Effect of Ishige okamurae Extract on Osteoclastogenesis In Vitro and In Vivo.

We demonstrated the effect of Ishige okamurae extract (IOE) on the receptor activator of nuclear factor-κB ligand (RANKL)-promoted osteoclastogenesis in RAW 264.7 cells and confirmed that IOE inhibited RANKL-induced tartrate-resistant acid phosphatase (TRAP) activity and osteoclast differentiation. IOE inhibited protein expression of TRAP, metallopeptidase-9 (MMP-9), the calcitonin receptor (CTR), and cathepsin K (CTK). IOE treatment suppressed the expression of activated T cell cytoplasmic 1 and activator protein-1, thus controlling the expression of osteoclast-related factors. Moreover, IOE significantly reduced RANKL-phosphorylated extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK). It also reduced the RANKL-induced phosphorylation of NF-κB and nuclear translocation of p65. IOE inhibited Dex-induced bone loss and osteoclast-related gene expression in zebrafish larvae. HPLC analysis shows that IOE consists of 3.13% and 3.42% DPHC and IPA, respectively. Our results show that IOE has inhibitory effects on osteoclastogenesis in vitro and in vivo and is a potential therapeutic for osteoporosis.

Inhibition of Polyphosphate Degradation in Synechocystis sp. PCC6803 through Inactivation of the phoU Gene.

Phosphorus is an essential but non-renewable nutrient resource critical for agriculture. Luxury phosphorus uptake allows microalgae to synthesize polyphosphate and accumulate phosphorus, but, depending on the strain of algae, polyphosphate may be degraded within 4 hours of accumulation. We studied the recovery of phosphorus from wastewater through luxury uptake by an engineered strain of Synechocystis sp. with inhibited polyphosphate degradation and the effect of this engineered Synechocystis biomass on lettuce growth. First, a strain (ΔphoU) lacking the phoU gene, which encodes a negative regulator of environmental phosphate concentrations, was generated to inhibit polyphosphate degradation in cells. Polyphosphate concentrations in the phoU knock-out strain were maintained for 24 h and then decreased slowly. In contrast, polyphosphate concentrations in the wild-type strain increased up to 4 h and then decreased rapidly. In addition, polyphosphate concentration in the phoU knockout strain cultured in semi-permeable membrane bioreactors with artificial wastewater medium was 2.5 times higher than that in the wild type and decreased to only 16% after 48 h. The biomass of lettuce treated with the phoU knockout strain (0.157 mg P/m2) was 38% higher than that of the lettuce treated with the control group. These results indicate that treating lettuce with this microalgal biomass can be beneficial to crop growth. These results suggest that the use of polyphosphate-accumulating microalgae as biofertilizers may alleviate the effects of a diminishing phosphorous supply. These findings can be used as a basis for additional genetic engineering to increase intracellular polyphosphate levels.

Interwoven MOF-Coated Janus Cells as a Novel Carrier of Toxic Proteins.

Two major issues in cell-mediated drug delivery systems (c-DDS) are the availability of free cell surfaces for the binding of the cells to the target or to their microenvironment and internalization of the cytotoxic drug. In this study, the Janus structure, MOF nanoparticles, and tannic acid (TA) are utilized to address these issues. Janus carrier cells coated with metal-organic frameworks (MOFs) are produced by asymmetrically immobilizing the nanoparticles of a MOF based on zinc with cytotoxic enzymes that are internally encapsulated on the surface of carrier cells. By maintaining the biological and structural features of regular living cells, the MOF-coated Janus cells developed in the present study preserve the intrinsic binding capacity of the cells to their microenvironment. Interconnected MOFs loaded onto the other face of the Janus cells cannot penetrate the cell. Therefore, the carrier cells are protected from the cytotoxic drug contained in MOFs. These MOF-Janus carrier cells are demonstrated to successfully eliminate three-dimensional (3D) tumor spheroids when a chemotherapeutic protein of proteinase K is released from the MOF nanoparticles in an acid environment. The ease with which the MOF-Janus carrier cells are prepared (in 15 min), and the ability to carry a variety of enzymes and even multiple ones should make the developed system attractive as a general platform for drug delivery in various applications, including combination therapy.

