- Organization of the Zone of Transition between the Pretectum and the Thalamus, with Emphasis on the Pretectothalamic Lamina. [Journal Article]
- Front Neuroanat 2016.:82.
The zone of transition between the pretectum, derived from prosomere 1, and the thalamus, derived from prosomere 2, is structurally complex and its understanding has been hampered by cytoarchitectural and terminological confusion. Herein, using a battery of complementary morphological approaches, including cytoarchitecture, myeloarchitecture and the expression of molecular markers, we pinpoint the features or combination of features that best characterize each nucleus of the pretectothalamic transitional zone of the rat. Our results reveal useful morphological criteria to identify and delineate, with unprecedented precision, several [mostly auditory] nuclei of the posterior group of the thalamus, namely the pretectothalamic lamina (PTL; formerly known as the posterior limitans nucleus), the medial division of the medial geniculate body (MGBm), the suprageniculate nucleus (SG), and the ethmoid, posterior triangular and posterior nuclei of the thalamus. The PTL is a sparsely-celled and fiber rich flattened nucleus apposed to the lateral surface of the anterior pretectal nucleus (APT) that marks the border between the pretectum and the thalamus; this structure stains selectively with the Wisteria floribunda agglutinin (WFA), and is essentially immunonegative for the calcium binding protein parvalbumin (PV). The MGBm, located medial to the ventral division of the MGB (MGBv), can be unequivocally identified by the large size of many of its neurons, its dark immunostaining for PV, and its rather selective staining for WFA. The SG, which extends for a considerable caudorostral distance and deviates progressively from the MGB, is characterized by its peculiar cytoarchitecture, the paucity of myelinated fibers, and the conspicuous absence of staining for calretinin (CR); indeed, in many CR-stained sections, the SG stands out as a blank spot. Because most of these nuclei are small and show unique anatomical relationships, the information provided in this article will facilitate the interpretation of the results of experimental manipulations aimed at the auditory thalamus and improve the design of future investigations. Moreover, the previously neglected proximity between the MGBm and the caudal region of the scarcely known PTL raises the possibility that certain features or roles traditionally attributed to the MGBm may actually belong to the PTL.
- High-resolution crystal structures of Colocasia esculenta tarin lectin. [JOURNAL ARTICLE]
- Glycobiology 2016 Aug 24.
Tarin, the Colocasia esculenta lectin from the superfamily of α-d-mannose-specific plant bulb lectins, is a tetramer of 47 kDa composed of two heterodimers. Each heterodimer possesses homologous monomers of ~11.9 (A chain) and ~12.7 (B chain) kDa. The structures of apo and carbohydrate-bound tarin were solved to 1.7 Å and 1.91 Å, respectively. Each tarin monomer forms a canonical β-prism II fold, common to all members of Galanthus nivalis agglutinin (GNA) family, which is partially stabilized by a disulfide bond and a conserved hydrophobic core. The heterodimer is formed through domain swapping involving the C-terminal β-strand and the β-sheet on face I of the prism. The tetramer is assembled through the dimerization of the B chains from heterodimers involving face II of each prism. The 1.91 Å crystal structure of tarin bound to Manα(1,3)Manα(1,6)Man reveals an expanded carbohydrate-binding sequence (QxDxNxVxYx4/6WX) on face III of the β-prism. Both monomers possess a similar fold, except for the length of the loop, which begins after the conserved tyrosine and creates the binding pocket for the α(1,6)-terminal mannose. This loop differs in size and amino-acid composition from 10 other β-prism II domain proteins, and may confer carbohydrate-binding specificity among members of the GNA-related lectin family.
- Nicotine self-administration remodels perineuronal nets in ventral tegmental area and orbitofrontal cortex in adult male rats. [JOURNAL ARTICLE]
- Addict Biol 2016 Aug 22.
