α-Lipoic acid alleviates myocardial injury and induces M2b macrophage polarization after myocardial infarction via HMGB1/NF-kB signaling pathway.

BACKGROUND: Myocardial infarction (MI) is a serious cardiovascular disease with a poor prognosis. Macrophages are the predominant immune cells in patients with MI and macrophage regulation during the different phases of MI has important consequences for cardiac recovery. Alpha-lipoic acid (ALA) plays a critical role in MI by modulating the number of cardiomyocytes and macrophages. METHODS: MI mice were generated by ligating the left anterior descending coronary artery. Macrophages were exposed to hypoxia to establish a hypoxia model and M1 polarization was induced by LPS and IFN-γ. Different groups of macrophages and MI mice were treated with ALA. The cardiomyocytes were treated with various macrophage supernatants and the cardiac function, cytokine levels, and pathology were also analyzed. Factors related to apoptosis, autophagy, reactive oxygen species (ROS), and the mitochondrial membrane potential (MMP) were assessed. Finally, the HMGB1/NF-κB pathway was identified. RESULTS: ALA promoted M2b polarization in normal cells and suppressed inflammatory cytokines during hypoxia. ALA inhibited ROS and MMP production in vitro. Supernatants containing ALA inhibited apoptosis and autophagy in hypoxic cardiomyocytes. Moreover, ALA suppressed the HMGB1/NF-κB pathway in macrophages, which may be a potential mechanism for attenuating MI. CONCLUSION: ALA alleviates MI and induces M2b polarization via the HMGB1/NF-κB pathway, impeding inflammation, oxidation, apoptosis, and autophagy, and might be a potential strategy for MI treatment.

Saturated fatty acids increase LPI to reduce FUNDC1 dimerization and stability and mitochondrial function.

Ectopic lipid deposition and mitochondrial dysfunction are common etiologies of obesity and metabolic disorders. Excessive dietary uptake of saturated fatty acids (SFAs) causes mitochondrial dysfunction and metabolic disorders, while unsaturated fatty acids (UFAs) counterbalance these detrimental effects. It remains elusive how SFAs and UFAs differentially signal toward mitochondria for mitochondrial performance. We report here that saturated dietary fatty acids such as palmitic acid (PA), but not unsaturated oleic acid (OA), increase lysophosphatidylinositol (LPI) production to impact on the stability of the mitophagy receptor FUNDC1 and on mitochondrial quality. Mechanistically, PA shifts FUNDC1 from dimer to monomer via enhanced production of LPI. Monomeric FUNDC1 shows increased acetylation at K104 due to dissociation of HDAC3 and increased interaction with Tip60. Acetylated FUNDC1 can be further ubiquitinated by MARCH5 for proteasomal degradation. Conversely, OA antagonizes PA-induced accumulation of LPI, and FUNDC1 monomerization and degradation. A fructose-, palmitate-, and cholesterol-enriched (FPC) diet also affects FUNDC1 dimerization and promotes its degradation in a non-alcoholic steatohepatitis (NASH) mouse model. We thus uncover a signaling pathway that orchestrates lipid metabolism with mitochondrial quality.

Comparison of Different Cooling Schemes for AlGaN/GaN High-Electron Mobility Transistors.

Cooling is important for AlGaN/GaN high-electron mobility transistors (HEMTs) performance. In this paper, the advantages and disadvantages of the cooling performance of three cooling schemes: remote cooling (R-cool), near-chip cooling (NC-cool), and chip-embedded cooling (CE-cool) are compared. The influences of distinct geometric parameters and operating conditions on thermal resistance are investigated. The results show that the thermal resistances of NC-cool and CE-cool are almost the same as each other. Decreasing microchannel base thickness (hb) significantly increases the thermal resistance of CE-cool, and when its thickness is less than a critical value, NC-cool exhibits superior cooling performance than CE-cool. The critical thickness increases when decreasing the heat source pitch (Ph) and the convective heat transfer coefficient (hconv) or increasing the thermal conductivity of the substrate (λsub). Moreover, increasing Ph or λsub significantly improves the thermal resistance of three cooling schemes. Increasing hconv significantly decreases the thermal resistances of NC-cool and CE-cool while hardly affecting the thermal resistance of R-cool. The influence of the boundary thermal resistance (TBR) on the thermal resistance significantly increases at higher λsub and larger hconv.