Recent advances in process engineering and upcoming applications of metal-organic frameworks.

Progress in metal-organic frameworks (MOFs) has advanced from fundamental chemistry to engineering processes and applications, resulting in new industrial opportunities. The unique features of MOFs, such as their permanent porosity, high surface area, and structural flexibility, continue to draw industrial interest outside the traditional MOF field, both to solve existing challenges and to create new businesses. In this context, diverse research has been directed toward commercializing MOFs, but such studies have been performed according to a variety of individual goals. Therefore, there have been limited opportunities to share the challenges, goals, and findings with most of the MOF field. In this review, we examine the issues and demands for MOF commercialization and investigate recent advances in MOF process engineering and applications. Specifically, we discuss the criteria for MOF commercialization from the views of stability, producibility, regulations, and production cost. This review covers progress in the mass production and formation of MOFs along with future applications that are not currently well known but have high potential for new areas of MOF commercialization.

Protective Effects of Oenothera biennis against Hydrogen Peroxide-Induced Oxidative Stress and Cell Death in Skin Keratinocytes.

BACKGROUND: Oenothera biennis (evening primrose) produces bioactive substances with a diverse range of pharmacological functions. However, it is currently unknown whether extract prepared from the aerial parts of O. biennis (APOB) can protect the skin against oxidative stress. OBJECTIVE: The aim of this study is to investigate the protective effects of APOB against oxidative stress-induced damage in human skin keratinocytes (HaCaT) and elucidate the underlying mechanisms. METHODS: We pretreated HaCaT cells with various concentrations of APOB or the antioxidant N-acetyl-L-cysteine before applying H2O2. We then compared the cell viability, intracellular reactive oxygen species (ROS) production, and DNA and mitochondrial damage between pretreated and untreated control cells using a range of assays, flow cytometry, and Western blot analysis and also examined the reducing power and DPPH free radical scavenging activity of APOB. RESULTS: APOB pretreatment significantly increased cell viability, effectively attenuated H2O2-induced comet tail formation, and inhibited H2O2-induced phosphorylation of the histone γH2AX, as well as the number of apoptotic bodies and Annexin V-positive cells. APOB was found to have high reducing power and DPPH radical scavenging activity and also exhibited scavenging activity against intracellular ROS accumulation and restored the loss of mitochondrial membrane potential caused by H2O2. APOB pretreatment almost totally reversed the enhanced cleavage of caspase-3, the degradation of poly (ADP-ribose)-polymerase (PARP), DNA fragmentation that usually occurs in the presence of H2O2, and increased the levels of heme oxygenase-1 (HO-1), a potent antioxidant enzyme that is associated with the induction of nuclear factor-erythroid 2-related factor 2 (Nrf2). CONCLUSIONS: APOB can protect HaCaT cells from H2O2-induced DNA damage and cell death by blocking cellular damage related to oxidative stress via a mechanism that affects ROS elimination and by activating the Nrf2/HO-1 signaling pathway.

Tailoring Nanocrystalline Metal-Organic Frameworks as Fluorescent Dye Carriers for Bioimaging.

Challenges exist in taking advantage of dye molecules for reliable and reproducible molecular probes in biomedical applications. In this study, we show how to utilize the dye molecules for bioimaging within protective carriers of nanocrystalline metal-organic frameworks (nMOFs) particles. Specifically, Resorufin and Rhodamine-6G having different molecular sizes were encapsulated within close-fitting pores of nMOF-801 and nUiO-67 particles, respectively. The resulting nanocrystalline particles have high crystallinity, uniform size, and morphology and preserve enhanced photoluminescence properties with exceptional stabilities in biomedical environment. The samples are further functionalized with a targeting agent and successfully work for fluorescence imaging of FL83B (human hepatocyte cell) and HepG2 (human hepatocellular carcinoma) without cytotoxicity.

