Regulation of TrkB activity by PTPN11 is mediated through transmembrane protein Caveolin in the inner retina

Empirical thesis.Bibliography: pages 69-89.Chapter 1. Introduction -- Chapter 2. Methods -- Chapter 3. Results -- Chapter 4. Discussion -- Conclusion -- References.Brain derived neurotrophic factor (BDNF) and its high affinity receptor tropomyosin-related kinase receptor B (TrkB) play a protective role in the survival of retinal ganglion cells (RGCs) in healthy and disease states. SH2 domain tyrosine phosphatase PTPN11 (Shp2) is a ubiquitously expressed tyrosine phosphatase that regulates TrkB receptor and its activation is mediated through its interactions with the adapter protein Caveolin (Cav-1). This study determines the involvement of Cav-1 in facilitating Shp2 mediated inner retinal effects by genetically upregulating Shp2 expression in Cav-1 knockout mice.Shp2 was over-expressed in mice retina (n=38) through intravitreal injection of recombinant adeno-associated virus vector (AAV2). Wildtype and Cav-1 transgenic mice were used and for each animal one eye was transduced with PTPN11 transgene whereas GFP expression in the contralateral eye was used as control. Retinal functional changes were assessed by electroretinogram (ERG) and scotopic threshold response (STR) recordings. In vivo imaging using optical coherence tomography (OCT) and histological analysis were used to evaluate structural changes in the retinal laminar structure.Shp2 upregulation resulted in significantly reduced STR amplitude in both WT and Cav-1+/- mice (p<0.001 and p<0.01 respectively). Cav-1-/- group on the other hand demonstrated only a slight loss of STR amplitudes upon Shp2 upregulation that was statistically insignificant. Cellular density in the ganglion cell layer (GCL) was also significantly reduced in WT and Het groups (p<0.001 and p<0.01 respectively). Shp2 upregulation did not result in a significant decrease in the GCL density in Cav-1-/- mice.Together, these results suggest a novel role of Cav-1 in mediating Shp2 actions in retina particularly RGCs. Loss of Cav-1 exerts a protective effect particularly on the inner retinal function and structure. Future studies will help establish the pathophysiological cross talk between these two proteins.Mode of access: World wide web1 online resource (ix, 89 pages) colour illustration

Iron Status in Diffuse Telogen Hair Loss among Women

The relationship between iron body status and different types of hair loss has been investigated in a number of studies, however, with relatively discrepant findings. Therefore we conducted an analytical case-control study to assess whether diffuse telogen hair loss in women of childbearing age (15 to 45 years old) is associated with iron deficiency. Using the analytical case-control methodology, we studied 30 consecutive women with documented diffuse telogen hair loss in comparison with 30 women without hair loss. Study subjects had no history of nutritional supplement intake or chronic underlying diseases, and had normal thyroid function and inflammatory profiles. Biochemical investigations were performed in all study women. The mean ferritin level and trasferrin saturation was statistically significantly lower in patients with diffuse telogen hair loss than in subjects without hair loss (16.3+/-12.6 vs. 60.3+/-50.1, ng/mL;

Association of Circulating Procoagulant Microvesicles with Painful Vaso-Occlusive Crisis in Sickle Cell Disease

Introduction: Thrombotic complication is one of the features of sickle cell disease (SCD), characterized by appearance of phosphatidylserine on the outer membrane of sickle-shaped red blood cells and most abundantly on membrane protrusions called microvesicles (MVs). However, the exact mechanism by which MVs may enhance coagulant activity in SCD patients has not been fully addressed. The aim of this study was to further investigate the procoagulant activity of circulating MVs in sickle cell crises. Materials and Methods: Subjects included in this cross-sectional study were 47 patients with SCD and 25 normal subjects with written informed consent obtained from all the participants. MV analysis was conducted by using CD61, CD235 alpha, and Annexin-V monoclonal antibodies. The coagulant activity of MVs was determined by an ELISA-based procoagulant activity assay. Results: The majority of MVs were originated from platelets (CD61+) and erythrocytes (CD235+). These MVs demonstrated significantly enhanced levels during the painful crisis when compared with the steady-state period (p &amp;lt; 0.001) and controls (p &amp;lt; 0.001). Also, the procoagulant activity of MVs was significantly higher in crisis compared to those of steady state (p &amp;lt; 0.001) and positively correlated with the number of Annexin-V+ MVs (p &amp;lt; 0.001). Significant correlations were found between erythrocyte-derived MVs with hemolysis marker (r = 0.51, p &amp;lt; 0.001) and the hemoglobin level (r = -0.63, p &amp;lt; 0.001). Conclusion: The numbers of platelet- and erythrocyte-derived MVs are related to painful crisis, and their quantification in SCD may be helpful for identifying cases at increased risk of thrombotic complications

