Targeting T cells alone has yielded limited success in the prevention of graft versus host disease (GvHD) after allogeneic blood and marrow transplantation (BMT). Dendritic cells (DCs) play a central role in alloreactivity and therefore represent a suitable target. Due to their immune modulatory effects and their inhibition of maturation and function of DC, proteasome inhibitors have prompted investigators to examine their potential role in the prevention of GvHD. Ixazomib is a specific and reversible proteasome inhibitor with rapid dissociation from 20S proteasome. It is orally bioavailable. We aimed to explore its effect on healthy volunteer DCs. DCs were isolated using EasySep Pan-DC Pre-Enrichment Cocktail with a purity over 90% (STEMCELL Technologies). DCs were treated with ixazomib at different concentrations for 4 hrs and then stimulated with Lipopolysaccharide (LPS) for 16 hrs. After the treatment, DCs were surface stained with antibodies against maturation markers and analyzed by flow cytometry. DC survival was evaluated with 7AAD staining and FACS analysis. To assess the effect of ixazomib on the production of pro-inflammatory cytokines, DCs were incubated with ixazomib at increasing concentration before or after addition of LPS. Pro-inflammatory cytokines in the supernatant of tissue culture were measured using EMD Millipore cytokine arrays. Experiments were done in triplicate at least. Unpaired T test was used for statistical analysis. P < 0.05 was considered significant. Ixazomib inhibited expression of 6 DC maturation markers including CD40, CD54, CD80, CD83, CD86 and CD197 (CCR-7). The inhibition started at a concentration of 10 nM and was dose related. Ixazomib also decreased the percentage of DCs simultaneously expressing multiple markers. DCs viability remained unchanged in comparison to control at a concentration of 10 nM and dropped to 68% and 43% on average with concentrations of 20 nM and 40 nM. There was no significant change in the cytokine production when ixazomib was added 4 hours after LPS. On the other hand, ixazomib significantly decreased production of IL-6 and IL-23 by DCs at the concentration of 20-30 nM. However, there was no significant change of TNF-β and INF-γ upon ixazomib treatments. In summary, ixazomib inhibits DC maturation with relative preservation of cell viability. On the other hand, ixazomib inhibits pro-inflammatory cytokine production in DCs only when it is added before LPS stimulation. This may have clinical implications and suggests that adequate inhibition of DCs before graft infusion might be essential to the prevention GvHD.
Toxicity determination based on electrochemically active bacteria (EAB) shows great prospects for early warning of water pollution. However, the lifetime and reusability of EAB in toxicity determination remain uncertain. This study performed continuous toxicity determination by using an automatic water toxicity determination system based on EAB. Results demonstrated that EAB are capable of rapid responses to common heavy metal pollutants and make a full recovery after refreshment. Despite changes in microbial communities, EAB maintain a similar current generation capacity and toxicity sensitivity even after 20 continuous toxicity tests. The main reason for the stable performance was unchanged gene functions, as the toxicity tests did not result in a decrease in genes related to current generation or an increase in genes related to resistance. This study first reported that EAB possess a prolonged lifetime and good reusability in water toxicity determination, providing a basis for the continuous determination for water toxicity and the online monitoring of industrial wastewater toxicity based on EAB.
Extracellular matrix (ECM) actively participates in normal cell regulation and in the process of tumor progression. The Rho GTPase Cdc42 has been shown to regulate cell-ECM interaction in conventional two-dimensional culture conditions by using dominant mutants of Cdc42 in immortalized cell lines that may introduce nonspecific effects. Here, we employ three-dimensional culture systems for conditional gene targeted primary mouse embryonic fibroblasts that better simulate the reciprocal and adaptive interactions between cells and surrounding matrix to define the role of Cdc42 signaling pathways in ECM organization. Cdc42 deficiency leads to a defect in global cell-matrix interactions reflected by a decrease in collagen gel contraction. The defect is associated with an altered cell-matrix interaction that is evident by morphologic changes and reduced focal adhesion complex formation. The matrix defect is also associated with a reduction in synthesis and activation of matrix metalloproteinase 9 (MMP9) and altered fibronectin deposition patterning. A Cdc42 mutant rescue experiment found that downstream of Cdc42, p21-activated kinase (PAK), but not Par6 or WASP, may be involved in regulating collagen gel contraction and fibronectin organization. Thus, in addition to the previously implicated roles in intracellular regulation of actin organization, proliferation, and vesicle trafficking, Cdc42 is essential in ECM remodeling in three dimensions. Extracellular matrix (ECM) actively participates in normal cell regulation and in the process of tumor progression. The Rho GTPase Cdc42 has been shown to regulate cell-ECM interaction in conventional two-dimensional culture conditions by using dominant mutants of Cdc42 in immortalized cell lines that may introduce nonspecific effects. Here, we employ three-dimensional culture systems for conditional gene targeted primary mouse embryonic fibroblasts that better simulate the reciprocal and adaptive interactions between cells and surrounding matrix to define the role of Cdc42 signaling pathways in ECM organization. Cdc42 deficiency leads to a defect in global cell-matrix interactions reflected by a decrease in collagen gel contraction. The defect is associated with an altered cell-matrix interaction that is evident by morphologic changes and reduced focal adhesion complex formation. The matrix defect is also associated with a reduction in synthesis and activation of matrix metalloproteinase 9 (MMP9) and altered fibronectin deposition patterning. A Cdc42 mutant rescue experiment found that downstream of Cdc42, p21-activated kinase (PAK), but not Par6 or WASP, may be involved in regulating collagen gel contraction and fibronectin organization. Thus, in addition to the previously implicated roles in intracellular regulation of actin organization, proliferation, and vesicle trafficking, Cdc42 is essential in ECM remodeling in three dimensions.
Electrochemically active bacteria (EAB) serve as the catalyst for microbial electrochemical systems, enabling simultaneous wastewater treatment and electricity generation. However, the increasing presence of toxic pollutants in wastewater may inhibit EAB metabolism and decrease EAB abundance, and the detailed effects and microbial adaptability remain unclear. The study used two toxic pollutants to simulate long-term and incremental toxic exposure. The study's results show that the current generation of EAB is significantly inhibited at the early stage of toxic exposure, but the inhibition becomes less severe in the middle stage and disappears by the end stage. As a result, EAB is able to resist concentrations of 12 mg/L HgSO4 or 240 mg/L CH2Cl2. This increased resistance is due to the adaptive response that occurs in EAB. The susceptible Geobacter population decreases while the resistant Geomonas population increases, but the total sum of the two genera remains unchanged. These findings provide insight into the adaptive mechanism of EAB to toxic exposure, which offers a basis for using EAB to treat toxic wastewater.
Silicon carbide (SiC) MOSFET module in HybridPACK™ drive package has been employed for mainstream automotive high power inverter applications. The short-circuit fault, however, threatens the reliability of the power module to a large extent. The tunnel magnetoresistance (TMR) sensor indicates short response delay, fair accuracy and low temperature drift for the short-circuit protection. The TMR sensor detect the short-circuit current by sensing the magnetic field in the sensitive direction. For the purpose of compact application and convenient current control, the TMR sensor is integrated on the surface of the power terminal. Concrete magnetic analysis with the assistance of finite element method (FEM) calculation is conducted by which means the short-circuit fault is to be detected in advance. A prototype based on SiC MOSFET module (STARPOWER MD29HTC120P6HE, 1200V/450A) is constructed to testify the TMR-based scheme. The short-circuit detection time of the TMR sensor is within 100ns and the SiC MOSFET module is protected in a safe manner.
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