Optimizing Gluten Extraction Using Eco-friendly Imidazolium-Based Ionic Liquids: Exploring the Impact of Cation Side Chains and Anions.

Gluten is a well-known food allergen globally, and it can induce immune responses in celiac- and nonceliac gluten-sensitive patients. The gliadin proteins from gluten have a special amino acid sequence that make it hydrophobic. One way to deal with gluten allergies is to provide a gluten-free diet. The hydrophobic characteristic of gliadin makes gliadin detection more difficult. An analyst needs to use an organic solvent or multiple processes to denature gluten for extraction. Although organic solvents can rapidly extract gluten in a sample, organic solvent also denatures the antibody and induces false biotest results without buffer dilute, and the accuracy will reduce with buffer dilute. An ionic liquid (IL) is a highly modifiable green chemical organic salt. The imidazolium has a cationic structure and is modified with different lengths (C = 0, 1, 3, 5, 7, 9, and 12) of carbon side chains with organic and inorganic anions [methanesulfonate (MSO), Cl-, F-, NO3-, HSO4-, and H2PO4-] to make different kinds of ILs for testing the solubility of gliadin. Different IL/water ratios were used to test the solubility of gluten. We measured the solubility of gliadin in different imidazolium ILs, and the kinetic curve of gliadin dissolved in 1% [C5DMIM][MSO]aq was conducted. We also used circular dichroism spectroscopy and an enzyme-linked immunosorbent assay to measure the gliadin structure and the effect of binding with an antibody after 1% [C5DMIM][MSO]aq treatment. An 2,3-bis-(2-methoxy-4- nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay was used to test the toxicity of [C5DMIM][MSO]aq in N2a cells. In our research, 1% [C5DMIM][MSO]aq produced a good solubility of gluten, and it could dissolve more than 3000 ppm of gluten in 5 min. [C5DMIM][MSO]aq did not break down the gluten structure and did not restrict antibody binding to gluten, and more importantly, [C5DMIM][MSO] did not exhibit cell toxicity. In this report, we showed that [C5DMIM][MSO] could be a good extraction solution applied for gluten detection.

Synthesis and Structure-Activity Relationship of Salvinal Derivatives as Potent Microtubule Inhibitors.

Salvinal is a natural lignan isolated from the roots of Salvia mitorrhiza Bunge (Danshen). Previous studies have demonstrated its anti-proliferative activity in both drug-sensitive and -resistant cancer cell lines, with IC50 values ranging from 4-17 µM. In this study, a series of salvinal derivatives was synthesized and evaluated for the structure-activity relationship. Among the twenty-four salvinal derivatives, six compounds showed better anticancer activity than salvinal. Compound 25 displayed excellent anticancer activity, with IC50 values of 0.13-0.14 µM against KB, KB-Vin10 (overexpress MDR/Pgp), and KB-7D (overexpress MRP) human carcinoma cell lines. Based on our in vitro microtubule depolymerization assay, compound 25 showed depolymerization activity in a dose-dependent manner. Our findings indicate that compound 25 is a promising anticancer agent with depolymerization activity that has potential for the management of malignance.

Dual DNA Transfection Using 1,6-Hexanedithiol-Conjugated Maleimide-Functionalized PU-PEI600 For Gene Correction in a Patient iPSC-Derived Fabry Cardiomyopathy Model.

Non-viral gene delivery holds promises for treating inherited diseases. However, the limited cloning capacity of plasmids may hinder the co-delivery of distinct genes to the transfected cells. Previously, the conjugation of maleimide-functionalized polyurethane grafted with small molecular weight polyethylenimine (PU-PEI600-Mal) using 1,6-hexanedithiol (HDT) could promote the co-delivery and extensive co-expression of two different plasmids in target cells. Herein, we designed HDT-conjugated PU-PEI600-Mal for the simultaneous delivery of CRISPR/Cas9 components to achieve efficient gene correction in the induced pluripotent stem cell (iPSC)-derived model of Fabry cardiomyopathy (FC) harboring GLA IVS4 + 919 G > A mutation. This FC in vitro model recapitulated several clinical FC features, including cardiomyocyte hypertrophy and lysosomal globotriaosylceramide (Gb3) deposition. As evidenced by the expression of two reporter genes, GFP and mCherry, the addition of HDT conjugated two distinct PU-PEI600-Mal/DNA complexes and promoted the co-delivery of sgRNA/Cas9 and homology-directed repair DNA template into target cells to achieve an effective gene correction of IVS4 + 919 G > A mutation. PU-PEI600-Mal/DNA with or without HDT-mediated conjugation consistently showed neither the cytotoxicity nor an adverse effect on cardiac induction of transfected FC-iPSCs. After the gene correction and cardiac induction, disease features, including cardiomyocyte hypertrophy, the mis-regulated gene expressions, and Gb3 deposition, were remarkably rescued in the FC-iPSC-differentiated cardiomyocytes. Collectively, HDT-conjugated PU-PEI600-Mal-mediated dual DNA transfection system can be an ideal approach to improve the concurrent transfection of non-viral-based gene editing system in inherited diseases with specific mutations.

