Molecular Characterization of Cultivated Bromeliad Accessions with Inter-Simple Sequence Repeat (ISSR) Markers

Bromeliads are of great economic importance in flower production; however little information is available with respect to genetic characterization of cultivated bromeliads thus far. In the present study, a selection of cultivated bromeliads was characterized via inter-simple sequence repeat (ISSR) markers with an emphasis on genetic diversity and population structure. Twelve ISSR primers produced 342 bands, of which 287 (~84%) were polymorphic, with polymorphic bands per primer ranging from 17 to 34. The Jaccard’s similarity ranged from 0.08 to 0.89 and averaged ~0.30 for the investigated bromeliads. The Bayesian-based approach, together with the un-weighted paired group method with arithmetic average (UPGMA)-based clustering and the principal coordinate analysis (PCoA), distinctly grouped the bromeliads from Neoregelia, Guzmania, and Vriesea into three separately clusters, well corresponding with their botanical classifications; whereas the bromeliads of Aechmea other than the recently selected hybrids were not well assigned to a cluster. Additionally, ISSR marker was proven efficient for the identification of hybrids and bud sports of cultivated bromeliads. The findings achieved herein will further our knowledge about the genetic variability within cultivated bromeliads and therefore facilitate breeding for new varieties of cultivated bromeliads in future as well

Conditional müller cell ablation causes independent neuronal and vascular pathologies in a novel transgenic model

Müller cells are the major glia of the retina that serve numerous functions essential to retinal homeostasis, yet the contribution of Müller glial dysfunction to retinal diseases remains largely unknown. We have developed a transgenic model using a portion of the regulatory region of the retinaldehyde binding protein 1 gene for conditional Müller cell ablation and the consequences of primary Müller cell dysfunction have been studied in adult mice. We found that selective ablation of Müller cells led to photoreceptor apoptosis, vascular telangiectasis, blood-retinal barrier breakdown and, later, intraretinal neovascularization. These changes were accompanied by impaired retinal function and an imbalance between vascular endothelial growth factor-A (VEGF-A) and pigment epithelium-derived factor. Intravitreal injection of ciliary neurotrophic factor inhibited photoreceptor injury but had no effect on the vasculopathy. Conversely, inhibition of VEGF-A activity attenuated vascular leak but did not protect photoreceptors. Our findings show that Müller glial deficiency may be an important upstream cause of retinal neuronal and vascular pathologies in retinal diseases. Combined neuropro-tective and anti-angiogenic therapies may be required to treat Müller cell deficiency in retinal diseases and in other parts of the CNS associated with glial dysfunction

Testing and Analysis Method of Low Remanence Materials for Magnetic Shielding Device

Magnetic shielding devices with a grid structure of multiple layers of highly magnetically permeable materials (such as permalloy) can achieve remanent magnetic fields at the nanotesla (nT) level or even lower. The remanence of the material inside the magnetic shield, such as the building materials used in the support structure, can cause serious damage to the internal remanence of the magnetic shield. Therefore, it is of great significance to detect the remanence of the materials used inside the magnetic shielding device. The existing test methods do not limit the test environment, the test process is vulnerable to additional magnetic field interference and did not consider the real results of the material in the weak magnetic environment. In this paper, a novel method of measuring the remanence of materials in a magnetic shielding cylinder is proposed, which prevents the interference of the earth’s magnetic field and reduces the measurement error. This method is used to test concrete components, composite materials and metal materials commonly applicated in magnetic shielding devices and determine the materials that can be used for magnetic shielding devices with 1 nT, 10 nT and 100 nT as residual magnetic field targets

The dysregulation of Retinal Binding Protein 3 in the Conditional Müller Cell Ablation mouse model

Abstract of a presentation at ARVO 2014 meeting, 4-8 May, Orlando, Florida

Effects of Ranibizumab and Aflibercept on Human Müller Cells and Photoreceptors under Stress Conditions

Anti-vascular endothelial growth factor (VEGF) therapy has revolutionized the treatment of retinal vascular diseases. However, constitutive VEGF also acts as a trophic factor on retinal nonvascular cells. We have studied the effects of aflibercept and ranibizumab on human Müller cells and photoreceptors exposed to starvation media containing various concentrations of glucose, with or without CoCl2-induced hypoxia. Cell survival was assessed by calcein-AM cell viability assays. Expression of heat shock proteins (Hsp) and redox proteins thioredoxin 1 and 2 (TRX1, TRX2) was studied by Western blots. The production of neurotrophic factors in Müller cells and interphotoreceptor retinoid-binding protein (IRBP) in photoreceptors was measured by enzymelinked immunosorbent assays. Aflibercept and ranibizumab did not affect the viability of both types of cells. Neither aflibercept nor ranibizumab affected the production of neurotrophic factors or expression of Hsp60 and Hsp90 in Müller cells. However, aflibercept but not ranibizumab affected the expression of Hsp60, Hsp9, TRX1 and TRX2 in photoreceptors. Aflibercept and ranibizumab both inhibited the production of IRBP in photoreceptors, aflibercept more so than ranibizumab. Our data indicates that the potential influence of aflibercept and ranibizumab on photoreceptors should be specifically monitored in clinical studies

Tyrosine phosphorylation of VE-cadherin and claudin-5 is associated with TGF-β1-induced permeability of centrally derived vascular endothelium