Alpha-lipoic acid impedes myocardial ischemia-reperfusion injury, myocardial apoptosis, and oxidative stress by regulating HMGB1 expression.

BACKGROUND: Inflammation, oxidative stress, and apoptosis contribute to myocardial ischemia/reperfusion injury (I/RI). Alpha-lipoic acid (ALA) plays a critical role in I/RI by impeding apoptosis and inflammation. Here, we aimed to explore the underlying mechanisms of ALA after I/RI. METHODS: The left anterior descending coronary artery (LAD) was ligated, and H9c2 cells were exposed to hypoxia/reoxygenation (H/R) to establish an I/RI model. Prior to this, H9c2 cells and rats were treated using an appropriate amount of ALA. The cardiac function, inflammatory factors, and myocardial pathology were assessed in vitro. We detected cell viability, apoptosis, and oxidative stress-related factors in vivo. Moreover, proteins of the HMGB1/TLR4/NF-κB signaling pathway were detected both in vivo and in vitro. RESULTS: We observed that ALA increased cell viability in vitro and decreased apoptosis in vitro and in vivo. ALA inhibited reactive oxygen species production, decreased malondialdehyde, and increased superoxide dismutase activity to resist oxidative stress in vitro. ALA also reduced the expression of inflammatory cytokines (IL-6, IL-1ß, and TNF-α) in vivo. ALA also suppressed the levels of the apoptotic protein, Bax, and increased the expression of the anti-apoptotic protein Bcl-2, in vitro and in vivo. Moreover, we observed that ALA significantly inhibited the cytoplasmic localization of HMGB1, which might attenuate MI/RI or H/R via HMGB1/TLR4/NF-κB pathway. CONCLUSION: ALA regulates HMGB1 translocation and attenuates I/R via the HMGB1/TLR4/NF-κB signaling pathway, thus impeding apoptosis, oxidation, and inflammation, and might be a potential target for myocardial ischemia/reperfusion injury.

Hypoxia-induced proteasomal degradation of DBC1 by SIAH2 in breast cancer progression.

DBC1 has been characterized as a key regulator of physiological and pathophysiological activities, such as DNA damage, senescence, and tumorigenesis. However, the mechanism by which the functional stability of DBC1 is regulated has yet to be elucidated. Here, we report that the ubiquitination-mediated degradation of DBC1 is regulated by the E3 ubiquitin ligase SIAH2 and deubiquitinase OTUD5 under hypoxic stress. Mechanistically, hypoxia promoted DBC1 to interact with SIAH2 but not OTUD5, resulting in the ubiquitination and subsequent degradation of DBC1 through the ubiquitin-proteasome pathway. SIAH2 knockout inhibited tumor cell proliferation and migration, which could be rescued by double knockout of SIAH2/CCAR2. Human tissue microarray analysis further revealed that the SIAH2/DBC1 axis was responsible for tumor progression under hypoxic stress. These findings define a key role of the hypoxia-mediated SIAH2-DBC1 pathway in the progression of human breast cancer and provide novel insights into the metastatic mechanism of breast cancer.

Macrophages-Related Genes Biomarkers in the Deterioration of Atherosclerosis.

