Morpho–physiological responses of Rocket (<Emphasis Type="Italic">Eruca sativa</Emphasis> L.) varieties to sodium sulfate (Na<Subscript>2</Subscript>SO<Subscript>4</Subscript>) stress: an experimental approach

Rocket (Eruca sativa L.) is a medicinal plant that belongs to the Brassicaceae family and was reported to be a tolerant plant under soil salinity as well as high genetic diversity among its varieties. Since morphological and physiological changes to sodium sulfate stress toward this plant have not been investigated yet, the present study was implemented to assess the response of rocket (Eruca sativa L.) varieties to sodium sulfate (Na2SO4) stress as well as the relationship among these traits. Two varieties of rocket plants, the Iranian and Italian ones, were subjected to four salinity (Na₂SO₄) treatments [0 (control), 15, 30 and 60 mM of Na₂SO₄ solution] and three growth stages (49, 65, and 74 days) in factorial experiment with completely randomized design and three replications were considered. Some morphologic traits such as grain yield were measured during the growth period. The results of the analysis of variances between the mentioned variables indicated a significant difference between the varieties in terms of K⁺, Na⁺, Na⁺/K⁺, leaf length, grain yield, organic and mineral matter. The results of correlation and regression of the amounts of K⁺ showed a linear relationship with the grain yield and its variations were not independent from the variations of grain yield. Eventually, it seems that the Italian variety was more tolerant and having better performance in comparison with the Iranian variety, in response to salt stress.     

Evaluation of known and novel inhibitors of Orai1-mediated store operated Ca2+ entry in MDA-MB-231 breast cancer cells using a Fluorescence Imaging Plate Reader assay

The Orai1 Ca²⁺ permeable ion channel is an important component of store operated Ca²⁺ entry (SOCE) in cells. It’s over-expression in basal molecular subtype breast cancers has been linked with poor prognosis, making it a potential target for drug development. We pharmacologically characterised a number of reported inhibitors of SOCE in MDA-MB-231 breast cancer cells using a convenient Fluorescence Imaging Plate Reader (FLIPR) assay, and show that the rank order of their potencies in this assay is the same as those reported in a wide range of published assays. The assay was also used in a screening project seeking novel inhibitors. Following a broad literature survey of classes of calcium channel inhibitors we used simplified ligand structures to query the ZINC on-line database, and following two iterations of refinement selected a novel Orai1-selective dichlorophenyltriazole hit compound. Analogues of this were synthesized and evaluated in the FLIPR assay to develop structure–activity relationships (SAR) for the three domains of the hit; triazole (head), dichlorophenyl (body) and substituted phenyl (tail). For this series, the results suggested the need for a lipophilic tail domain and an out-of-plane twist between the body and tail domains.     

Metal ion specificities for folding and cleavage activity in the Schistosoma hammerhead ribozyme

The effects of various metal ions on cleavage activity and global folding have been studied in the extended Schistosoma hammerhead ribozyme. Fluorescence resonance energy transfer was used to probe global folding as a function of various monovalent and divalent metal ions in this ribozyme. The divalent metals ions Ca²⁺, Mg²⁺, Mn²⁺, and Sr²⁺ have a relatively small variation (less than sixfold) in their ability to globally fold the hammerhead ribozyme, which contrasts with the very large difference (>10,000-fold) in apparent rate constants for cleavage for these divalent metal ions in single-turnover kinetic experiments. There is still a very large range (>4600-fold) in the apparent rate constants for cleavage for these divalent metal ions measured in high salt (2 M NaCl) conditions where the ribozyme is globally folded. These results demonstrate that the identity of the divalent metal ion has little effect on global folding of the Schistosoma hammerhead ribozyme, whereas it has a very large effect on the cleavage kinetics. Mechanisms by which the identity of the divalent metal ion can have such a large effect on cleavage activity in the Schistosoma hammerhead ribozyme are discussed.     

Differences and similarities between mesenchymal stem cell and endothelial progenitor cell immunoregulatory properties against T cells

Bone-marrow-derived mesenchymal stem cells and endothelial progenitor cells have some interesting biological properties that make them unique for cell therapy of degenerative and cardiovascular disorders. Although both cell populations have been already studied and used for their regenerative potentials, recently their special immunoregulatory features have brought much more attention. Mesenchymal stem cells and endothelial progenitor cells have both proangiogenic functions and have been shown to suppress the immune response, particularly T cell proliferation, activation, and cytokine production. This makes them suitable choices for allogeneic stem cell transplantation. Nevertheless, these two cells do not have equal immunoregulatory activities. Many elements including their extraction sources, age/passage, expression of different markers, secretion of bioactive mediators, and some others could change the efficiency of their immunosuppressive function. However, to our knowledge, no publication has yet compared mesenchymal stem cells and endothelial progenitor cells for their immunological interaction with T cells. This review aims to specifically compare the immunoregulatory effect of these two populations including their Tcell suppression, deactivation, cytokine production, and regulatory T cells induction capacities. Moreover, it evaluates the implications of the tumor necrosis factor alpha-tumor necrosis factor receptor 2 axis as an emerging immune checkpoint signaling pathway controlling most of their immunological properties.     

