Iron transport in the genus Marinobacter

Marinobacter belong to the class of Gammaproteobacteria and these motile, halophilic or halotolerent bacteria are widely distributed throughout the world’s oceans having been isolated from a wide variety of marine environments. They have also been identified as members of the bacterial flora associated with other marine organisms. Here, using a combination of natural products chemistry and genomic analysis, we assess the nature of the siderophores produced by this genus and their potential relationship to phylogeny and lifestyle/ecological niche of this diverse group of organisms. Our analysis shows a wide level of diversity in siderophore based iron uptake systems among this genus with three general strategies: (1) production and utilization of native siderophores in addition to utilization of a variety of exogenous ones, (2) production and utilization of native siderophores only, (3) lack of siderophore production but utilization of exogenous ones. They all share the presence of at least one siderophore-independent iron uptake ABC transport systems of the FbpABC iron metal type and lack the ability for direct transport of ferrous iron. Siderophore production and utilization can be correlated with phylogeny and thus it forms a type of chemotaxonomic marker for this genus.     

Incorporation of SPION-casein core-shells into silk-fibroin nanofibers for cardiac tissue engineering

Mimicking the structure of extracellular matrix (ECM) of myocardium is necessary for fabrication of functional cardiac tissue. The superparamagnetic iron oxide nanoparticles (SPIONs, Fe₃O₄), as new generation of magnetic nanoparticles (NPs), are highly intended in biomedical studies. Here, SPION NPs (1 wt%) were synthesized and incorporated into silk-fibroin (SF) electrospun nanofibers to enhance mechanical properties and topography of the scaffolds. Then, the mouse embryonic cardiac cells (ECCs) were seeded on the scaffolds for in vitro studies. The SPION NPs were studied by scanning electron microscope (SEM), X-ray diffraction (XRD), and transmission electron microscope (TEM). SF nanofibers were characterized after incorporation of SPIONs by SEM, TEM, water contact angle measurement, and tensile test. Furthermore, cytocompatibility of scaffolds was confirmed by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. SEM images showed that ECCs attached to the scaffolds with elongated morphologies. Also, the real-time PCR and immunostaining studies approved upregulation of cardiac functional genes in ECCs seeded on the SF/SPION-casein scaffolds including GATA-4, cardiac troponin T, Nkx 2.5, and alpha-myosin heavy chain, compared with the ones in SF. In conclusion, incorporation of core-shells in SF supports cardiac differentiation, while has no negative impact on ECCs' proliferation and self-renewal capacity.     

Genetic Deletion of Mst1 Alters T Cell Function and Protects against Autoimmunity

Mammalian sterile 20-like kinase 1 (Mst1) is a MAPK kinase kinase kinase which is involved in a wide range of cellular responses, including apoptosis, lymphocyte adhesion and trafficking. The contribution of Mst1 to Ag-specific immune responses and autoimmunity has not been well defined. In this study, we provide evidence for the essential role of Mst1 in T cell differentiation and autoimmunity, using both genetic and pharmacologic approaches. Absence of Mst1 in mice reduced T cell proliferation and IL-2 production in vitro, blocked cell cycle progression, and elevated activation-induced cell death in Th1 cells. Mst1 deficiency led to a CD4⁺ T cell development path that was biased toward Th2 and immunoregulatory cytokine production with suppressed Th1 responses. In addition, Mst1^−/− B cells showed decreased stimulation to B cell mitogens in vitro and deficient Ag-specific Ig production in vivo. Consistent with altered lymphocyte function, deletion of Mst1 reduced the severity of experimental autoimmune encephalomyelitis (EAE) and protected against collagen-induced arthritis development. Mst1^−/− CD4⁺ T cells displayed an intrinsic defect in their ability to respond to encephalitogenic antigens and deletion of Mst1 in the CD4⁺ T cell compartment was sufficient to alleviate CNS inflammation during EAE. These findings have prompted the discovery of novel compounds that are potent inhibitors of Mst1 and exhibit desirable pharmacokinetic properties. In conclusion, this report implicates Mst1 as a critical regulator of adaptive immune responses, Th1/Th2-dependent cytokine production, and as a potential therapeutic target for immune disorders.     