Comparison of commonly used ICR stocks and the characterization of Korl:ICR.

Mouse is a commonly used animal in life science studies and is classified as outbred if genetically diverse and inbred if genetically homogeneous. Outbred mouse stocks, are used in toxicology, oncology, infection and pharmacology research. The National Institute of Food and Drug Safety Evaluation (NIFDS; former the Korea National Institute of Health) have bred ICR mice for more than 50 years. We investigated to provide users with information and promote accountability to the Korl:ICR. To secure the indigenous data, biological characteristics of Korl:ICR were identified by comparing with other ICR stocks. This domestic ICR stock was denominated as 'Korl:ICR'. Phylogenetic analysis using SNPs indicated that the population stratification of the Korl:ICR was allocated different area with other ICR. In addition, we measured litter size, body weight, body length, various organ weight, hematology and clinical blood chemistry of the Korl:ICR compared to other ICR. Otherwise, there are no significant differences among the biological phenotypes of Korl:ICR and other ICR. These results suggest that as a genetically indigenous source colony, the Korl:ICR is seperated (or independent) stock with other ICR. Also, we confirmed that there is no difference among the Korl:ICR and other ICR on biological phenotypes. Therefore, the Korl:ICR source colony might be a new stock in distinction from other ICR, it is a good milestone in securing ownership of the national laboratory animal resource. The NIFDS expects that the Korl:ICR mice will be useful animal resource for our domestic researchers.

Oral administration of red ginseng powder fermented with probiotic alleviates the severity of dextran-sulfate sodium-induced colitis in a mouse model.

Red ginseng is a well-known alternative medicine with anti-inflammatory activity. It exerts pharmacological effects through the transformation of saponin into metabolites by intestinal microbiota. Given that intestinal microflora vary among individuals, the pharmacological effects of red ginseng likely vary among individuals. In order to produce homogeneously effective red ginseng, we prepared probiotic-fermented red ginseng and evaluated its activity using a dextran sulfate sodium (DSS)-induced colitis model in mice. Initial analysis of intestinal damage indicated that the administration of probiotic-fermented red ginseng significantly decreased the severity of colitis, compared with the control and the activity was higher than that induced by oral administration of ginseng powder or probiotics only. Subsequent analysis of the levels of serum IL-6 and TNF-α, inflammatory biomarkers that are increased at the initiation stage of colitis, were significantly decreased in probiotic-fermented red ginseng-treated groups in comparison to the control group. The levels of inflammatory cytokines and mRNAs for inflammatory factors in colorectal tissues were also significantly decreased in probiotic-fermented red ginseng-treated groups. Collectively, oral administration of probiotic-fermented red ginseng reduced the severity of colitis in a mouse model, suggesting that it can be used as a uniformly effective red ginseng product.

Fucoidan inhibits LPS-induced inflammation in vitro and during the acute response in vivo.

Studies have been focused on natural products with antibacterial and anti-inflammatory activities, such as fucoidan. Many in vivo studies have evaluated the effect of fucoidan on tumor growth, diabetes, obesity, ischemia reperfusion, and oxidative stress. However, the effects of fucoidan on bacteria-induced gingival inflammation and periodontitis have not been reported. We previously characterized the anti-inflammatory effect of fucoidan in vitro. Here, we confirmed the anti-inflammatory activity of fucoidan in a macrophage cell line in terms of its inhibition of the expression of inflammatory mediators and pro-inflammatory cytokines. Additionally, we confirmed the ability of fucoidan to inhibit gingival inflammation, expression of pro-inflammatory cytokines, and neutrophil recruitment in the gingival tissue of mice injected with LPS prepared from P. gingivalis. Interestingly, however, fucoidan did not inhibit the expression of pro-inflammatory cytokines in a P. gingivalis-infected mouse model of periodontitis. Additionally, fucoidan treatment did not lead to clearance of P. gingivalis or improvement of P. gingivalis infection-mediated bone loss in the periodontitis model. We conclude that fucoidan exerts anti-inflammatory effects in vitro and in vivo, together with a limited antibacterial effect in vivo.