An Ex Vivo Model for Anti-Angiogenic Drug Testing on Intact Microvascular Networks

New models of angiogenesis that mimic the complexity of real microvascular networks are needed. Recently, our laboratory demonstrated that cultured rat mesentery tissues contain viable microvascular networks and could be used to probe pericyte-endothelial cell interactions. The objective of this study was to demonstrate the efficacy of the rat mesentery culture model for anti-angiogenic drug testing by time-lapse quantification of network growth. Mesenteric windows were harvested from adult rats, secured in place with an insert, and cultured for 3 days according to 3 experimental groups: 1) 10% serum (angiogenesis control), 2) 10% serum + sunitinib (SU11248), and 3) 10% serum + bevacizumab. Labeling with FITC conjugated BSI-lectin on Day 0 and 3 identified endothelial cells along blood and lymphatic microvascular networks. Comparison between day 0 (before) and 3 (after) in networks stimulated by 10% serum demonstrated a dramatic increase in vascular density and capillary sprouting. Growing networks contained proliferating endothelial cells and NG2+ vascular pericytes. Media supplementation with sunitinib (SU11248) or bevacizumab both inhibited the network angiogenic responses. The comparison of the same networks before and after treatment enabled the identification of tissue specific responses. Our results establish, for the first time, the ability to evaluate an anti-angiogenic drug based on time-lapse imaging on an intact microvascular network in an ex vivo scenario.     

Association of two groups of phytoplasma with various symptoms in some wooden and herbaceous plants

During 2015–2016, wooden and herbaceous plants growing in parks, boulevards, fields, gardens and forests in Khuzestan province, southwestern Iran, were visually inspected for symptoms resembling phytoplasma. Fifty‐one symptomatic samples from nine different species and one symptomless sample from each plant were collected. Leaf midribs, petioles and the parts of stem cambium were separated and freeze‐dried. Total DNA was extracted using CTAB‐based method and tested for phytoplasma using a nested PCR assay. The expected size amplicons of 16S rDNA were sequenced and compared to those of reference phytoplasmas by BLASTn search and phylogenetic analysis. The consensus 16S rDNA sequence of the detected phytoplasma in narrow cattail related to reference phytoplasma group 16SrVI, “Candidatus Phytoplasma trifolii” while in the other plants were related to reference phytoplasma subgroup 16SrII–D, “Candidatus Phytoplasma aurantifolia.” All isolates showed 98%–99% sequence identity to members of their reference groups. To our knowledge, this is the first report of “Candidatus Phytoplasma aurantifolia”‐related strains infecting the plants of Acacia salicina, Alternanthera ficoidea, Melaleuca citrine, Citrus aurantium throughout the world and Celosia christata in Iran. Furthermore, this study is the first to report the association of a “Candidatus Phytoplasma trifolii”‐related strain with Typha angustifolia worldwide.     

An SAR study of hydroxy-trifluoromethylpyrazolines as inhibitors of Orai1-mediated store operated Ca2+ entry in MDA-MB-231 breast cancer cells using a convenient Fluorescence Imaging Plate Reader assay

The proteins Orai1 and STIM1 control store-operated Ca²⁺ entry (SOCE) into cells. SOCE is important for migration, invasion and metastasis of MDA-MB-231 human triple negative breast cancer (TNBC) cells and has been proposed as a target for cancer drug discovery. Two hit compounds from a medium throughput screen, displayed encouraging inhibition of SOCE in MDA-MB-231 cells, as measured by a Fluorescence Imaging Plate Reader (FLIPR) Ca²⁺ assay. Following NMR spectroscopic analysis of these hits and reassignment of their structures as 5-hydroxy-5-trifluoromethylpyrazolines, a series of analogues was prepared via thermal condensation reactions between substituted acylhydrazones and trifluoromethyl 1,3-dicarbonyl arenes. Structure-activity relationship (SAR) studies showed that small lipophilic substituents at the 2- and 3-positions of the RHS and 2-, 3- and 4-postions of the LHS terminal benzene rings improved activity, resulting in a novel class of potent and selective inhibitors of SOCE.     

Disturbance-free rapid solution exchange for magnetic tweezers single-molecule studies

Single-molecule manipulation technologies have been extensively applied to studies of the structures and interactions of DNA and proteins. An important aspect of such studies is to obtain the dynamics of interactions; however the initial binding is often difficult to obtain due to large mechanical perturbation during solution introduction. Here, we report a simple disturbance-free rapid solution exchange method for magnetic tweezers single-molecule manipulation experiments, which is achieved by tethering the molecules inside microwells (typical dimensions–diameter (D): 40–50 μm, height (H): 100 μm; H:D∼2:1). Our simulations and experiments show that the flow speed can be reduced by several orders of magnitude near the bottom of the microwells from that in the flow chamber, effectively eliminating the flow disturbance to molecules tethered in the microwells. We demonstrate a wide scope of applications of this method by measuring the force dependent DNA structural transitions in response to solution condition change, and polymerization dynamics of RecA on ssDNA/SSB-coated ssDNA/dsDNA of various tether lengths under constant forces, as well as the dynamics of vinculin binding to α-catenin at a constant force (< 5 pN) applied to the α-catenin protein.