Investigation of glycosylation in flagellin biosynthesis and LPS O-antigen biosynthesis in Aeromonas caviae

Aeromonas species are Gram-negative facultative anaerobic bacteria which are widespread in fresh water and salt water. Aeromonas caviae and Aeromonas hydrophila are two kinds of mesophilic aeromonads which belong to genus Aeromonas and are emerging as major pathogens in humans. Flagella are important pathogenic factors of bacteria and there are two kinds of flagella discovered in A. caviae and A. hydrophila which are polar flagella and lateral flagella (Rabaan et al. 2001; Tabei et al. 2009; Canals et al. 2006a). The flagella of Aeromonas are glycosylated and investigation of glycosylation in flagellin biosynthesis is valuable for the understanding of pathogenicity of Aeromonas species. The aim of this project is to investigate the glycosylation during flagellin biosynthesis in A. caviae. It has been discovered that the flagella of A. caviae are glycosylated 6 to 8 times by Pse5Ac7Ac which is under the control of Maf1 which is known as flagellin glycosyl-transferase (Parker et al. 2012). Theoretically, Maf1 of A. caviae Sch3N has the ability to transfer activated pseudaminic acid (CMP- Pse5Ac7Ac) to the hydroxyl group of serine and threonine residues in the central immunogenic D2/D3 domain of flagellin of A. caviae Sch3N (Parker et al. 2012; Tabei et al. 2009). The details of how Maf1 interacts with flagellin will be explored in this project. CMP-Pse5Ac7Ac which is the substrate for glycosylation is generated from UDP-GlcNAc. The biosynthesis of CMP-Pse5Ac7Ac is under the control of flm locus including flmA, flmB, neuA, neuB, flmD. The pseudaminic acid biosynthetic pathway has been confirmed in Campylobacter jejuni and related enzymes are homologous proteins of A. caviae (Schoenhofen et al. 2006). The proteins encoded by the flm genes will be applied in this project to investigate the biosynthesis of CMP- Pse5Ac7Ac. In addition, there is evidence that the LPS O-antigen incorporates with Pse5Ac7Ac. The substrate of O-antigen glycosylation is also CMP-Pse5Ac7Ac (Tomás 2012). The connections between glycosylation in LPS O-antigen biosynthesis and flagellin biosynthesis will be investigated in this project

Biocatalytic Transfer of Pseudaminic Acid (Pse5Ac7Ac) Using Promiscuous Sialyltransferases in a Chemoenzymatic Approach to Pse5Ac7Ac-Containing Glycosides

Pseudaminic acid (Pse5Ac7Ac) is a nonmammalian sugar present on the cell surface of a number of bacteria including Pseudomonas aeruginosa, Campylobacter jejuni, and Acinetobacter baumannii. However, the role Pse5Ac7Ac plays in host–pathogen interactions remains underexplored, particularly compared to its ubiquitous sialic acid analogue Neu5Ac. This is primarily due to a lack of access to difficult to prepare Pse5Ac7Ac glycosides. Herein, we describe the in vitro biocatalytic transfer of an activated Pse5Ac7Ac donor onto glycosyl acceptors, enabling the enzymatic synthesis of Pse5Ac7Ac-containing glycosides. In a chemoenzymatic approach, chemical synthesis initially afforded access to a late-stage Pse5Ac7Ac biosynthetic intermediate, which was subsequently converted to the desired CMP-glycosyl donor in a one-pot two-enzyme process using biosynthetic enzymes. Finally, screening a library of 13 sialyltransferases (SiaT) with the unnatural substrate enabled the identification of a promiscuous inverting SiaT capable of turnover to afford β-Pse5Ac7Ac-terminated glycosides.</p

Cellular and Extracellular Components in Tumor Microenvironment and Their Application in Early Diagnosis of Cancers

Tumors are surrounded by complex environmental components, including blood and lymph vessels, fibroblasts, endothelial cells, immune cells, cytokines, extracellular vesicles, and extracellular matrix. All the stromal components together with the tumor cells form the tumor microenvironment (TME). In addition, extracellular physical and chemical factors, including extracellular pH, hypoxia, elevated interstitial fluid pressure, and fibrosis, are closely associated with tumor progression, metastasis, immunosuppression, and drug resistance. Cellular and extracellular components in TME contribute to nearly all procedures of carcinogenesis. By summarizing the recent work in this field, we make a comprehensive review on the role of cellular and extracellular components in the process of carcinogenesis and their potential application in early diagnosis of cancer. We hope that a systematic review of the diverse aspects of TME will help both research scientists and clinicians in this field

Coal and rock identification method based on Kalman optimal estimation of load data of rocker arm pin axle of shearer

