Integrative network pharmacology and experimental verification to reveal the anti-inflammatory mechanism of ginsenoside Rh4.

Inflammation is an innate immune response to infection, and it is the main factor causing bodily injury and other complications in the pathological process. Ginsenoside Rh4 (G-Rh4), a minor ginsenoside of Panax ginseng C. A. Meyer and Panax notoginseng, has excellent pharmacological properties. However, many of its major pharmacological mechanisms, including anti-inflammatory actions, remain unrevealed. In this study, network pharmacology and an experimental approach were employed to elucidate the drug target and pathways of G-Rh4 in treating inflammation. The potential targets of G-Rh4 were selected from the multi-source databases, and 58 overlapping gene symbols related to G-Rh4 and inflammation were obtained for generating a protein-protein interaction (PPI) network. Molecular docking revealed the high affinities between key proteins and G-Rh4. Gene ontology (GO) and pathway enrichment analyses were used to analyze the screened core targets and explore the target-pathway networks. It was found that the JAK-STAT signaling pathway, TNF signaling pathway, NF-κB signaling pathway, and PI3K-Akt signaling pathway may be the key and main pathways of G-Rh4 to treat inflammation. Additionally, the potential molecular mechanisms of G-Rh4 predicted from network pharmacology analysis were validated in RAW264.7 cells. RT-PCR, Western blot, and ELISA analysis indicated that G-Rh4 significantly inhibited the production of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-1ß, as well as inflammation-related enzymes in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Moreover, in vitro experiments evaluated that Ginsenoside Rh4 exerts anti-inflammatory effects via the NF-κB and STAT3 signaling pathways. It is believed that our study will provide the basic scientific evidence that G-Rh4 has potential anti-inflammatory effects for further clinical studies.

(20S) Ginsenoside Rh2 Exerts Its Anti-Tumor Effect by Disrupting the HSP90A-Cdc37 System in Human Liver Cancer Cells.

(20S) ginsenoside Rh2 (G-Rh2), a major bioactive metabolite of ginseng, effectively inhibits the survival and proliferation of human liver cancer cells. However, its molecular targets and working mechanism remain largely unknown. Excitingly, we screened out heat shock protein 90 alpha (HSP90A), a key regulatory protein associated with liver cancer, as a potential target of (20S) G-Rh2 by phage display analysis and mass spectrometry. The molecular docking and thermal shift analyses demonstrated that (20S) G-Rh2 directly bound to HSP90A, and this binding was confirmed to inhibit the interaction between HSP90A and its co-chaperone, cell division cycle control protein 37 (Cdc37). It is well-known that the HSP90A-Cdc37 system aids in the folding and maturation of cyclin-dependent kinases (CDKs). As expected, CDK4 and CDK6, the two G0-G1 phase promoting kinases as well as CDK2, a key G1-S phase transition promoting kinase, were significantly downregulated with (20S) G-Rh2 treatment, and these downregulations were mediated by the proteasome pathway. In the same condition, the cell cycle was arrested at the G0-G1 phase and cell growth was inhibited significantly by (20S) G-Rh2 treatment. Taken together, this study for the first time reveals that (20S) G-Rh2 exerts its anti-tumor effect by targeting HSP90A and consequently disturbing the HSP90A-Cdc37 chaperone system. HSP90A is frequently overexpressed in human hepatoma cells and the higher expression is closely correlated to the poor prognosis of liver cancer patients. Thus, (20S) G-Rh2 might become a promising alternative drug for liver cancer therapy.

LncRNA NR2F2-AS1 induces epithelial-mesenchymal transition of non-small cell lung cancer by modulating BVR/ATF-2 pathway via regulating miR-545-5p/c-Met axis.

