Effect of Paeoniae Radix Rubra (Paeonia lactiflora Pall.) extract on mucin secretion, gene expression in human airway epithelial cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Paeoniae Radix Rubra (PRR), the root of Paeonia lactiflora Pall., is a traditional Chinese medicine which has the effects of regulating various inflammatory diseases, treating blood stasis, and enhancing blood circulation. AIM OF THE STUDY: This study examined whether Paeoniae Radix rubra extract (PRRE) and Paeoniflorin (PF) affect mucin production, gene expression including MUC5AC, and protein expression related to the ERK pathway induced by TNF-α from human airway epithelial cells. MATERIALS AND METHODS: NCI-H292 cells induced by TNF-α were treated with each agent. MUC5AC mucin gene expression and mucin protein production were measured by reverse transcription polymerase chain reaction, staining, and enzyme-linked immunosorbent assay. Western blot was used to investigate the cell signaling pathways. RESULTS: PRRE and PF inhibited the production of MUC5AC mucin protein and gene expression in TNF-α-induced H292 cells. In Western blot, PRRE was involved in protein expression related to the ERK pathway. CONCLUSIONS: Overall, PRRE effectively inhibited the MUC5AC, and inflammatory cytokines expression caused by TNF-α, which was closely involved in the ERK pathway. PRRE may have the potential for treating mucus producing respiratory inflammation.

Anti-Inflammatory Effect of Protopine through MAPK and NF-κB Signaling Regulation in HepG2 Cell.

Protopine is a substance used for hemostasis with an anti-inflammatory action and is one of the substances that are actively undergoing experiments to confirm their utility as anticancer agents. This study examined the molecular changes in the cellular signaling pathways associated with inflammatory responses in phorbol 12-myristate 13 acetate (PMA)-induced human hepatocellular carcinoma cell line (Hep G2). The inhibition of PMA-induced phosphorylation of I-κB in HepG2, the effect of protopine on the MAPK signals, the inhibition of COX-2 activity, and the inhibition of MMP-9 as a medium of inflammatory response were evaluated by Western blot and qPCR. The effect of protopine on the survival rates in HepG2 cells was evaluated and found to be stable to a processing concentration of up to 40µM. Subsequent Western blot analyses showed that protopine blocks the transfer of the MAPKs cell signals induced by PMA and the transfer of the subunit of the nuclear factor-kappa B (NF-κB) to the nucleolus. Protopine inhibited the kappa alpha (I-κBα) phosphorylation in the cytosol and blocked PMA-induced inflammation via COX-2 activity inhibition. The expression of MMP-9 at the gene and protein levels, which is associated with cell migration and metastasis, was reduced by protopine.

Inhibition of MEK-ERK pathway enhances oncolytic vaccinia virus replication in doxorubicin-resistant ovarian cancer.

Oncolytic vaccinia virus (OVV) has been reported to induce cell death in various types of cancer; however, the oncolytic activity of OVV in drug-resistant ovarian cancer remains limited. In the present study, we established doxorubicin-resistant ovarian cancer cells (A2780-R) from the A2780 human ovarian cancer cell line. Both A2780 and A2780-R cells were infected with OVV to explore its anticancer effects. Interestingly, OVV-infected A2780-R cells showed reduced viral replication and cell death compared with A2780 cells, suggesting their resistance against OVV-induced oncolysis; to understand the mechanism underlying this resistance, we explored the involvement of protein kinases. Among protein kinase inhibitors, PD0325901, an MEK inhibitor, significantly augmented OVV replication and cell death in A2780-R cells. PD0325901 treatment increased the phosphorylation of STAT3 in A2780-R cells. Moreover, cryptotanshinone, a STAT3 inhibitor, abrogated PD0325901-stimulated OVV replication. Furthermore, trametinib, a clinically approved MEK inhibitor, increased OVV replication in A2780-R cells. Transcriptomic analysis showed that the MEK inhibitor promoted OVV replication via increasing STAT3 activation and downregulating the cytosolic DNA-sensing pathway. Combined treatment with OVV and trametinib attenuated A2780-R xenograft tumor growth. These results suggest that pharmacological inhibition of MEK reinforces the oncolytic efficacy of OVV in drug-resistant ovarian cancer.