Anxiolytic effect of an extract of Salvia miltiorrhiza Bunge (Danshen) in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia miltiorrhiza Bunge (Danshen), a traditional Chinese medicine, has demonstrated in modern studies for its pharmacological activities in treatments of CNS disorders like insomnia, dysphoria. However, its application on anxiolytic effect from the ethanol extract of Salvia miltiorrhiza Bunge (SMEtOH) has not yet been reported. MATERIALS AND METHODS: This study investigated the anxiolytic effect of the SMEtOH using the elevated plus-maze test (EPM) and the hole-board test (HBT) with diazepam and buspirone as positive controls. Also, the spontaneous locomotor activity of mice had been investigated in the open field. Further, we have illustrated the anxiolytic mechanisms of SMEtOH with its influencing upon GABAergic and/or serotonergic nervous systems via a method that SMEtOH was co-administered with flumazenil, a benzodiazepine (BZD) antagonist, or a drug (WAY-100635), a selective 5HT1A receptor antagonist. RESULTS: In hole-board test, results presented that SMEtOH increased head-dip counts and duration time. On the other hand, a decrease in spontaneous locomotor activity was observed. In the EPM test, SMEtOH increased the percentage of open-arm entries and the percentage of time spent in open arms. However, when SMEtOH co-administered with flumazenil or WAY-100635, the anxiolytic effect of SMEtOH was significantly counteracted. CONCLUSION: From these results, we can conclude that the anxiolytic mechanism of SMEtOH is exerted through an activation of the BZD and 5HT1A receptors.

Using cationic polyurethane-short branch PEI as microRNA-driven nano-delivery system for stem cell differentiation.

BACKGROUND: Non-viral gene delivery, such as using biodegradable polyurethane short-branch polyethylenimine (PU-PEI), has been considered a potentially safer gene delivery system in comparison to conventional virus systems. METHODS: The polycationization of DNA complexes protects DNA from nuclease degradation, and these DNA complexes are nanoscale in size to enter the cell through endocytosis. RESULTS: Due to the net positive surface charge of the cell, these polyplexes efficiently bind to the cell through electrostatic interactions with negatively charged membrane components. Cationic PU-PEI has been shown to be non-cytotoxic and has a high transfection efficiency, making it a practical gene delivery material in diseases. CONCLUSION: We developed a PU-PEI nanomedicine-based platform to efficiently deliver microRNA in promoting differentiation capacity of stem cells, especially on induced pluripotent stem cells.

Treatment of ß-thujaplicin counteracts di(2-ethylhexyl)phthalate (DEHP)-exposed vascular smooth muscle activation, inflammation and atherosclerosis progression.

The initiation of atherosclerosis involves up-regulation of molecules such as E-selectin, VCAM-1, and ICAM-1. The progression of atherosclerosis is linked to proliferation and migration of vascular smooth muscle cell via MMP-2 and MMP-9 activities. However, the etiology of atherosclerosis concerning plasticizers is unknown. We evaluated ß-thujaplicin in preventing the development of atherosclerosis in a model induced by pro-inflammatory cytokines. Moreover, we established a new atherosclerosis model in vascular smooth muscle cells (VSMC) exposed to a common contact plasticizer, di(2-ethylhexyl)phthalate (DEHP). SEVC4-10 endothelial cells were treated with 50% RAW conditioned medium and A7r5 VSMC was treated with the plasticizer, with/without ß-thujaplicin (4 or 12 µM). Production of E-selectin, ICAM-1, and VCAM-1 in SEVC4-10 cells as well as MMP-2/MMP-9 (both expression and activity) in VSMC were monitored. Results showed that the conditioned medium induced E-selectin and ICAM were significantly prevented by ß-thujaplicin. However, inhibition on the production of VCAM by ß-thujaplicin was only seen in a concentration of 12 µM. Both concentrations of ß-thujaplicin also significantly prevented DEHP-induced MMP-2 and MMP-9 expression and activities. Evidence uncovers that ß-thujaplicin has additional factors in amelioration of atherosclerosis and corroborates that ß-thujaplicin is a strong candidate in preventing the initiation and progression of atherosclerosis.

