Purification of Therapeutic Antibodies Using the Ca2+-Dependent Phase-Transition Properties of Calsequestrin.

Affinity chromatography utilizing specific interactions between therapeutic proteins and bead-immobilized capturing agents is a standard method for protein purification, but its scalability is limited by long purification times, activity loss by the capturing molecules and/or purified protein, and high costs. Here, we report a platform for purifying therapeutic antibodies via affinity precipitation using the endogenous calcium ion-binding protein, calsequestrin (CSQ), which undergoes a calcium ion-dependent phase transition. In this method, ZZ-CSQ fusion proteins with CSQ and an affinity protein (Z domain of protein A) capture antibodies and undergo multimerization and subsequent aggregation in response to calcium ions, enabling the antibody to be collected by affinity precipitation. After robustly validating and optimizing the performance of the platform, the ZZ-CSQ platform can rapidly purify therapeutic antibodies from industrial harvest feedstock with high purity (>97%) and recovery yield (95% ± 3%). In addition, the ZZ-CSQ platform outperforms protein A-based affinity chromatography (PAC) in removing impurities, yielding ∼20-fold less DNA and ∼4.8-fold less host cell protein (HCP) contamination. Taken together, this platform is rapid, recyclable, scalable, and cost-effective, and it shows antibody-purification performance superior or comparable to that of the standard affinity chromatography method.

Fucosterol inhibits adipogenesis through the activation of AMPK and Wnt/ß-catenin signaling pathways.

Fucosterol is a sterol constituent primarily derived from brown algae. Recently, the antiadipogenic effect of fucosterol has been reported; however, its molecular mechanism remains to be studied. Fucosterol effectively upregulated the phosphorylations of both adenosine monophosphate (AMP)-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC), and downregulated the expression levels of lipogenesis-related factors. Moreover, fucosterol activated the major components of the Wnt/ß-catenin signaling pathway, including ß-catenin, disheveled 2 (DVL2), and cyclin D1 (CCND1), whereas it inactivated glycogen synthase kinase 3ß (p-GSK3ß) by stimulating its phosphorylation. In the presence or absence of fucosterol, the adipogenic transcriptional factors [peroxisome proliferator activated-receptor γ (PPARγ), CCAAT/enhancer binding protein α (C/EBPα), and sterol regulatory element binding protein-1c (SREBP-1c)] were upregulated by the inhibition of AMPK by compound C or the knockdown of ß-catenin by siRNA. Overall, these data demonstrate that fucosterol prevents adipogenesis by mediating both AMPK- and Wnt/ß-catenin-signaling pathways.

5,7-Dimethoxyflavone Attenuates Obesity by Inhibiting Adipogenesis in 3T3-L1 Adipocytes and High-Fat Diet-Induced Obese C57BL/6J Mice.

The antiobesity effect of 5,7-dimethoxyflavone (DMF) was evaluated in 3T3-L1 adipocytes and high-fat diet (HFD)-induced obese C57BL/6J mice. The accumulation of lipid droplets and triglycerides in adipocytes was dose dependently suppressed by DMF through inhibition of adipogenesis. DMF downregulated the adipogenic transcription factors (peroxisome proliferator-activated receptor [PPAR]γ, CCAAT/enhancer binding protein [C/EBP]α, and sterol regulatory element-binding protein-1c [SREBP-1c]) and lipid synthesis enzymes (fatty acid synthase [FAS], acetyl-CoA carboxylase [ACC], lipoprotein lipase [LPL], and HMG-CoA reductase [HMGR]). AMP-activated protein kinase (AMPK) and AMPK related lipolytic proteins in differentiated adipocytes were activated by DMF. In the animal model, oral administration of DMF (50 mg/kg/day for 6 weeks) significantly decreased body weight gain without affecting food intake. Elevated serum levels of total cholesterol and low-density lipoprotein cholesterol were suppressed by DMF. Fat pad masses were reduced in DMF-treated obese mice, as evidenced by reduced adipocyte size. DMF altered the expression of adipogenic transcription factors in epididymal fat tissue. In addition, DMF attenuated HFD-induced nonalcoholic fatty liver disease by decreasing hepatic triglyceride accumulation. Overall, these results suggest that DMF is a potential natural agent for attenuating obesity and other obesity-related metabolic syndromes.