Non-small cell lung cancer (NSCLC) is one type of the most common cancers, which results in the major death worldwide. This study focuses on the understanding of the molecular mechanism of lncRNA NR2F2-AS1 and its regulation on epithelial-mesenchymal transition (EMT) in the development of NSCLC. Expressions of lncRNA NR2F2-AS1, miR-545-5p, c-Met, biliverdin reductase (BVR), ATF-2 and EMT-related markers in NSCLC tissues and cells were measured by western blotting and RT-qPCR assays. The impact of lncRNA NR2F2-AS1 and miR-545-5p on the cell proliferation, migration, invasion and EMT were analyzed by CCK-8, colony formation, wound healing and transwell assays. The interactions among lncRNA NR2F2-AS1, miR-545-5p and c-Met predicted by bioinformatic analysis were evaluated through dual luciferase reporter assay and fluorescence in situ hybridization (FISH). After generating tumor xenografts, immunohistochemistry was utilized to measure the expression of Ki-67 and EMT-related proteins in vivo. Our results showed that lncRNA NR2F2-AS1, c-Met, BVR and ATF-2 were overexpressed while miR-545-5p was silenced in NSCLC tissues and cells. Silencing of lncRNA NR2F2-AS1 or upregulating miR-545-5p significantly inhibited the cell proliferation, migration, invasion and EMT process. The EMT process could be inhibited by suppressing c-Met/BVR/ATF-2 axis. The tumor xenograft experiments demonstrated that the tumor growth and EMT process were significantly inhibited by silencing lncRNA NR2F2-AS1 or overexpression of miR-545-5p in vivo. LncRNA NR2F2-AS1 promoted the NSCLC development through suppressing miR-545-5p to activate EMT process through c-Met/BVR/ATF-2 axis. Our study indicated that lncRNA NR2F2-AS1 and miR-545-5p could be used as potential therapeutic targets to improve NSCLC treatment.

(20S) Ginsenoside Rh2 Inhibits STAT3/VEGF Signaling by Targeting Annexin A2.

Signal transducers and activators of transcription 3 (STAT3) acts as a transcriptional signal transducer, converting cytokine stimulation into specific gene expression. In tumor cells, aberrant activation of the tyrosine kinase pathway leads to excessive and continuous activation of STAT3, which provides further signals for tumor cell growth and surrounding angiogenesis. In this process, the tumor-associated protein Annexin A2 interacts with STAT3 and promotes Tyr705 phosphorylation and STAT3 transcriptional activation. In this study, we found that (20S) ginsenoside Rh2 (G-Rh2), a natural compound inhibitor of Annexin A2, inhibited STAT3 activity in HepG2 cells. (20S) G-Rh2 interfered with the interaction between Annexin A2 and STAT3, and inhibited Tyr705 phosphorylation and subsequent transcriptional activity. The inhibitory activity of STAT3 leaded to the negative regulation of the four VEGFs, which significantly reduced the enhanced growth and migration ability of HUVECs in co-culture system. In addition, (20S)G-Rh2 failed to inhibit STAT3 activity in cells overexpressing (20S)G-Rh2 binding-deficient Annexin A2-K301A mutant, further proving Annexin A2-mediated inhibition of STAT3 by (20S)G-Rh2. These results indicate that (20S)G-Rh2 is a potent inhibitor of STAT3, predicting the potential activity of (20S)G-Rh2 in targeted therapy applications.

Hispidin inhibits LPS-induced nitric oxide production in BV-2 microglial cells via ROS-dependent MAPK signaling.

Neuroinflammation is associated with many neurodegenerative diseases. Abnormal activation of microglial cells in the central nervous system (CNS) is a major characteristic of neuroinflammation. Nitric oxide (NO) free radicals are produced by activated microglia and prolonged presence of large quantities of NO in the CNS can lead to neuroinflammation and disease. Hispidin is a polyphenol derived from Phellinus linteus (a valuable medicinal mushroom) with strong antioxidant, anticancer and antidiabetic properties. A previous study demonstrated that hispidin significantly inhibited NO production via lipopolysaccharide (LPS)-induced RAW264.7 macrophages. Therefore, the present study used MTT assay was used to detect the effect of hispdin on cell viability. Griess reagent analysis was used to measure NO production. Reverse transcription-semi quantitative PCR and western blotting were used to evaluate the effects of hispdin on iNOS mRNA and MAPK/ERK/JNK protein levels. Fluorescence microscopy and flow cytometry were used to detect the effects of hispdin on the production of ROS and phagocytosis of cells. The present results indicated that hispidin could significantly inhibit the increase of NO production and iNOS expression in BV-2 microglial cells stimulated by LPS. The inhibitory effect of hispidin on NO production was similar to that of S-methylisothiourea sulfate, an iNOS inhibitor. Signaling studies demonstrated that hispidin markedly suppresses LPS-induced mitogen activated protein kinases and JAK1/STAT3 activation, although not the NF-κB signaling pathway. The present observations in LPS-stimulated BV-2 microglial cells indicated that hispidin might serve as a therapeutic candidate for the treatment of NO-induced neuroinflammation and, potentially, as a novel iNOS inhibitor.