Development of Perphenazine-Loaded Solid Lipid Nanoparticles: Statistical Optimization and Cytotoxicity Studies

Objective. Perphenazine (PPZ), as a typical antipsychotic medical substance, has the same effectiveness compared to atypical antipsychotic medications for the treatment of schizophrenia. Despite the lipophilic essence, PPZ encounters limited bioavailability caused by the first-pass metabolism following oral administration. In the present study, PPZ-containing solid lipid nanoparticles (PPZ-SLNs) were prepared and optimized based on different factors, including lipid and surfactant amount, to develop appropriate and safe novel oral dosage forms of PPZ. Methods. The solvent emulsification-evaporation method was utilized to form SLNs by using soybean lecithin, glycerol monostearate (GMS), and Tween 80. Statistical optimization was done by the Box-Behnken design method to achieve formulation with optimized particle size, entrapment efficiency, and zeta potential. Also, transmission electron microscopy, in vitro release behavior, differential scanning calorimetry (DSC), and powder X-ray diffractometry (P-XRD) studies and cytotoxicity studies were assessed. Results. Optimization exhibited the significant effect of various excipients on SLN characteristics. Our finding indicated that the mean particle size, zeta potential, and entrapment efficiency of optimized PPZ-SLN were, respectively, 104±3.92 nm, −28±2.28 mV, and 83%±1.29. Drug release of PPZ-SLN was observed to be greater than 90% for 48 h that emphasized a sustained-release pattern. The DSC and P-XRD studies revealed the amorphous state of PPZ-SLN. FTIR spectra showed no incompatibility between the drug and the lipid. Performing cytotoxicity studies indicated no significant cytotoxicity on HT-29 cell culture. Conclusion. Our study suggests that PPZ-SLNs can make a promising vehicle for a suitable therapy of schizophrenia for the oral drug delivery system

Effect of C/N Ratio and Media Optimization through Response Surface Methodology on Simultaneous Productions of Intra- and Extracellular Inulinase and Invertase from Aspergillus niger ATCC 20611

The study is to identify the extraction of intracellular inulinase (exo- and endoinulinase) and invertase as well as optimization medium composition for maximum productions of intra- and extracellular enzymes from Aspergillus niger ATCC 20611. From two different methods for extraction of intracellular enzymes, ultrasonic method was found more effective. Response surface methodology (RSM) with a five-variable and three-level central composite design (CCD) was employed to optimize the medium composition. The effect of five main reaction parameters including sucrose, yeast extract, NaNO3, Zn+2, and Triton X-100 on the production of enzymes was analyzed. A modified quadratic model was fitted to the data with a coefficient of determination (R2) more than 0.90 for all responses. The intra-extracellular inulinase and invertase productions increased in the range from 16 to 8.4 times in the optimized medium (10% (w/v) sucrose, 2.5% (w/v) yeast extract, 2% (w/v) NaNO3, 1.5 mM (v/v) Zn+2, and 1% (v/v) Triton X-100) by RSM and from around 1.2 to 1.3 times greater than in the medium optimized by one-factor-at-a-time, respectively. The results of bioprocesses optimization can be useful in the scale-up fermentation and food industry

Computational analysis unravels novel destructive single nucleotide polymorphisms in the non-synonymous region of human caveolin gene

The caveolin (Cav) family of proteins comprises key constituents of caveolar structures in cell membranes, which are involved in receptor-independent endocytosis and cellular signalling pathways. Three isoforms of caveolin viz. cav-1, cav-2 and cav-3 encoded by CAV1, CAV2 and CAV3 genes respectively, have been reported to be expressed in various tissues. Genetic polymorphism in CAV has been identified as associated with the development of pathological changes in the cardiovascular system, chronic kidney disease as well as neurodegenerative diseases of the brain and retina such as Alzheimer\u27s disease, Parkinson\u27s disease and glaucoma. In this study, we investigate and characterise various polymorphisms associated with CAV1, CAV2 and CAV3 by using a combination of in silico algorithms such as SIFT, Polyphen 2.0, I-Mutant, PROVEAN, PANTHER, SNP&Go, PhD-SNP, MutPred and SNPEffects. Three-dimensional comparative modelling was performed using Phyre2 server, ab initio modelling, using the I-TASSER and RaptorX program. The predicted models were evaluated using Ramachandran plot to establish the accuracy of the models generated. The resulting mutant and wild type proteins obtained were energy minimized in Swiss Deep Viewer and evaluated. The study has identified two of the non-synonymous single nucleotide polymorphism (nsSNP) in CAV3 gene that may have a damaging effect on the protein stability. The surface residues in the wild type and mutant forms highlight different accessible surface area (ASA) of amino acid residues in the corresponding proteins. Our analysis predicted that none of the known nsSNPs have a negative effect on the CAV1 and CAV2 protein structures. Phylogenetic analysis using ConSurf further identified that most of the disease-associated nsSNPs were within the conserved regions in human cav3