Breakdown of the inner blood–retinal barrier and the blood–brain barrier is associated with changes in tight and adherens junction-associated proteins that link vascular endothelial cells. This study aimed to test the hypothesis that transforming growth factor (TGF)-β1 increases the paracellular permeability of vascular endothelial monolayers through tyrosine phosphorylation of VE-cadherin and claudin-5. Bovine retinal and human brain capillary endothelial cells were grown as monolayers on coated polycarbonate membranes. Paracellular permeability was studied by measuring the equilibration of 14C-inulin or fluorescence-labelled dextran. Changes in VE-cadherin and claudin-5 expression were studied by immunocytochemistry (ICC) and quantified by cell-based enzyme linked immunosorbent assays (ELISA). Tyrosine phosphorylation of VE-cadherin and claudin-5 was studied by ICC, immunoprecipitation and Western blotting. We found that exposure of endothelial cells to TGF-β1 caused a dose-dependent increase in paracellular permeability as reflected by increases in the equilibration of 14C-inulin. This effect was enhanced by the tyrosine phosphatase inhibitor orthovanadate and attenuated by the tyrosine kinase inhibitor lavendustin A. ICC and cell-based ELISA revealed that TGF-β1 induced both dose- and time-dependent decreases in VE-cadherin and claudin-5 expression. Assessment of cell viability indicated that changes in these junction-associated proteins were not due to endothelial death or injury. ICC revealed that tyrosine phosphorylation of endothelial monolayers was greatly enhanced by TGF-β1 treatment, and immunoprecipitation of cell lysates showed increased tyrosine phosphorylation of VE-cadherin and claudin-5. Our results suggest that tyrosine phosphorylation of VE-cadherin and claudin-5 is involved in the increased paracellular permeability of central nervous system-derived vascular endothelium induced by TGF-β1

Systemic administration of erythropoietin inhibits retinopathy in RCS rats.

OBJECTIVE: Royal College of Surgeons (RCS) rats develop vasculopathy as photoreceptors degenerate. The aim of this study was to examine the effect of erythropoietin (EPO) on retinopathy in RCS rats. METHODS: Fluorescein angiography was used to monitor retinal vascular changes over time. Changes in retinal glia and vasculature were studied by immunostaining. To study the effects of EPO on retinal pathology, EPO (5000 IU/kg) was injected intraperitoneally in 14 week old normal and RCS rats twice a week for 4 weeks. Changes in the retinal vasculature, glia and microglia, photoreceptor apoptosis, differential expression of p75 neurotrophin receptor (p75NTR), pro-neurotrophin 3 (pro-NT3), tumour necrosis factor-α (TNFα), pigment epithelium derived factor (PEDF) and vascular endothelial growth factor-A (VEGF-A), the production of CD34(+) cells and mobilization of CD34(+)/VEGF-R2(+) cells as well as recruitment of CD34(+) cells into the retina were examined after EPO treatment. RESULTS: RCS rats developed progressive capillary dropout and subretinal neovascularization which were accompanied by retinal gliosis. Systemic administration of EPO stabilized the retinal vasculature and inhibited the development of focal vascular lesions. Further studies showed that EPO modulated retinal gliosis, attenuated photoreceptor apoptosis and p75NTR and pro-NT3 upregulation, promoted the infiltration of ramified microglia and stimulated VEGF-A expression but had little effect on TNFα and PEDF expression. EPO stimulated the production of red and white blood cells and CD34(+) cells along with effective mobilization of CD34(+)/VEGF-R2(+) cells. Immunofluorescence study demonstrated that EPO enhanced the recruitment of CD34+ cells into the retina. CONCLUSIONS: Our results suggest that EPO has therapeutic potentials in treatment of neuronal and vascular pathology in retinal disease. The protective effects of EPO on photoreceptors and the retinal vasculature may involve multiple mechanisms including regulation of retinal glia and microglia, inhibition of p75NTR-pro-NT3 signaling together with stimulation of production and mobilization of bone marrow derived cells

Identification of novel diabetes impaired miRNA-transcription factor co-regulatory networks in bone marrow-derived Lin-/VEGF-R2+ endothelial progenitor cells

Endothelial progenitor cells (EPCs) are a group of rare cells that play an important role in the repair of injured vascular endothelial cells and assist in reperfusion of ischemic tissue. Decreased production and/or loss of function of EPCs are associated with diabetic vasculopathy. The molecular mechanisms by which diabetes impairs EPCs remain unclear. We conducted microarray experiments followed by integrative regulatory analysis on cells isolated from Akita diabetic mice (18-weeks after onset of diabetes) and age-matched non-diabetic controls. Two types of cells were isolated from mice bone marrow; Lin+ cells and Lin-/VEGF-R2+ EPCs. RNA was hybridized to mouse WG-6 V2 beadchips followed by comprehensive gene network analysis and computational validation of the obtained results. In total, 80 genes were exclusively DE between non-diabetic Lin-/VEGF-R2+ EPCs and diabetic Lin-/VEGF-R2+ EPCs, of which the 3 genes Clcnka, Pik3c2a, and Ptf1a are known to be associated with diabetic complications. Further analysis led to the establishment of a TF-miRNA mediated regulatory network specific to diabetic Lin-/VEGF-R2+ EPCs and to identify 11 central-hub TFs (Tbp, Ahr, Trp53, Gata1, Foxo1, Foxo4, Yy1, Max, Pparg, Myc, Cebpa), and 2 miRNAs (mir-139-5p, mir-709) that might act as putative genomic drivers of diabetic pathogenesis in Lin-/VEGF-R2+ EPCs. Moreover, we identified multiple TF-miRNA co-regulatory network motifs for which we validated their contribution to diabetic Lin-/VEGF-R2+ EPCs in terms of statistical significance and relevance to biological evidence. Our findings suggest that diabetic Lin-/VEGF-R2+ EPCs have specifically altered signature genes and miRNAs that render their capacity to proliferate and differentiate