GCS overexpression is associated with multidrug resistance of human HCT-8 colon cancer cells

<p>Abstract</p> <p>Purpose</p> <p>Multidrug resistance is one of the main impediments to the successful treatment of colon cancer. Glucosylceramide synthase (GCS) which is related to multidrug resistance (MDR) can reduce the level of ceramide and can help cells escape from the ceramide-induced cell apoptosis. However, the underlying mechanism is still unclear.</p> <p>Methods</p> <p>The cell proliferation and cell toxicity were measured with Cell Counting Kit-8 (CCK-8). The mRNA levels of GCS and MDR1 were detected by semiquantitative reverse transcription-PCR amplification, the protein levels of GCS, caspase-3 and P-gp proteins were indicated by Western blotting. The apoptosis rates of cells were measured with flow cytometry.</p> <p>Results</p> <p>The relative mRNA levels of GCS in HCT-8, HCT-8/VCR, HCT-8/VCR- sh-mock and HCT-8/VCR-sh-GCS were 71.4 ± 1.1%, 95.1 ± 1.2%, 98.2 ± 1.5%, and 66.6 ± 2.1% respectively. The mRNA levels of MDR1 were respectively 61.3 ± 1.1%, 90.5 ± 1.4%, 97.6 ± 2.2% and 56.1 ± 1.2%. The IC50 of Cisplatin complexes were respectively 69.070 ± 0.253 μg/ml, 312.050 ± 1.46 μg/ml, 328.741 ± 5.648 μg/ml, 150.792 ± 0.967 μg/ml in HCT-8, HCT-8/VCR, HCT-8/VCR-sh-mock and HCT-8/VCR-sh-GCS. The protein levels of caspase-3 were 34.2 ± o.6%, 93.0 ± 0.7%, 109.09 ± 0.7%, 42.7 ± 1.3% respectively. The apoptosis rates of cells were 8.77 ± 0.14%, 12.75 ± 0.54%, 15.39 ± 0.41% and 8.49 ± 0.23% respectively.</p> <p>Conclusion</p> <p>In conclusion, our research indicated that suppression of GCS restores the sensitivity of multidrug resistance colon cancer cells to drug treatment.</p

NBMOD: Find It and Grasp It in Noisy Background

Grasping objects is a fundamental yet important capability of robots, and many tasks such as sorting and picking rely on this skill. The prerequisite for stable grasping is the ability to correctly identify suitable grasping positions. However, finding appropriate grasping points is challenging due to the diverse shapes, varying density distributions, and significant differences between the barycenter of various objects. In the past few years, researchers have proposed many methods to address the above-mentioned issues and achieved very good results on publicly available datasets such as the Cornell dataset and the Jacquard dataset. The problem is that the backgrounds of Cornell and Jacquard datasets are relatively simple - typically just a whiteboard, while in real-world operational environments, the background could be complex and noisy. Moreover, in real-world scenarios, robots usually only need to grasp fixed types of objects. To address the aforementioned issues, we proposed a large-scale grasp detection dataset called NBMOD: Noisy Background Multi-Object Dataset for grasp detection, which consists of 31,500 RGB-D images of 20 different types of fruits. Accurate prediction of angles has always been a challenging problem in the detection task of oriented bounding boxes. This paper presents a Rotation Anchor Mechanism (RAM) to address this issue. Considering the high real-time requirement of robotic systems, we propose a series of lightweight architectures called RA-GraspNet (GraspNet with Rotation Anchor): RARA (network with Rotation Anchor and Region Attention), RAST (network with Rotation Anchor and Semi Transformer), and RAGT (network with Rotation Anchor and Global Transformer) to tackle this problem. Among them, the RAGT-3/3 model achieves an accuracy of 99% on the NBMOD dataset. The NBMOD and our code are available at https://github.com/kmittle/Grasp-Detection-NBMOD