The shearer's coal and rock identification technology is the basis of intelligent control. The existing coal and rock identification method for the site environment and testing equipment requirements are higher. The actual fully mechanized working face is difficult to meet the necessary conditions. In order to solve the above problems, a coal and rock identification method based on the Kalman optimal estimation of the shearer rocker pin axle load data is proposed. On the basis of not increasing external auxiliary instruments and equipment, the rocker pin axle sensor is used to replace the existing pin axle to sense the coal and rock load, which can better adapt to the environment. By measuring the strain data of the rocker pin axle sensor located at the connection between the rocker arm and the connecting frame, the Kalman optimal estimation method is used to reduce the noise of the load data. The load intervals of the shearer under different working conditions such as cutting coal and rock are separated from each other. By judging the interval of the real-time load value, the coal and rock interface can be identified. The identification of coal and rock is verified on a fully mechanized experimental platform. The load of the rocker pin axle at the upper end of the coal wall side along the shearer traction direction is analyzed in three stages: no-load, cutting the coal wall and cutting the rock. The results show that before the load data is processed, there is overlap between the load intervals of cutting the coal wall and cutting the rock, and the coal and rock interface identification cannot be completed accurately. After the load data is processed by the Kalman optimal estimation algorithm, the load intervals under no-load, cutting the coal wall and cutting the rock conditions are separated from each other. In addition, the load interval length of each working condition is shortened by 65.6%-83.3%, and the mean square deviation is reduced by 66.5%-72.9%. The data fluctuation is smaller, which effectively improves data identification. In practical engineering applications, the expected load stress range in the coal seam cutting state can be set according to the method. Once this range is exceeded, it is judged that this is not a cutting the coal wall state and thus coal and rock identification is achieved

Identification of Molecular Regulatory Features and Markers for Acute Type A Aortic Dissection

Background. Acute type A aortic dissection (ATAAD) is one of the most lethal cardiovascular diseases, and its molecular mechanism remains unclear. Methods. Differentially expressed genes (DEGs) between ATAAD and control were detected by limma R package in GSE52093, GSE153434, GSE98770, and GSE84827, respectively. The coexpression network of DEGs was identified by the WGCNA package. Enrichment analysis was performed for module genes that were positively correlated with ATAAD using clusterProfiler R package. In addition, differentially methylated markers between aortic dissection and control were identified by ChAMP package. After comparing with ATAAD-related genes, a protein-protein interaction (PPI) network was established based on the STRING database. The genes with the highest connectivity were identified as hub genes. Finally, differential immune cell infiltration between ATAAD and control was identified by ssGSEA. Results. From GSE52093 and GSE153434, 268 module genes were obtained with consistent direction of differential expression and high correlation with ATAAD. They were significantly enriched in T cell activation, HIF-1 signaling pathway, and cell cycle. In addition, 2060 differentially methylated markers were obtained from GSE84827. Among them, 77 methylation markers were ATAAD-related DEGs. Using the PPI network, we identified MYC, ITGA2, RND3, BCL2, and PHLPP2 as hub genes. Finally, we identified significantly differentially infiltrated immune cells in ATAAD. Conclusion. The hub genes we identified may be regulated by methylation and participate in the development of ATAAD through immune inflammation and oxidative stress response. The findings may provide new insights into the molecular mechanisms and therapeutic targets for ATAAD

A Circulating miRNA-Based Scoring System Established by WGCNA to Predict Colon Cancer

Introduction. Circulation microRNAs (miRNAs) perform as potential diagnostic biomarkers of many kinds of cancers. This study is aimed at identifying circulation miRNAs as diagnostic biomarkers in colon cancer. Methods. We conducted a weighted gene coexpression network analysis (WGCNA) in miRNAs to find out the expression pattern among circulation miRNAs by using a “WGCNA” package in R. Correlation analysis was performed to find cancer-related modules. Differentially expressed miRNAs (DEmiRs) in colon cancer were identified by a “limma” package in R. Hub gene analysis was conducted for these DEmiRs in the cancer-related modules by the “closeness” method in cytoscape software. Then, logistic regression was performed to identify the independent risk factors, and a scoring system was constructed based on these independent risk factors. Then, we use data from the GEO database to confirm the reliability of this scoring system. Results. A total of 9 independent coexpression modules were constructed based on the expression levels of 848 miRNAs by WGCNA. After correlation analysis, green (cor=0.77, p=3×10‐25) and yellow (cor=0.65, p=6×10‐16) modules were strongly correlated with cancer development. 20 hub genes were found after hub gene analysis in these DEmiRs by cytoscape. Among all these hub genes, hsa-miR-23a-3p (OR=2.6391, p=6.23×10‐5) and hsa-miR-663a (OR=1.4220, p=0.0069) were identified as an independent risk factor of colon cancer by multivariate regression. Furthermore, a scoring system was built to predict the probability of colon cancer based on both of these miRNAs, the area under the curve (AUC) of which was 0.828. Data from GSE106817 and GSE112264 was used to confirm this scoring system. And the AUC of them was 0.980 and 0.917, respectively. Conclusion. We built a scoring system based on circulation hub miRNAs found by WGCNA to predict the development of colon cancer

Analysis of the efficacy of drilling decompression autologous bone marrow and allogeneic bone grafting in the treatment of HIV-positive patients with early osteonecrosis of the femoral head

