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Objective: To explore the regulatory effect of Pien Tze Huang (PZH) on targeting partner of NOB1 (PNO1) and it's down-stream mediators in colorectal cancer (CRC) cells. Methods: Quantitative polymerase chain reaction was performed to determine mRNA levels of PNO1, TP53, and CDKN1A. Western blotting was performed to determine protein levels of PNO1, p53, and p21. HCT-8 cells were transduced with a lentivirus over-expressing PNO1. Colony formation assay was used to detect cell survival in PNO1 overexpression of HCT-8 cells after PZH treatment. Cell-cycle distribution, cell viability and cell apoptosis were performed to identify the effect of PNO1 overexpression on cell proliferation and apoptosis of HCT-8 cells after PZH treatment. Xenograft BALB/c nude mice bearing HCT116 cells transduced with sh-PNO1 or sh-Ctrl lentivirus were evaluated. Western blot assay was performed to detect PNO1, p53, p21 and PCNA expression in tumor sections. Terminal deoxynucleotidyl transferase dUTP nick end labling (TUNEL) assay was used to determine the apoptotic cells in tissues. Results: PZH treatment decreased cell viability, down-regulated PNO1 expression, and up-regulated p53 and p21 expressions in HCT-8 cells (P<0.05). PNO1 overexpression attenuated the effects of PZH treatment, including the expression of p53 and p21, cell growth, cell viability, cell cycle arrest and cell apoptosis in vitro (P<0.05). PNO1 knockdown eliminated the effects of PZH treatment on tumor growth, inhibiting cell proliferation inhibition and apoptosis induction in vivo (P<0.05). Similarly, PNO1 knockdown attenuated the effects of PZH treatment on the down-regulation of PNO1 and up-regulation of p53 and p21 in vivo (P<0.05). Conclusion: The mechanism by which PZH induces its CRC anti-proliferative effect is at least in part by regulating the expression of PNO1 and its downstream targets p53 and p21. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the protective function of Babao Dan (BBD) on 5-flurouracil (5-FU)-induced intestinal mucositis (IM) and uncover the underlying mechanism. Methods: A total of 18 male mice were randomly divided into 3 groups by a random number table, including control, 5-FU and 5-FU combined BBD groups, 6 mice in each group. A single intraperitoneal injection of 5-FU (150 mg/kg) was performed in 5-FU and 5-FU combined BBD groups on day 0. Mice in 5-FU combined BBD group were gavaged with BBD (250 mg/kg) daily from day 1 to 6. Mice in the control group were gavaged with saline solution for 6 days. The body weight and diarrhea index of mice were recorded daily. On the 7th day, the blood from the heart of mice was collected to analyze the proportional changes of immunological cells, and the mice were subsequently euthanized by mild anesthesia with 2% pentobarbital sodium. Colorectal lengths and villus heights were measured. Intestinal-cellular apoptosis and proliferation were evaluated by Tunel assay and immunohistochemical staining of proliferating cell nuclear antigen, respectively. Immunohistochemistry and Western blot were performed to investigate the expressions of components in Wnt/β-catenin pathway (Wnt3, LRP5, β-catenin, c-Myc, LRG5 and CD44). Results: BBD obviously alleviated 5-FU-induced body weight loss and diarrhea, and reversed the decrease in the number of white blood cells, including monocyte, granulocyte and lymphocyte, and platelet (P<0.01). The shortening of colon caused by 5-FU was also reversed by BBD (P<0.01). Moreover, BBD inhibited apoptosis and promoted proliferation in jejunum tissues so as to reduce the intestinal mucosal damage and improve the integrity of villus and crypts. Mechanically, the expression levels of Wnt/β -catenin mediators such as Wnt3, LRP5, β-catenin were upregulated by BBD, activating the transcription of c-Myc, LRG5 and CD44 (P<0.01). Conclusions: BBD attenuates the adverse effects induced by 5-FU via Wnt/β-catenin pathway, suggesting it may act as a potential agent against chemotherapy-induced intestinal mucositis. [ABSTRACT FROM AUTHOR]