Background: The macrophages are involved in all stages of cardiovascular diseases, demonstrating the correlation between inflammation, atherosclerosis, and myocardial infarction (MI). Here, we aim to investigate macrophages-related genes in the deterioration of atherosclerosis. Methods: GSE41571 was downloaded and the abundance of immune cells was estimated by utilizing the xCell. By utilizing the limma test and correlation analysis, differentially expressed macrophages-related genes (DEMRGs) were documented. The functional pathways and the protein-protein interaction (PPI) network were analyzed and the hub DEMRGs were obtained. The hub DEMRGs and their interactions were analyzed using NetworkAnalyst 3.0 and for validation, the expressions of hub DEMRGs were analyzed using the GSE135055 and GSE116250 datasets as well as atherosclerosis and MI mice model. Results: A total of 509 differentially expressed genes (DEGs) were correlated with the abundance of macrophages and were identified as DEMRGs (Pearson correlation coefficients (PCC) > 0.6), which were mainly enriched in extracellular structure organization, lysosomal membrane, MHC protein complex binding, and so on. After screening out, 28 hub DEMRGs were obtained with degrees ≥20, including GNAI1 (degree = 113), MRPS2 (degree = 56), HCK (degree = 45), SOCS3 (degree = 40), NET1 (degree = 28), and so on. After validating using Gene Expression Omnibus (GEO) datasets and the atherosclerosis and MI mice model, eight proteins were validated using ApoE-/- and C57 mice. The expression levels of proteins, including SYNJ2, NET1, FZD7, LCP2, HCK, GNB2, and PPP4C were positively correlated to left ventricular ejection fraction (LVEF), while that of EIF4EBP1 was negatively correlated to LVEF. Conclusion: The screened hub DEMRGs, SYNJ2, NET1, FZD7, LCP2, HCK, GNB2, EIF4EBP1, and PPP4C, may be therapeutic targets for treatment and prediction in the patients with plaque progression and MI recurrent events. The kit of the eight hub DEMRGs may test plaque progression and MI recurrent events and help in the diagnosis and treatment of MI-induced heart failure (HF), thus decreasing mortality and morbidity.

MrPEX33 is involved in infection-related morphogenesis and pathogenicity of Metarhizium robertsii.

Peroxisomes, being indispensable organelles, play an important role in different biological processes in eukaryotes. PEX33, a filamentous fungus-specific peroxin of the docking machinery of peroxisomes, is involved in the virulence and development of other fungal pathogens. However, it is not clear whether PEX33 is necessary for the pathogenicity and development of an insect pathogenic fungus. In the present study, we report the presence of homologs of PEX33, namely MrPEX33 (MAA_05331), in the entomopathogenic fungus, Metarhizium robertsii. An M. robertsii transgenic strain expressing the fusion protein with MrPEX33-GFP and mCherry-PTS1 showed that MrPEX33 localizes to peroxisomes. The results also demonstrated that MrPEX33 is involved in the peroxisomal import pathway by peroxisomal targeting signals. Targeted gene deletion of MrPEX33 led to a significant decline in the asexual sporulation capacity, which was accompanied by downregulation of several conidiation-associated genes, such as wetA, abaA, and brlA. More importantly, our bioassay results showed that the virulence of ∆MrPEX33 mutants, against Galleria mellonella through cuticle infection, was greatly reduced. This was further accompanied by a significant drop in appressorium formation and cuticle penetration. Additionally, ∆MrPEX33 mutants showed a significant decrease in tolerance to cell wall integrity and oxidative stress. Taken together, our results suggest that MrPEX33 is involved in the cuticle infection-related morphogenesis and pathogenicity. KEY POINTS: • MrPEX33 is a specific peroxin of the docking machinery of peroxisomes. • MrPEX33 localizes to peroxisomes and is involved in the import of matrix proteins. • MrPEX33 is involved in the pathogenicity associated with cuticle infections.

Melatonin suppresses ER stress-dependent proapoptotic effects via AMPK in bone mesenchymal stem cells during mitochondrial oxidative damage.

