Solanum Tuberosum Lectin Research Articles

Proteome Analysis of Serum Purified Using Solanum tuberosum and Lycopersicon esculentum Lectins.

Serum and plasma exhibit a broad dynamic range of protein concentrations, posing challenges for proteome analysis. Various technologies have been developed to reduce this complexity, including high-abundance depletion methods utilizing antibody columns, extracellular vesicle enrichment techniques, and trace protein enrichment using nanobead cocktails. Here, we employed lectins to address this, thereby extending the scope of biomarker discovery in serum or plasma using a novel approach. We enriched serum proteins using 37 different lectins and subjected them to LC-MS/MS analysis with data-independent acquisition. Solanum tuberosum lectin (STL) and Lycopersicon esculentum lectin (LEL) enabled the detection of more serum proteins than the other lectins. STL and LEL bind to N-acetylglucosamine oligomers, emphasizing the significance of capturing these oligomer-binding proteins when analyzing serum trace proteins. Combining STL and LEL proved more effective than using them separately, allowing us to identify over 3000 proteins from serum through single-shot proteome analysis. We applied the STL/LEL trace-protein enrichment method to the sera of systemic lupus erythematosus model mice. This revealed differences in >1300 proteins between the systemic lupus erythematosus model and control mouse sera, underscoring the utility of this method for biomarker discovery.

Alzheimer’s Amyloid β Peptide Induces Angiogenesis in an Alzheimer’s Disease Model Mouse through Placental Growth Factor and Angiopoietin 2 Expressions

Increased angiogenesis, especially the pathological type, has been documented in Alzheimer's disease (AD) brains, and it is considered to be activated due to a vascular dysfunction-mediated hypoxic condition. To understand the role of the amyloid β (Aβ) peptide in angiogenesis, we analyzed its effects on the brains of young APP transgenic AD model mice. Immunostaining results revealed that Aβ was mainly localized intracellularly, with very few immunopositive vessels, and there was no extracellular deposition at this age. Solanum tuberosum lectin staining demonstrated that compared to their wild-type littermates, the vessel number was only increased in the cortex of J20 mice. CD105 staining also showed an increased number of new vessels in the cortex, some of which were partially positive for collagen4. Real-time PCR results demonstrated that placental growth factor (PlGF) and angiopoietin 2 (AngII) mRNA were increased in both the cortex and hippocampus of J20 mice compared to their wild-type littermates. However, vascular endothelial growth factor (VEGF) mRNA did not change. Immunofluorescence staining confirmed the increased expression of PlGF and AngII in the cortex of the J20 mice. Neuronal cells were positive for PlGF and AngII. Treatment of a neural stem cell line (NMW7) with synthetic Aβ1-42 directly increased the expression of PlGF and AngII, at mRNA levels, and AngII at protein levels. Thus, these pilot data indicate that pathological angiogenesis exists in AD brains due to the direct effects of early Aβ accumulation, suggesting that the Aβ peptide regulates angiogenesis through PlGF and AngII expression.

Pancreatic Cyst Glycopatterns as Potential Biomarkers for MCN and SCN

Background & Aims: The pathophysiologies of malignant pancreatic cystic lesions, including mucinous cystic neoplasm (MCN), intraductal papillary mucinous neoplasm (IPMN), and mucinous cystic adenocarcinoma (MCA), are not fully understood, and no diagnostic or predictive biomarkers have been established. Glycosylation modifications on as many as 70% of all human proteins can reflect various pathological changes with high sensitivity. However, little is known about the alterations of glycosylation and glycoproteins in MCN. Methods: This study is based on the results of surgical pathology and cystic fluid cytology from 211 cases diagnosed with pancreatic cystic tumours. Liquid samples were obtained through endoscopic ultrasonography-guided fine needle aspiration (EUS-FNA). Of the 75 cases, the cystic fluid of 35 cases was previously analysed by a lectin microarray, which was used to characterize the altered glycosylation between the MCN and SCN groups. We observed an increase in trimers and tetramers of GlcNAc, core (GlcNAc), multivalent sialic acid, Galβ1-3GalNAc, Terminal GalNAc, αGalNAc, Tn antigen, and GalNAcα1-3((Fucα1-2)) Gal (blood group A antigen) in the MCN group (P < 0.05). Of the remaining 40 cases, 20 were from the MCN group and 20 were from the SCN group; the accuracy of this distinction was verified. Moreover, altered glycan patterns on the surface of pancreatic cystic lesions were verified by lectin immunohistochemistry. This study is based on the results of increased binding signals in the MCN group by solanum tuberosum lectin (STL) and Bauhinia purpurea lectin (BPL), which bind glycoproteins, and the results of magnetic particle conjugateassisted LC-MS/MS analyses and bioinformatics analysis. The characteristics of lectin binding glycoproteins between STL and BPL demonstrated altered glycopatterns between the MCN and SCN groups. Each protein was evaluated in triplicate. Result: The ability of STLs and BPLs to bind glycoproteins in MCN/SCN cystic fluid was determined via quantitative analysis. Through bioinformatics analysis and STRING protein interaction diagrams, we concluded that the glycoproteins LAMC1, YWHAG, YWHAQ, YWHAZ, S100A11, HSPA5, LDHA, ACTB, KRT18, and ACTB were increased in MCN.Conclusions: We identified a panel of molecular markers and clinical features. These markers and features may be correlated with a significant increase in the mucin secretion of malignant epithelial cells. Funding Statement: This work was supported by the National Natural Science Foundation of Beijing (No.7182066). Declaration of Interests: The authors declare no conflict of interest. Ethics Approval Statement: This prospective study was approved by the Department of Chinese PLA General Hospital Ethics Committee (Ethics NO. S2014-108-01).

