Potential Applications in Sewage Bioremediation of the Highly Efficient Pyridine-Transforming Paenochrobactrum sp.

Applied Biochemistry and Microbiology - A pyridine-transforming strain P2 was isolated from sewage collected from Guangzhou oil stain field(China).According to the system analysis, it was...     

A doubly auxotrophic CHO-K1 cell line for the production of recombinant monoclonal antibodies

Chinese hamster ovary (CHO) cells are the most widely used mammalian hosts for recombinant protein production due to their hardiness, ease of transfection, and production of glycan structures similar to those in natural human monoclonal antibodies. To enhance the usefulness of CHO-K1 cells we developed a new selection system based on double auxotrophy. We used CRISPR-Cas9 to knockout the genes that encode the bifunctional enzymes catalyzing the last two steps in the de novo synthesis of pyrimidines and purines (uridine monophosphate synthase and 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/IMP cyclohydrolase [ATIC], respectively). Survival of these doubly auxotrophic cells depends on the provision of sources of purines and pyrimidines or on the transfection and integration of open reading frames encoding these two enzymes. We successfully used one such double auxotroph (UA10) to select for stable transfectants carrying (a) the recombinant tumor necrosis factor-α receptor fusion protein etanercept and (b) the heavy and light chains of the anti-Her2 monoclonal antibody trastuzumab. Transfectant clones produced these recombinant proteins in a stable manner and in substantial amounts. The availability of this double auxotroph provides a rapid and efficient selection method for the serial or simultaneous transfer of genes for multiple polypeptides of choice into CHO cells using readily available purine- and pyrimidine-free commercial media.     

Pristimerin Induces Autophagy‐Mediated Cell Death in K562 Cells through the ROS/JNK Signaling Pathway

Chronic myeloid leukemia (CML) is a lethal malignancy, and the progress toward long‐term survival has stagnated in recent decades. Pristimerin, a quinone methide triterpenoid isolated from the Celastraceae and Hippocrateaceae families, is well‐known to exert potential anticancer activities. In this study, we investigated the effects and the mechanisms of action on CML. We found that pristimerin inhibited cell proliferation of K562 CML cells by causing G1 phase arrest. Furthermore, we demonstrated that pristimerin triggered autophagy and apoptosis. Intriguingly, pristimerin‐induced cell death was restored by an autophagy inhibitor, suggesting that autophagy is cross‐linked with pristimerin‐induced apoptosis. Further studies revealed that pristimerin could produce excessive reactive oxygen species (ROS), which then induce JNK activation. These findings provide clear evidence that pristimerin might be clinical benefit to patients with CML.     

Bringing Conducting Polymers to High Order: Toward Conductivities beyond 105 S cm−1 and Thermoelectric Power Factors of 2 mW m−1 K−2

Here, an effective design strategy of polymer thermoelectric materials based on structural control in doped polymer semiconductors is presented. The strategy is illustrated for two archetypical polythiophenes, e.g., poly(2,5‐bis(3‐dodecyl‐2‐thienyl)thieno[3,2‐b]thiophene) (C₁₂‐PBTTT) and regioregular poly(3‐hexylthiophene) (P3HT). FeCl₃ doping of aligned films results in charge conductivities up to 2 × 10⁵ S cm^?1 and metallic‐like thermopowers similar to iodine‐doped polyacetylene. The films are almost optically transparent and show strongly polarized near‐infrared polaronic bands (dichroic ratio >10). The comparative study of structure–property correlations in P3HT and C₁₂‐PBTTT identifies three conditions to obtain conductivities beyond 10⁵ S cm^?1: i) achieve high in‐plane orientation of conjugated polymers with high persistence length; ii) ensure uniform chain oxidation of the polymer backbones by regular intercalation of dopant molecules in the polymer structure without disrupting alignment of π‐stacked layers; and iii) maintain a percolating nanomorphology along the chain direction. The highly anisotropic conducting polymer films are ideal model systems to investigate the correlations between thermopower S and charge conductivity σ. A scaling law S ∝ σ^?1/4 prevails along the chain direction, but a different S ∝ ?ln(σ) relation is observed perpendicular to the chains, suggesting different charge transport mechanisms. The simultaneous increase of charge conductivity and thermopower along the chain direction results in a substantial improvement of thermoelectric power factors up to 2 mW m^?1 K^?2 in C₁₂‐PBTTT.     