In vivo photothermal treatment by the peritumoral injection of macrophages loaded with gold nanoshells.

Photothermal treatment methods have been widely studied for their target specificity and potential for supplementing the limitations of conventional surgical treatments. In this study, we conducted in vivo photothermal treatments using macrophages containing nanoshells as live vectors. We injected macrophages at the peritumoral sites and observed that they had penetrated into the tumor approximately 48 hours after injection. Afterwards, we irradiated with a near-infrared laser for 2 minutes at 1 W/cm(2), causing cancer cell death. Our study identified the optimal conditions of the photothermal treatment and confirmed the feasibility of its use in in vivo treatments.

In-situ observation for growth of hierarchical metal-organic frameworks and their self-sequestering mechanism for gas storage.

Although structures with the single functional constructions and micropores were demonstrated to capture many different molecules such as carbon dioxide, methane, and hydrogen with high capacities at low temperatures, their feeble interactions still limit practical applications at room temperature. Herein, we report in-situ growth observation of hierarchical pores in pomegranate metal-organic frameworks (pmg-MOFs) and their self-sequestering storage mechanism, not observed for pristine MOFs. Direct observation of hierarchical pores inside the pmg-MOF was evident by in-situ growth X-ray measurements while self-sequestering storage mechanism was revealed by in-situ gas sorption X-ray analysis and molecular dynamics simulations. The results show that meso/macropores are created at the early stage of crystal growth and then enclosed by micropore crystalline shells, where hierarchical pores are networking under self-sequestering mechanism to give enhanced gas storage. This pmg-MOF gives higher CO2 (39%) and CH4 (14%) storage capacity than pristine MOF at room temperature, in addition to fast kinetics with robust capacity retention during gas sorption cycles, thus giving the clue to control dynamic behaviors of gas adsorption.

Fermented red ginseng extract inhibits cancer cell proliferation and viability.

Red ginseng (Panax ginseng C.A. Meyer) is the most widely recognized medicinal herb due to its remedial effects in various disorders, such as cancers, diabetes, and heart problems. In this study, we investigated the anticancer effect of fermented red ginseng extract (f-RGE; provided by Jeonju Biomaterials Institute, Jeonju, South Korea) in a parallel comparison with the effect of nonfermented red ginseng extract (nf-RGE; control) on several cancer cell lines--MCF-7 breast cancer cells, HepG2 hepatocellular carcinoma cells, and reprogrammed MCF-7 cells (mimicking cancer stem cells). Cells were cultured at various concentrations of RGE (from 0.5 up to 5 mg/mL) and their viabilities and proliferative properties were examined. Our data demonstrate the following: (1) nf-RGE inhibited cell viability at ≥1 mg/mL for MCF-7 cells and ≥2 mg/mL for HepG2 cells, (2) in the presence of a carcinogenic agent, 12-O-tetradecanoylphorbol-13-acetate (TPA), nf-RGE treatment in combination with paclitaxel synergistically decreased MCF-7 as well as HepG2 cell viability, (3) f-RGE (which contained a greater level of Rg3 content) more effectively decreased the viability of MCF-7 and HepG2 cells compared to nf-RGE, and (4) f-RGE appeared more potent for inhibiting cancerous differentiation of reprogrammed MCF-7 cells in a synergistic fashion with paclitaxel, especially in the presence of TPA, compared to nf-RGE. These findings suggest that f-RGE treatment may be more effective for decreasing cancer cell survival by inducing apoptotic cell death and also presumably for preventing cancer stem cell differentiation compared to nf-RGE.

Real-time phase-contrast imaging of photothermal treatment of head and neck squamous cell carcinoma: an in vitro study of macrophages as a vector for the delivery of gold nanoshells.