Helicobacter pylori SabA binding gangliosides of human stomach

ABSTRACT Adhesion of Helicobacter pylori to the gastric mucosa is a prerequisite for the pathogenesis of H. pylori related diseases. In this study, we investigated the ganglioside composition of human stomach as the target for attachment mediated by H. pylori SabA (sialic acid binding adhesin). Acid glycosphingolipids were isolated from human stomach and separated into subfractions, which were characterized by mass spectrometry and by binding of antibodies, bacteria, and Solanum tuberosum lectin. H. pylori SabA binding gangliosides were characterized as Neu5Acα3-neolactohexaosylceramide and Neu5Acα3-neolactooctaosylceramide, while the other acid human stomach glycosphingolipids characterized (sulfatide and the gangliosides GM3, GD3, GM1, Neu5Acα3-neolactotetraosylceramide, GD1a and GD1b) were not recognized by the bacteria. Defining H. pylori binding glycosphingolipids of the human gastric mucosa will be useful to specifically target this microbe-host interaction for therapeutic intervention.

Lectins identify distinct populations of coelomocytes in Strongylocentrotus purpuratus.

Coelomocytes represent the immune cells of echinoderms, but detailed knowledge about their roles during immune responses is very limited. One major challenge for studying coelomocyte biology is the lack of reagents to identify and purify distinct populations defined by objective molecular markers rather than by morphology-based classifications that are subjective at times. Glycosylation patterns are known to differ significantly between cell types in vertebrates, and furthermore they can vary depending on the developmental stage and activation states within a given lineage. Thus fluorescently labeled lectins that recognize distinct glycan structures on cell surface proteins are routinely used to identify discrete cell populations in the vertebrate immune system. Here we now employed a panel of fifteen fluorescently-labeled lectins to determine differences in the glycosylation features on the surface of Strongylocentrotus purpuratus coelomocytes by fluorescence microscopy and flow cytometry. Eight of the lectins (succinylated wheat germ agglutinin, Len culinaris lectin, Pisum sativum agglutinin, Saphora japonica agglutinin, Solanum tuberosum lectin, Lycopersicon esculentum lectin, Datura stramonium lectin, Vicia villosa lectin) showed distinct binding patterns to fixed and live cells of three major coelomocyte classes: phagocytic cells, red spherule cells, and vibratile cells. Importantly, almost all lectins bound only to a subgroup of cells within each cell type. Lastly, we established fluorescently-labeled lectin-based fluorescence activated cell sorting as a strategy to purify distinct S. purpuratus coelomocyte (sub-)populations based on molecular markers. We anticipate that this will become a routine approach in future studies focused on dissecting the roles of different coelomocytes in echinoderm immunity.

Neurovascular Specifications intheAlzheimer-Like Brain of Mice Affected by Focal Cerebral Ischemia: Implications for Future Therapies.