Nicotine, a major psychoactive component of tobacco smoke, alters gamma-aminobutyric acid (GABA) modulation of dopamine neurons in the ventral tegmental area (VTA). Changes in structural neuroplasticity can occur in GABAergic parvalbumin (PRV) positive neurons, which are enveloped by structures of the extracellular matrix called perineuronal nets (PNNs). In the current study, rats were trained to self-administer intravenous nicotine (0.03 mg/kg/infusion) for 21 days in 1-hour daily sessions with an incrementing fixed ratio requirement; a control group received saline infusions. At either 45 minutes or 72 hours after the last session, immunofluorescence measurements for PNNs, PRV and c-Fos were conducted. In VTA, nicotine self-administration reduced the number of PRV+ cells surrounded by PNNs at 45 minutes, as well as reducing the intensity of PNNs, suggesting a remodeling of GABA interneurons in this region; the number of PRV+ cells surrounded by PNNs was also reduced at 72 hours. A similar reduction of PNNs occurred in orbitofrontal cortex (OFC) but not in medial prefrontal cortex (prelimbic or infralimbic), 45 minutes after the last session; PNNs were not detected in nucleus accumbens (shell or core). The reduction of PNNs in VTA and OFC was unrelated to c-Fos + cells, as the percent of wisteria floribunda agglutinin + cells co-expressing c-Fos was decreased in OFC but not in VTA. Thus, nicotine self-administration remodeled PNNs surrounding GABA interneurons in VTA and its indirect connections to OFC, suggesting a new possible molecular target where nicotine-induced neuroplasticity takes place. PNN manipulations may prevent or reverse the different stages of tobacco addiction.
- Use of an Intravascular Warming Catheter during Off-Pump Coronary Artery Bypass Surgery in a Patient with Severe Cold Hemagglutinin Disease. [Journal Article]
- Tex Heart Inst J 2016 Aug; 43(4):363-6.
Cold hemagglutinin disease with broad thermal amplitude and high titers presents challenges in treating cardiac-surgery patients. Careful planning is needed to prevent the activation of cold agglutinins and the agglutination of red blood cells as the patient's temperature drops during surgery. We describe our approach to mitigating cold agglutinin formation in a 77-year-old man with severe cold hemagglutinin disease who underwent off-pump coronary artery bypass surgery without the use of preoperative plasmapheresis. This experience shows that the use of an intravascular warming catheter can maintain normothermia and prevent the activation and subsequent formation of cold agglutinins. To our knowledge, this is the first reported use of this technique in a patient with cold hemagglutinin disease. The chief feature in this approach is the use of optimal thermal maintenance-rather than the more usual decrease in cold-agglutinin content by means of therapeutic plasma exchange.
- The Total and Lectin-binding Proteome of Spherulin from Coccidioides posadasii. [JOURNAL ARTICLE]
- J Proteome Res 2016 Aug 22.
Coccidioides is a virulent dimorphic fungus that causes coccidioidomycosis (Valley Fever) in mammals, including humans. Although the genome has been sequenced, a proteomic analysis does not exist. To address this gap in proteomic knowledge, we generated the proteome of Spherulin (a well-studied lysate of fungal spherules) and identified 1390 proteins. Some of the proteins included glycosylation enzymes which led us to hypothesize that fungal glycosylation patterns may be different from that of mammals and could be exploited to detect Coccidioides in tissues. We performed lectin-based immunohistochemistry on formalin-fixed paraffin-embedded human patients' lung tissues. GSL-II (Griffonia simplificonia lectin II) and sWGA (succinylated wheat germ agglutinin) lectins bound specifically to endospores and spherules in infected lungs. To identify lectin-binding glycoproteins in Spherulin, we performed lectin-affinity chromatography followed by LC-MS/MS. A total of 195 glycoproteins from Spherulin bound to GSL-II, 224 glycoproteins bound to sWGA, and 145 glycoproteins bound to both lectins. This is the first report of the specific reactivity of GSL-II and sWGA lectins to Coccidioides endospores and spherules in infected human tissues and the first listing of the Coccidioidal proteome from Spherulin using sequences present in three Coccidioides databases: RefSeq, SwissProt and The Broad Institute's Coccidioides Genome project. .