Control of Cutaneous Leishmaniasis Using Geographic Information Systems from 2010 to 2014 in Khuzestan Province, Iran - Fig 1

<p>(A) Incidence of cutaneous leishmaniasis (CL) using geographical information system in (GIS) 2010, (B) GIS showed a new focus of CL in 2011 compared to 2010 (arrow), (C) The highest incidence of CLin 2012 with 1363 patients of CL has shown in Khuzestan Province with GIS mapping, (D) Geographical distribution mapping of CL incidence in 2013, E: The incidence rate of CL was diminished after control programs. Ten distinct had no reported of the disease in 2014.</p

Bexarotene modulates retinoid-x-receptor expression and is protective against neurotoxic endoplasmic reticulum stress response and apoptotic pathway activation

Retinoid X-receptors (RXRs) are members of the ligand-dependent transcription factor family of nuclear receptors that have gained recent research focus as potential targets for neurodegenerative disorders. Bexarotene is an RXR pharmacological agonist that is shown to be neuroprotective through its effects in promoting amyloid beta (Aβ) uptake by the glial cells in the brain. This study aimed to evaluate the dose-dependent effects of bexarotene on RXR expression in SH-SY5Y neuroblastoma cells and validate the drug effects in the brain in vivo. The protein expression studies were carried out using a combination of various drug treatment paradigms followed by expression analysis using Western blotting and immunofluorescence. Our study demonstrated that bexarotene promoted the expression of RXR α, β and γ isoforms at optimal concentrations in the cells and in the mice brain. Interestingly, a decreased RXR expression was identified in Alzheimer\u27s disease mouse model and in the cells that were treated with Aβ. Bexarotene treatment not only rescued the RXR expression loss caused by Aβ treatment (p \u3c 0.05) but also protected the cells against Aβ-induced ER stress (p \u3c 0.05) and pro-apoptotic BAD protein activation (p \u3c 0.05). In contrast, higher concentrations of bexarotene upregulated the ER stress proteins and led to BAD activation. Our study revealed that these downstream neurotoxic effects of high drug concentrations could be prevented by pharmacological targeting of the TrkB receptor. The ER stress and BAD activation induced by high concentrations of bexarotene were rescued by the TrkB agonist, 7,8 dihydroxyflavone (p \u3c 0.05) while TrkB inhibitor CTX-B treatment further exacerbated these effects. Together, these findings suggest a cross-talk of TrkB signalling with downstream effects of bexarotene toxicity and indicate that therapeutic targeting of RXRs could prevent the Aβ-induced molecular neurotoxic effects

PTPN11 induces endoplasmic stress and apoptosis in SH-SY5Y cells

PTPN11 is associated with regulation of growth factor signaling pathways in neuronal cells. Using SH-SY5Y neuroblastoma cells, we showed that adeno-associated virus (AAV)-mediated PTPN11 upregulation was associated with TrkB antagonism, reduced neuritogenesis and enhanced endoplasmic reticulum (ER) stress response leading to apoptotic changes. Genetic knock-down of PTPN11 on the other hand leads to increased TrkB phosphorylation in SH-SY5Y cells. ER stress response induced by PTPN11 upregulation was alleviated pharmacologically by a TrkB agonist. Conversely the enhanced ER stress response induced by TrkB receptor antagonism was ameliorated by PTPN11 suppression, providing evidence of cross-talk of PTPN11 effects with TrkB actions. BDNF treatment of neuronal cells with PTPN11 upregulation also resulted in reduced expression of ER stress protein markers. This study provides evidence of molecular interactions between PTPN11 and the TrkB receptor in SH-SY5Y cells. The results reinforce the role played by PTPN11 in regulating neurotrophin protective signaling in neuronal cells and highlight that PTPN11 dysregulation promotes apoptotic activation. Based on these findings we suggest that blocking PTPN11 could have potential beneficial effects to limit the progression of neuronal loss in neurodegenerative disorders