Biochemical and Physiological Responses of Cucumis sativus Cultivars to Different Combinations of Low-Temperature and High Humidity

Low-temperature and high humidity are typical environmental factors in the plastic tunnel and solar greenhouse during the cold season that restricts plant growth and development. Herein, we investigated the impact of different combinations of low-temperature and high humidity (day/night: T1 15/10 °C?+?95%, T2 12/8 °C?+?95%, and T3 9/5 °C?+?95%) along with a control (CK 25/18 °C?+?80%) on cucumber cultivars viz: Zhongnong37 (ZN37: resistant) and Shuyanbailv (SYB: sensitive). The low-temperature and high humidity stresses increased electrolyte leakage (EL), malondialdehyde (MDA), hydrogen peroxide (H₂O₂) and intercellular concentration of carbon dioxide (Ci), and reduced morphological indices, relative water content (RWC), net photosynthesis rate (Pn), stomatal conductance (Gs), transpiration rate (E) and leaf pigments in both cultivars as compared to control (CK). Superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), and glutathione reductase (GR) were decreased in cv. SYB under stress conditions as compared to cv. ZN37. Low-temperature and high humidity treatments showed an increase in proline and soluble protein content in cv. ZN37 as compared to cv. SYB. Abscisic acid (ABA) and jasmonic acid (JA) were augmented while indole-3-acetic acid (IAA), zeatin (ZT), zeatin riboside (ZR), and gibberellic acid (GA) were decreased in both cultivars. Under T3 (9/5 °C?+?95%), Pn, protoporphyrin, and ZT were extremely decreased by 71.3%, 74.3%, and 82.4%, respectively, in cv. SYB compared to control. Moreover, principal component analysis (PCA) based on physiochemical traits confirmed that cv. ZN37 had the strongest correlation with antioxidant enzymes, proline, and soluble protein content than cv. SYB under low-temperature and high humidity treatments. Our results suggest that a stress-tolerant cultivar mitigates stress damage in cucumber transplants by regulating photosynthetic efficiency, antioxidant capacity and hormonal profile when compared to a stress-sensitive cultivar.

     

Exploring the Therapeutic Properties of Alga-Based Silver Nanoparticles: Anticancer,Antibacterial, and Free Radical Scavenging Capabilities

The current study was designed to evaluate the antioxidant, anticancer and antimicrobial activities of silver nanoparticles (AgNPs) biosynthesized by Spirulina platensis extract. The biosynthesized silver nanoparticles were characterized using Fourier transform infrared (FT-IR) analysis, scanning electron microscopy (SEM), transmission electron microscopy (TEM) and X-ray diffraction (XRD) analysis. The antioxidant activity of the biosynthesized AgNPs were determined via DPPH radical scavenging assay while its anticancer activity was determined using the MTT assay. The antimicrobial activity of the biosynthesized AgNPs were analyzed by disc diffusion method. Spirulina platensis acts as a reducing and capping agent. The efficacy of silver nanoparticles (AgNPs) in inhibiting the growth of Gram-negative bacteria, specifically Acetobacter, Klebsiella, Proteus vulgaris, and Pseudomonas aeruginosa, was assessed by the utilisation of the diffusion method. The study aimed to evaluate the efficacy of biosynthesized silver nanoparticles (AgNPs) against many strains of Pseudomonas aeruginosa bacteria. The findings of the study revealed that when administered in doses of 50 μl, 75 μl, and 100 μl, the largest observed zone of inhibition corresponded to measurements of 10.5 mm, 14 mm, and 16 mm, respectively. A zone of inhibition with dimensions of 8 mm, 10.5 mm, and 12 mm was detected during testing against Acetobacter at concentrations of 50 μl, 75 μl, and 100 μl, respectively. The findings also indicate that there is a positive correlation between the concentration of AgNP and the DPPH scavenging ability of silver nanoparticles. The percentage of inhibition observed at concentrations of 500 μg/ml, 400 μg/ml, 300 μg/ml, 200 μg/ml, and 100 μg/ml were recorded as 80±1.98, 61±1.98, 52±1.5, 42±1.99, and 36±1.97, respectively. In addition, it was observed that the silver nanoparticles exhibited the greatest antioxidant activity at a concentration of 500 g/ml, with a measured value of 80.89±1.99. The IC-50 values, representing the inhibitory concentration required to achieve 50 % inhibition, were found to be 8.16, 19.15, 30.14, 41.13, and 63.11 at inhibition levels of 36±1.97, 42±1.99, 52±1.5, 61±1.98, and 80±1.98, respectively.     