Molecular genetic analysis of phosphomannomutase genes in Triticum monococcum

AbstractIn higher plants, phosphomannomutase (PMM) is essential for synthesizing the antioxidant ascorbic acid through the Smirnoff–Wheeler pathway. Previously, we characterized six PMM genes (TaPMM-A1, A2, B1, B2, D1 and D2) in common wheat (Triticum aestivum, AABBDD). Here, we report a molecular genetic analysis of PMM genes in Triticum monococcum (AmAm), a diploid wheat species whose Am genome is closely related to the A genome of common wheat. Two distinct PMM genes, TmPMM-1 and TmPMM-2, were found in T. monococcum. The coding region of TmPMM-1 was intact and highly conserved. In contrast, two main TmPMM-2 alleles were identified, with TmPMM-2a possessing an intact coding sequence and TmPMM-2b being a pseudogene. The transcript level of TmPMM-2a was much higher than that of TmPMM-2b, and a bacterially expressed TmPMM-2a recombinant protein displayed relatively high PMM activity. In general, the total transcript level of PMM was substantially higher in accessions carrying TmPMM-1 and TmPMM-2a than those harboring TmPMM-1 and TmPMM-2b. However, total PMM protein and activity levels did not differ drastically between the two genotypes. This work provides new information on PMM genes in T. monococcum and expands our understanding on Triticeae PMM genes, which may aid further functional and applied studies of PMM in crop plants

Research on the development and testing methods of physical education and agility training equipment in universities

IntroductionBecause of the problems of insufficient funds and traditional training methods in college sports agile training, an agile training system based on a wireless ad hoc network was developed to evaluate the effect of improving the sensitive quality of ordinary college students. Based on the ESP-MESH network, the lower computer realizes automatic networking between devices and tests the performance of the mesh network. Fourteen male college students received 9 weeks of agility training, with seven students in each of two groups: traditional agility training and agile equipment training. The researchers evaluated the performance of both groups in rapid disguise, body coordination, changing movements, and predictive decision-making.ResultsThere was no significant difference between the groups before training, but there were significant differences in the four abilities after training (p &lt; 0.01). The experimental group had significant differences in rapid direction change and physical coordination (p &lt; 0.05), and in changing movement and predictive decision-making ability (p &lt; 0.01).ConclusionBoth traditional training and agile equipment training improve the agility quality of college students, and the latter shows better results in certain abilities. However, limited by other physical qualities, the improvement of motor changes and predictive decision-making ability is not as obvious as the other two abilities

Gut microbiota and acylcarnitine metabolites connect the beneficial association between estrogen and lipid metabolism disorders in ovariectomized mice

Decreased estrogen level is one of the main causes of lipid metabolism disorders and coronary heart disease in women after menopause. Exogenous estradiol benzoate is effective to some extent in alleviating lipid metabolism disorders caused by estrogen deficiency. However, the role of gut microbes in the regulation process is not yet appreciated. The objective of this study was to investigate the effects of estradiol benzoate supplementation on lipid metabolism, gut microbiota, and metabolites in ovariectomized (OVX) mice and to reveal the importance of gut microbes and metabolites in the regulation of lipid metabolism disorders. This study found that high doses of estradiol benzoate supplementation effectively attenuated fat accumulation in OVX mice. There was a significant increase in the expression of genes enriched in hepatic cholesterol metabolism and a concomitant decrease in the expression of genes enriched in unsaturated fatty acid metabolism pathways. Further screening of the gut for characteristic metabolites associated with improved lipid metabolism revealed that estradiol benzoate supplementation influenced major subsets of acylcarnitine metabolites. Ovariectomy significantly increased the abundance of characteristic microbes that are significantly negatively associated with acylcarnitine synthesis, such as Lactobacillus and Eubacterium ruminantium group bacteria, while estradiol benzoate supplementation significantly increased the abundance of characteristic microbes that are significantly positively associated with acylcarnitine synthesis, such as Ileibacterium and Bifidobacterium spp. The use of pseudosterile mice with gut microbial deficiency greatly facilitated the synthesis of acylcarnitine due to estradiol benzoate supplementation and also alleviated lipid metabolism disorders to a greater extent in OVX mice. IMPORTANCE Our findings establish a role for gut microbes in the progression of estrogen deficiency-induced lipid metabolism disorders and reveal key target bacteria that may have the potential to regulate acylcarnitine synthesis. These findings suggest a possible route for the use of microbes or acylcarnitine to regulate disorders of lipid metabolism induced by estrogen deficiency