- Sustained mitogenic effect on K562 human chronic myelogenous leukemia cells by dietary lectin, jacalin. [JOURNAL ARTICLE]
- Glycoconj J 2016 Aug 19.
Dietary lectins have been shown to affect the proliferation of human cancer cell lines. The anti-proliferative effects of lectins from varied sources have been extensively studied and in some cases, the underlying mechanism has been explored. Except for peanut agglutinin (PNA), the mitogenic effects of no other lectins have been studied in detail. In the present study, we have shown that jacalin, lectin purified from jackfruit (Artocarpus integrifolia) seeds act as a mitogen for K562, the Bcr-Abl expressing erythroleukemia cell line (K562) and the effect was found to be dose dependent. K562 cells remained in the proliferative state for a longer period even after the withdrawal of jacalin stimulation, thus jacalin was found to induce sustained mitogenic effect on K562 cells. Further, conditioned media from K562 cells treated with jacalin were observed to have the similar mitogenic effect even in the presence of galactose. Importantly, galactose which is a known ligand for jacalin will interact with functionally active jacalin present in the conditioned media and neutralise its effect. In addition, jacalin treatment also resulted in increased mRNA expression levels of pro-inflammatory cytokines including IL-1β, IL-6 and IFN-γ. Our results indicate that jacalin induces secretion of soluble molecules, which maybe responsible for this observed increased proliferation of K562 cells.
- Identification of sperm equatorial segment protein 1 in the acrosome as the primary binding target of peanut agglutinin (PNA) in the mouse testis. [JOURNAL ARTICLE]
- Histochem Cell Biol 2016 Aug 18.
Peanut agglutinin (PNA), a plant lectin protein that recognizes the galactose β (1 -> 3) N-acetylgalactosamine carbohydrate sequence, has been widely used as a sperm acrosome-specific marker; however, the acrosomal glycoproteins that specifically bind to PNA have yet to be identified. We herein purified and identified PNA-binding glycoproteins in the mouse testis using biotinylated PNA and streptavidin-coupled magnetic beads, and liquid chromatography-tandem mass spectrometry (LC-MS/MS), respectively. In six repeated experiments, sperm equatorial segment protein 1 (SPESP1) was detected most frequently as a PNA-binding glycoprotein, followed by dipeptidase 3, proacrosin-binding protein, and acrosin prepropeptide. The identification of SPEPS1 in the testis lysate and its PNA-bound fraction was verified with lectin and Western blot analyses, and the co-localization of PNA and SPEPS1 in acrosomes was confirmed with lectin- and immunohistochemistry. Since the PNA reactivity of sperm acrosomes was observed not only in normal mice, but also in SPESP1-deficient mice, although at lower levels, PNA was also considered to bind to other candidate glycoproteins. The present study identified SPESP1 in the acrosome as the primary binding target of PNA in the mouse testis. Further defining the specific lectin-glycoprotein relationships in individual cells will enhance the value of lectin histochemistry.
- Spatially Directed Proteomics of the Human Lens Outer Cortex Reveals an Intermediate Filament Switch Associated With the Remodeling Zone. [Journal Article]
- Invest Ophthalmol Vis Sci 2016 Aug 1; 57(10):4108-14.