Multidrug resistance (MDR) is a critical reason for cancer chemotherapy failure. Babaodan (BBD) is a famous traditional Chinese patent medicine reported to have antigastric cancer activity. However, the roles and molecular mechanisms of the reversal of MDR of gastric cancer by BBD have not been well described until now. Therefore, the purpose of this study was to elucidate further the role of BBD in reversing the MDR of gastric cancer cells and its specific regulatory mechanism via in vitro experiments. To verify our results, MTT, Doxorubicin (DOX) staining, Rhodamin123 (Rho123) staining, DAPI staining, Annexin V-FITC, propidium iodide (PI), Cyto-ID, and western blot assays were performed. To determine whether BBD triggers apoptosis and autophagy through the PI3K/AKT/mTOR signaling, we also applied 3-methyladenine (3-MA), chloroquine (CQ), and 740Y-P (an activator of PI3K). The results showed that BBD reversed the MDR and induced apoptosis and autophagy of SGC7901/DDP cells. Pathway analyses suggested BBD inhibits PI3K/AKT/mTOR pathway activity and subsequent apoptosis-autophagy induction. Inhibition of autophagy with 3-MA and chloroquine (CQ) was performed to confirm that BBD promoted autophagy. PI3K agonist, 740Y-P, further verified BBD inhibition of PI3K/AKT/mTOR pathway activation. In conclusion, BBD may reverse the MDR of gastric cancer cells, induce apoptosis, and promote autophagy via inactivation of the PI3K/AKT/mTOR signaling pathway. [ABSTRACT FROM AUTHOR]

Background. Qing Guang An Granule (QGAG), a Chinese patent medicine, has been used clinically to treat glaucoma for more than 20 years. Objective. To explore the possible mechanism of treatment of QGAG in glaucoma by using network pharmacology and molecular docking in this study. Methods. Active compounds and targets of each herb in QGAG were retrieved via the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP). Glaucoma-related targets were acquired from OMIM and DisGeNET database. Key targets of QGAG against glaucoma were acquired by overlapping the above targets via the Venn diagram. Using the DAVID, Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the key targets were performed. The docking process was performed using the AutoDock 4.2.6 and AutoDock Vina 1.1.2. Results. The 55 active compounds and 173 targets were obtained and constructed a compound-target network. The 20 key targets of QGAG in treating glaucoma were acquired, and these targets are involved in the apoptotic process, cellular response to hypoxia, negative regulation of cell growth, and ovarian follicle development. The main pathways are p53, HIF-1, PI3K-Akt, and neurotrophin signaling pathway. Conclusion. QGAG may exert a protective effect by acting on the optic nerve at a molecular and systemic level. This study can provide a certain basis for future researches on exploring the QGAG in treating glaucoma and provide new ideas for developing new drugs. [ABSTRACT FROM AUTHOR]

The use of 5-fluorouracil (5-FU) has been proven benefits, but it also has adverse events in colorectal cancer (CRC) chemotherapy. In this study, we explored the mechanism of 5-FU resistance by bioinformatics analysis of the NCBI public dataset series GSE81005. Fifteen hub genes were screened out of 582 different expressed genes. Modules of the hub genes in protein-protein interaction networks gathered to TOP2α showed a decrease in HCT-8 cells but an increase in 5-FU-resistant HCT-8/5-FU cells with 5-FU exposure. Downregulation of TOP2α with siRNA or miR-494 transfection resulted in an increase of cytotoxicity and decrease of cell colonies to 5-FU for HCT-8/5-FU cells. Moreover, we found that an ethanol extract of Spica Prunellae (EESP), which is a traditional Chinese medicine with clinically beneficial effects in various cancers, was able to enhance the sensitivity of 5-FU in HCT-8/5-FU cells and partly reverse the 5-FU resistance effect. It significantly helped suppress cell growth and induced cell apoptosis in HCT-8/5-FU cells with the expression of TOP2α being significantly suppressed, which increased by 5-FU. Consistently, miR-494, which reportedly regulates TOP2α, exhibited reverse trends in EESP/5-FU combination treatment. These results suggested that Spica Prunellae may be beneficial in the treatment of 5-FU-resistant CRC patients. [ABSTRACT FROM AUTHOR]