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) have been used as important cell-based tools for clinical applications. Oxidative stress-induced apoptosis causes a low survival rate after transplantation, and the underlying mechanisms remain unknown. The endoplasmic reticulum (ER) and mitochondria are vital organelles regulated by adenosine monophosphate (AMP)-activated protein kinase (AMPK), especially during oxidative stress injury. Melatonin exerts an antioxidant effect by scavenging free radicals. Here, we aimed to explore whether cytoprotective melatonin relieves ER stress-mediated mitochondrial dysfunction through AMPK in BMSCs after oxidative stress injury. METHODS: Mouse BMSCs were isolated and exposed to H2O2 in the absence or presence of melatonin. Thereafter, cell damage, oxidative stress levels, mitochondrial function, AMPK activity, ER stress-related proteins, and apoptotic markers were measured. Additionally, the involvement of AMPK and ER stress in the melatonin-mediated protection of BMSCs against H2O2-induced injury was investigated using pharmacologic agonists and inhibitors. RESULTS: Melatonin improved cell survival and restored mitochondrial function. Moreover, melatonin intimately regulated the phosphorylation of AMPK and molecules associated with ER stress pathways. AMPK activation and ER stress inhibition following melatonin administration improved the mitochondrial membrane potential (MMP), reduced mitochondria-initiated oxidative damage, and ultimately suppressed apoptotic signaling pathways in BMSCs. Cotreatment with N-acetyl-L-cysteine (NAC) significantly enhanced the antioxidant effect of melatonin. Importantly, pharmacological AMPK activation/ER stress inhibition promoted melatonin-induced cytoprotection, while pharmacological AMPK inactivation/ER stress induction conferred resistance to the effect of melatonin against H2O2 insult. CONCLUSIONS: Our data also reveal a new, potentially therapeutic mechanism by which melatonin protects BMSCs from oxidative stress-mediated mitochondrial apoptosis, possibly by regulating the AMPK-ER stress pathway.

Activation of Paraventricular Melatonin Receptor 2 Mediates Melatonin-Conferred Cardioprotection Against Myocardial Ischemia/Reperfusion Injury.

Previous studies have shown that melatonin (Mel) can effectively ameliorate myocardial ischemia/reperfusion (MI/R) injury, but the mechanism is yet to be fully elucidated. Mel receptors are expressed in the paraventricular nucleus (PVN), which is also involved in regulating cardiac sympathetic nerve activity. The aim of this study was to examine whether Mel receptors in the PVN are involved in the protective effects of Mel against MI/R injury. The results of quantitative polymerase chain reaction, western blot, and immunofluorescence assays indicated that Mel receptor 2 (MT2) expression in the PVN was upregulated after MI/R. Intraperitoneal administration of Mel significantly improved post-MI/R cardiac function and reduced the infarct size, whereas shRNA silencing of MT2 in the PVN partially blocked this effect. Intraperitoneal administration of Mel reduced sympathetic nerve overexcitation caused by MI/R, whereas shRNA silencing of MT2 in the PVN partially diminished this effect. Furthermore, enzyme-linked immunosorbent assay and western blot results indicated that intraperitoneal administration of Mel lowered the levels of inflammatory cytokines in the PVN after MI/R injury, whereas the application of sh-MT2 in the PVN reduced this effect of Mel. Mel significantly reduced the levels of NF-κB after astrocyte oxygen and glucose deprivation/reoxygenation injury, and this effect was offset when MT2 was silenced. The above experimental results suggest that MT2 in the PVN partially mediated the protective effects of Mel against MI/R injury, and its underlying mechanisms may be related to postactivation amelioration of PVN inflammation and reduction of cardiac sympathetic nerve overexcitation.

A novel dielectric barrier discharge instrument with asymmetrical electrodes for generating different patterns simultaneously.

We report on a novel dielectric barrier discharge (DBD) instrument with asymmetrical electrodes. It consists of three water electrodes, one big electrode faces to two small electrodes placed side by side, any one of two small electrodes connects with a high-voltage diode which can control on or off of applied power. Using this DBD experimental setup, the phenomenon of forming two kinds of different patterns at the different parts of discharge gap simultaneously is investigated for the first time. The spatio-temporal characteristic of these different patterns limited by the boundary which contains two connected same circles is studied. It is shown that although the power source is applied on one part of the discharge gap at a half period because of existence of high voltage photodiode, the discharge occurs at both areas in each half period of the applied voltage. Results have prospective applications for studying the discharge mechanism and different characteristic of positive and negative charges in forming of pattern.