Alzheimer's disease (AD), the most frequent type of dementia, is a prototypical neurodegenerative disease, but shares with stroke certain common risk factors. Consequently, how vascular pathology may modulate AD pathogenesis has gained scientific attention. Therefore, aside from typical features of AD (e.g., amyloid-β, tau hyperphosphorylation, and cholinergic dysfunction), changes within the 'neurovascular unit' (NVU) are of particular interest. This study focused on cholinergic, choline acetyltransferase (ChAT)-immunopositive, and tyrosine hydroxylase (TH)-containing neurons in association with the vasculature to explore the neurovascular complex of the AD brain affected by stroke. Wild-type and triple-transgenic (3xTg) mice of different ages underwent unilateral permanent focal cerebral ischemia. Histochemical analyses comprised diverse neuronal and vascular NVU components, and markers of AD. Immunofluorescence labeling confirmed the existence of Aβ deposits and phospho-tau together with glial reactions and morphologically altered endothelia, visualized by Solanum tuberosum lectin. Twenty-four hours after ischemia induction, immunoreactivities for ChAT and TH declined in the ischemia-affected striatum and, at least in part, in the ischemic border zone and ipsilateral neocortex. Correlation analyses indicated simultaneous degeneration of neuronal and vascular components. A trend for more severe affection of ChAT was observed in younger as compared with older mice. The present findings suggest complex interactions within the NVU of the AD-like brain affected by ischemia, comprising alterations of the cholinergic system in conjunction with vascular pathology. Hence, it may be worthwhile to explore the impact of a cellular stabilization approach on vascular and glial elements in AD in terms of a potential disease-alleviating strategy.

Sorafenib induced alteration of protein glycosylation in hepatocellular carcinoma cells.

Sorafenib is a multikinase inhibitor and is effective in treating hepatocellular carcinoma (HCC). However, it remains unknown whether sorafenib induces the alteration of protein glycosylation. The present study treated HCC MHCC97L and MHCC97H cells with a 50% inhibitory concentration of sorafenib. Following this treatment, alteration of protein glycosylation was detected using a lectin microarray. Compared with the controls, the binding capacity of glycoproteins extracted from sorafenib-treated HCC cells to the lectins Bauhinia purpurea lectin, Dolichos biflorus agglutinin, Euonymus europaeus lectin, Helix aspersa lectin, Helix pomatia lectin, Jacalin, Maclura pomifera lectin and Vicia villosa lectin were enhanced; while, the binding capacities to the lectins Caragana arborescens lectin, Lycopersicon esculentum lectin, Limulus polyphemus lectin, Maackia amurensis lecin I, Phaseolus vulgaris leucoagglutinin, Ricinus communis agglutinin 60, Sambucus nigra lectin and Solanum tuberosum lectin were reduced (spot intensity median/background intensity median ≥2, P<0.05). This difference in glycoprotein binding capacity indicates that cells treated with sorafenib could increase α-1,3GalNAc/Gal, β-1,3 Gal, GalNAcα-Ser/Thr(Tn) and α-GalNAc structures and decrease GlcNAc, sialic acid, tetra-antennary complex-type N-glycan and β-1,4Gal structures. These results were additionally confirmed by lectin blotting. Expression levels of signaling molecules including erythroblastosis 26–1 (Ets-1), extracellular signal-related kinases (ERK) and phosphorylated-ERK were measured by western blotting. There was a reduction in the expression of Ets-1 and ERK phosphorylation in sorafenib or 1,4-Diamino-2,3-dicyano-1,4-bis (2-aminophenylthio) butadiene treated cells suggesting that sorafenib may reduce the expression levels of Ets-1 by blocking the Ras/Raf/mitogen activated protein kinase signaling pathway. In the present study, it was clear that sorafenib could inhibit the proliferation of HCC cells and alter protein glycosylation. The findings of this study may lead to providing a novel way of designing new anti-HCC drugs.

Carbohydrate Analogue Microarrays for Identification of Lectin-Selective Ligands.

Fifty-five mono- and disaccharide analogues were prepared and used for the construction of microarrays to uncover lectin-selective ligands. The microarray study showed that two disaccharide analogues, 28' and 44', selectively bind to Solanum tuberosum lectin (STL) and wheat germ agglutinin (WGA), respectively. Cell studies indicated that 28' and 44' selectively block the binding of STL and WGA to mammalian cells, unlike the natural ligand LacNAc, which suppresses binding of both STL and WGA to cells.

Solanum tuberosum lectin inhibits Ehrlich ascites carcinoma cells growth by inducing apoptosis and G2/M cell cycle arrest.