Topographic regulation of neuronal intermediate filaments by phosphorylation, role of peptidyl-prolyl isomerase 1: significance in neurodegeneration

The neuronal cytoskeleton is tightly regulated by phosphorylation and dephosphorylation reactions mediated by numerous associated kinases, phosphatases and their regulators. Defects in the relative kinase and phosphatase activities and/or deregulation of compartment-specific phosphorylation result in neurodegenerative disorders. The largest family of cytoskeletal proteins in mammalian cells is the superfamily of intermediate filaments (IFs). The neurofilament (NF) proteins are the major IFs. Aggregated forms of hyperphosphorylated tau and phosphorylated NFs are found in pathological cell body accumulations in the central nervous system of patients suffering from Alzheimer’s disease, Parkinson’s disease, and Amyotrophic Lateral Sclerosis. The precise mechanisms for this compartment-specific phosphorylation of cytoskeletal proteins are not completely understood. In this review, we focus on the mechanisms of neurofilament phosphorylation in normal physiology and neurodegenerative diseases. We also address the recent breakthroughs in our understanding the role of different kinases and phosphatases involved in regulating the phosphorylation status of the NFs. In addition, special emphasis has been given to describe the role of phosphatases and Pin1 in phosphorylation of NFs.     

Chemoreceptors in signalling complexes: shifted conformation and asymmetric coupling

Bacterial chemotaxis is mediated by signalling complexes of chemoreceptors, histidine kinase CheA and coupling protein CheW. Interactions in complexes profoundly affect the kinase. We investigated effects of these interactions on chemoreceptors by comparing receptors alone and in complexes. Assays of initial rates of methylation indicated that signalling complexes shifted receptor conformation towards the methylation-on, higher-ligand-affinity, kinase-off state, tuning receptors for greater sensitivity. In contrast, transmembrane and conformational signalling within chemoreceptors was essentially unaltered, consistent with other evidence identifying receptor dimers as the fundamental units of such signalling. In signalling complexes, coupling of ligand binding to kinase activity is cooperative and the dynamic range of kinase control expanded > 100-fold by receptor adaptational modification. We observed no cooperativity in influence of ligand on receptor conformation, only on kinase activity. However, receptor modification generated increased dynamic range in a stepwise fashion, partly in coupling ligand to receptor conformation and partly in coupling receptor conformation to kinase activity. Thus, receptors and kinase were not equivalently affected by interactions in signalling complexes or by ligand binding and adaptational modification, indicating asymmetrical coupling between them. This has implications for mechanisms of precise adaptation. Coupling might vary, providing a previously unappreciated locus for sensory control.     

Establishment and Biological Characterization of a Panel of Glioblastoma Multiforme (GBM) and GBM Variant Oncosphere Cell Lines

Objective

Human tumor cell lines form the basis of the majority of present day laboratory cancer research. These models are vital to studying the molecular biology of tumors and preclinical testing of new therapies. When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity. Using a modified neural stem cell culture technique, here we report the characterization of GBM cell lines including GBM variants.

Methods

Tumor tissue samples were obtained intra-operatively and cultured in neural stem cell conditions containing growth factors. Tumor lines were characterized in vitro using differentiation assays followed by immunostaining for lineage-specific markers. In vivo tumor formation was assayed by orthotopic injection in nude mice. Genetic uniqueness was confirmed via short tandem repeat (STR) DNA profiling.

Results

Thirteen oncosphere lines derived from GBM and GBM variants, including a GBM with PNET features and a GBM with oligodendroglioma component, were established. All unique lines showed distinct genetic profiles by STR profiling. The lines assayed demonstrated a range of in vitro growth rates. Multipotency was confirmed using in vitro differentiation. Tumor formation demonstrated histologic features consistent with high grade gliomas, including invasion, necrosis, abnormal vascularization, and high mitotic rate. Xenografts derived from the GBM variants maintained histopathological features of the primary tumors.

Conclusions

We have generated and characterized GBM suspension lines derived from patients with GBMs and GBM variants. These oncosphere cell lines will expand the resources available for preclinical study.