Stimuli-Responsive Adaptive Nanotoxin to Directly Penetrate the Cellular Membrane by Molecular Folding and Unfolding.

Biological nanomachines, including proteins and nucleic acids whose function is activated by conformational changes, are involved in every biological process, in which their dynamic and responsive behaviors are controlled by supramolecular recognition. The development of artificial nanomachines that mimic the biological functions for potential application as therapeutics is emerging; however, it is still limited to the lower hierarchical level of the molecular components. In this work, we report a synthetic machinery nanostructure in which actuatable molecular components are integrated into a hierarchical nanomaterial in response to external stimuli to regulate biological functions. Two nanometers core-sized gold nanoparticles are covered with ligand layers as actuatable components, whose folding/unfolding motional response to the cellular environment enables the direct penetration of the nanoparticles across the cellular membrane to disrupt intracellular organelles. Furthermore, the pH-responsive conformational movements of the molecular components can induce the apoptosis of cancer cells. This strategy based on the mechanical motion of molecular components on a hierarchical nanocluster would be useful to design biomimetic nanotoxins.

Salidroside, 8(E)-Nuezhenide, and Ligustroside from Ligustrum japonicum Fructus Inhibit Expressions of MMP-2 and -9 in HT 1080 Fibrosarcoma.

A phenyl ethanoid, salidroside (SAL), and two secoiridoids, 8(E)-nuezhenide (NZD) and ligustroside (LIG), were isolated from fruits of Ligustrumjaponicum, used as traditional folk medicine, and their chemical structures were elucidated by the comparison of spectral data with published literature. Matrix metalloproteinases (MMPs) are major enzymes that play crucial roles in the metastasis and invasive behavior of tumors. In particular, MMP-2 and MMP-9, regulated by the MAPK signaling pathways, including p38, ERK and JNK, are known to play a key role in the degradation of the basement membrane. In the present study, the effects of SAL, NZD and LIG on the expression of MMP-2 and -9 were examined in phorbol 12-myristate 13-acetate (PMA)-induced HT 1080 cells. All the compounds significantly lowered the amount of MMP-2 and MMP-9 released, as determined by gelatin zymography and ELISA. In addition, the mRNA and protein expression levels of MMP-2 and MMP-9 were significantly suppressed, as measured by RT-PCR and Western blotting. According to the Western blotting assay, SAL and LIG effectively reduced the expression of MMP-2 in a dose-dependent manner. NZD lowered the expression of MMP-9 in a similar way. The phosphorylation of p38, ERK and JNK was also significantly suppressed by these compounds. These findings suggest that all the compounds regulate the release and expression of MMP-2 and MMP-9 via MAPK signaling pathways.

Noninvasive Precision Screening of Prostate Cancer by Urinary Multimarker Sensor and Artificial Intelligence Analysis.

Screening for prostate cancer relies on the serum prostate-specific antigen test, which provides a high rate of false positives (80%). This results in a large number of unnecessary biopsies and subsequent overtreatment. Considering the frequency of the test, there is a critical unmet need of precision screening for prostate cancer. Here, we introduced a urinary multimarker biosensor with a capacity to learn to achieve this goal. The correlation of clinical state with the sensing signals from urinary multimarkers was analyzed by two common machine learning algorithms. As the number of biomarkers was increased, both algorithms provided a monotonic increase in screening performance. Under the best combination of biomarkers, the machine learning algorithms screened prostate cancer patients with more than 99% accuracy using 76 urine specimens. Urinary multimarker biosensor leveraged by machine learning analysis can be an important strategy of precision screening for cancers using a drop of bodily fluid.

Combination of Scutellaria baicalensis and Metformin Ameliorates Diet-Induced Metabolic Dysregulation in Mice via the Gut-Liver-Brain Axis.