Recently, a lectin was purified from the potato cultivated in Bangladesh locally known as Sheel. In the present study cytotoxicity of the lectin against Ehrlich ascites carcinoma (EAC) cells was studied by MTT assay in vitro in RPMI-1640 medium and 8.0-36.0% cell growth inhibition was observed at the range of 2.5-160μg/ml protein concentration when incubated for 24h. The lectin-induced apoptosis in EAC cells was confirmed by fluorescence and optical microscope. The apoptotic cell death was also confirmed by using caspase inhibitors. Cells growth inhibition caused by the lectin (36%) was remarkably decreased to 7.6 and 22.3% respectively in the presence of caspase-3 and -8 inhibitors. RT-PCR was used to evaluate the expression of apoptosis-related genes Bcl-X, p53, and Bax. An intensive expression of Bcl-X gene was observed in untreated control EAC cells with the disappeared of the gene in Sheel-treated EAC cells. At the same time, Bax gene expression appeared only in Sheel-treated EAC cells and the expression level of the p53 gene was increased remarkable after the treatment of EAC cells with the lectin. The lectin showed strong agglutination activity against EAC cells. Flow cytometry was used to study the cell cycle phases of EAC cells and it was observed that the lectin arrested the G2/M phase. In conclusion, Sheel lectin inhibited EAC cells growth by inducing apoptosis.

Microgels as vehicles for effective delivery of hydrophilic drugs to the brain

Event Abstract Back to Event Microgels as vehicles for effective delivery of hydrophilic drugs to the brain Madeline Simpson1, Justin Piazza2, Yogesh K. Katare2, Sharnpreet K. Kooner2, Niels B. Smeets1, Ram K. Mishra2 and Todd Hoare1 1 McMaster University, Department of Chemical Engineering, Canada 2 McMaster University, Department of Psychiatry and Behavioural Neuroscience, Canada Introduction: The treatment of central nervous system (CNS) disorders is challenging due to difficulties in drug transport across the blood-brain barrier (BBB). Nanoparticle-based therapies are of interest as they can be designed for specific targets, to control drug release, and lower the required dosage for therapeutic intervention. Degradable microgels (hydrogel-based nanoparticles) composed of poly(oligoethylene glycol methacrylate) (POEGMA), are promising delivery vehicles which have demonstrated protein-repellent properties[1]. In comparison to “hard” nanoparticles widely cited to cross the BBB with diameters of <150 nm[2], we show that larger elastic microgels can be transported across the BBB, which can improve drug loading and biodistribution of the hydrophilic drug 3(R)-[(2(S)-pyrrolidinylcarbonyl)amino]-2-oxo-1-pyrrolidineacetamide) (PAOPA). This drug is a positive allosteric modulator that selectively targets the dopamine D2 receptor and has been demonstrated to reduce the negative symptoms of schizophrenia in animal models[3]. Materials and Methods: POEGMA-based microgels were synthesized by precipitation polymerization of diethylene glycol methacrylate, acrylic acid (AA) and 2-hydroxyethyl disulfide dimethacrylate (degradable crosslinker) with various quantities of sodium dodecyl sulfate (SDS). The AA groups were functionalized using carbodiimide chemistry with Solanum tuberosum lectin (STL) which enables binding to the nasal epithelium for intranasal delivery. Microgel characterisation was carried out by light scattering (size), transmission electron microscopy (morphology) and conductometric titration (functional group content). PAOPA was passively loaded, with excess drug removed by centrifugation. The in vitro drug release was assessed using HPLC, and microgel cytotoxicity was determined by an MTT assay using RPMI 2650 nasal septum and SH-SY5Y dopamine 2D receptor-transduced neuronal cells. Rhodamine-labelled microgels were injected into the peritoneum of male Sprague-Dawley rats to assess the in vivo biodistribution of different microgel sizes. Blood and tissues were recovered after one hour and analyzed using fluorescence to quantify microgel biodistribution. Results and Discussion: Microgels with sizes of 99±2 nm, 152±3 nm, and 249±4 nm can be prepared with low polydipsersity (<0.1). PAOPA was loaded with an encapsulation efficiency of 30±2%, independent of size. The diffusion of PAOPA from the two smallest microgels occured over four days, with 50% of the drug still retained after 24 hours (Figure 1) while the largest microgel prolonged release to five days. The biodistribution studies with the rhodamine-labelled microgels demonstrated that the smallest microgels were the most effective at crossing the BBB; however the largest microgels were transported in significant numbers despite their larger size than the 150 nm limit cited while also accumulating less in major non-target organs (Figure 2). In vivo efficacy studies as a function of microgel size in a rat schitzophrenia model will be presented. Conclusion: POEGMA microgels are effective delivery vehicles for the transport of hydrophilic drugs across the BBB. Due to their elastic nature, larger microgels may be advantageous as the compressibility may enable the delivery of bigger, longer circulating, and more highly drug loaded particles across tight biological barriers. Canadian Institutes for Health Research (CIHR); Natural Sciences and Engineering Research Council of Canada (NSERC)

