependent manner. ConclusionJYXD may inhibit the proliferation and reduce the stemness of HGC-27 cells by regulating the aerobic glycolysis.

Baicalein promotes laryngeal cancer cell death and inhibits invasion via miR-125b-5p/IRF4 axis / 中国临床药理学与治疗学

AIM: To investigate the mechanism of baicalin-induced apoptosis in human laryngeal cancer cells. METHODS: AMC-HN-8 cells were selected for the study, and baicalin was applied to the cells at different concentrations (0, 10, 30, 100, and 300 μmol/L), and the half-inhibitory concentration (IC50) was measured by the CCK-8 method. Bax, cleaved-caspase-3, Cyto-c, IRF4 protein expression by protein blotting (Western blot); miR-125b-5p and IRF4 expression by RT-qPCR. Dual-luciferase reporter gene validation of Targetscan prediction (binding of miR-125b-5p to IRF4-3'UTR); apoptosis and necrosis inhibitors explore the way baicalein induces death in laryngeal cancer cells. AMC-HN-8 was then divided into blank group, baicalein (IC50), miR-125b-5p inhibitor group, baicalein + inhibitor NC group, baicalein+miR-125b-5p inhibitor group, and cell invasion and clone formation assays to detect cell invasion and proliferation ability, respectively. Apoptosis was detected by flow cytometry. RESULTS: Baicalein inhibited the proliferation of AMC-HN-8 cells in a dose-dependent manner with an IC50 value of 47.31 μmol/L. Compared with the blank group, 47.31 μmol/L baicalin induced apoptosis and inhibited cell invasion, while upregulating the expression of miR-125b-5p and suppressing the mRNA and protein levels of IRF4. The luciferase results showed that the miR-125b-5p mimic was able to inhibit the activity of the IRF4-3'UTR promoter relative to the NC mimic (mimic) group. Baicalein induces laryngeal cancer cell death in an apoptotic manner. In addition, the combination of 47.31 μmol/L baicalin and miR-125b-5p inhibitor affected the behavior of AMC-HN-8 cells, showing that compared with the blank group, the baicalin group showed a decrease in the number of cell clones, weakened invasion ability, and increased apoptosis; the miR - 125b-5p inhibitor group showed an increase in the number of cell clones, enhanced invasion ability and decreased apoptosis. The baicalin+ inhibitor NC group was consistent with baicalin, with no significant effect of inhibitor NC on cell behavior. The cloning, invasion, and apoptosis of cells in the baicalin+miR-125b-5p inhibitor group were intermediate between the baicalin and miR-125b-5p inhibitor groups. CONCLUSION: Baicalin inhibits the proliferation of AMC-HN-8 cells, and the mechanism may be related to miR-125b-5p targeting to inhibit the expression of IRF4, inducing the pro-apoptotic proteins Bax, cleaved-caspase3, and Cyto-c, and inhibiting the apoptosis suppressor protein Bcl-2 thereby inducing apoptosis.

Molecular mechanism of a Matijin-su derivative for inhibiting proliferation, invasion and migration of prostate cancer cells / 中国药理学通报

Aim To investigate the effects of HXL130 on the proliferation, invasion and migration of prostate cancer PC3 cells and its molecular mechanism. Methods MTT assay was used to detect the effect of HXL130 on the proliferation of prostate cancer PC3 cells. Hoechst 33258 staining and flow cytometry were used to detect the effects on apoptosis and cell cycle of cancer cells. Transwell was used to detect the effects of compounds on the invasion and migration of cancer cells. Proteomic sequencing was employed to detect differentially expressed proteins (DEPs) induced by compound treatment of cancer cells. Bioinformatics was used to analyze the functions of DEPs and the related signaling pathways regulated by DEPs, and Western blot was used to verify the result. Results The survival rate of PC3 cells decreased with the increase of HXL130 concentration and treatment time. HXL130 could significantly induce cell apoptosis and block G

Daidzein affects proliferation and apoptosis in non-small cell lung cancer cells:role of p53 signaling pathway / 中国药理学通报