Objective: To investigate the molecular mechanisms by which Qianliening Capsule (前列宁胶囊, QC) treats benign prostatic hyperplasia (BPH). Methods: Human prostate stromal cell line WPMY-1 was treated with 0, 1, 3 and 5 mg/mL of QC for 24, 48 and 72 h, respectively, in the presence of 10 ng/mL basic fibroblast growth factor (bFGF). The viability of WPMY-1 cells was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell morphology was observed by phase-contrast microscopy. 4′,6-diamidino-2-phenylindole (DAPI) staining and fluorescence activated cell sorting (FACS) analysis with Annexin-V/propidium iodide (PI) staining were performed to determine cell apoptosis. The loss of mitochondrial membrane potential was examined by FACS analysis with 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolyl-carbocyarine iodide (JC-1) staining. Activation of caspase-3 and -9 was evaluated by colorimetric assay. The mRNA and protein expression levels of Bcl-2 and Bax were measured by reverse transcription polymerase chain reaction (RT-PCR) and Western blotting, respectively. Results: Upon bFGF stimulation, the viability of WPMY-1 cells was increased to 122%-118% compared with the control cells ( P <0.05). However, treatment with 1-5 mg/mL of QC for 24, 48 and 72 h decreased the viability of bFGF-stimulated cells to 80%-92%, 59%-82%, 36%-62% compared with the untreated cells ( P <0.05). In addition, QC treatment reduced WPMY-1 cell density in a dose-dependent manner. Moreover, QC treatment dose-dependently induced the loss of plasma membrane asymmetry, the nuclear condensation and fragmentation, collapse of mitochondrial membrane potential, activation of caspase-9 and caspase-3, and increase of pro-apoptotic Bax/Bcl-2 ratio. Conclusion: Promoting mitochondrion-dependent apoptosis of prostate stromal cells might be one of the mechanisms by which QC treats BPH. [ABSTRACT FROM AUTHOR]

In this study, hepatocellular carcinoma (HCC) mouse xenograft model, MTT assay, colony formation, nuclear staining, and Annexin-V/PI staining assays were used to evaluate the effect of Qingjie Fuzheng granules (QFG) on cell proliferation and apoptosis of HCC cell in vivo and in vitro. Furthermore, Western blotting was performed to detect the expression of Fas, FasL, Bcl-2, Bax, and the activation of caspase-3/-8/-9. The results showed that QFG reduced tumor weight (P < .05) but had no effect on body weight gain in HCC mice in vivo. QFG significantly reduced HCC cell viability and attenuated cell proliferation in a dose-dependent manner (P < .05). QFG increased the expression of Fas, FasL, and Bax (P < .05). QFG downregulated the expression of Bcl-2 and promoted the activation of caspase-8, -9, and -3 (P < .05). These results suggested that QFG possessed anticancer effects, and the mechanisms of action may involve the death receptor pathway and mitochondrion-dependent pathway-mediated apoptosis. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Pien-Tze-Huang (PZH) is a famous formula of traditional Chinese medicine used to treating stroke. However, the protective effect of PZH and its mechanisms in acute ischemic stroke remain to be explored. Aim of the study To investigate the protective effect of PZH on neuronal apoptosis in acute cerebral ischemic injury rats and explore its underlying mechanisms. Materials and methods The effects of PZH were studied in acute ischemic stroke rats induced by transient middle cerebral artery occlusion, and the mitochondria-mediated apoptotic proteins including cytochrome C (Cyt C), Bax, Bcl-xl, P53, caspase-3, and caspase-9 as well as AKT and glycogen synthase kinase-3 beta (GSK-3β) were assessed. Results Four days of PZH treatment (180 mg/kg) could significantly reduce cerebral infarct volume, improve neurological deficit, attenuate inflammatory response, and inhibit neuronal apoptosis in acute ischemic stroke rats. Moreover, PZH treatment significantly decreased cytosolic Cyt C, Bax, P53, cleaved caspase-3, and cleaved caspase-9 levels, but elevated mitochondrial Cyt C and Bcl-xl levels. PZH treatment also increased phosphorylation of AKT and GSK-3β. Conclusion PZH potently protects the brain from cerebral ischemia/reperfusion injury in vivo, and inhibiting mitochondria-mediated neuronal apoptosis as well as attenuating inflammatory responses may be involved in this effect. This study provides experimental basis of PZH in treating acute cerebral ischemic stroke, which would provide some novel insights for its prevention and treatment of ischemic stroke. [ABSTRACT FROM AUTHOR]

