Molecularly imprinted composite cryogels for hemoglobin depletion from human blood

A molecularly imprinted composite cryogel (MICC) was prepared for depletion of hemoglobin from human blood prior to use in proteome applications. Poly(hydroxyethyl methacrylate) based MICC was prepared with high gel fraction yields up to 90%, and characterized by Fourier transform infrared spectrophotometer, scanning electron microscopy, swelling studies, flow dynamics and surface area measurements. MICC exhibited a high binding capacity and selectivity for hemoglobin in the presence of immunoglobulin G, albumin and myoglobin. MICC column was successfully applied in fast protein liquid chromatography system for selective depletion of hemoglobin for human blood. The depletion ratio was highly increased by embedding microspheres into the cryogel (93.2%). Finally, MICC can be reused many times with no apparent decrease in hemoglobin adsorption capacity. Copyright © 2014 John Wiley & Sons, Ltd.     

Identification of anticancer drugs for hepatocellular carcinoma through personalized genome‐scale metabolic modeling

Genome‐scale metabolic models (GEMs) have proven useful as scaffolds for the integration of omics data for understanding the genotype–phenotype relationship in a mechanistic manner. Here, we evaluated the presence/absence of proteins encoded by 15,841 genes in 27 hepatocellular carcinoma (HCC) patients using immunohistochemistry. We used this information to reconstruct personalized GEMs for six HCC patients based on the proteomics data, HMR 2.0, and a task‐driven model reconstruction algorithm (tINIT). The personalized GEMs were employed to identify anticancer drugs using the concept of antimetabolites; i.e., drugs that are structural analogs to metabolites. The toxicity of each antimetabolite was predicted by assessing the in silico functionality of 83 healthy cell type‐specific GEMs, which were also reconstructed with the tINIT algorithm. We predicted 101 antimetabolites that could be effective in preventing tumor growth in all HCC patients, and 46 antimetabolites which were specific to individual patients. Twenty‐two of the 101 predicted antimetabolites have already been used in different cancer treatment strategies, while the remaining antimetabolites represent new potential drugs. Finally, one of the identified targets was validated experimentally, and it was confirmed to attenuate growth of the HepG2 cell line.     

Chiral separation-based ligand exchange by open-tubular capillary electrochromatography

Chiral ligand-exchange enantioseparation of aliphatic and aromatic amino acids was successfully performed using a new open-tubular zwitterionic column with tentacle-type polymer stationary phase. The polymeric stationary phase was prepared using 3-chloro-2-hydroxypropyl methacrylate (HPMA-Cl), a new reactive monomer. The preparation procedure of the open-tubular column included silanization, in situ graft polymerization with HPMA-Cl, and l-histidine (l-His) modification. l-His was used as a chiral ligand-exchange selector and copper(II) as a central ion. Successful enantioseparation of six pairs of amino acid enantiomers was achieved with a buffer of 5 mM CuSO₄, 20 mM (NH₄)₂SO₄ at pH 3.0.     

A Review on Saponin Biosynthesis and its Transcriptomic Resources in Medicinal Plants

Plant Molecular Biology Reporter - Saponins are naturally occurring glycosides which are produced by various plant species with diverse biological properties. The surface-active properties of...     

Gene Expression Profiling and Secretome Analysis Differentiate Adult-Derived Human Liver Stem/Progenitor Cells and Human Hepatic Stellate Cells

Adult-derived human liver stem/progenitor cells (ADHLSC) are obtained after primary culture of the liver parenchymal fraction. The cells are of fibroblastic morphology and exhibit a hepato-mesenchymal phenotype. Hepatic stellate cells (HSC) derived from the liver non-parenchymal fraction, present a comparable morphology as ADHLSC. Because both ADHLSC and HSC are described as liver stem/progenitor cells, we strived to extensively compare both cell populations at different levels and to propose tools demonstrating their singularity.ADHLSC and HSC were isolated from the liver of four different donors, expanded in vitro and followed from passage 5 until passage 11. Cell characterization was performed using immunocytochemistry, western blotting, flow cytometry, and gene microarray analyses. The secretion profile of the cells was evaluated using Elisa and multiplex Luminex assays.Both cell types expressed α-smooth muscle actin, vimentin, fibronectin, CD73 and CD90 in accordance with their mesenchymal origin. Microarray analysis revealed significant differences in gene expression profiles. HSC present high expression levels of neuronal markers as well as cytokeratins. Such differences were confirmed using immunocytochemistry and western blotting assays. Furthermore, both cell types displayed distinct secretion profiles as ADHLSC highly secreted cytokines of therapeutic and immuno-modulatory importance, like HGF, interferon-γ and IL-10.Our study demonstrates that ADHLSC and HSC are distinct liver fibroblastic cell populations exhibiting significant different expression and secretion profiles.     

How visual stimulus effects the time perception? The evidence from time perception of emotional videos

Time perception is defined as a subjective judgment on the elapsed time of an event. It can change according to both external and internal factors. There are two main paradigms of time perception; retrospective time perception (RTP) and prospective time perception (PTP). Two paradigms differ from each other according to whether the subject has knowledge on the importance of passage of time in the given task. Since RTP paradigm studies are harder to conduct, studies on RTP paradigm is far fewer than studies on PTP. Thus in the current study, both RTP and PTP paradigms are investigated. Also, time perception is discussed in relation to internal clock model and cognitive load. Emotional motion videos are used to create cognitive load and manipulate internal clock. Results showed the effect of emotion on time perception. Another major finding is that shorter videos are perceived longer whereas longer videos are perceived shorter as in accordance with Vierordt’s Law. However, there was no difference between RTP and PTP paradigms. These results indicate that emotional videos change our internal clock while a number of changes in a motion video create cognitive load causing disturbance of time perception.     

Cell‐free production of a therapeutic protein: Expression,purification, and characterization of recombinant streptokinase using a CHO lysate

The use of cell‐free systems to produce recombinant proteins has grown rapidly over the past decade. In particular, cell‐free protein synthesis (CFPS) systems based on mammalian cells provide alternative methods for the production of many proteins, including those that contain disulfide bonds, glycosylation, and complex structures such as monoclonal antibodies. In the present study, we show robust production of turbo green fluorescent protein (tGFP) and streptokinase in a cell‐free system using instrumented mini‐bioreactors for highly reproducible protein production. We achieved recombinant protein production (～600 μg/ml of tGFP and 500 μg/ml streptokinase) in 2.5 hr of expression time, comparable to previously reported yields for cell‐free protein expression. Also, we demonstrate the use of two different affinity tags for product capture and compare those to a tag‐free self‐cleaving intein capture technology. The intein purification method provided a product recovery of 86%, compared with 52% for conventionally tagged proteins, while resulting in a 30% increase in total units of activity of purified recombinant streptokinase compared with conventionally tagged proteins. These promising beneficial features combined with the intein technology makes feasible the development of dose‐level production of therapeutic proteins at the point‐of‐care.