Danshen improves survival of patients with advanced lung cancer and targeting the relationship between macrophages and lung cancer cells.

In traditional Chinese medicine, Salvia miltiorrhiza Bunge (danshen) is widely used in the treatment of numerous cancers. However, its clinical effort and mechanism in the treatment of advanced lung cancer are unclear. In our study, the in vivo protective effort of danshen in patients with advanced lung cancer were validated using data from the National Health Insurance Research Database in Taiwan. We observed in vitro that dihydroisotanshinone I (DT), a bioactive compound in danshen, exerts anticancer effects through many pathways. First, 10 µM DT substantially inhibited the migration ability of lung cancer cells in both macrophage and macrophage/lung cancer direct mixed coculture media. Second, 10 µM DT repressed the phosphorylation of signal transducer and activator of transcription 3 (STAT3), the protein expression of S-phase kinase associated protein-2 (Skp2), and the mRNA levels of STAT3-related genes, including chemokine (C-C motif) ligand 2 (CCL2). In addition, 10 µM DT suppressed the macrophage recruitment ability of lung cancer cells by reducing CCL2 secretion from both macrophages and lung cancer cells. Third, 20 µM DT induced apoptosis in lung cancer cells. Furthermore, DT treatment significantly inhibited the final tumor volume in a xenograft nude mouse model. In conclusion, danshen exerts protective efforts in patients with advanced lung cancer. These effects can be attributed to DT-mediated interruption of the cross talk between lung cancer cells and macrophages and blocking of lung cancer cell proliferation.

Bacteria-Templated NiO Nanoparticles/Microstructure for an Enzymeless Glucose Sensor.

The bacterial-induced hollow cylinder NiO (HCNiO) nanomaterial was utilized for the enzymeless (without GOx) detection of glucose in basic conditions. The determination of glucose in 0.05 M NaOH solution with high sensitivity was performed using cyclic voltammetry (CV) and amperometry (i-t). The fundamental electrochemical parameters were analyzed and the obtained values of diffusion coefficient (D), heterogeneous rate constant (ks), electroactive surface coverage (Г), and transfer coefficient (alpha-α) are 1.75 × 10(-6) cm²/s, 57.65 M(-1)·s(-1), 1.45 × 10(-10) mol/cm², and 0.52 respectively. The peak current of the i-t method shows two dynamic linear ranges of calibration curves 0.2 to 3.5 µM and 0.5 to 250 µM for the glucose electro-oxidation. The Ni(2+)/Ni(3+) couple with the HCNiO electrode and the electrocatalytic properties were found to be sensitive to the glucose oxidation. The green chemistry of NiO preparation from bacteria and the high catalytic ability of the oxyhydroxide (NiOOH) is the good choice for the development of a glucose sensor. The best obtained sensitivity and limit of detection (LOD) for this sensor were 3978.9 µA mM(-1)·cm(-2) and 0.9 µM, respectively.

Possible Mechanisms of Di(2-ethylhexyl) Phthalate-Induced MMP-2 and MMP-9 Expression in A7r5 Rat Vascular Smooth Muscle Cells.