Photothermal treatment (PTT) using nanoparticles has gained attention as a promising alternative therapy for malignant tumors. One strategy for increasing the selectivity of PTT is the use of macrophages as a cellular vector for delivering nanoparticles. The aim of the present study is to examine the use of macrophages as a cellular vector for efficient PTT and determine the appropriate irradiation power and time of a near-infrared (NIR) laser using real-time phase-contrast imaging. Thermally induced injury and death of cancer cells were found to begin at 44°C to 45°C, which was achieved using the PTT effect with gold nanoshells (NS) and irradiation with a NIR laser at a power of 2 W for 5 min. The peritoneal macrophage efficiently functioned as a cellular vector for the NS, and the cancer cells surrounding the NS-loaded macrophages selectively lost their cellular viability after being irradiated with the NIR laser.

Heterogeneous nuclear ribonucleoprotein C1/C2 controls the metastatic potential of glioblastoma by regulating PDCD4.

MicroRNAs (miRNAs) have been implicated in the pathogenesis and progression of brain tumors. miR-21 is one of the most highly overexpressed miRNAs in glioblastoma multiforme (GBM), and its level of expression correlates with the tumor grade. Programmed cell death 4 (PDCD4) is a well-known miR-21 target and is frequently downregulated in glioblastomas in accordance with increased miR-21 expression. Downregulation of miR-21 or overexpression of PDCD4 can inhibit metastasis. Here, we investigate the role of heterogeneous nuclear ribonucleoprotein C1/C2 (hnRNPC) in the metastatic potential of the glioblastoma cell line T98G. hnRNPC bound directly to primary miR-21 (pri-miR-21) and promoted miR-21 expression in T98G cells. Silencing of hnRNPC lowered miR-21 levels, in turn increasing the expression of PDCD4, suppressing Akt and p70S6K activation, and inhibiting migratory and invasive activities. Silencing of hnRNPC reduced cell proliferation and enhanced etoposide-induced apoptosis. In support of a role for hnRNPC in the invasiveness of GBM, highly aggressive U87MG cells showed higher hnRNPC expression levels and hnRNPC abundance in tissue arrays and also showed elevated levels as a function of brain tumor grade. Taken together, our data indicate that hnRNPC controls the aggressiveness of GBM cells through the regulation of PDCD4, underscoring the potential usefulness of hnRNPC as a prognostic and therapeutic marker of GBM.

Cosmetic efficacy evaluation of an anti-acne cream using the 3D image analysis system.

BACKGROUND/PURPOSE: Acne is a skin disease which accompanies pathological and morphological changes. Although acne severity is scored by clinicians based on pathological status, aesthetic aspect of acne symptom is also concerned by patients. This study was conducted to examine the usefulness of a 3D image analysis method for the cosmetic efficacy evaluation of an anti-acne cream. METHODS: Twenty-one healthy volunteers with acne lesions were recruited for the study and treated with an 'anti-acne cream' for 4 weeks. Acne symptoms on the facial skin were graded by the visual evaluation of photographs taken before and after the treatment. Skin color of acne lesions was and measured by a spectrophotometer. In addition, a 3D image analysis system was used to quantify skin surface roughness and acne volumes. RESULTS: Both visual and spectrophotometric assessments of acne lesions provided similar results indicating that the cream treatment improved acne symptoms significantly. The 3D image analysis of acne lesions confirmed that the cream treatment decreased skin surface roughness and acne volumes. CONCLUSION: The current study demonstrated that the 3D image-based analysis of the skin may be useful for the quantification of acne symptoms of cosmetic relevance.

Identification and characterization of the acid phosphatase HppA in Helicobacter pylori.