MrArk1, an actin-regulating kinase gene, is required for endocytosis and involved in sustaining conidiation capacity and virulence in Metarhizium robertsii.

Actin-regulating kinase (Ark) plays an important role in controlling endocytosis, which has been shown to be involved in the development and virulence of several fungal pathogens. However, it remains unclear whether Ark1 is required for the development and pathogenicity of an entomopathogenic fungus. Here, MrArk1 (MAA_03415), a homologue of yeast Ark1, was characterized in the insect pathogenic fungus, Metarhizium robertsii. Disruption of MrArk1 led to defects in endocytosis and a marked reduction (58%) in conidiation capacity. The reduced conidiation level was accompanied by repression of several key conidiation-related genes, including brlA, abaA, and wetA. Additionally, the deletion mutant showed a significant decrease in its tolerance to heat shock, but not to UV-B irradiation. Bioassays demonstrated attenuated virulence for the deletion mutant against Galleria mellonella via normal cuticle infection, accompanied by suppressed appressorium formation and reduced transcript levels of several genes involved in cuticle penetration. Taken together, our results indicate that MrArk1 is involved in the heat tolerance, sporulation, and virulence of M. robertsii, and thus is an important factor for sustaining the fungal potential against insect pests.

TRIM72 contributes to cardiac fibrosis via regulating STAT3/Notch-1 signaling.

Cardiac fibrosis is a pathophysiological process characterized by excessive deposition of extracellular matrix. We developed a cardiac hypertrophy model using transverse aortic constriction (TAC) to uncover mechanisms relevant to excessive deposition of extracellular matrix in mouse myocardial cells. TAC caused upregulation of Tripartite motif protein 72 (TRIM72), a tripartite motif-containing protein that is critical for proliferation and migration. Importantly, in vivo silencing of TRIM72 reversed TAC-induced cardiac fibrosis, as indicated by markedly increased left ventricular systolic pressure and decreased left ventricular end-diastolic pressure. TRIM72 knockdown also attenuated deposition of fibrosis marker collagen type I and α-smooth muscle actin (α-SMA). In an in vitro study, TRIM72 was similarly upregulated in cardiac fibroblasts. Knockdown of TRIM72 markedly suppressed collagen type I and α-SMA expression and significantly decreased the proliferation and migration of cardiac fibroblasts. However, TRIM72 overexpression markedly increased collagen type I and α-SMA expression and increased the proliferation and migration of cardiac fibroblasts. Further study demonstrated that TRIM72 increased phosphorylated STAT3 in cardiac fibroblasts. TRIM72 knockdown in cardiac fibroblasts resulted in increased expression of Notch ligand Jagged-1 and its downstream gene and Notch-1 intracellular domain. Inhibition of Notch-1 abrogated sh-TRIM72-induced cardiac fibrosis. Together, our results support a novel role for TRIM72 in maintaining fibroblast-to-myofibroblast transition and suppressing fibroblast growth by regulating the STAT3/Notch-1 pathway.

Honokiol induces endoplasmic reticulum stress-mediated apoptosis in human lung cancer cells.