The Anti-Tumor Effect and Underlying Apoptotic Mechanism of Ginsenoside Rk1 and Rg5 in Human Liver Cancer Cells.

Ginsenoside Rk1 and Rg5 are minor ginseng saponins that have received more attention recently because of their high oral bioavailability. Each of them can effectively inhibit the survival and proliferation of human liver cancer cells, but the underlying mechanism remains largely unknown. Network pharmacology and bioinformatics analysis demonstrated that G-Rk1 and G-Rg5 yielded 142 potential targets, and shared 44 putative targets associated with hepatocellular carcinoma. Enrichment analysis of the overlapped genes showed that G-Rk1 and G-Rg5 may induce apoptosis of liver cancer cells through inhibition of mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signal pathways. Methyl thiazolyl tetrazolium (MTT) assay was used to confirm the inhibition of cell viability with G-Rk1 or G-Rg5 in highly metastatic human cancer MHCC-97H cells. We evaluated the apoptosis of MHCC-97H cells by using flow cytometry and 4',6-diamidino-2-phenylindole (DAPI) staining. The translocation of Bax/Bak led to the depolarization of mitochondrial membrane potential and release of cytochrome c and Smac. A sequential activation of caspase-9 and caspase-3 and the cleavage of poly(ADP-ribose) polymerase (PARP) were observed after that. The levels of anti-apoptotic proteins were decreased after treatment of G-Rk1 or G-Rg5 in MHCC-97H cells. Taken together, G-Rk1 and G-Rg5 promoted the endogenous apoptotic pathway in MHCC-97H cells by targeting and regulating some critical liver cancer related genes that are involved in the signal pathways associated with cell survival and proliferation.

Peroxiredoxin II with dermal mesenchymal stem cells accelerates wound healing.

Peroxiredoxin II (Prx II) is involved in proliferation, differentiation, and aging in various cell types. However, Prx II-mediated stem cell regulation is poorly understood. Here, dermal mesenchymal stem cells (DMSCs), cell-growth factor-rich conditioned medium from DMSCs (DMSC-CM), and DMSC-derived exosomes (DMSC-Exos) were used to explore the regulatory role of Prx II in DMSC wound healing. Following treatment, wound healing was significantly decelerated in Prx II-/- DMSCs than in Prx II+/+ DMSCs. In vitro stimulation with 10 µM H2O2 significantly increased apoptosis in Prx II-/- DMSCs compared with Prx II+/+ DMSCs. The mRNA expression levels of EGF, b-FGF, PDGF-B, and VEGF did not significantly differ between Prx II-/- and Prx II+/+ DMSCs. Fibroblasts proliferated comparably when treated with Prx II+/+ DMSC-CM or Prx II-/- DMSC-CM. Wound healing was significantly higher in the Prx II-/- DMSC-Exos-treated group than in the Prx II+/+ DMSCs-Exos-treated group. Moreover, microRNA (miR)-21-5p expression levels were lower and miR-221 levels were higher in Prx II-/- DMSCs than in Prx II+/+ DMSCs. Therefore, our results indicate that Prx II accelerated wound healing by protecting DMSCs from reactive oxygen species-induced apoptosis; however, Prx II did not regulate cell/growth factor secretion. Prx II potentially regulates exosome functions via miR-21-5p and miR-221.