Aim To investigate the effects of daidzeinDD on the proliferation and apoptosis of non-small cell lung cancer cells,with a focus on the possible role of the p53 signaling pathway in this regard. Methods CCK-8 method and flow cytometry were used to detect the effects of soy isoflavone crude extract and DD on the viability and apoptosis of HELF and H1299 cells. Gene microarray was used to detect the changes in gene expression after treatment of H1299 cells with DD. GSEA and differential analysis were used to screen the major pathways and key genes. RT-qPCR and Western blot were performed to verify the differences in mRNA and protein expression of key genesp53 and CASP9 in the major pathways. After p53 inhibitor Pifithrin-α inhibited the expression of p53,the effect of DD on p53 mRNA and protein expression levels was examined,and the proliferative effect on H1299 cells was observed. Results Soy isoflavone crude extract and DD promoted proliferation and inhibited apoptosis of normal lung cells and inhibited proliferation and promoted apoptosis of lung cancer cells. p53 signaling pathway was significantly enriched in the DD-treated groupNES=1.78,P=0.000,and the expressions of p53 and CASP9 genes were found to be significantly up-regulated in the treated group. Compared with the control group,mRNA expression of CASP9 and p53 significantly increased in both HELF and H1299 cells treated with DDP<0.05,and p53 protein expression also increased in HELF cellsP<0.05. After inhibition of p53 expression,DD significantly increased the mRNA expression of p53 in H1299 and HELF cellsP<0.05 and also markedly increased the expression of p53 protein in H1299 cellsP<0.05,and it was observed that DD inhibited the proliferation of lung cancer cells. Conclusions DD inhibits the proliferation and promotes the apoptosis of lung cancer H1299 cells,and the mechanism mainly involves the p53 signaling pathway.

The effects of curcumin on the biological behavior of colorectal cancer cells through the JAK/STAT3 and RAS/MAPK/NF-κB pathways / Los efectos de la curcumina en el comportamiento biológico de las células del cáncer colorrectal mediante las vías JAK/STAT3 y RAS/MAPK/NF-KB

Abstract The purpose of this work was to investigate the effects of curcumin on the biological behavior of colorectal cancer cells through the JAK/STAT3 and RAS/MAPK/NF-κB pathways. Human colorectal cancer HCT116 cells were cultured and divided into a control group and low, medium and high-dose curcumin groups (n =5). HCT116 colorectal cancer cells became long-growing cells after incubation and culture at 37°C. The control group was treated with 15μL phosphate-buffered saline, and the low-dose, medium-dose and high-dose curcumin groups were treated with 20, 40 and 80μmol/L curcumin, respectively. All groups were treated with relevant drug intervention, digested and centrifuged for 48h, washed twice with a PBS solution, centrifuged at 1000 rpm for 3 min, and the cells precipitated. The proliferation, apoptosis and growth cycle of cells in each group were observed, and the expressions of the JAK/STAT3 and RAS/MAPK/NF-κB pathways and related proteins in each group were studied. Compared with the curcumin low-dose and medium-dose groups, the proliferation ability of the curcumin high-dose group was significantly decreased (P<0.05). When the low-dose and medium-dose curcumin groups were compared with the high-dose curcumin group, the apoptosis ability was significantly increased (P<0.05). When the low-dose and medium-dose curcumin groups were compared, the growth ratio of the G0/G1 phase in the high-dose curcumin group was significantly increased, and the percentage of the S phase was significantly decreased (P<0.05). Compared with the curcumin low-dose and medium-dose groups, the expression of JAK-STAT3 and RAS/MAPK/NF-κB pathway in the curcumin high-dose group was significantly decreased (P<0.05). The protein expressions of STAT3, RAS, P-P38 and P65 in the curcumin high-dose group were significantly lower than those in the curcumin low-dose and medium-dose groups (P<0.05). Curcumin can inhibit the expression of JAK/STAT3 and RAS/MAPK/NF-κB pathways, block the growth cycle, and inhibit the proliferation and induce apoptosis of colorectal cancer cells, providing a new idea for the clinical treatment of colorectal cancer.