A Novel Intranasal Spray Device for the Administration of Nanoparticles to Rodents

Experimental intranasal (IN) delivery of nanoparticle (NP) drug carriers is typically performed using a pipette with or without anesthesia, a technique that may be a poor simulation of practical IN administration of drug-loaded NPs in humans. Existing IN spray devices suffer from drawbacks in terms of variability in dose-control and spray duration as well as the application of nonuniform pressure fields when a NP-formulated drug is aerosolized. Furthermore, existing spray devices require large volumes that may not be available or may be prohibitively expensive to prepare. In response, we have developed a novel pneumatically driven IN spray device for the administration of NPs, which is capable of administering extremely small quantities (50–100 μl) of NP suspension in a fine spray that disperses the NPs uniformly onto the tissue. This device was validated using haloperidol-loaded Solanum tuberosum lectin (STL)-functionalized, poly(ethylene glycol)–block-poly(d,l-lactic-co-glycolic acid) (PEG–PLGA) NPs targeted for delivery to the brain for schizophrenia treatment. A pneumatic pressure of 100 kPa was found to be optimal to produce a spray that effectively aerosolizes NP suspensions and delivers them evenly to the olfactory epithelium. IN administration of STL-functionalized NPs using the IN spray device increased brain tissue haloperidol concentrations by a factor of 1.2–1.5× compared to STL-functionalized NPs administered IN with a pipette. Such improved delivery enables the use of lower drug doses and thus offers both fewer local side-effects and lower costs without compromising therapeutic efficacy.

Haloperidol-loaded intranasally administered lectin functionalized poly(ethylene glycol)–block-poly(d,l)-lactic-co-glycolic acid (PEG–PLGA) nanoparticles for the treatment of schizophrenia

Lectin-functionalized, polyethylene glycol–block-poly-(d,l)-lactic-co-glycolic acid nanoparticles loaded with haloperidol were prepared with narrow size distributions and sizes <135nm. The nanoparticles exhibited high Solanum tuberosum lectin (STL) conjugation efficiencies, encapsulation efficiencies, and drug loading capacities. The in vitro release of haloperidol was 6–8% of the loaded amount in endo-lysosomal conditions over 96h, demonstrating minimal drug leakage and the potential for the efficient drug transport to the targeted brain tissue. The haloperidol released upon erosion was successful in displacing [3H] N-propylnorapomorphine and binding to bovine striatal dopamine D2 receptors. Both haloperidol-loaded nanoparticle formulations were found to be highly effective at inducing catalepsy. Intranasal administration of STL-functionalized nanoparticles increased the brain tissue haloperidol concentrations by 1.5–3-fold compared to non-STL-functionalized particles and other routes of administration. This formulation demonstrates promise in the reduction of the drug dose necessary to produce a therapeutic effect with antipsychotic drugs for the treatment of schizophrenia.

Characterization of glycans of CD4+T cells in HAM/TSP

The outermost structure of cells is galectin-glycan lattice, that is shown to have many roles in cell-cell interactions. Recently, biofilm-like extracellular viral assemblies were shown to mediate HTLV-1 cell-to-cell transmission. Here, we tried to characterize the glycans on the CD4+T cells in HAM. CD4+T cells from four respective cases of HAM, AC(asymptomatic carriers), and NC(negative control) were subjected to lectin array analysis. Similarly, glycans liberated from membrane proteins of CD4+T cells from six respective cases of HAM and NC analyzed by MALDI-TOF MS. We found that UDA(Urtica dioica agglutinin) and STL(Solanum tuberosum lectin (Potato)) lectins, that recognize N-glycan polylactosamine which consist of repeats of the disaccharide betaGal(1-4)betaGlcNAc(1-3), were significantly high in HAM in lectin array analysis. Interestingly, UDA was reported to inhibit cell-to-cell infection in vitro. On the other hand, MALDI-TOF MS analysis found several O-glycans in HAM. Candidates verified in GlycoSuite database were mainly O-glycans attach to MUC1 and Leukosialin(CD43) as carrier proteins. These proteins were reported to play roles in ATL and cell-to-cell infection as well. These glycans and carrier proteins may be therapeutic target of HAM.