The compound 2-benzylthio-5,8-dimethoxynaphthalene-1,4-dione leads to apoptotic cell death by increasing the cellular reactive oxygen species levels in Ras-mutated liver cancer cells.

The aim of the present study was to verify the pro-apoptotic anticancer potential of several 5,8-dimethoxy-1,4-phthoquinone (DMNQ) derivatives in Ras-mediated tumorigenesis. MTT assays were used to detect cellular viability and flow cytometry was performed to assess intracellular reactive oxygen species (ROS) levels and apoptosis. The expression levels of proteins were detected via western blotting. Among the 12 newly synthesized DMNQ derivatives, 2-benzylthio-5,8-dimethoxynaphthalene-1,4-dione (BZNQ; component #1) significantly reduced cell viability both in mouse NIH3T3 embryonic fibroblasts cells (NC) and H-RasG12V transfected mouse NIH3T3 embryonic fibroblasts cells (NR). Moreover, BZNQ resulted in increased cytotoxic sensitivity in Ras-mutant transfected cells. Furthermore, the reactive oxygen species (ROS) levels in H-RasG12V transfected HepG2 liver cancer cells (HR) were significantly higher compared with the levels in HepG2 liver cancer cells (HC) following BZNQ treatment, which further resulted in increased cellular apoptosis. Eliminating cellular ROS using an ROS scavenger N-acetyl-L-cysteine markedly reversed BZNQ-induced cellular ROS accumulation and cell apoptosis in HC and HR cells. Western blotting results revealed that BZNQ significantly downregulated H-Ras protein expression and inhibited the Ras-mediated downstream signaling pathways such as protein kinase B, extracellular signal-related kinase and glycogen synthase kinase phosphorylation and ß-catenin protein expression. These results indicated that the novel DMNQ derivative BZNQ may be a therapeutic drug for Ras-mediated liver tumorigenesis. The results of the current study suggest that BZNQ exerts its effect by downregulating H-Ras protein expression and Ras-mediated signaling pathways.

Peroxiredoxin I deficiency increases keratinocyte apoptosis in a skin tumor model via the ROS-p38 MAPK pathway.

Keratinocyte hyperproliferation is an essential link in skin cancer pathogenesis. Peroxiredoxin I (Prx I) is known to regulate cancer cell proliferation, differentiation, and apoptosis, but its role in skin cancer remains unclear. This study aimed to elucidate the role and mechanism of Prx I in skin cancer pathogenesis. Dimethylbenz[a]anthracene (DMBA) and 12-O-tetradecanoyl-phorbol-13-acetate (TPA) were used to create a skin tumor model of the initiation/promotion stage of cancer. The role of Prx I in H2O2-induced keratinocyte apoptosis was also investigated. After DMBA/TPA treatment, Prx I deficiency was significantly associated with less skin tumors, lower Bcl-2 expression, and higher p-p38 and cleaved caspase-3 expressions in Prx I knockout tumors than in wild-type controls. H2O2 stimulation caused more cellular apoptosis in Prx I knockdown HaCaT cells than in normal HaCaT cells. The signaling study revealed that Bcl-2, p-p38, and cleaved caspase-3 expressions were consistent with the results in the tumors. In conclusion, the deletion of Prx I triggered the DMBA/TPA-induced skin tumor formation in vivo and in vitro by regulating the reactive oxygen species (ROS)-p38 mitogen-activated protein kinase (MAPK) pathway. These findings provide a theoretical basis for treating skin cancer.

Peroxiredoxin II Inhibits Alcohol-induced Apoptosis in L02 Hepatocytes Through AKT/ß-Catenin Signaling Pathway.