Resumen El objetivo del presente trabajo fue investigar los efectos de la curcumina en el comportamiento biológico de las células del cáncer colorrectal mediante el estudio de las vías JAK/STAT3 y RAS/MAPK/NF-KB. Las células del cáncer colorrectal humano HCT116 se cultivaron y dividieron en un grupo control y en grupos con dosis baja, media y alta (n = 5) de curcumina. Las células de cáncer colorrectal HCT116 se convirtieron en células de crecimiento prolongado después de la incubación y cultivo a 37°C. El grupo de control se trató con 15 μL de solución tampón fosfato salina (PBS) y los grupos de curcumina de dosis baja, media y alta se trataron con 20, 40 y 80 μmol/L de curcumina, respectivamente. Todos los grupos fueron tratados con la intervención farmacológica pertinente, digeridos y centrifugados durante 48 horas, lavados dos veces con solución de PBS, centrifugados a 1000 rpm durante 3 minutos, y las células precipitadas. Se observaron la proliferación, la apoptosis y el ciclo de crecimiento de las células de cada grupo, y fueron estudiados las expresiones de las vías JAK/STAT3, RAS/MAPK/NF-KB y proteínas relacionadas en cada grupo. Comparado con los grupos de la dosis baja y media de la curcumina, disminuyó obviamente la capacidad de proliferación del grupo de la dosis alta de la curcumina (P<0,05). Comparado con los grupos de la dosis baja y media de la curcumina, aumentó de modo significativo la capacidad de la apoptosis del grupo de la dosis alta de la curcumina (P<0,05). Comparado con los grupos de la curcumina de dosis baja y media, aumentó obviamente la proporción del crecimiento de la fase G0/G1 en el grupo de la curcumina de dosis alta y el porcentaje de la fase S disminuyó considerablemente (P<0,05). Las expresiones proteicas STAT3, RAS, P-P38 y P65 en el grupo de la dosis alta de la curcumina fueron evidentemente más bajas que las de los grupos de la dosis baja y media de la curcumina (P<0.05). La curcumina puede inhibir la expresión de las vías JAK/STAT3 y RAS/MAPK/NF-KB, bloquear el ciclo del crecimiento y luego inhibir la proliferación e inducir apoptosis de las células del cáncer colorrectal, lo que brinda una nueva idea para el tratamiento clínico del cáncer colorrectal.

Vitamina C y su acción en células cancerígenas

RESUMEN Introducción: La vitamina C es una sustancia que desde hace ya varios años ha suscitado debate debido a la cantidad de usos que se han demostrado. En algunos casos, las utilidades han ido desde profilaxis y acortamiento de la duración de resfriados, hasta estudios de acción en enfermedades, tales como el cáncer; los mecanismos de acción de esta han sido evaluados en forma de monoterapia o en combinación con quimioterapia para demostrar o descartar su utilidad en el cáncer. Objetivo: Demostrar si los efectos de la vitamina C fueron efectivos y si su uso, solo o en combinación con quimioterapia, es de utilidad. Método: Se realizó una investigación de tipo descriptiva documental, realizada con artículos científicos en el periodo comprendido desde 2016 hasta 2021, con un análisis detallado de los resultados del uso de la vitamina C y su posible efecto sobre los diferentes tipos de cáncer. Fueron buscados en las bases de datos de SciELO, Scopus y Medline. Resultados: La información hallada fue organizada según concentraciones plasmáticas de vitamina C y su acción en células cancerosas, dosis evaluadas de la vitamina C, mecanismos de acción en relación a células cancerígenas, desequilibrio redox, efecto específico en cánceres, vitamina C y cáncer de mama. Conclusiones: En la revisión realizada se evidencia que la vitamina C tiene un efecto benéfico en los cánceres hematopoyéticos, como: leucemia, melanoma, cáncer de mama o ciertos tipos de cáncer colorrectal y que disminuyen los efectos adversos producidos por medicamentos quimioterapéuticos.

ABSTRACT Introduction: Vitamin C is a water-soluble substance that has been in debate since a long time ago due to its wide demonstrated use. In some cases, its usage have ranged from prophylaxis and shortening the duration of colds, to studies of its action in diseases, such as cancer; Its mechanisms of action have been evaluated in the form of monotherapy or in combination with the chemotherapy to demonstrate or rule out its usefulness in cancer. Objective: To demonstrate whether the effects of vitamin C were effective and whether its use, alone or in combination with chemotherapy, is useful. Method: A documentary descriptive research was carried out, supported with scientific articles (period 2016 to 2021) which analyze in detail the results of the use of vitamin C and its possible effect on different types of cancer. Results: The information found was organized according to plasma concentrations of vitamin C, its action on cancer cells, evaluated doses of vitamin C, mechanisms of action in relation to cancer cells, redox imbalance, specific effect on cancers, vitamin C and breast cancer. Conclusions: The review shows that the use of vitamin C has a beneficial effect on hematopoietic cancers, such as leukemia, melanoma, breast cancer or certain types of colorectal cancer, and also reduces the adverse effects produced by chemotherapeutic drugs.