Activation of MET signaling by HDAC6 offers a rationale for a novel ricolinostat and crizotinib combinatorial therapeutic strategy in diffuse large B‐cell lymphoma

Some histone deacetylases (HDACs) promote tumor cell growth and pan‐ or selective HDAC inhibitors are active in some cancers; however, the pivotal HDAC enzyme and its functions in human diffuse large B‐cell lymphoma (DLBCL) remain largely unknown. Using NanoString nCounter assays, we profiled HDAC mRNA expression and identified HDAC6 as an upregulated HDAC family member in DLBCL tissue samples. We then found that HDAC6 plays an oncogenic role in DLBCL, as evidenced by its promotion of cell proliferation in vitro and tumor xenograft growth in vivo. Mechanistically, the interaction between HDAC6 and HR23B downregulated HR23B expression, thereby reducing the levels of casitas B‐lineage lymphoma (c‐Cbl), an E3 ubiquitin ligase for hepatocyte growth factor receptor (MET), which resulted in the inhibition of MET ubiquitination‐dependent degradation. In addition, enhanced HDAC6 expression and decreased HR23B expression were correlated with poor overall survival rates among patients with DLBCL. Taken together, these results establish an HDAC6–HR23B–MET axis and indicate that HDAC6 is a potent promoter of lymphomagenesis in DLBCL. Thus, a therapeutic strategy based on HDAC6 inhibitors in combination with MET inhibitors is promising. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/146400/1/path5108_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/146400/2/path5108.pd

Structural basis for the assembly and quinone transport mechanisms of the dimeric photosynthetic RC-LH1 supercomplex

The reaction center (RC) and light-harvesting complex 1 (LH1) form a RC–LH1 core supercomplex that is vital for the primary reactions of photosynthesis in purple phototrophic bacteria. Some species possess the dimeric RC–LH1 complex with a transmembrane polypeptide PufX, representing the largest photosynthetic complex in anoxygenic phototrophs. However, the details of the architecture and assembly mechanism of the RC–LH1 dimer are unclear. Here we report seven cryo-electron microscopy (cryo-EM) structures of RC–LH1 supercomplexes from Rhodobacter sphaeroides. Our structures reveal that two PufX polypeptides are positioned in the center of the S-shaped RC–LH1 dimer, interlocking association between the components and mediating RC–LH1 dimerization. Moreover, we identify another transmembrane peptide, designated PufY, which is located between the RC and LH1 subunits near the LH1 opening. PufY binds a quinone molecule and prevents LH1 subunits from completely encircling the RC, creating a channel for quinone/quinol exchange. Genetic mutagenesis, cryo-EM structures, and computational simulations provide a mechanistic understanding of the assembly and electron transport pathways of the RC–LH1 dimer and elucidate the roles of individual components in ensuring the structural and functional integrity of the photosynthetic supercomplex

Structural basis for the assembly and electron transport mechanisms of the dimeric photosynthetic RC–LH1 supercomplex

AbstractThe reaction center (RC) and light-harvesting complex 1 (LH1) form a RC–LH1 core supercomplex that is vital for the primary reactions of photosynthesis in purple photosynthetic bacteria. Some species possess the dimeric RC–LH1 complex with an additional polypeptide PufX, representing the largest photosynthetic complex in anoxygenic phototrophs. However, the details of the architecture and assembly mechanism of the RC–LH1 dimer are unclear. Here we report seven cryo-electron microscopy (cryo-EM) structures of RC–LH1 supercomplexes from Rhodobacter sphaeroides. Our structures reveal that two PufX polypeptides are positioned in the center of the S-shaped RC–LH1 dimer, interlocking association between the components and mediating RC–LH1 dimerization. Moreover, we identify a new transmembrane peptide, designated PufY, which is located between the RC and LH1 subunits near the LH1 opening. PufY binds a quinone molecule and prevents LH1 subunits from completely encircling the RC, creating a channel for quinone/quinol exchange. Genetic mutagenesis, cryo-EM structures, and computational simulations enable a mechanistic understanding of the assembly and electron transport pathways of the RC–LH1 dimer and elucidate the roles of individual components in ensuring the structural and functional integrity of the photosynthetic supercomplex.</jats:p