BACKGROUND: Peroxiredoxin II (PRDX2) performs unique roles in cells. It can reduce peroxides through cysteine residues, and helps prevent the effects of oxidative stress on cells. It is closely related to the occurrence and development of various diseases, especially alcoholic liver injury and even liver cancer. The metabolism of alcohol in hepatocytes leads to the increase in the levels of reactive oxygen species (ROS), oxidative stress, injury, and apoptosis. Therefore, this study focused on the investigating the protection conferred by PRDX2 against alcohol-induced apoptosis of hepatocytes. MATERIALS AND METHODS: PRDX2 inhibition of alcohol-induced apoptosis in L02 hepatocytes was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, fluorescence microscopy, flow cytometry, western blotting and hematoxylin and eosin staining. RESULTS: The results showed that the levels of reactive oxygen species, protein kinase B, ß-catenin, B-cell lymphoma-2 (BCL2), BCL-XL, BCL2-associated X, cleaved caspase-3, and cleaved poly (ADP-ribose) polymerase in PRDX2-silenced cells were increased significantly after the treatment of cells with ethanol. Similar results were obtained in an in vivo Prdx2-knockout mouse model of alcoholic liver injury. Therefore, PRDX2 may regulate the phosphorylation of the AKT signal protein by eliminating reactive oxygen species from cells, and it inhibits the downstream mitochondria-dependent apoptosis pathway, and, thereby, the apoptosis of cells. CONCLUSION: Thus, PRDX2 may be a potential molecular target for the prevention and treatment of alcoholic liver injury.

Peroxiredoxin I deficiency increases pancreatic ß­cell apoptosis after streptozotocin stimulation via the AKT/GSK3ß signaling pathway.

Apoptosis of pancreatic ß­cells is involved in the pathogenesis of type I and II diabetes. Peroxiredoxin I (Prx I) serves an important role in regulating cellular apoptosis; however, the role of Prx I in pancreatic ß­cell apoptosis is not completely understood. In the present study, the role of peroxiredoxin 1 (Prx I) during streptozotocin (STZ)­induced apoptosis of pancreatic ß­cells was investigated. The expression level of Prx I was decreased by STZ treatment in a time­dependent manner, and apoptosis of Prx I knockdown MIN6 cells was increased by STZ stimulation, compared with untransduced MIN6 cells. Furthermore, an intraperitoneal injection of STZ increased pancreatic islet damage in Prx I knockout mice, compared with wild­type and Prx II knockout mice. AKT and glycogen synthase kinase (GSK)­3ß phosphorylation significantly decreased following Prx I knockdown in MIN6 cells. However, phosphorylated ß­catenin and p65 levels significantly increased after STZ stimulation, compared with untransduced cells. The results of the present study indicate that deletion of Prx I mediated STZ­induced pancreatic ß­cell death in vivo and in vitro by regulating the AKT/GSK­3ß/ß­catenin signaling pathway, as well as NF­κB signaling. These findings provide a theoretical basis for treatment of pancreatic damage.

Picrasma quassioides Extract Elevates the Cervical Cancer Cell Apoptosis Through ROS-Mitochondrial Axis Activated p38 MAPK Signaling Pathway.

BACKGROUND/AIM: Picrasma quassioides (P. quassioides) is used in traditional Asian medicine widely for the treatment of anemopyretic cold, eczema, nausea, loss of appetite, diabetes mellitus, hypertension etc. In this study we aimed to understand the effect of P. quassioides ethanol extract on SiHa cervical cancer cell apoptosis. MATERIALS AND METHODS: The P. quassioides extract-induced apoptosis was analyzed using the MTT assay, fluorescence microscopy, flow cytometry and western blotting. RESULTS: P. quassioides extract induced cellular apoptosis by increasing the accumulation of cellular and mitochondrial reactive oxygen species (ROS) levels and inhibiting ATP synthesis. Pretreatment with N-Acetylcysteine (NAC), a classic antioxidant, decreased the intracellular ROS production and inhibited apoptosis. In addition, the P38 MAPK signaling pathway is a key in the apoptosis of SiHa cells induced by the P. quassioides extract. CONCLUSION: The P. quassioides extract exerts its anti-cancer properties on SiHa cells through ROS-mitochondria axis and P38 MAPK signaling. Our data provide a new insight for P. quassioides as a therapeutic strategy for cervical cancer treatment.