Proliferation and migration of vascular smooth muscle cells (VSMC) are important in the development and/or progression of many cardiovascular diseases, including atherosclerosis. Evidence shows that matrix metalloproteinase (MMP)-2 and MMP-9 are related to the pathogenesis of atherosclerosis. The expressions of MMP-2 and MMP-9 in atherosclerosis are regulated via various pathways, such as p38 mitogen activated protein kinase (MAPK), extracellular signal regulated kinase 1 and 2 (ERK1/2), Akt, and nuclear factor kappa (NF-κB). Di(2-ethylhexyl) phthalate (DEHP) has been shown to induce atherosclerosis by increasing tumor necrosis factor (TNF)-α, interleukin (IL)-6, and intercellular adhesion molecule (ICAM) productions. However, whether DEHP poses any effects on MMP-2 or MMP-9 expression in VSMC has not yet been answered. In our studies, rat aorta VSMC was treated with DEHP (between 2 and 17.5 ppm) and p38 MAPK, ERK1/2, Akt, NF-κB, and MMP-2 and MMP-9 proteins and activities were measured. Results showed that the presence of DEHP can induce higher MMP-2 and MMP-9 expression than the controls. Similar results on MMP-regulating proteins, i.e., p38 MAPK, ERK1/2, Akt, and NF-κB, were also observed. In summary, our current results have showed that DEHP can be a potent inducer of atherosclerosis by increasing MMP-2 and MMP-9 expression at least through the regulations of p38 MAPK, ERK1/2, Akt, and NF-κB.

Role of medial abrasion phenomenon in the pathogenesis of knee osteoarthritis.

Osteoarthritis of the knee affects a large population worldwide and is associated with an extremely high economic burden largely attributable to the effects of disability, comorbid disease, and the expense of treatment. Since the initiating events that result in the cartilage degradation are poorly understood, there has been very limited success in demonstrating disease modification in clinical trials of potential therapies. Medial plica related medial abrasion phenomenon has recently been identified to have close relationship with medial compartment osteoarthritis. We hypothesized that this abrasion phenomenon will elicit lifelong interplay between pathologic medial plica and the facing medial femoral condyle and might play a role in the pathogenesis of knee osteoarthritis by both physical and chemical effects. After evaluating current evidence, we designed a study to prove that the concentrations of total protein, cartilage degrading related cytokines (tumor necrosis factor-α and interleukin-1ß) and enzyme (matrix metalloproteinase-3) are higher in the medial compartment of the knee having the phenomenon of medial abrasion. The accumulating data and findings about medial abrasion phenomenon might be important for the understanding of the pathogenesis or progression of this common disease. We hope that our hypothesis will stimulate further studies verifying if medial abrasion phenomenon plays more roles in the pathogenesis of knee osteoarthritis. Further clinical observations for its appropriate treatment based on this hypothesis are also mandatory for the benefits of patients.

Reduction of adhesion molecule production and alteration of eNOS and endothelin-1 mRNA expression in endothelium by Euphorbia hirta L. through its beneficial ß-amyrin molecule.

The inflammatory reaction in large blood vessels involves up-regulation of vascular adhesion molecules such as endothelial cell selectin (E-selectin), soluble vascular cell adhesion molecule (sVCAM)-1, and soluble intercellular adhesion molecule (sICAM)-1. These vascular dysfunctions are associated with the development of atherosclerosis. ß-Amyrin, an active component of Euphorbia hirta L., has potent anti-inflammatory effects. So far, its preventive effects against the expression of inflammatory mediator-induced adhesion molecules have not been investigated. Endothelial cells (SVEC4-10 cell line) were treated with 50% RAW conditioned media (i.e., normal SVEC4-10 culture media contains 50% of lipopolysaccharide-activated macrophage culture media) without or with ß-amyrin (0.6 and 0.3 µM). The production levels of E-selectin, sICAM-1, and sVCAM-1 in the SVEC4-10 cells were measured with ELISA assay kits. Under the same treatment conditions, expression of endothelin (ET)-1 and endothelial type of NO synthase (eNOS) mRNA were analyzed by RT-PCR and agarose gel. With ß-amyrin, the 50% RAW conditioned media-induced E-selectin, sICAM-1, and sVCAM-1 levels as well as ET-1 gene expression were all suppressed. ß-Amyrin treatment also restored the 50% RAW conditioned media-suppressed eNOS mRNA expression. These data indicate that ß-amyrin is potentially useful in preventing chronic inflammation-related vascular diseases.

Fabrication of piezoelectric components for a tunable and efficient device for DNA delivery into mammalian cells.