Intranasal nanoparticles of basic fibroblast growth factor for brain delivery to treat Alzheimer's disease

Disabilities caused by neurodegeneration have become one of the main causes of mortality in elderly population, with drug distribution to the brain remaining one of the most difficult challenges in the treatment of the central nervous system (CNS) diseases due to the existence of blood-brain barrier. Lectins modified polyethylene glycol-polylactide-polyglycolide (PEG-PLGA) nanoparticles could enhance the drug delivery to the brain following intranasal administration. In this study, basic fibroblast growth factor (bFGF) was entrapped in nanoparticles conjugated with Solanum tuberosum lectin (STL), which selectively binds to N-acetylglucosamine on the nasal epithelial membrane for its brain delivery. The resulting nanoparticles had uniform particle size and negative zeta potential. The brain distribution of the formulations following intranasal administration was assessed using radioisotopic tracing method. The areas under the concentration-time curve of (125)I-bFGF in the olfactory bulb, cerebrum, and cerebellum of rats following nasal application of STL modified nanoparticles (STL-bFGF-NP) were 1.79-5.17 folds of that of rats with intravenous administration, and 0.61-2.21 and 0.19-1.07 folds higher compared with intranasal solution and unmodified nanoparticles, respectively. Neuroprotective effect was evaluated using Mirror water maze task in rats with intracerebroventricular injection of β-amyloid25-35 and ibotenic acid. The spatial learning and memory of Alzheimer's disease (AD) rats in STL-bFGF-NP group were significantly improved compared with AD model group, and were also better than other preparations. The results were consistent with the value of choline acetyltransferase activity of rat hippocampus as well as the histological observations of rat hippocampal region. The histopathology assays also confirmed the in vivo safety of STL-bFGF-NP. These results clearly indicated that STL-NP was a promising drug delivery system for peptide and protein drugs such as bFGF to enter the CNS and play the therapeutic role.

Histological and Lectin Histochemical Studies on the Olfactory and Respiratory Mucosae of the Sheep

ABSTRACTThe olfactory and respiratory mucosae of the Corriedale sheep were examined using lectin histochemistry in order to clarify the histochemical and glycohistochemical differences between these two tissues. The olfactory epithelium was stained with 13 lectins out of 21 lectins examined, while the respiratory epithelium was positive to 16 lectins. The free border of both of the olfactory and respiratory epithelia was stained with 12 lectins: Wheat germ agglutinin (WGA), succinylated-wheat germ agglutinin (s-WGA), Lycopersicon esculentum lectin (LEL), Solanum tuberosum lectin (STL), Datura stramonium lectin (DSL), Soybean agglutinin (SBA), Bandeiraea simplicifolia lectin-I (BSL-I), Ricinus communis agglutinin-I (RCA-120), Erythrina cristagalli lectin (ECL), Concanavalin A (Con A), Phaseolus vulgaris agglutinin-E (PHA-E) and Phaseolus vulgaris agglutinin-L (PHA-L). The associated glands of the olfactory mucosa, Bowman’s glands, were stained with 13 lectins. While both the goblet cells and mucous nasal glands were stained with 8 lectins; five of them (WGA, s-WGA, STL, Vicia villosa agglutinin (VVA) and ECL) were mutually positive among the Bowman’s glands, mucous nasal glands and the goblet cells. These findings indicate that the glycohistochemical characteristics of the free borders of both olfactory and respiratory epithelia are similar to each other, suggesting that secretions from the Bowman’s glands and those of the goblet cells and mucous nasal glands are partially exchanged between the surface of two epithelia to contribute the functions of the respiratory epithelium and the olfactory receptor cells, respectively.

Stroke-induced opposite and age-dependent changes of vessel-associated markers in co-morbid transgenic mice with Alzheimer-like alterations