Anti-tumor Properties of Picrasma quassioides Extracts in H-RasG12V Liver Cancer Are Mediated Through ROS-dependent Mitochondrial Dysfunction.

BACKGROUND: Picrasma quassioides (PQ) is a traditional Asian herbal medicine with anti-tumor properties that can inhibit the viability of HepG2 liver cancer cells. H-Ras is often mutated in liver cancer, however, the effect of PQ treatment on H-Ras mutated liver cancer is unclear. This study aimed to investigate the role of PQ on ROS accumulation and mitochondrial dysfunction in H-ras mutated HepG2 (HepG2G12V) cells. MATERIALS AND METHODS: PQ ethanol extract-induced HepG2G12V apoptosis was analyzed by the MTT assay, fluorescence microscopy, flow cytometry and western blotting. RESULTS: PQ treatment affected cell migration and colony formation in HepG2G12V cells. Cleaved-caspase-3, cleaved-caspase-9 and BCL2 associated agonist of cell death (BAD) expression levels were increased, while the levels of B-cell lymphoma-extra large (Bcl-xL) were decreased with PQ treatment. PQ treatment led to a reduction of H-Ras expression levels in liver cancer cells, thus reducing their abnormal proliferation. Furthermore, it led to increased expression levels of Peroxiredoxin VI, which regulates the redox signal in cells. CONCLUSION: Taken together these results provide a new functional significance for the role of PQ in treating HepG2G12V liver cancer.

Specific Interaction With Human Serum Albumin Reduces Ginsenoside Cytotoxicity in Human Umbilical Vein Endothelial Cells.

Human serum albumin (HSA) is an important component of plasma, which has the functions of maintaining colloid osmotic pressure and capillary membrane stability, promoting blood circulation, and anti-oxidation. Three-dimensional structure of HSA determines its ability to bind and transport hormones and other substances. In this study we examined the interactions between HSA and ginsenoside Rg3, Rg5, Rk1, Rh2, and Rh4, which are the main cytotoxic ginsenosides extracted from red ginseng. Heat transfer generated by the specific interaction between HSA and each ginsenoside was measured using isothermal titration calorimetry (ITC) assay, which demonstrated that all these 5 ginsenosides bound to HSA with binding constants of 3.25, 1.89, 6.04, 2.07, and 5.17 × 105 M-1, respectively. Molecular docking also displayed that these ginsenosides interact with HSA at different sites of the HSA surface. Importantly, cell viability assay showed that the cytotoxicity of these ginsenosides reduced significantly at the presence of HSA in human vascular endothelial cells (HUVEC). Taken together, this study reveals the mechanism by which these ginsenosides are transported in vivo by not causing damage in vascular endothelium, and also suggests HSA might be an ideal carrier help to transport and execute these ginsenoside functions in human body.

(20S)G-Rh2 Inhibits NF-κB Regulated Epithelial-Mesenchymal Transition by Targeting Annexin A2.

(1) Background: Epithelial-mesenchymal transition (EMT) is an essential step for cancer metastasis; targeting EMT is an important path for cancer treatment and drug development. NF-κB, an important transcription factor, has been shown to be responsible for cancer metastasis by enhancing the EMT process. Our previous studies showed that (20S)Ginsenoside Rh2 (G-Rh2) inhibits NF-κB activity by targeting Anxa2, but it is still not known whether this targeted inhibition of NF-κB can inhibit the EMT process. (2) Methods: In vivo (20S)G-Rh2-Anxa2 interaction was assessed by cellular thermal shift assay. Protein interaction was determined by immuno-precipitation analysis. NF-κB activity was determined by dual luciferase reporter assay. Gene expression was determined by RT-PCR and immuno-blot. EMT was evaluated by wound healing and Transwell assay and EMT regulating gene expression. (3) Results: Anxa2 interacted with the NF-κB p50 subunit, promoted NF-κB activation, then accelerated mesenchymal-like gene expression and enhanced cell motility; all these cellular processes were inhibited by (20S)G-Rh2. In contrast, these (20S)G-Rh2 effect were completely eliminated by overexpression of Anxa2-K301A, an (20S)G-Rh2-binding-deficient mutant of Anxa2. (4) Conclusion: (20S)G-Rh2 inhibited NF-κB activation and related EMT by targeting Anxa2 in MDA-MB-231 cells.