We fabricated three piezoelectric components (PZT) that can produce ultrasonic waves with various generated power in order to improve the delivery of DNA molecule and polymer/DNA complexes into cells. Two cationic polymers (PEI and PDMAEMA) were interacted with DNA to form nano-scaled DNA/polymer complexes with/without the help of PZT devices. The application of PZT devices under optimal conditions helped to avoid cytotoxicity and greatly increased the transfection (DNA delivery) efficiency of these complexes in mammalian cells. The cytotoxicity and transfection efficiency were found to be correlated with the PZT-generated power, waveforms and duration of ultrasonic treatment. There was no observable cytotoxicity in our experimental models and, a maximum transfection efficiency 700% greater than that of polymer/DNA complexes without applying ultrasound was achieved. The transfection efficiency of plain polymer/DNA complexes (without PZT treatment) corresponded to a 630-fold increase in comparison to the naked DNA. The waveforms of generated ultrasound greatly influenced the transfection efficiency, while cytotoxicity was not significantly affected. This means that, for optimal DNA delivery, duration of the peak voltage (Vmax/Div) also plays a role. In addition, the generated waves from PZT do not cause dissociation of polymer/DNA complexes or a change in the particle sizes of these complexes. In conclusion, these results suggest that the operation of PZT devices can be a tunable/safe way to greatly improve DNA delivery for gene therapy.

Chlorella 11-peptide inhibits the production of macrophage-induced adhesion molecules and reduces endothelin-1 expression and endothelial permeability.

The inflammation process in large vessels involves the up-regulation of vascular adhesion molecules such as endothelial cell selectin (E-selectin), intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) which are also known as the markers of atherosclerosis. We have reported that Chlorella 11-peptide exhibited effective anti-inflammatory effects. This peptide with an amino sequence Val-Glu-Cys-Tyr-Gly-Pro-Asn-Arg-Pro-Gln-Phe was further examined for its potential in preventing atherosclerosis in this study. In particular, the roles of Chlorella 11-peptide in lowering the production of vascular adhesion molecules, monocyte chemoattractant protein (MCP-1) and expression of endothelin-1 (ET-1) from endothelia (SVEC4-10 cells) were studied. The production of E-selectin, ICAM-1, VCAM-1 and MCP-1 in SVEC4-10 cells was measured with ELISA. The mRNA expression of ET-1 was analyzed by RT-PCR and agarose gel. Results showed that Chlorella 11-peptide significantly suppressed the levels of E-selectin, ICAM, VCAM, MCP-1 as well as ET-1 gene expression. The inhibition of ICAM-1 and VCAM-1 production by Chlorella 11-peptide was reversed in the presence of protein kinase A inhibitor (H89) which suggests that the cAMP pathway was involved in the inhibitory cause of the peptide. In addition, this peptide was shown to reduce the extent of increased intercellular permeability induced by combination of 50% of lipopolysaccharide (LPS)-activated RAW 264.7 cells medium and 50% normal SEVC cell culture medium (referred to as 50% RAW-conditioned medium). These data demonstrate that Chlorella 11-peptide is a promising biomolecule in preventing chronic inflammatory-related vascular diseases.

Polyurethane-based drug delivery systems.

Polyurethanes (PUs) are formed by a reaction between isocyanates and diols to yield polymers with urethane bonds (-NH-COO-) in their main chain. A great variety of building blocks is commercially available that allows the chemical and physical properties of PUs to be tailored to their target applications, particularly for the biomedical and pharmaceutical fields. This article reviews the synthesis and characterization of PUs and PU-copolymers, as well as their in vitro and in vivo biodegradability and biocompatibility. Particular emphasis is placed on the use of PUs for the controlled release of drugs and for the (targeted) delivery of biotherapeutics.

Protective effects of Chlorella-derived peptide against UVC-induced cytotoxicity through inhibition of caspase-3 activity and reduction of the expression of phosphorylated FADD and cleaved PARP-1 in skin fibroblasts.

UVC irradiation induces oxidative stress and leads to cell death through an apoptotic pathway. This apoptosis is caused by activation of caspase-3 and formation of poly(ADP-ribose) polymerase-1 (PARP-1). In this study, the underlying mechanisms of Chlorella derived peptide (CDP) activity against UVC-induced cytotoxicity were investigated. Human skin fibroblasts were treated with CDP, vitamin C, or vitamin E after UVC irradiation for a total energy of 15 J/cm². After the UVC exposure, cell proliferation and caspase-3 activity were measured at 12, 24, 48, and 72 h later. Expression of phosphorylated FADD and cleaved PARP-1 were measured 16 h later. DNA damage (expressed as pyrimidine (6-4) pyrimidone photoproducts DNA concentration) and fragmentation assay were performed 24 h after the UVC exposure. Results showed that UVC irradiation induced cytotoxicity in all groups except those treated with CDP. The caspase-3 activity in CDP-treated cells was inhibited from 12 h onward. Expression of phosphorylated FADD and cleaved PARP-1 were also reduced in CDP-treated cells. Moreover, UVC-induced DNA damage and fragmentation were also prevented by the CDP treatment. This study shows that treatment of CDP provides protective effects against UVC-induced cytotoxicity through the inhibition of caspase-3 activity and the reduction of phosphorylated FADD and cleaved PARP-1 expression.