Scutellaria baicalensis (SB), a herbal medicine, is commonly used to treat metabolic diseases, while Metformin (MF) is a widely used drug for type 2 diabetes. The purpose of this study was to investigate whether co-treatment of SB with MF could produce a potential therapeutic effect on high-fat and high-fructose diet (HFFD)-induced metabolic dysregulation. First, we optimized the dose of SB (100, 200, 400, and 800[Formula: see text]mg/kg) with MF (200[Formula: see text]mg/kg) in HFFD-induced C57BL6J mice. Next, the optimized dose of SB (400[Formula: see text]mg/kg) was co-administered with MF (50, 100, and 200[Formula: see text]mg/kg) in a similar animal model to find the effective combinations of SB and MF. Metabolic markers were determined in serum and tissues using different assays, histology, gene expression, and gut microbial population. The SB and MF co-treatment significantly decreased the body, liver, and VAT weights. The outcome of OGTT was improved, and the fasting insulin, HbA1c, TG, TC, LDL-c, AST, and ALT were decreased, while HDL-c was significantly increased. Histological analyses revealed maintained the integrity of liver, adipose tissue, and intestine prevented lipid accumulation in the liver and intestine and combated neuronal damage in the brain. Importantly, controlled the expression of PPAR[Formula: see text], and IL-6 genes in the liver, and expression of BDNF, Glut1, Glut3, and Glut4 genes in the brain. Treatment-specific gut microbial segregation was observed in the PCA chart. Our findings indicate that SB and MF co-treatment is an effective therapeutic approach for HFFD-induced metabolic dysregulation which is operated through the gut-liver-brain axis.

Wirelessly controlled, bioresorbable drug delivery device with active valves that exploit electrochemically triggered crevice corrosion.

Implantable drug release platforms that offer wirelessly programmable control over pharmacokinetics have potential in advanced treatment protocols for hormone imbalances, malignant cancers, diabetic conditions, and others. We present a system with this type of functionality in which the constituent materials undergo complete bioresorption to eliminate device load from the patient after completing the final stage of the release process. Here, bioresorbable polyanhydride reservoirs store drugs in defined reservoirs without leakage until wirelessly triggered valve structures open to allow release. These valves operate through an electrochemical mechanism of geometrically accelerated corrosion induced by passage of electrical current from a wireless, bioresorbable power-harvesting unit. Evaluations in cell cultures demonstrate the efficacy of this technology for the treatment of cancerous tissues by release of the drug doxorubicin. Complete in vivo studies of platforms with multiple, independently controlled release events in live-animal models illustrate capabilities for control of blood glucose levels by timed delivery of insulin.

Chemically or surgically induced thyroid dysfunction altered gut microbiota in rat models.

Thyroid hormones are essential for the regulation of energy homeostasis and metabolic processes. However, the relationship between thyroid function and host gut microbial communities is not properly understood. To determine whether and how gut microbiota is associated with thyroid function, metagenomics analysis of the bacterial population in fecal samples of rat models of hyperthyroidism (induced by levothyroxine) and hypothyroidism (induced by propylthiouracil or thyroidectomy) was conducted through 16S rRNA gene sequencing. Our results revealed that all thyroid dysfunction models were definitely established and gut microbial composition varied according to different thyroid functional status. The relative abundance of Ruminococcus was significantly higher in the hyperthyroidism group (HE) vs both the normal and hypothyroidism groups (HO) while S24-7 was significantly higher in the HO group. The population of Prevotellaceae and Prevotella were significantly lower in the HO group vs the normal. Firmicutes and Oscillospira were significantly higher in the SHO (surgery-induced hypothyroidism) group, while Prevotellaceae and Prevotella showed lower abundance in the SHO group than the SHAM group. Present results suggest that thyroid functions may have the potential to influence the profile of gut microbiota and could be used as foundation to investigate interaction mechanism between thyroid and gut microbiome.

6-Acetyl-2,2-Dimethylchroman-4-One Isolated from Artemisia princeps Suppresses Adipogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stromal Cells via Activation of AMPK.

Obesity is a world-wide health concern with increasing mortality and morbidity rates. Development of novel therapeutic agents for obesity from phytochemicals may lead to the effective prevention and control of obesity and obesity-related complications. 6-acetyl-2,2-dimethylchroman-4-one (1) was isolated from a dietary plant, Artemisia princeps. The antiobesity effect of compound 1 was determined in human bone marrow-derived mesenchymal stromal cells (hBM-MSCs) induced to differentiate into adipocytes. Treatment with compound 1 resulted in decreased lipid accumulation and expression of key adipogenic markers, proliferator-activated receptor-γ, CCAAT/enhancer-binding protein-α, and sterol regulatory element-binding transcription factor 1. It was also shown that compound 1 downregulated the adipogenesis-induced p38 and JNK MAPK activation, while upregulating adipogenesis inhibitory ß-catenin-dependent Wnt10b pathway. Compound 1 was also able to stimulate adenosine monophosphate-activated protein kinase phosphorylation, which was suggested to be the underlying mechanism that resulted in inhibition of adipogenesis in hBM-MSCs. In conclusion, 6-acetyl-2,2-dimethylchroman-4-one was identified as a bioactive constituent of A. princeps that exerts antiobesity properties via suppressing adipocyte formation.