RESUMO Introdução: A vitamina C é uma substância que há vários anos desperta o debate devido ao número de usos que foram demonstrados. Em alguns casos, os benefícios vão desde a profilaxia e redução da duração dos resfriados, até estudos de ação em doenças, como o câncer; os mecanismos de ação deste foram avaliados como monoterapia ou em combinação com quimioterapia para provar ou descartar sua utilidade no câncer. Objetivo: Demonstrar se os efeitos da vitamina C foram eficazes e se seu uso, isoladamente ou em combinação com a quimioterapia, é útil. Método: Foi realizada uma pesquisa documental descritiva, realizada com artigos científicos no período de 2016 a 2021, com análise detalhada dos resultados do uso da vitamina C e seu possível efeito nos diferentes tipos de câncer. Eles foram pesquisados nas bases de dados SciELO, Scopus e Medline. Resultados: As informações encontradas foram organizadas de acordo com as concentrações plasmáticas de vitamina C e sua ação nas células cancerígenas, doses avaliadas de vitamina C, mecanismos de ação em relação às células cancerígenas, desequilíbrio redox, efeito específico sobre cânceres, vitamina C e câncer de mama. Conclusões: Na revisão realizada, fica evidente que a vitamina C tem efeito benéfico sobre os cânceres hematopoiéticos, como: leucemia, melanoma, câncer de mama ou certos tipos de câncer colorretal e que reduz os efeitos adversos produzidos pelos quimioterápicos.

Study on the mechanism of curcumol inhibiting the proliferation of breast cancer cells T 47D / 中国药房

OBJECTIVE To study the mechanism of curcumol inhibiting the pro liferation of breast cancer cells T 47D. METHODS MTT assay was used to detect the inhibitory effects of different doses of curcumol （0，6.25，12.5，25，50，100 μg/mL）on the proliferation of T 47D cells. After treated with curcumol （12.5，25，50，100 μg/mL），the morphology of T 47D cells was observed by inverted phase contrast microscope. The cell cycle and the levels of reactive oxygen species （ROS）were detected by flow cytometry. Quantitative real-time PCR （qRT-PCR）was used to detect the expressions of proliferating cell nuclear antigen （PCNA），cell cycle regular p 21 and cyclin-dependent kinase 2（CDK2）mRNA. Western blot assay was used to detect the protein expression of CDK 2，CDK6，Cyclin D ，PCNA，nucler transcription factor E 2-related factor （Nrf2）and Kelch-like ECH associated protein 1（Keap1）. Breast cancer cells T 47D were divided into 2 groups，one group was given different doses of curcumol ，and another group was given curcumol 33 μg/mL for 6，12，24，48 h. After the optimal oxidation time and administration concentration were determined according to the results of the above two groups ，the blank control group ，N-acetylcysteine（NAC）group（ROS antioxidant NAC alone ），curcumol group （curcumol alone ），curcumol combined with NAC group （pretreatment with ROS antioxidant NAC ，and then adding into curcumol ）. Cell cycle and fluorescence intensity of ROS were detected. RESULTS Curcumol could significantly increase the inhibitory rate of the proliferation of T 47D cells （P＜0.05 or P＜0.01），and showed a certain dose and time dependent trend. Curcumol blocked the ， cycle in the G 1 phase and significantly increased the level of ROS （P＜0.05 or P＜0.01）；ROS antioxidant NAC could significantly reverse above inductive effect of curcumol （P＜ 0.01）. qRT-PCR showed that curcumol down-regulated the com expression of PCNA and CDK 2 mRNA and up-regulated the expression of p 21 mRNA（P＜0.05 or P＜0.01）. Western blot assay showed that curcumol significantly down-regulated the edu.cn protein expression of Keap 1，Nrf2，CDK2，CDK6 and Cyclin D（P＜0.05，P＜0.01）；ROS antioxidant NAC could reverse the down-regulation effects of curcumol on the expression of these proteins（P＜0.05 or P＜0.01）. CONCLUSIONS Curcumol may induce oxidative stress and cell arrest in G 1 phase to inhibit the proliferation of T 47D cells.