Non-viral delivery of RNA interference targeting cancer cells in cancer gene therapy.

RNA interference (RNAi) is a collection of small RNA-directed mechanisms that result in sequence-specific inhibition of gene expression. RNAi delivery has demonstrated promising efficacy in the treatment of genetic disorders in cancer. Although viral vectors are currently the most efficient systems for gene therapy, potent immunogenicity, mutagenesis, and the biohazards of viral vectors remain their major risks. Various non-viral delivery vectors have been developed to provide a safer approach for gene delivery, including polymers, peptides, liposomes, and nanoparticles. However, some concerns and challenges of these non-viral gene delivery approaches remain to be overcome. In this review, we summarize the recent progress in the development of non-viral systems delivering RNAi and the currently available preclinical and clinical data, and discuss the challenges and future directions in cancer therapy.

A one-step process in preparation of cationic nanoparticles with poly(lactide-co-glycolide)-containing polyethylenimine gives efficient gene delivery.

A one-step preparation of nanoparticles with poly(lactide-co-glycolide) (PLGA) pre-modified with polyethylenimine (PEI) is better in requirements for DNA delivery compared to those prepared in a two-step process (preformed PLGA nanoparticles and subsequently coated with PEI). The particles were prepared by emulsification of PLGA/ethyl acetate in an aqueous solution of PVA and PEI. DLS, AFM and SEM were used for the size characteristics. The cytotoxicity of PLGA/PEI nanoparticles was detected by MTT assay. The transfection activity of the particles was measured using pEGFP and pß-gal plasmid DNA. Results showed that the PLGA/PEI nanoparticles were spherical and non-porous with a size of about 0.2 µm and a small size distribution. These particles had a positive zeta potential demonstrating that PEI was attached. Interestingly, the zeta potential of the particles (from one-step procedure) was substantially higher than that of two-step process and is ascribed to the conjugation of PEI to PLGA via aminolysis. The PLGA/PEI nanoparticles were able to bind DNA and the formed complexes had a substantially lower cytotoxicity and a higher transfection activity than PEI polyplexes. In conclusion, given their small size, stability, low cytotoxicity and good transfection activity, PLGA/PEI-DNA complexes are attractive gene delivery systems.

Preventive effects of ß-thujaplicin against UVB-induced MMP-1 and MMP-3 mRNA expressions in skin fibroblasts.

Solar UV radiation damages human skin by affecting skin tone and resiliency and leads to premature aging (photoaging). The skin damage is caused by the activation of the AP-1 transcription factor, which increases matrix metalloproteinase (MMP) expression and collagen degradation. An increase of interleukin (IL)-6 is also correlated with the activation of MMP-1 expression. ß-thujaplicin has shown both acaricidal and antimicrobial activities. Also, ß-thujaplicin has been shown to be protective against apoptosis due to the oxidative effects of UV irradiation. However, the effect of ß-thujaplicin on UVB-induced MMPs had not been investigated. In this study, after UVB exposure, MMP-1 and IL-6 production in human skin fibroblasts was examined in the presence of ß-thujaplicin, vitamin C, and vitamin E. The expression of MMP-1, MMP-3, tissue inhibitor of metalloproteinase (TIMP-1, TIMP-3) and procollagen mRNA was also investigated. Results showed that UVB-induced MMP-1 production was suppressed by the ß-thujaplicin treatment in a dose-dependent manner, but not by vitamin C and vitamin E. ß-thujaplicin also prevented the up-regulation of MMP-1 and MMP-3 mRNA. Moreover, the UVB-suppressed procollagen gene expression was restored to normal by ß-thujaplicin. Neither UVB nor ß-thujaplicin affected the mRNA expression of TIMP-1 and TIMP-3. The IL-6 production induced by UVB was lower in ß-thujaplicin treated fibroblasts than in the controls. In conclusion, this study shows the capability of ß-thujaplicin in preventing MMP-1 production due to UVB irradiation via inhibition of MMP gene expression. Importantly, the UVB-suppressed procollagen gene expression can be restored to normal by ß-thujaplicin. These findings indicate that ß-thujaplicin is a promising and potent agent to inhibit UVB-induced MMP-1 and MMP-3 gene expression in skin fibroblasts.