Unimolecular Polypeptide Micelles via Ultrafast Polymerization of N-Carboxyanhydrides.

Polypeptide micelles are widely used as biocompatible nanoplatforms but often suffer from their poor structural stability. Unimolecular polypeptide micelles can effectively address the structure instability issue, but their synthesis with uniform structure and well-controlled and desired sizes remains challenging. Herein we report the convenient preparation of spherical unimolecular micelles through dendritic polyamine-initiated ultrafast ring-opening polymerization of N-carboxyanhydrides (NCAs). Synthetic polypeptides with exceptionally high molecular weights (up to 85 MDa) and low dispersity (D < 1.05) can be readily obtained, which are the biggest synthetic polypeptides ever reported. The degree of polymerization was controlled in a vast range (25-3200), giving access to nearly monodisperse unimolecular micelles with predictable sizes. Many NCA monomers can be polymerized using this ultrafast polymerization method, which enables the incorporation of various structural and functional moieties into the unimolecular micelles. Because of the simplicity of the synthesis and superior control over the structure, the unimolecular polypeptide micelles may find applications in nanomedicine, supermolecular chemistry, and bionanotechnology.

Beneficial effect of immunobiotic strains on attenuation of Salmonella induced inflammatory response in human intestinal epithelial cells.

Probiotic bacteria have the ability to modulate host immune responses and have potent therapeutic functional effects against several diseases, including inflammatory diseases. However, beneficial effects of probiotics are strain specific and their interactions with host immune cells to modulate inflammatory response are largely unknown. Intestinal epithelial cells (IECs), which are the first line of defense against invading pathogens, and connects between commensals/probiotics and immune system; therefore, in this study, we used human IECs to assess the probiotic effects of three selected Lactobacillus strains in vitro. An HT-29 colonic epithelial cell and HT-29/blood mononuclear cells co-culture system were stimulated with Lactobacillus followed by Salmonella for different hours, after which the mRNA level of cytokines, ß-defensin-2 and negative regulators for TLR signaling and protein levels of ZO-1 and IκB-α were analyzed by real-time polymerase chain reaction and western blot analysis. L. brevis decreased Salmonella induced IL-6, IL-8, MCP-1 and IL-1ß levels, whereas L. pentosus suppressed IL-6 and MCP-1 in HT-29 cells. Moreover, L. brevis was able to increase the mRNA levels of A20, Tollip, SIGIRR and IRAKM, while L. pentosus reduced the levels of A20, and IRAKM in response to Salmonella. In addition, decrease in protein level of TNF-α and increase in mRNA level of IL-10 was observed in L. brevis and L. pentosus treated HT-29 cells. Lactobacillus strains were differentially modulated ZO-1 and p-IκB-α in HT-29 cells treated with Salmonella. Overall, the results of this study indicate that Lactobacillus strains attenuate Salmonella induced inflammatory responses through beneficial modulation of TLR negative regulators and the NF-κB pathway.

Combined effects of Scutellaria baicalensis with metformin on glucose tolerance of patients with type 2 diabetes via gut microbiota modulation.