Novel Evolved Immunoglobulin (Ig)-Binding Molecules Enhance the Detection of IgM against Hepatitis C Virus

Detection of specific antibodies against hepatitis C virus (HCV) is the most widely available test for viral diagnosis and monitoring of HCV infections. However, narrowing the serologic window of anti-HCV detection by enhancing anti-HCV IgM detection has remained to be a problem. Herein, we used LD5, a novel evolved immunoglobulin-binding molecule (NEIBM) with a high affinity for IgM, to develop a new anti-HCV enzyme-linked immunosorbent assay (ELISA) using horseradish peroxidase-labeled LD5 (HRP-LD5) as the conjugated enzyme complex. The HRP-LD5 assay showed detection efficacy that is comparable with two kinds of domestic diagnostic kits and the Abbott 3.0 kit when tested against the national reference panel. Moreover, the HRP-LD5 assay showed a higher detection rate (55.9%, 95% confidence intervals (95% CI) 0.489, 0.629) than that of a domestic diagnostic ELISA kit (Chang Zheng) (53.3%, 95% CI 0.463, 0.603) in 195 hemodialysis patient serum samples. Five serum samples that were positive using the HRP-LD5 assay and negative with the conventional anti-HCV diagnostic ELISA kits were all positive for HCV RNA, and 4 of them had detectable antibodies when tested with the established anti-HCV IgM assay. An IgM confirmation study revealed the IgM reaction nature of these five serum samples. These results demonstrate that HRP-LD5 improved anti-HCV detection by enhancing the detection of anti-HCV IgM, which may have potential value for the early diagnosis and screening of hepatitis C and other infectious diseases

Hydrogen Peroxide Acts on Sensitive Mitochondrial Proteins to Induce Death of a Fungal Pathogen Revealed by Proteomic Analysis

How the host cells of plants and animals protect themselves against fungal invasion is a biologically interesting and economically important problem. Here we investigate the mechanistic process that leads to death of Penicillium expansum, a widespread phytopathogenic fungus, by identifying the cellular compounds affected by hydrogen peroxide (H2O2) that is frequently produced as a response of the host cells. We show that plasma membrane damage was not the main reason for H2O2-induced death of the fungal pathogen. Proteomic analysis of the changes of total cellular proteins in P. expansum showed that a large proportion of the differentially expressed proteins appeared to be of mitochondrial origin, implying that mitochondria may be involved in this process. We then performed mitochondrial sub-proteomic analysis to seek the H2O2-sensitive proteins in P. expansum. A set of mitochondrial proteins were identified, including respiratory chain complexes I and III, F1F0 ATP synthase, and mitochondrial phosphate carrier protein. The functions of several proteins were further investigated to determine their effects on the H2O2-induced fungal death. Through fluorescent co-localization and the use of specific inhibitor, we provide evidence that complex III of the mitochondrial respiratory chain contributes to ROS generation in fungal mitochondria under H2O2 stress. The undesirable accumulation of ROS caused oxidative damage of mitochondrial proteins and led to the collapse of mitochondrial membrane potential. Meanwhile, we demonstrate that ATP synthase is involved in the response of fungal pathogen to oxidative stress, because inhibition of ATP synthase by oligomycin decreases survival. Our data suggest that mitochondrial impairment due to functional alteration of oxidative stress-sensitive proteins is associated with fungal death caused by H2O2