Lycium barbarum L. and Salvia miltiorrhiza Bunge (Gouqi and Danshen, LS) are traditional herbs for the treatment of retinal degeneration in China. LS have been integrated into pharmacopoeia and health care system of many countries around the world. However, the mechanisms by which LS protect retina are not fully clarified. We aimed at exploration of the effect of LS on retinal pigment epithelium (RPE) cells apoptosis as well as the endoplasmic reticulum (ER) stress mechanisms. ARPE-19 cells were exposed to tunicamycin to induce ER stress, followed by LS treatment for 24 h. The cell morphology was photographed using the Incucyte S3 instrument, and the potential cytotoxic effect and viability were evaluated by CCK-8 assays. The Annexin V-FITC/PI staining and TUNEL assay were conducted to detect cells apoptotic. Western blot and digital PCR were used to detected related protein and gene expression. The ARPE-19 cells are increased in number and aligned after treating with LS. 1 mg/ml is the LS high dose group dose and treatment with LS increased cell vitality. LS significantly inhibit ARPE-19 cells apoptosis. Moreover, LS were markedly decreased the expression levels of ER stress-related factors in the ARPE-19 cells. This study reveals that LS relieve ARPE-19 cells apoptosis by inhibiting ER stress, and here we can speculate that LS have a certain protective effect on retina. [Display omitted] • Lycium barbarum L. and Salvia miltiorrhiza Bunge against tunicamycin-induced endoplasmic reticulum stress cell model was first explored. [ABSTRACT FROM AUTHOR]

Kuanxiong aerosol (KXA) is a common clinical drug based on Fangxiang Wentong (FXWT) therapy in the treatment of angina pectoris. However, the pharmacological mechanism of KXA in the prevention and treatment of myocardial injury (MI) is not clear. The purpose of this study was to explore the protective effect of KXA on isoproterenol (ISO)-induced MI in rats. The study included male Wistar Kyoto rats (age: 6 weeks). The rats were randomly divided into the following 5 groups (n = 6 per group): control group, ISO group, isosorbide mononitrate (ISMN) group (5 mg/kg), KXA-L group (0.1 mL/kg), and KXA-H group (0.3 mL/kg). The rats in the last three groups were given intragastric administration for 14 days, and rats in control group and ISO group were given the same amount of normal saline daily. ISO (120 mg/kg) was used to induce MI on the 13th and 14th days. We assessed electrocardiograms (ECGs), myocardial specific enzymes, histopathological changes, and apoptosis. We found that KXA reduced the increase in the ST-segment amplitude (elevation or depression) and the levels of myocardial marker enzymes induced by ISO in MI rats, improved the pathological changes in myocardial tissue, and reduced cardiomyocyte apoptosis. At the same time, KXA significantly inhibited the up-regulation of caspase-3 and Bax expression and down-regulation of Bcl-2 expression induced by ISO. RNA sequencing showed that 90 up-regulated genes induced by ISO were down-regulated after KXA treatment, whereas 27 down-regulated genes induced by ISO were up-regulated after KXA treatment. In addition, KEGG pathway enrichment analysis showed that the mitogen-activated protein kinase (MAPK) signaling pathway may be an important target of KXA in the treatment of ISO-induced MI in rats. The results of RNA sequencing verified by Western blot analysis showed that KXA significantly inhibited the activation of the ISO-induced MAPK pathway. KXA improves cardiac function in MI rats by inhibiting apoptosis mediated by the MAPK signaling pathway. Image 1 [ABSTRACT FROM AUTHOR]