Metformin is a widely prescribed antidiabetic agent, whereas Scutellaria baicalensis (SB) is a commonly used medicinal herb for treatment of type 2 diabetes (T2D). Gut microbiota is involved in pathophysiology of metabolic diseases including T2D, and intestinal microbiota may be one of the important therapeutic targets for the ailment. This study was conducted to investigate the effects of SB combined with metformin on treatment of T2D while evaluating changes in the gut microbiota composition. Patients with T2D were randomized into control and treatment groups. Subjects who had already been prescribed metformin were allotted to additional SB (3.52 g/day) group or placebo group. The initial treatment session was 8 wk, and after washout period for 4 wk they were crossed over to the opposite treatment for another 8 wk. The influence of SB and placebo on the intestinal microbiota was analyzed by MiSeq system based on 16S rRNA gene. Glucose tolerance was lower in the SB group than the placebo group. Similarly, the relative RNA expression of TNF-α was significantly reduced after SB treatment. SB treatment influenced the gut microbiota, especially Lactobacillus and Akkermansia, which showed remarkable increases after SB treatment. Some subjects showed high liver enzyme levels after SB treatment, and their microbiota composition at baseline differed with subjects whose liver enzymes were not affected. We also predicted that selenocompound metabolism was increased and naphthalene degradation was decreased after SB treatment. These results suggest that SB with metformin treatment may improve the glucose tolerance and inflammation and influence the gut microbiota community in T2D.

Mixing oil and water with ionic liquids: bicontinuous microemulsions under confinement.

We report the structural transition of a phosphonium ionic liquid-based microemulsion from the bulk to nanoconfined between atomically flat micas. Upon the nanoconfinement, we observed a firmly surface-adsorbed ionic liquid film that stabilizes the nanoconfined microemulsion. Further confinement (<11 nm) induces rearrangements in the microemulsion culminating into two well-ordered layers with slow dynamics.

Immunobiotic Strains Modulate Toll-Like Receptor 3 Agonist Induced Innate Antiviral Immune Response in Human Intestinal Epithelial Cells by Modulating IFN Regulatory Factor 3 and NF-κB Signaling.

Many studies have demonstrated that immunobiotics with immunoregulatory functions improve the outcomes of several bacterial and viral infections by modulating the mucosal immune system. However, the precise mechanisms underlying the immunoregulatory and antiviral activities of immunobiotics have not yet been elucidated in detail. The present study was conducted to determine whether selected lactic acid bacteria (LAB) modulate toll-like receptor 3 (TLR3) agonist polyinosinic:polycytidylic acid (PolyI:C) induced viral response in human intestinal epithelial cells (IECs). PolyI:C increased the expression of interferon-ß (IFN-ß), interleukin-6 (IL-6), interleukin-8 (IL-8), monocyte chemoattractant protein (MCP-1), and interleukin-1ß (IL-1ß) in HCT116 cells, and these up-regulations were significantly altered when cells were pre-stimulated with LAB isolated from Korean fermented foods. LAB strains were capable to up-regulate IFN-ß but down-regulated IL-6, IL-8, MCP-1, and IL-1ß mRNA levels as compared with PolyI: C. HCT-116 cell treatment with LABs beneficially modulated the mRNA levels of C-X-C motif chemokine 10 (CXCL-10), 2-5A oligoadenylate synthetase 1 (OSA1), myxovirus resistance protein (MxA), TLR3, and retinoic acid inducible gene-I (RIG-I), and TLR negative regulators. In addition, LABs increased IFN-ß, IFN-α, and interleukin-10 (IL-10) and decreased tumor necrosis factor-α (TNF-α) and IL-1ß protein/mRNA levels in THP-1 cells. LABs also protected the cells by maintaining tight-junction proteins (zonula occludens-1 and occludin). The beneficial effects of these LABs were mediated via modulation of the interferon regulatory factor 3 (IRF3) and nuclear factor-kappa B (NF-κB) pathways. Overall, the results of this study indicate that immunobiotics have potent antiviral and anti-inflammatory activities that may use as antiviral substitutes for human and animal applications.

Immunobiotics Beneficially Modulate TLR4 Signaling Triggered by Lipopolysaccharide and Reduce Hepatic Steatosis In Vitro.

Hepatic inflammation and injury may result from the translocation of pathological bacteria and their proinflammatory mediators. Probiotics attenuate hepatic diseases related to inflammation by exhibiting immunoregulatory effects. Therefore, this study was conducted to evaluate lipid reduction and immunoregulatory potentials of probiotic bacteria in vitro. HepG2 cells treated with total cellular fluid (TCF) of LABs reduced lipid accumulation. Moreover, cells responded to lipopolysaccharide (LPS) by producing higher levels of IL-6, IL-8, MCP-1, and TNF-α. TCF of LABs treatment showed remarkably diminished levels of the expression of these cytokines via modulation of the expression of TLR-negative regulators, as well as MAPK and NF-κB pathways. Moreover, heat-killed LABs were able to diminish TGF-ß, IL-1ß, and IL-6 and to increase IL-10 and TLR4 levels in THP-1 cells. LABs also decreased the protein level of TNF-α. These results demonstrated that immunobiotics exhibit potent immunoregulatory activity and may be used as effective therapeutic agents to alleviate inflammatory response.