Deletion of Peroxiredoxin II Inhibits the Growth of Mouse Primary Mesenchymal Stem Cells Through Induction of the G0/G1 Cell-cycle Arrest and Activation of AKT/GSK3ß/ß-Catenin Signaling.

BACKGROUND/AIM: Dermal mesenchymal stem cells (DMSCs) are pluripotent stem cells found in the skin which maintain the thickness of the dermal layer and participate in skin wound healing. MATERIALS AND METHODS: The MTT assay was performed to detect cell proliferation and cell-cycle progression and cell-surface markers were assessed by flow cytometry. The levels of proteins in related signaling pathways were detected by western blotting assay and the translocation of ß-catenin into the nucleus were detected by immunofluorescence. Red oil O staining was performed to examine the differentiational ability of DMSCs. RESULTS: Knockout of PRDX2 inhibited DMSC cell growth, and cell-cycle arrest at G0/G1 phase; p16, p21 and cyclin D1 expression levels in Prdx2 knockout DMSCs were significantly increased. Furthermore, AKT phosphorylation were significantly increased in Prdx2 knockout DMSCs, GSK3ß activity were inhibited, result in ß-Catenin accumulated in the nucleus. CONCLUSION: In conclusion, these results demonstrated that PRDX2 plays a pivotal role in regulating the proliferation of DMSCs, and this is closely related to the AKT/glycogen synthase kinase 3 beta/ß-catenin signaling pathway.

Peroxiredoxin II Maintains the Mitochondrial Membrane Potential against Alcohol-Induced Apoptosis in HT22 Cells.

Excessive alcohol intake can significantly reduce cognitive function and cause irreversible learning and memory disorders. The brain is particularly vulnerable to alcohol-induced ROS damage; the hippocampus is one of the most sensitive areas of the brain for alcohol neurotoxicity. In the present study, we observed significant increasing of intracellular ROS accumulations in Peroxiredoxin II (Prx II) knockdown HT22 cells, which were induced by alcohol treatments. We also found that the level of ROS in mitochondrial was also increased, resulting in a decrease in the mitochondrial membrane potential. The phosphorylation of GSK3ß (Ser9) and anti-apoptotic protein Bcl2 expression levels were significantly downregulated in Prx II knockdown HT22 cells, which suggests that Prx II knockdown HT22 cells were more susceptible to alcohol-induced apoptosis. Scavenging the alcohol-induced ROS with NAC significantly decreased the intracellular ROS levels, as well as the phosphorylation level of GSK3ß in Prx II knockdown HT22 cells. Moreover, NAC treatment also dramatically restored the mitochondrial membrane potential and the cellular apoptosis in Prx II knockdown HT22 cells. Our findings suggest that Prx II plays a crucial role in alcohol-induced neuronal cell apoptosis by regulating the cellular ROS levels, especially through regulating the ROS-dependent mitochondrial membrane potential. Consequently, Prx II may be a therapeutic target molecule for alcohol-induced neuronal cell death, which is closely related to ROS-dependent mitochondria dysfunction.

Peroxiredoxin V Reduces ß-Lapachone-induced Apoptosis of Colon Cancer Cells.