AIMS: Honokiol is a hydroxylated biphenyl natural product and displays potent antitumor activity against several cancers including prostate cancer, melanoma, leukemia, and colorectal cancer. The present study was to investigate the in vitro activity of honokiol against A549 and 95-D human lung cancer cells. MAIN METHODS: A549 and 95-D cells were used with honokiol treatment. Cell viability was determined by CCK-8 assay. The cell migration and apoptosis were evaluated by wound healing assay and TUNEL staining method respectively. The expressions of ER-related proteins were analyzed by western blot and the CHOP siRNA was used to downregulate the CHOP expression. KEY FINDINGS: The results demonstrated that treatment of A549 and 95-D cells with honokiol significantly reduced cell viability in a dose- and time-dependent manner. Furthermore, honokiol treatment decreased cell migration and enhanced cell apoptosis, which is accompanied by the upregulation of the expressions of ER stress-induced apoptotic signaling molecules such as GRP78, phosphorylated PERK, phosphorylated eIF2α, CHOP, Bcl-2, Bax, and cleaved Caspase 9. Honokiol treatment-induced increase of ER stress-related signaling molecules and apoptotic proteins in A549 and 95-D cells were reversed by CHOP siRNA. SIGNIFICANCE: Collectively, we conclude that ER stress may participate in the action of the anticancer activity of honokiol in A549 and 95-D cells and induction of ER stress-related apoptosis may represent a novel therapeutic intervention for human lung cancer.

Berberine ameliorates lipopolysaccharide-induced acute lung injury via the PERK-mediated Nrf2/HO-1 signaling axis.

A fundamental element of acute lung injury (ALI) is the inflammatory response, which can affect the entire respiratory system, including the respiratory tract and alveoli. Berberine has gained attention because of its anti-inflammatory effects. Nuclear factor-erythroid 2-related factor 2 (Nrf2) and endoplasmic reticulum (ER) stress are involved in lung injury. Nrf2 also acts as a protein kinase-like ER kinase (PERK) substrate in heart disease. Therefore, this study investigated the effect of berberine against lipopolysaccharide (LPS)-induced ALI and the role of the PERK-mediated Nrf2/HO-1 signaling axis. Berberine promoted Nrf2 nuclear translocation and phosphorylation in vitro. After LPS stimulation, this effect was further enhanced, whereas inflammatory factor (IL-6 and IL-8) release and reactive oxygen species generation were significantly decreased. Berberine effectively alleviated lung injury by reducing lung edema and neutrophil infiltration. Berberine also significantly reduced histopathological inflammatory changes via inhibition of ER stress and activation of Nrf2 signaling. Thapsigargin-induced ER stress and small interference RNA (siRNA)-mediated Nrf2 inhibition abrogated the protective effects of berberine in vitro, whereas siRNA-mediated suppression of ER stress and sulforaphane-induced Nrf2 activation further improved those effects. Importantly, ER stress induction led to Nrf2 activation, whereas PERK depletion partly reduced the level of Nrf2 phosphorylation and translocation in LPS-induced cells. Therefore, berberine inhibits LPS-induced ALI through the PERK-mediated Nrf2/HO-1 signaling axis.

Icariin protects cardiomyocytes against ischaemia/reperfusion injury by attenuating sirtuin 1-dependent mitochondrial oxidative damage.

BACKGROUND AND PURPOSE: Icariin, a major active ingredient in traditional Chinese medicines, is attracting increasing attention because of its unique pharmacological effects against ischaemic heart disease. The histone deacetylase, sirtuin-1, plays a protective role in ischaemia/reperfusion (I/R) injury, and this study was designed to investigate the protective role of icariin in models of cardiac I/R injury and to elucidate the potential involvement of sirtuin-1. EXPERIMENTAL APPROACH: I/R injury was simulated in vivo (mouse hearts), ex vivo (isolated rat hearts) and in vitro (neonatal rat cardiomyocytes and H9c2 cells). Prior to I/R injury, animals or cells were exposed to icariin, with or without inhibitors of sirtuin-1 (sirtinol and SIRT1 siRNA). KEY RESULTS: In vivo and in vitro, icariin given before I/R significantly improved post-I/R heart contraction and limited the infarct size and leakage of creatine kinase-MB and LDH from the damaged myocardium. Icariin also attenuated I/R-induced mitochondrial oxidative damage, decreasing malondialdehyde content and increasing superoxide dismutase activity and expression of Mn-superoxide dismutase. Icariin significantly improved mitochondrial membrane homeostasis by increasing mitochondrial membrane potential and cytochrome C stabilization, which further inhibited cell apoptosis. Sirtuin-1 was significantly up-regulated in hearts treated with icariin, whereas Ac-FOXO1 was simultaneously down-regulated. Importantly, sirtinol and SIRT1 siRNA either blocked icariin-induced cardioprotection or disrupted icariin-mediated mitochondrial homeostasis. CONCLUSIONS AND IMPLICATIONS: Pretreatment with icariin protected cardiomyocytes from I/R-induced oxidative stress through activation of sirtuin-1 /FOXO1 signalling.