Leaves of Persimmon (Diospyros kaki Thunb.) Ameliorate N-Methyl-N-nitrosourea (MNU)-Induced Retinal Degeneration in Mice.

The purpose of the study was to investigate the protective effects of the ethanol extract of Diospyros kaki (EEDK) persimmon leaves to study N-methyl-N-nitrosourea (MNU)-induced retinal degeneration in mice. EEDK was orally administered after MNU injection. Retinal layer thicknesses were significantly increased in the EEDK-treated group compared with the MNU-treated group. The outer nuclear layer was preserved in the retinas of EEDK-treated mice. Moreover, EEDK treatment reduced the MNU-dependent up-regulation of glial fibrillary acidic protein (GFAP) and nestin expression in Müller and astrocyte cells. EEDK treatment also inhibited MNU-dependent down-regulation of rhodopsin expression. Quercetin exposure significantly attenuated the negative effects of H2O2 in R28 cells, suggesting that quercetin can act in an antioxidative capacity. Thus, EEDK may be considered as an agent for treating or preventing degenerative retinal diseases, such as retinitis pigmentosa and age-related macular degeneration.

Antihyperglycemic and Anti-Inflammatory Effects of Standardized Curcuma xanthorrhiza Roxb. Extract and Its Active Compound Xanthorrhizol in High-Fat Diet-Induced Obese Mice.

Xanthorrhizol, a natural compound isolated from Curcuma xanthorrhiza Roxb. (Java turmeric), has been reported to possess antioxidant and anticancer properties; however, its effects on metabolic disorders remain unknown. The aim of the present study was to evaluate the effects of xanthorrhizol (XAN) and C. xanthorrhiza extract (CXE) with standardized XAN on hyperglycemia and inflammatory markers in high-fat diet- (HFD-) induced obese mice. Treatment with XAN (10 or 25 mg/kg/day) or CXE (50 or 100 mg/kg/day) significantly decreased fasting and postprandial blood glucose levels in HFD-induced obese mice. XAN and CXE treatments also lowered insulin, glucose, free fatty acid (FFA), and triglyceride (TG) levels in serum. Epididymal fat pad and adipocyte size were decreased by high doses of XAN (26.6% and 20.1%) and CXE (25.8% and 22.5%), respectively. XAN and CXE treatment also suppressed the development of fatty liver by decreasing liver fat accumulation. Moreover, XAN and CXE significantly inhibited production of inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-1ß (IL-1ß), and C-reactive protein (CRP) in adipose tissue (27.8-82.7%), liver (43.9-84.7%), and muscle (65.2-92.5%). Overall, these results suggest that XAN and CXE, with their antihyperglycemic and anti-inflammatory activities, might be used as potent antidiabetic agents for the treatment of type 2 diabetes.

Kirenol stimulates osteoblast differentiation through activation of the BMP and Wnt/ß-catenin signaling pathways in MC3T3-E1 cells.

Kirenol has been reported to possess anti-oxidant, anti-inflammatory, anti-allergic, anti-adipogenic, and anti-arthritic activities; however, its effect on osteoblast differentiation has not yet been reported. The aim of the present study was to evaluate the effect of kirenol on osteoblast differentiation through activation of the bone morphogenetic protein (BMP) and Wnt/ß-catenin signaling pathways in MC3T3-E1 cells. Kirenol markedly promoted alkaline phosphatase (ALP) activity and mineralization. Kirenol not only increased the expression of osteoblast differentiation markers, such as ALP, type I collagen (ColA1), and osteopontin (OPN), but also increased the expression of osteoprotegerin/receptor activator of nuclear factor kappa B ligand (OPG/RANKL) ratio. The effects of kirenol on osteoblast differentiation were accompanied by stimulating the expression of the BMP and Wnt/ß-catenin signaling pathways, including BMP2, runt-related transcription factor 2 (Runx2), osterix (Osx), low density lipoprotein receptor related protein 5 (LRP5), disheveled 2 (DVL2), ß-catenin, cyclin D1 (CCND1), and phosphorylated glycogen synthase kinase 3ß (GSK3ß). In addition, kirenol up-regulated the expression of ß-catenin, CCND1, ALP, and ColA1 which were down-regulated by siRNA knockdown of ß-catenin. Overall, these results demonstrate that kirenol is capable of promoting osteoblast differentiation in MC3T3-E1 cells through activation of the BMP and Wnt/ß-catenin signaling pathways, suggesting that it is a potential candidate target for treating or preventing osteoporosis.