Evaluation of MMP Inhibitors Isolated from Ligustrum japonicum Fructus.

The current study investigated the ability of two secoiridoids, GL-3 (1) and oleonuezhenide (2), isolated from the fruits of Ligustrum japonicum to inhibit MMP-2 and -9 activity in phorbol 12-myristate 13-acetate (PMA)-induced HT-1080 human fibrosarcoma cells. Both compounds 1 and 2 were able to exert lowered gelatin digestion activity for MMP-2 and -9 tested by gelatin zymography via suppressing the release of MMPs to culture medium according to ELISA results. Treatment with compounds was also able to suppress the expression of both mRNA and protein levels of MMP-2 and -9. Action mechanism behind the MMP inhibitory effect of the compounds was suggested to be via MAPK pathway indicated by decreased levels of phosphorylated p38, ERK and JNK proteins evaluated employing immunoblotting. Compound 1 was shown to be slightly more active to inhibit MMP-2 and -9, however, compound 2 showed more regular dose-dependency during inhibition. In conclusion, this study suggested that GL-3 and oleonuezhenide were notable natural origin potent MMP inhibitors and could serve as lead compounds for development of anti-invasive MMP inhibitors against tumor metastasis.

Quantitative Single-Cell Analysis of Isolated Cancer Cells with a Microwell Array.

The heterogeneous nature of tumor-cell populations suggests that quantitative analysis at the single-cell level may provide better insights into cancer biology. Specifically, detection of multiple biomarkers from a single cell offers important initial information about cellular behavior. However, conventional approaches limit biomarker detection at the single-cell level. Here, we fabricated a polymer microwell array to capture single cells from prostate-cancer cell lines and quantitatively analyzed the expression of three different cancer-related biomarkers, CD44, EpCAM, and PSMA, without a membrane protein-extraction step. The resulting information on cell-surface biomarker distributions was compared with that from other standard analytical techniques. Interestingly, a large variation in CD44-expression levels was observed when the cell-proliferation cycle was modulated. On the other hand, the expression levels of EpCAM in three different cell lines are consistent among the different analytical methods with the exception of the microarray, where it has a different substrate material to adhere to. This observation clearly emphasizes that biomarker choice and environmental control are critical for properly understanding the single-cell state.

Functional capabilities of probiotic strains on attenuation of intestinal epithelial cell inflammatory response induced by TLR4 stimuli.

Intestinal epithelial cells (IECs) respond to intruders and their cellular molecules by displaying inflammatory state that can be abrogated by probiotics. However, the molecular mechanisms underlying the beneficial activity of probiotic strains have yet to be elucidated. This study was conducted to investigate whether probiotic strains have immunoregulatory effects in IECs, and how they respond to bacterial lipopolysaccharide (LPS) in vitro. Caco2 cells were stimulated with LABs and followed by LPS. The expression of different cytokines that involved in toll-like receptor (TLR) signaling was analyzed. Caco2 cells that were exposed to LPS showed upregulated expression of IL-6, CXCL8, CCL2, and BPI that were counteracted by LAB strains through the modulation of TLR negative regulators (A20, Tollip, SIGIRR, and IRAKM), p38 MAPK and p65 NF-κB signaling. Lactobacillus plantarum, L. farciminis, and L. pentosus unveiled better activity as compared to other strains. Moreover, LAB strains were the potent inducers of immunoregulatory cytokines in coculture system. The expression of IL-10 and TGF-ß were found to be higher as compared with LPS. Conversely, TNF-α at the protein level was dampened by LAB strains in both the apical and basolateral compartments. Collectively, our results demonstrated that the selected LAB strains exert profound immunoregulatory effects in response to LPS on IECs; however, further studies in vivo and in clinical settings are important to corroborate these effects. © 2018 BioFactors, 45(2):223-235, 2019.