A complex pattern with hexagonal lattice and white-eye stripe in dielectric barrier discharge.

A novel type of white-eye pattern in a dielectric barrier discharge system has been investigated in this paper. It is a superposition of a hexagonal lattice and a white-eye stripe in appearance and evolves from a white-eye square grid state with the applied voltage increasing. Its spatio-temporal dynamics obtained by an intensified charge-coupled device shows that it consists of three transient rectangular sublattices. The spatiotemporally resolved evolutions of the molecular vibrational temperature and electron density of the pattern are measured by optical emission spectra. The evolution of surface charge distribution is given and its effect on the self-organized pattern formation is discussed.

Scutellaria barbata D. Don inhibits migration and invasion of colorectal cancer cells via suppression of PI3K/AKT and TGF-ß/Smad signaling pathways.

Metastasis is one of the most aberrant behaviors of cancer cells. Patients with cancers, including colorectal cancer (CRC), have a higher risk of tumor recurrence and cancer-related mortality once metastasis is diagnosed. Existing treatment strategies fail to cure cancer mostly due to the onset of metastasis. Therefore, metastasis remains a challenge in cancer treatment. Some complementary and alternative medical therapies using traditional Chinese medicine have been demonstrated to be clinically effective in cancer treatment. Scutellaria barbata D. Don (SB) is a promising medicinal herb. It was previously reported that the ethanol extract of SB (EESB) is able to promote apoptosis, and inhibit cell proliferation and angiogenesis in human colon cancer cells. However, the anticancer effect of SB and the underlying mechanism require further investigation, particularly its role against metastasis. To further elucidate the antimetastatic effect of SB, MTT and Transwell assays were used in the present study to evaluate the effect of EESB on the proliferation, migration and invasion of the CRC cell line HCT-8. In addition, western blot analysis was performed to detect the expression of matrix metalloproteinases (MMPs), cadherins and other metastasis-associated proteins. EESB significantly reduced HCT-8 cell viability and attenuated the migration and invasion ability of HCT-8 cells in a dose-dependent manner. In addition, EESB decreased the expression of MMP-1, MMP-2, MMP-3/10, MMP-9 and MMP-13, and proteins in the phosphoinositide 3-kinase (PI3K)/AKT and transforming growth factor (TGF)-ß/Smad pathways, but not the epithelial-mesenchymal transition (EMT)-related factors E-cadherin and N-cadherin. In conclusion, the results suggested that SB inhibits CRC cell metastasis via the suppression of PI3K/AKT and TGF-ß/Smad signaling pathways, which may represent a mechanism by which SB exerts an anticancer effect.

Hedyotis diffusa Willd suppresses metastasis in 5­fluorouracil­resistant colorectal cancer cells by regulating the TGF­ß signaling pathway.