Effect and Mechanism of Berberine Hydrochloride Against Lung Cancer Cells in Vitro Based on Mevalonate Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the efficacy and mechanism of berberine hydrochloride （BBH） against lung cancer cells through the mevalonate （MVA） pathway. MethodHuman lung cancer A549 cells and mouse Lewis lung carcinoma （LLC） cells were used as research subjects. Cell proliferation and cell counting kit-8 （CCK-8） assay were performed to detect the inhibitory effect of BBH （10， 20， 30， 40， 50 μmol·L-1） on the proliferation of the two kinds of cells （48 h）. Then cell scratch assay was used to explore the influence of BBH （40 μmol·L-1） on the migration of A549 and LLC cells （24， 48 h）， and colony formation assay was conducted to compare the colony formation ability of the cells under different concentrations of BBH （10， 20， 40 μmol·L-1）. Moreover， the effects of BBH （40 μmol·L-1） on the content of acetyl-coenzyme A （A-CoA） and total cholesterol （TC） in A549 and LLC cells were determined by kit assay. AutoDock Vina was used for the dock of BBH and MVA pathway regulatory protein， sterol regulatory element-binding protein 2 （SREBP2）. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to observe the effects of BBH （40 μmol·L-1） on the mRNA expression of nine genes related to the MVA pathway in A549 and LLC cells： hydroxymethylglutaryl-CoA synthase 1 （HMGCS1）， hydroxymethylglutaryl-CoA Reductase （HMGCR）， mevalonate kinase （MVK）， phosphomevalonate kinase （PMVK）， mevalonate 5-pyrophosphate decarboxylase （MVD）， farnesyl diphosphate synthase （FDPS）， squalene epoxidase （SQLE）， farnesyl-diphosphate farnesyltransferase 1 （FDFT1）， and geranylgeranyl diphosphate synthase 1 （GGPS1）. Western blot was performed to clarify the effects of BBH （40 μmol·L-1） on the expression of three key proteins of the MVA pathway： HMGCS1， HMGCR， and FDFT1. The Cancer Genome Atlas （TCGA） database was searched to analyze the relationship between HMGCS1， HMGCR， FDFT1 and transcription gene SREBF2 in non-small cell lung cancer （NSCLC）. ResultCompared with the conditions in the control group， the proliferation， migration， and colony formation of A549 and LLC cells in the BBH group were decreased （P<0.01）， while the cell apoptosis rate was increased （P<0.01）. Molecular docking showed that BBH had good binding activity with SREBP2. In addition， the content of A-CoA and TC of the MVA pathway was reduced （P<0.01）. BBH down-regulated the mRNA expression of HMGCS1， HMGCR， MVK， PMVK， MVD， FDPS， SQLE， FDFT1， and GGPS1 in A549 and LLC cells （P<0.01）， and lowered the levels of HMGCS1， HMGCR， and FDFT1 proteins （P<0.05， P<0.01）. In NSCLC patients， HMGCS1， HMGCR， and FDFT1 were highly correlated with SREBF2 （R=0.54， R=0.57， and R=0.48）. ConclusionBBH can inhibit the proliferation， migration， and colony formation of A549 and LLC cells and promote cell apoptosis， which may be related to the regulation of MVA pathway by BBH binding to SREBP2.

Anti-tumor Effect of Draconis Sanguis Petroleum Ether Fraction on Human Gastric Cancer HGC-27 and MGC-803 Cells / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Draconis Sanguis petroleum ether fraction （DSPEF） on the proliferation， apoptosis， migration， and autophagy of human gastric cancer HGC-27 and MGC-803 cells， and preliminarily elucidate its molecular mechanism. MethodCell counting kit-8 （CCK-8） assay was used to detect the effect of DSPEF at different concentrations （0， 20， 40， 60， 80 mg·L-1） on the proliferation of HGC-27 and MGC-803 cells after 24， 48， 72 h. Hoechst staining and flow cytometry were used to explore the effects of DSPEF at different concentrations on the apoptosis and apoptosis rate of HGC-27 and MGC-803 cells after 48 h treatment， respectively. The wound healing assay and acridine orange staining were used to investigate the effects of DSPEF on the migration and autophagy of HGC-27 and MGC-803 cells， respectively. Western blot was used to detect the expression levels of signaling pathway-related proteins in HGC-27 and MGC-803 cells treated with DSPEF for 48 h. ResultCompared with the control group， DSPEF（30 mg·L-1） inhibited the proliferation and migration of HGC-27 and MGC-803 cells in a concentration- and time-dependent manner （P<0.05）， and induced the apoptosis （P<0.01） and autophagy of HGC-27 and MGC-803 cells. DSPEF （60 mg·L-1） down-regulated the protein levels of phosphorylated mammalian target of rapamycin （p-mTOR） （P<0.05， P<0.01） and down-regulated phospho-signal transducer and activator of transcription 3 （p-STAT3） in HGC-27 and MGC-803 cells （P<0.01）， suggesting that DSPEF presumedly inhibited the proliferation and migration of human gastric cancer HGC-27 and MGC-803 cells and induced their apoptosis and autophagy by inhibiting the mTOR/STAT3 signaling pathway. ConclusionThe down-regulation of the mTOR/STAT3 signaling pathway may be involved in the anti-gastric cancer effect of DSPEF. This study is expected to provide a reference for the investigation of the anti-tumor effect of Draconis Sanguis.