Caffeic Acid Phenethyl Ester Loaded in Microemulsions: Enhanced In Vitro Activity against Colon and Breast Cancer Cells and Possible Cellular Mechanisms

Food Biophysics - Caffeic acid phenethyl ester (CAPE) has high cytotoxicity against various cancer cells but has low water solubility and poor bioavailability. The objective of this work was to...     

Expression Profiling and Validation of Potential Reference Genes During <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis> Embryogenesis

Differential expression of genes is crucial to embryogenesis. The analysis of gene expression requires appropriate references that should be minimally regulated during the embryonic development. To select the most stable genes for gene normalization, the expression profiles of eight commonly used reference genes (ACTB, GAPDH, rpL17, α-Tub, EF1-α, UbcE, B2M, and 18S rRNA) were examined during Japanese flounder (Paralichthys olivaceus) embryonic development using quantitative real-time polymerase chain reaction. It was found that all seven mRNA genes appeared to be developmentally regulated and exhibited significant variation of expression. However, further analyses revealed the stage-specific expression stability. Hence when normalization using these mRNA genes, the differential and stage-related expression should be considered. 18S rRNA gene, on the other hand, showed the most stable expression and could be recommended as a suitable reference gene during all embryonic developmental stages in P. olivaceus. In summary, our results provided not only the appropriate reference gene for embryonic development research in P. olivaceus, but also possible guidance to reference gene selection for embryonic gene expression analyses in other fish species.     

Involvement of the acid sphingomyelinase pathway in uva-induced apoptosis

The sphingomyelin-ceramide pathway is an evolutionarily conserved ubiquitous signal transduction system that regulates many cell functions including apoptosis. Sphingomyelin (SM) is hydrolyzed to ceramide by different sphingomyelinases. Ceramide serves as a second messenger in mediating cellular effects of cytokines and stress. In this study, we find that acid sphingomyelinase (SMase) activity was induced by UVA in normal JY lymphoblasts but was not detectable in MS1418 lymphoblasts from Niemann-Pick type D patients who have an inherited deficiency of acid SMase. We also provide evidence that UVA can induce apoptosis by activating acid SMase in normal JY cells. In contrast, UVA-induced apoptosis was inhibited in MS1418 cells. Exogenous SMase and its product, ceramide (10-40 micrometer), induced apoptosis in JY and MS1418 cells, but the substrate of SMase, SM (20-80 micrometer), induced apoptosis only in JY cells. These results suggest that UVA-induced apoptosis by SM is dependent on acid SMase activity. We also provide evidence that induction of apoptosis by UVA may occur through activation of JNKs via the acid SMase pathway.     

The Successful Diagnosis and Typing of Systemic Amyloidosis Using A Microwave-Assisted Filter-Aided Fast Sample Preparation Method and LC/MS/MS Analysis

Laser microdissection followed by mass spectrometry has been successfully used for amyloid typing. However, sample contamination can interfere with proteomic analysis, and overnight digestion limits the analytical throughput. Moreover, current quantitative analysis methods are based on the spectrum count, which ignores differences in protein length and may lead to misdiagnoses. Here, we developed a microwave-assisted filter-aided sample preparation (maFASP) method that can efficiently remove contaminants with a 10-kDa cutoff ultrafiltration unit and can accelerate the digestion process with the assistance of a microwave. Additionally, two parameters (P- and D-scores) based on the exponentially modified protein abundance index were developed to define the existence of amyloid deposits and those causative proteins with the greatest abundance. Using our protocol, twenty cases of systemic amyloidosis that were well-typed according to clinical diagnostic standards (training group) and another twenty-four cases without subtype diagnoses (validation group) were analyzed. Using this approach, sample preparation could be completed within four hours. We successfully subtyped 100% of the cases in the training group, and the diagnostic success rate in the validation group was 91.7%. This maFASP-aided proteomic protocol represents an efficient approach for amyloid diagnosis and subtyping, particularly for serum-contaminated samples.