Colorectal cancer (CRC) is one of the most common malignant tumors of the digestive tract, and threatens the survival and health of patients with CRC. Chemotherapy remains one of the main therapeutic approaches for patients with CRC; however, drug resistance limits the long­term use. CRC cells with multi­drug resistance (MDR) exhibit increased survival times and metastatic potential, which may lead to the recurrence and metastasis of CRC. In addition, MDR is one of the major causes of chemotherapy failure in clinical treatment. Hedyotis diffusa Willd (HDW) has been used in the treatment of inflammation­associated diseases and malignant tumors, including CRC. The authors previously demonstrated that HDW could reverse MDR in CRC cells; however, its underlying mechanism, particularly in MDR­associated metastasis, remains to be elucidated. In the present study, the drug­resistant CRC cell line HCT­8/5­fluorouracil (5­FU) was used to investigate the effect of HDW on the growth and metastasis of cancer cells. Cell viability was assessed using the MTT assay. Cell adhesion potential was evaluated using adhesion experiments. Cell migration was assessed using wound healing and Transwell assays. The mRNA and protein expression levels of crucial factors in the transforming growth factor­ß (TGF­ß) signaling pathway, including TGF­ß, Mothers against decapentaplegic homolog 4 (SMAD4), neural (N)­cadherin, and epithelial (E)­cadherin, were analyzed using the reverse transcription­semi­quantitative polymerase chain reaction and western blotting, respectively. The results demonstrated that the HCT­8/5­FU cell line was more resistant to 5­FU and thus can be used as the resistant cell model. HDW was able to inhibit the viability, and adhesive, migratory and invasion potential of the HCT­8/5­FU cells. In addition, HDW was able to downregulate the expression of TGF­ß, SMAD4 and N­cadherin, and upregulate E­cadherin, at the gene and protein level. In conclusion, the results demonstrated that HDW may suppress the metastasis of 5­FU­resistant CRC cells via regulation of the TGF­ß signaling pathway, which was also considered to be one of the underlying mechanisms of its anti­CRC effect.

Scutellaria barbata D. Don inhibits 5-fluorouracil resistance in colorectal cancer by regulating PI3K/AKT pathway.

5-Fluorouracil (5-FU) resistance or multidrug resistance (MDR) has become a major obstacle in clinical treatment of cancers including colorectal cancer (CRC). Aberrant activation of phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT) pathway may lead to unlimited growth and chemoresistance in CRC cells, which thus could be a promising therapeutic target. As a long-term used traditional Chinese folk-medicine, Scutellaria barbata D. Don (SB) processes specific anticancer activity, but its activity against cancer chemoresistance is less known. Therefore, using a 5-FU-resistant CRC cell line HCT-8/5-FU, in this study we evaluated the therapeutic efficacy of the ethanol extracts of SB (EESB) against 5-FU resistance and explored the possible molecular mechanisms. We found that EESB significantly suppressed proliferation and promoted apoptosis in HCT-8/5-FU cells. Additionally, EESB displayed remarkable effect enhancing the retention of the ATP-binding cassette (ABC) transporter substrate, rhodamine­123 (Rh­123) in HCT-8/5-FU cells. Furthermore, EESB obviously downregulated the expression of cyclin D1, Bcl-2 and ABCG2, while upregulated p21 and Bax expression. Moreover, EESB showed a prominent suppressive effect on the activation of PI3K/AKT pathway. The findings suggested that Scutellaria barbata D. Don was able to inhibit chemoresistance in colorectal cancer by suppression of the PI3K/AKT pathway.

Spica Prunellae extract suppresses the growth of human colon carcinoma cells by targeting multiple oncogenes via activating miR-34a.

Spica Prunellae is the spike of the herb Prunella vulgaris L. in traditional Chinese medicine which is often used for the treatment of various cancers including colorectal cancer. In the present study, we found that a key tumor suppressor, microRNA-34a (miR-34a) is involved in the antitumor activity for Spica Prunellae. Human colon carcinoma HCT-8 cells treated with an ethanol extract of Spica Prunellae (EESP) had significantly decreased cell proliferation and viability, in a dose-dependent manner. Flow cytometry analysis with Annexin V/PI staining analysis revealed that EESP treatment could induce apoptosis of HCT-8 cells. The level of miR-34a was upregulated in HCT-8 cells following EESP treatment, whereas expression levels of its target genes Notch1, Notch2 and Bcl-2 were downregulated. Inhibition of miR-34a rescued the expression of these target genes. These results revealed that Spica Prunellae can suppress the growth of HCT-8 cells by targeting Notch1, Notch2 and Bcl-2 via activation of miR-34a.