Abstract Objective To investigate the efficacy of treating patients with HIV-positive osteonecrosis of the femoral head using drilled decompression autologous bone marrow and allogeneic bone grafting. Methods 40 patients (44 hips) with early osteonecrosis of the femoral head treated by drilling decompression autologous bone marrow and allogeneic bone grafting since October 2015 were retrospectively analyzed, among which 20 patients (24 hips) were HIV-positive patients with early osteonecrosis of the femoral head, 16 males and 4 females, age 22–43 years, average 39.6 ± 10.18 years, and 20 patients (20 hips) in the same period HIV-negative early osteonecrosis of the femoral head patients, 13 males and 7 females, aged 48–78 years, mean 63.50 ± 7.94 years were negative controls. General information including ARCO stage, Harris score, VAS score, hematological indexes including CD4+ T lymphocyte count, and HIV viral load was recorded for all patients before surgery. All patients were operated on by drilling and decompression of the necrotic area, harvesting autologous iliac bone marrow with allogeneic bone, and bone grafting through the decompression channel. The patients were followed up regularly at 6, 12, and 24 months after surgery and annually thereafter, and the repair of the necrotic femoral head was observed by reviewing the frontal and lateral X-ray, CT or MRI of the hip joint, and the complications and functional recovery of the hip joint was counted and compared between the two groups. Results All patients were followed up, and the ARCO stages in the HIV-positive group were stage I 2 hips, stage IIA 6 hips, stage IIB 8 hips, stage IIC 6 hips, and stage III 2 hips, with a follow-up time of 12 to 60 months and a mean of 24.6 months. In the negative control group, there were 3 hips in ARCO stage I, 7 hips in stage IIA, 5 hips in stage IIB, 3 hips in stage IIC, and 2 hips in stage III, and the follow-up time ranged from 13 to 62 months, with an average of 24.8 months. The Harris score and VAS score of the hip in both groups improved significantly at 6 months postoperatively compared with those before surgery (P < 0.001). The difference between the Harris score of the hip in the positive group at 24 months postoperatively compared with that at 6 months postoperatively was statistically significant, but the VAS score at 24 months postoperatively compared with that at 6 months postoperatively was not statistically significant. In the negative group, there was no statistically significant difference in the Harris score and VAS score of the hip at 24 months postoperatively compared with those at 6 months postoperatively. In the positive group, there was a trend of continuous increase in hip BMD from the beginning of the postoperative period (P < 0.001). There was no statistically significant difference between the negative group and the positive group at the 24 months postoperatively follow-up except for the Harris score, which was statistically significant (P < 0.001), and the VAS score, which was statistically insignificant. At the 24 months postoperatively follow-up, patients in both groups had good recovery of hip function, and no complications such as vascular and nerve injury and fracture occurred during the perioperative period and follow-up period, and no complications related to incisional infection and pulmonary infection occurred during hospitalization. Conclusion The treatment of early HIV-positive osteonecrosis of the femoral head patients with autologous bone marrow and allogeneic bone grafting by drilling and decompression to remove the tissue in the necrotic area of the femoral head can effectively stop the process of osteonecrosis of the femoral head and promoting femoral head repair in HIV-positive patients is a safe and effective method for treating HIV-positive patients with early osteonecrosis of the femoral head, and can effectively delay or postpone total hip replacement in patients

Carrier Transport and Molecular Displacement Modulated dc Electrical Breakdown of Polypropylene Nanocomposites

Dielectric energy storage capacitors have advantages such as ultra-high power density, extremely fast charge and discharge speed, long service lifespan and are significant for pulsed power system, smart power grid, and power electronics. Polypropylene (PP) is one of the most widely used dielectric materials for dielectric energy storage capacitors. It is of interest to investigate how to improve its electrical breakdown strength by nanodoping and the influencing mechanism of nanodoping on the electrical breakdown properties of polymer nanocomposites. PP/Al2O3 nanocomposite dielectric materials with various weight fraction of nanoparticles are fabricated by melt-blending and hot-pressing methods. Thermally stimulated current, surface potential decay, and dc electrical breakdown experiments show that deep trap properties and associated molecular chain motion are changed by incorporating nanofillers into polymer matrix, resulting in the variations in conductivity and dc electrical breakdown field of nanocomposite dielectrics. Then, a charge transport and molecular displacement modulated electrical breakdown model is utilized to simulate the dc electrical breakdown behavior. It is found that isolated interfacial regions formed in nanocomposite dielectrics at relatively low loadings reduce the effective carrier mobility and strengthen the interaction between molecular chains, hindering the transport of charges and the displacement of molecular chains with occupied deep traps. Accordingly, the electrical breakdown strength is enhanced at relatively low loadings. Interfacial regions may overlap in nanocomposite dielectrics at relatively high loadings so that the effective carrier mobility decreases and the interaction between molecular chains may be weakened. Consequently, the molecular motion is accelerated by electric force, leading to the decrease in electrical breakdown strength. The experiments and simulations reveals that the influence of nanodoping on dc electrical breakdown properties may origin from the changes in the charge transport and molecular displacement characteristics caused by interfacial regions in nanocomposite dielectrics