Cationic polyurethanes-short branch PEI-mediated delivery of Mir145 inhibited epithelial-mesenchymal transdifferentiation and cancer stem-like properties and in lung adenocarcinoma.

The high invasiveness and frequent recurrence of lung adenocarcinoma (LAC) are major reasons for treatment failures and poor prognoses. Alterations in microRNAs (miRNAs) expression have been shown in lung cancers. Recent reports have demonstrated that tumors contain a small subpopulation of cancer stem cells (CSCs) that possesses self-renewing capacity and is responsible for tumor malignancy including metastasis, relapse, and chemoradioresistance. However, a miRNAs-based therapeutic approach in LAC-associated CSCs (LAC-CSCs) is still blurred. Using miRNA/mRNA-microarray and Quantitative RT-PCR, we found that the expression of miR145 is negatively correlated with the levels of Oct4/Sox2/Fascin1 in LAC patient specimens, and an Oct4(high)Sox2(high)Fascin1(high)miR145(low) phenotype predicted poor prognosis. We enriched LAC-CSCs by side population sorting or identification of CD133 markers and found that LAC-CSCs exhibited low miR145 and high Oct4/Sox2/Fascin1 expression, CSC-like properties, and chemoradioresistance. To clarify the role of miR145, we used a polyurethane-short branch-polyethylenimine (PU-PEI) as the vehicle to deliver miR145 into LAC-CSCs. PU-PEI-mediated miR145 delivery reduced CSC-like properties, and improved chemoradioresistance in LAC-CSCs by directly targeting Oct4/Sox2/Fascin1. Importantly, the repressive effect of miR145 on tumor metastasis was mediated by inhibiting the epithelial-mesenchymal transdifferentiation (EMT) and metastastic ability, partially by regulating Oct4/Sox2/Fascin1, Tcf4, and Wnt5a. Finally, in vivo study showed that PU-PEI-mediated miR145 delivery to xenograft tumors reduced tumor growth and metastasis, sensitized tumors to chemoradiotherapies, and prolonged the survival times of tumor-bearing mice. Our results demonstrated that miR145 acts as a switch regulating lung CSC-like and EMT properties, and provide insights into the clinical prospect of miR145-based therapies for malignant lung cancers.

Inhibition of cancer stem cell-like properties and reduced chemoradioresistance of glioblastoma using microRNA145 with cationic polyurethane-short branch PEI.

Glioblastomas (GBMs) are the most common primary brain tumors with poor prognosis. CD133 has been considered a putative marker of cancer stem cells (CSCs) in malignant cancers, including GBMs. MicroRNAs (miRNAs), highly conserved small RNA molecules, may target oncogenes and have potential as a therapeutic strategy against cancer. However, the role of miRNAs in GBM-associated CSCs remains mostly unclear. In this study, our miRNA/mRNA-microarray and RT-PCR analysis showed that the expression of miR145 (a tumor-suppressive miRNA) is inversely correlated with the levels of Oct4 and Sox2 in GBM-CD133(+) cells and malignant glioma specimens. We demonstrated that miR145 negatively regulates GBM tumorigenesis by targeting Oct4 and Sox2 in GBM-CD133(+). Using polyurethane-short branch polyethylenimine (PU-PEI) as a therapeutic-delivery vehicle, PU-PEI-mediated miR145 delivery to GBM-CD133(+) significantly inhibited their tumorigenic and CSC-like abilities and facilitated their differentiation into CD133(-)-non-CSCs. Furthermore, PU-PEI-miR145-treated GBM-CD133(+) effectively suppressed the expression of drug-resistance and anti-apoptotic genes and increased the sensitivity of the cells to radiation and temozolomide. Finally, the in vivo delivery of PU-PEI-miR145 alone significantly suppressed tumorigenesis with stemness, and synergistically improved the survival rate when used in combination with radiotherapy and temozolomide in orthotopic GBM-CD133(+)-transplanted immunocompromised mice. Therefore, PU-PEI-miR145 is a novel therapeutic approach for malignant brain tumors.