The pathophysiological concept of ischaemic stroke was recently expanded to a more comprehensive perspective, focussing on the vasculature as well as peri- and juxtavascular cells including astrocytes. Increasing evidence also supports a role of the vasculature in Alzheimer's disease (AD), but causal relationships are poorly understood. The purpose of this study was to examine vascular alterations due to cerebral ischaemia in aged wildtype (WT) mice and in the triple-transgenic (3xTg) mouse model of AD. Three- and 12-month-old WT and 3xTg mice underwent permanent middle cerebral artery occlusion. One day after ischaemia onset, expression of collagen IV and laminin as basement membrane constituents, and Solanum tuberosum lectin (STL) as endothelial marker was quantified in the ischaemic neocortex, striatum and hippocampus. Further, CD31- and aquaporin-4-immunoreactivity served for coverage of endothelium and astrocyte endfeet. Ischaemia resulted in strong upregulation of collagen IV and laminin in the neocortex of 3-month-old WT and 3xTg mice, while STL appeared unaffected. Quantification confirmed collagen IV upregulation in the ischaemic neocortex of 3- and 12-month-old WT and 3xTg mice, whereas striatal changes were limited to young WT mice. However, collagen IV expression in the hippocampus appeared nearly unaltered. Qualitative and quantitative data evidenced more severe degeneration of endothelial cells and astrocyte endfeet in 3xTg mice. In conclusion, this study supports the critical impact of the vasculature in the aged and AD brain by showing an age- and genetic background-dependent response of basement membranes to cerebral ischaemia, and a pronounced endothelial and astrocytic degeneration in the AD-like brain.

β-Amyloid, Cholinergic Transmission, and Cerebrovascular System - A Developmental Study in a Mouse Model of Alzheimer's Disease

The majority of patients suffering from Alzheimer's disease (AD) demonstrate cerebral vascular changes and impaired regulation of cerebral blood flow, which has been assumed to play an important role in AD pathogenesis (vascular hypothesis of AD). There is strong evidence that both β-amyloid (Aβ) oligomers and plaques contribute to vascular injuries and functional impairments of the neurovascular unit. Vice versa, Aβ lesions can be triggered by hypertension and ischemic brain injury, while Aβ aggregates appear to have anti-angiogenic properties. Cholinergic dysfunction may result in impaired cerebral blood flow with consequences on normal function of the neurovascular unit including processing of the amyloid precursor protein (APP). To characterize in vivo the developmental relationship between Aβ formation and deposition, cortical cholinergic innervation and cerebrovascular abnormalities, transgenic Tg2576 mice that overexpress the Swedish double mutation of human APP, and demonstrate significant cerebral cortical deposition of Aβ plaques at ages from 9 months onwards, were considered as an appropriate animal model. Using the somatosensory cortex as a representative region, serial cryocut sections, were obtained from mice at ages ranging from 4 up to 18 months. These were subjected to immunohistochemistry to label vascular endothelial cells (anti-glucose transporter 1 (GluT1) immunostaining), cholinergic nerve terminals (anti-vesicular acetylcholine transporter (VAChT) immunostaining) and β-amyloid plaques (thioflavin S, and/or Solanum tuberosum lectin staining). This was followed by a thorough quantitative evaluation of the age-related spatial relationship between cerebral cortical capillaries, Aβ plaques and cholinergic terminals, using computer-assisted image analysis. The density of cholinergic terminals estimated by evaluation of VAChT immunohistochemistry in somatosensory cortical sections of wild type mice did not change with aging regardless of the cortical layer examined, while in cortical layers II/III and IV of somatosensory cortex of transgenic Tg2576 mice age-related decreases in cholinergic fiber densities were assessed. However, quantitative morphometric analysis demonstrated an age-related reduction in the number of varicosities on cholinergic fibers, particularly in layer IV, in both transgenic Tg2576 mice and non-transgenic littermates. Cholinergic innervation of microvessels in the somatosensory cortex decreased with aging in both Tg2576 mice and non-transgenic littermates, as revealed by estimating the ratio of the number of cholinergic vascular contacts and total length of blood vessel. There was no significant difference in the perivascular cholinergic innervation in areas that demonstrated significant plaque load and those with no plaque deposits regardless of the cortical layer examined. The density of blood vessels estimated in the somatosensory cortex of transgenic mice by anti-GluT-1 immunohistochemistry did not differ to that obtained in wild type mice before the onset of plaque deposition (younger than 10 months). However, in aged, 18-month-old Tg2576 mice, demonstrating high plaque loads, decreased blood vessel densities, particularly in layer IV of the somatosensory cortex, were observed. The data obtained in this study strongly support the idea of an age-related interplay between Aβ accumulation, cholinergic dysfunction, and vascular impairments. However, it remains to be elucidated as to which processes play a causative role and which events are secondary. A potential mechanism is provided by the vascular hypothesis of AD. Aging-, and life-style-associated damage of the brain microvasculature may affect Aβ clearance and perivascular drainage, promoting cerebrovascular Aβ deposition, inducing partial loss of cholinergic vascular innervation and changes in vascular function, angiogenesis and upregulation of the vesicular endothelial growth factor (VEGF) with consequences on APP processing and Aβ accumulation.