BACKGROUND/AIM: Peroxiredoxin (Prx) V has been known as an antioxidant enzyme which scavenges intracellular reactive oxygen species (ROS). Also, Prx V has been shown to mediate cell apoptosis in various cancers. However, the mechanism of Prx V-induced apoptosis in colon cancer cells remains unknown. Thus, in this study we analyzed the effects of Prx V in ß-lapachone-induced apoptosis in SW480 human colon cancer cells. MATERIALS AND METHODS: ß-lapachone-induced apoptosis was analyzed by the MTT assay, western blotting, fluorescence microscopy, Annexin V staining and flow cytometry. RESULTS: Overexpression of Prx V, significantly decreased ß-lapachone-induced cellular apoptosis and Prx V silencing increased ß-lapachone-induced cellular apoptosis via modulating ROS scavenging activity compared to mock SW480 cells. In addition, to further explore the mechanism of Prx V regulated ß-lapachone-induced SW480 cells apoptosis, the Wnt/ß-catenin signaling was studied. The Wnt/ ß-catenin signaling pathway was found to be induced by ß-lapachone. CONCLUSION: Prx V regulates SW480 cell apoptosis via scavenging ROS cellular levels and mediating the Wnt/ß-catenin signaling pathway, which was induced by ß-lapachone.

Ginsenoside compound K inhibits nuclear factor-kappa B by targeting Annexin A2.

BACKGROUND: Ginsenoside compound K(C-K), a major metabolite of ginsenoside, exhibits anticancer activity in various cancer cells and animal models. A cell signaling study has shown that C-K inhibited nuclear factor-kappa B (NF-κB) pathway in human astroglial cells and liver cancer cells. However, the molecular targets of C-K and the initiating events were not elucidated. METHODS: Interaction between C-K and Annexin A2 was determined by molecular docking and thermal shift assay. HepG2 cells were treated with C-K, followed by a luciferase reporter assay for NF-кB, immunofluorescence imaging for the subcellular localization of Annexin A2 and NF-кB p50 subunit, coimmunoprecipitation of Annexin A2 and NF-кB p50 subunit, and both cell viability assay and plate clone formation assay to determine the cell viability. RESULTS: Both molecular docking and thermal shift assay positively confirmed the interaction between Annexin A2 and C-K. This interaction prevented the interaction between Annexin A2 and NF-кB p50 subunit and their nuclear colocalization, which attenuated the activation of NF-кB and the expression of its downstream genes, followed by the activation of caspase 9 and 3. In addition, the overexpression of Annexin A2-K320A, a C-K binding-deficient mutant of Annexin A2, rendered cells to resist C-K treatment, indicating that C-K exerts its cytotoxic activity mainly by targeting Annexin A2. CONCLUSION: This study for the first time revealed a cellular target of C-K and the molecular mechanism for its anticancer activity.

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The typhoon outbreaks of 1986 caused many gaps in the cold temperate coniferous forest and Betula errmanii forest on the western and southern slopes in the Changbai Mountain. In 2017, a research area at the 1600-1800 m altitude in Changbai Mountain was established to explore the damage and changes of forest vegetation disturbed by the typhoon in cold temperate zone and to reveal the regulation of damage on vegetation and post-disaster change as well as the driving factors. The remote sensing data were used to classify research area based on the degree of vegetation damage and post-disaster change. A total of 40 plots were set up for vegetation survey. According to the damage degree of forest structure, three levels of severity of damage to vegetation in the cold temperate zone of Changbai Mountain were classified including gently, moderately, and severely damaged, in which moderately damaged area was largest, followed by gently damaged area and severely damaged area. Tree damage significantly differed among three level areas, with 20%, 50% and 85% reduction of the abundance of dominant trees in gently, moderately and severely damaged areas, respectively. The wind resistance ability of B. errmanii was higher than that of Picea jezoensis. The wind resistance ability of B. errmanii with larger diameter grade was higher than that of those with smaller diameter grade. The severity of damage to vegetation was strongly correlated with slope, with lower severity on the steeper slop area. From 1987 to 2017, the vegetation of wind disaster area significantly changed. The study area could be classified into three levels based on the degree of changing: fast, medium, and slow, with largest area in medium changed, followed by slowly and fast changed. The degree of vegetation changes was strongly correlated with altitude, with slower change at higher altitude area. The recovery rate of trees was slow, with P. jezoensis recovering slightly quicker than B. errmanii. The vegetation change mainly occurred in shrub and herb layers. Shrub layer recovered better than the herb layer in the fast changed area, while herb layer recovered better than the shrub layer in the medium changed area. In the slowly changed area, herb layer was generally low and dense with varying shrub layer.