To quantify protein changes in the morphologically distinct remodeling zone (RZ) and adjacent regions of the human lens outer cortex using spatially directed quantitative proteomics.Lightly fixed human lens sections were deparaffinized and membranes labeled with fluorescent wheat germ agglutinin (WGA-TRITC). Morphology directed laser capture microdissection (LCM) was used to isolate tissue from four distinct regions of human lens outer cortex: differentiating zone (DF), RZ, transition zone (TZ), and inner cortex (IC). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) of the plasma membrane fraction from three lenses (21-, 22-, and 27-year) revealed changes in major cytoskeletal proteins including vimentin, filensin, and phakinin. Peptides from proteins of interest were quantified using multiple reaction monitoring (MRM) mass spectrometry and isotopically-labeled internal peptide standards.Results revealed an intermediate filament switch from vimentin to beaded filament proteins filensin and phakinin that occurred at the RZ. Several other cytoskeletal proteins showed significant changes between regions, while most crystallins remained unchanged. Targeted proteomics provided accurate, absolute quantification of these proteins and confirmed vimentin, periplakin, and periaxin decrease from the DF to the IC, while filensin, phakinin, and brain acid soluble protein 1 (BASP1) increase significantly at the RZ.Mass spectrometry-compatible fixation and morphology directed laser capture enabled proteomic analysis of narrow regions in the human lens outer cortex. Results reveal dramatic cytoskeletal protein changes associated with the RZ, suggesting that one role of these proteins is in membrane deformation and/or the establishment of ball and socket joints in the human RZ.
- Termiticidal lectins from Myracrodruon urundeuva (Anacardiaceae) cause midgut damages when ingested by Nasutitermes corniger (Isoptera; Termitidae) workers. [JOURNAL ARTICLE]
- Pest Manag Sci 2016 Aug 16.
Myracrodruon urundeuva is a hardwood tree whose bark, heartwood, and leaf contain lectins (MuBL, MuHL and MuLL, respectively) with termiticidal activity against Nasutitermes corniger. In this work, the effects of these lectins on the midgut of N. corniger workers were evaluated.The insects were supplied with an artificial diet containing the lectins at their respective LC50 (previously determined). Forty-eight hours after the treatment, the midguts were dissected and fixed for histopathology analyses. Toluidine blue-stained midguts from lectin-treated workers showed disorganization, with presence of debris in the lumen and absence of the brush border. Fluorescence microscopy revealed that the numbers of digestive and proliferating cells were lower in lectin-treated individuals than in the control, and caspase-3 staining confirmed occurrence of cell apoptosis. Enteroendocrine cells were not seen in the treated individuals. The midguts from treated insects showed greater staining for peroxidase than the control, suggesting that the lectins caused oxidative stress. Staining with wheat germ agglutinin conjugated to FITC revealed that the lectins interfered with the integrity of the peritrophic matrix.This study showed that termiticidal lectins from M. urundeuva cause severe injuries, oxidative stress and cell death in the midgut of N. corniger workers.
- A Chemiluminescent Protein Microarray Method for Determining the Seroglycoid Fucosylation Index. [Journal Article]
- Sci Rep 2016.:31132.
The Lens culinaris agglutinin-reactive fraction of AFP (AFP-L3) is widely used to screen for hepatocellular carcinoma (HCC) in Japan and China. We developed a chemiluminescent protein microarray for determining the AFP-L3/AFP index (the ratio of AFP-L3 to total AFP, AFP-L3%) by fixing AFP-specific antibodies and Lens culinaris lectin on aldehyde-coated glass slides. Serum samples were tested for AFP using an enzyme-linked immunosorbent assay (ELISA) to validate the microarray. AFP-L3 was detected using Hotgen Biotech glycosyl capture spin column pretreatment technology and ELISA. When the AFP cut-off value was set to 20 ng/ml, the protein microarray displayed 89.74% sensitivity and 100% specificity for HCC diagnosis, and the ELISA displayed 87.17% sensitivity and 100% specificity. When the AFP-L3% cut-off value was set to 0.1, the protein microarray displayed 56.41% sensitivity and 100% specificity for HCC diagnosis, and the ELISA displayed 53.84% sensitivity and 100% specificity. The ROC curve for the HCC diagnosis showed that the AFP area under the ROC curve (AUC = 0.996; 95% CI: 0.986-1.005) was much higher than that of AFP-L3 (AUC = 0.857; 95% CI: 0.769-0.94) and AFP-L3% (AUC = 0.827; CI: 0.730-0.924). The microarray assay used in this study is a highly sensitive, accurate, and efficient assay for the determination of the AFP-L3%.