Glycoproteomic analysis of tissues from patients with colon cancer using lectin microarrays and nanoLC-MS/MS

The current study evaluated the glycoproteomic profile of tissues from colon cancer patients. The lectin microarray was first performed to compare the glycoprotein profiles between colon cancer and matched normal tissues. Level of N-acetylglucosamine (GlcNAc) that Solanum tuberosum lectin (STL) bound was found to be elevated in colon cancer, which was verified through lectin histochemistry. The subsequent glycoproteomic analysis based on STL enrichment of glycoproteins followed by label-free quantitative nano liquid chromatography-mass spectrometry/mass spectrometry (nanoLC-MS/MS) analysis identified 72 proteins in high confidence. Among these proteins, 17 were exclusively detected in cancer tissues, and 14 were significantly upregulated in tumor tissues. Annexin A1 and HSP90β were chosen for further investigation by immunoprecipitation coupled with lectin blots, western blots and tissue microarrays. Both Annexin A1 and HSP90β were GlcNAcylated, and their protein expressions were elevated in colon cancer, compared to normal tissues. Moreover, specific changes of GlcNAc abundances in Annexin A1 and HSP90β suggested that tumor-specific glycan patterns could serve as candidate biomarkers of colon cancer for distinguishing cancer patients from healthy individuals.

Lectin Histochemical Studies on the Vomeronasal Organ of the Sheep

The vomeronasal organ of sheep was examined using lectin histochemistry in order to compare the types and amounts of the glycoconjugates among various components of the vomeronasal sensory and non-sensory epithelia. In the vomeronasal sensory epithelium, Dolichos biflorus agglutinin (DBA) stained particular cells, located at the same level as the vomeronasal receptor cells, while the distribution, shape and number of the stained cells did not correspond to those of the vomeronasal receptor cells. Datura stramonium lectin (DSL), Concanavalin A (Con A), Phaseolus vulgaris agglutinin-E (PHA-E) and Phaseolus vulgaris agglutinin-L (PHA-L) labeled the basal cells of both vomeronasal sensory and non-sensory epithelia. While, Wheat germ agglutinin (WGA), Succinylated-wheat germ agglutinin (s-WGA), Lycopersicon esculentum lectin (LEL), Solanum tuberosum lectin (STL) and Ricinus communis agglutinin-I (RCA-120) labeled the basal cells of the sensory epithelium, and Bandeiraea simplicifolia lectin-I (BSL-I) stained the basal cells of the non-sensory epithelium, respectively. Seventeen lectins labeled the free border of both vomeronasal sensory and non-sensory epithelia, while Sophora japonica agglutinin (SJA), Jacalin and Pisum sativum agglutinin (PSA) labeled neither free border of the sensory nor that of non-sensory epithelia. The expression pattern of glycoconjugate was similar, but not identical, in the free border between the sensory and non-sensory epithelia. These results indicate that there are dissimilar features in the type and amount of glycoconjugates between the vomeronasal sensory and non-sensory epithelia, and at the same time, among the various cell types either in the vomeronasal sensory or non-sensory epithelium.

Histological and Lectin Histochemical Studies on the Main and Accessory Olfactory Bulbs in the Japanese Striped Snake, &lt;i&gt;Elaphe quadrivirgata&lt;/i&gt;

The main and accessory olfactory bulbs were examined by histological methods and lectin histochemistry in the Japanese striped snake. As the results, the histological properties are similar between the main olfactory bulb and the accessory olfactory bulb. In lectin histochemistry, 21 lectins used in this study showed similar binding patterns in the main olfactory bulb and the accessory olfactory bulb. In detail, 15 lectins stained these olfactory bulbs with similar manner, and 6 lectins did not stain them at all. Two lectins, Lycopersicon esculentum lectin (LEL) and Solanum tuberosum lectin (STL), stained the nerve and glomerular layers and did not stain any other layers in both olfactory bulbs. Four lectins, Soybean agglutinin (SBA), Vicia villosa agglutinin (VVA), Peanut agglutinin (PNA) and Phaseolus vulgaris agglutinin-L (PHA-L) stained the nerve and glomerular layers more intensely than other layers in both olfactory bulbs. In addition, VVA showed the dot-like stainings in the glomeruli of both olfactory bulbs. These findings suggest that the degree of development and the properties of glycoconjugates are similar between the main olfactory bulb and the accessory olfactory bulb in the Japanese striped snake.