Kirenol inhibits adipogenesis through activation of the Wnt/ß-catenin signaling pathway in 3T3-L1 adipocytes.

Kirenol, a natural diterpenoid compound, has been reported to possess anti-oxidant, anti-inflammatory, anti-allergic, and anti-arthritic activities; however, its anti-adipogenic effect remains to be studied. The present study evaluated the effect of kirenol on anti-adipogenesis through the activation of the Wnt/ß-catenin signaling pathway. Kirenol prevented intracellular lipid accumulation by down-regulating key adipogenesis transcription factors [peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding proteins α (C/EBPα), and sterol regulatory element binding protein-1c (SREBP-1c)] and lipid biosynthesis-related enzymes [fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC)], as well as adipocytokines (adiponectin and leptin). Kirenol effectively activated the Wnt/ß-catenin signaling pathway, in which kirenol up-regulated the expression of low density lipoprotein receptor related protein 6 (LRP6), disheveled 2 (DVL2), ß-catenin, and cyclin D1 (CCND1), while it inactivated glycogen synthase kinase 3ß (GSK3ß) by increasing its phosphorylation. Kirenol down-regulated the expression levels of PPARγ and C/EBPα, which were up-regulated by siRNA knockdown of ß-catenin. Overall, kirenol is capable of inhibiting the differentiation and lipogenesis of 3T3-L1 adipocytes through the activation of the Wnt/ß-catenin signaling pathway, suggesting its potential as natural anti-obesity agent.

Optically bifacial thin-film wire-grid polarizers with nano-patterns of a graded metal-dielectric composite layer.

We report on the concept of a thin film wire-grid polarizer (WGP) with optically dual characteristics by introducing a nano-patterned graded metal-dielectric composite-material layer. The Ti-SiO(2) composite layer with a depth profile of a gradually-varied composition ratio shows an absorptive feature due to the elimination of an optical interface between a metal and a glass substrate, while the metal side of the WGP gives a reflective character. The unprecedented optically-bifacial thin-film WGP with the 144 nm-period straight-line patterns of a 100 nm-thick Ti-SiO(2) composite layer and a 185 nm-thick Al layer shows the exceptionally low reflectance below 15 % from the absorptive side and the high polarization extinction ratio (PER) of over 500 at 550 nm, which is acceptable for use as various display applications such as AMOLEDs and LCDs.

Weak-microcavity organic light-emitting diodes with improved light out-coupling.

We propose and demonstrate weak-microcavity organic light-emitting diode (OLED) displays with improved light-extraction and viewing-angle characteristics. A single pair of low- and high-index layers is inserted between indium tin oxide (ITO) and a glass substrate. The electroluminescent (EL) efficiencies of discrete red, green, and blue weak-microcavity OLEDs are enhanced by 56%, 107%, and 26%, respectively, with improved color purity. Moreover, full-color passive-matrix bottom-emitting OLED displays are fabricated by employing low-index layers of two thicknesses. As a display, the EL efficiency of white color was 27% higher than that of a conventional OLED display.

Far-field radiation of photonic crystal organic light-emitting diode.

Utilizing the near- to far-field transformation based on the 3-D finite difference time domain (FDTD) method and Fourier transformation, the far-field profile of a photonic crystal organic light emitting diode is studied to understand the viewing angle dependence. The measured far-field profiles agree well with those of the simulation. The enhancement of the extraction efficiency in excess of 60% is observed for the optimized photonic crystal pattern.