An acid phosphatase (HppA) activated by NH4Cl was purified 192- and 34-fold from the periplasmic and membrane fractions of Helicobacter pylori, respectively. SDS-polyacrylamide gel electrophoresis revealed that HppA from the latter appears to be several kilodaltons larger in molecular mass than from the former by about 24 kDa. Under acidic conditions (pH< or =4.5), the enzyme activity was entirely dependent on the presence of certain mono- and/or divalent metal cations (e.g., K+, NH4 +, and/or Ni2+). In particular, Ni2+ appeared to lower the enzyme's Km for the substrates, without changing Vmax. The purified enzyme showed differential specificity against nucleotide substrates with pH; for example, the enzyme hydrolyzed adenosine nucleotides more rapidly at pH 5.5 than at pH 6.0, and vice versa for CTP or TTP. Analyses of the enzyme's N-terminal sequence and of an HppA- H. pylori mutant revealed that the purified enzyme is identical to rHppA, a cloned H. pylori class C acid phosphatase, and shown to be the sole bacterial 5'-nucleotidase uniquely activated by NH4Cl. In contrast to wild type, HppA- H. pylori cells grew more slowly. Strikingly, they imported Mg2+ at a markedly lowered rate, but assimilated urea rapidly, with a subsequent increase in extracellular pH. Moreover, mutant cells were much more sensitive to extracellular potassium ions, as well as to metronidazole, omeprazole, or thiophenol, with considerably lowered MIC values, than wild-type cells. From these data, we suggest that the role of the acid phosphatase HppA in H. pylori may extend beyond 5'-nucleotidase function to include cation-flux as well as pH regulation on the cell envelope.

Effect of ascorbic acid on bone marrow-derived mesenchymal stem cell proliferation and differentiation.

Mesenchymal stem cells (MSCs) derived from bone marrow are an important tool in tissue engineering and cell-based therapies because of their multipotent capacity. Majority of studies on MSCs have investigated the roles of growth factors, cytokines, and hormones. Antioxidants such as ascorbic acid can be used to expand MSCs while preserving their differentiation ability. Moreover, ascorbic acid can also stimulate MSC proliferation without reciprocal loss of phenotype and differentiation potency. In this study, we evaluated the effects of ascorbic acid on the proliferation, differentiation, extracellular matrix (ECM) secretion of MSCs. The MSCs were cultured in media containing various concentrations (0-500 microM) of L-ascorbate-2-phosphate (Asc-2-P) for 2 weeks, following which they were differentiated into adipocytes and osteoblasts. Ascorbic acid stimulated ECM secretion (collagen and glycosaminoglycan) and cell proliferation. Moreover, the phenotypes of the experimental groups as well as the differentiation potential of MSCs remained unchanged. The apparent absence of decreased cell density or morphologic change is consistent with the toxicity observed with 5-250 microM concentrations of Asc-2-P. The results demonstrate that MSC proliferation or differentiation depends on ascorbic acid concentration.

Spectrophotometric determination of peptide transport with chromogenic peptide mimetics.

A spectrophotometric assay to determine peptide transport has been developed. Using two chromogenic peptide mimetics, L-phenylalanyl-L-2-sulfanilylglycine (PSG) and L-phenylalanyl-L-3-thiaphenylalanine (PSP), the peptide transport patterns in individual cell species can be evaluated effectively. After the addition of PSG to a HeLa cell suspension, sulfanilic acid accumulated progressively inside, but not outside, the cells, demonstrating that PSG was transported wholly intact. The addition of PSP to the same cell suspension was followed immediately by extracellular thiophenol production. Measurement of the rate of thiophenol release thereby provided direct determination of PSP transport. The thiophenol release was consistent with Michaelis-Menten kinetics, with a K(m) of 0.016 mM and a V(max) of 5.07 nmol/min (1 x 10(6) cells/ml, pH 7.4, 37 degrees C). The resulting kinetic constants estimated were in agreement with values determined by single-substrate enzyme kinetics. Using PSP, transport kinetics of various dipeptides was examined by competitive spectrophotometry. As a result, dipeptides tested could be ranked in order of kinetic power for their transport.