Presence of erythroglycan on human K-562 chronic myelogenous leukemia-derived cells

Total glycopeptides from human K-562 cells, labeled metabolically with [3H]glucosamine or [3H]mannose, were prepared by extracting the cells with organic solvents to remove lipids and by digesting the residue with pronase. 3H-labeled glycopeptides were fractionated on Sephadex G-50 revealing a high molecular weight fraction (M(r) = 7,000 to 11,000), comprising approximately 10% of the [3H]glucosamine and 25% of the [3H]mannose label. Digestion of this glycopeptide fraction with endo-β-galactosidase from Escherichia freundii, specific for a repeating structure of Gal(β1 → 4)GlcNAc(β1→3), results in the following four products as resolved by Bio-Gel P-2 gel filtration: 1) a disaccharide with the structure β-2-deoxy-2-acetamidoglucosyl → β-galactose; 2) a trisaccharide with the structure β-galactosyl → β-2-deoxy-2-acetamidoglucosyl → β-galactose; 3) a tetrasaccharide with the sequence α-N-acetylneuraminyl → β-galactosyl → β-2-deoxy-2-acetamidoglucosyl → β-galactose; and 4) a larger, complex fragment which contains mannose and β-2-deoxy-2-acetamidoglucose and which is probably the protein linkage region. In addition, visualization of radiolabeled glycoproteins by fluorography on polyacrylamide gels revealed a 105,000-dalton \u27Band 3\u27-like glycoprotein and other bands that were sensitive to endo-β-galactosidase. These results indicate that the K-562 cell line bears a glycopeptide, erythroglycan, which has been found on erythrocytes, and that this polymer is expressed mainly in the fetal form as a linear chain

Synthesis of the branched form of erythroglycan by Friend GM 979 erythroleukemic cells

Mouse GM979 erythroleukemic cells were found to synthesize the branched form of erythroglycan, a large polylactosamine structure known to be attached to band 3 and possibly to other proteins on human erythrocytes. Total protein-derived oligosaccharides from GM979 cells, labeled metabolically with [3H]glucosamine, [3H]mannose, or [3H]galactose, were prepared by hydrazinolysis after extracting the lipids. The 3H-labeled oligosaccharides were fractionated on Sephadex G-50 revealing 10-25% of each labeled product as a high molecular weight fraction (M(r)=10,000). Digestion of this [3H]glucosamine fraction with endo-β-galactosidase from Escherichia freundii, specific for the repeating structure of Galβ(1→4)GlcNAcβ(1→3), resulted in the following four products upon Bio-Gel P-2 Gel filtration; 1) a disaccharide with the structure GlcNAcβ(1→3)Gal,2) a trisaccharide with the structure Galβ(1→4)GlcNAcβ(1→3)βGal,3) a tetrasaccharide with the sequence Fucα(1 → 2)Galβ(1 → 4)GlcNacβ(1→3)Gal, and 4) a large complex fragment which contained mannose, glucosamine, galactose, and fucose (presumably the protein linkage region). Methylation linkage analysis of the large complex fraction shows mainly the presence of 4-substituted and terminal N-acetylglucosamine; 3,6-substituted, 6-substituted, 2-substituted, and 2,3-substituted galactose. The GM979 cell erythroglycan is only 30% susceptible to endo-β-galactosidase degradation probably because of the branched galactose residues, whereas the linear form of erythroglycan from human K562 cells is 60% susceptible. The branched residues in GM979 cell saccharides indicate that this mouse cell line bears an arborized erythroglycan-like glycopeptide similar to those found on human adult erythrocytes, and thus may be a source for the enzyme which transfers an N-acetylglucosamine residue to a 3-linked galactose to form a 3,6-disubstituted galactose

<i>Leishmania donovani<i/>-derived lipophosphoglycan plus BCG induces a Th1 type immune response but does not protect Syrian golden hamsters (<i>Mesocricetus auratus<i/>) and BALB/c mice against <i>Leishmania donovani<i/>

The efficacy of Leishmania donovani-derived lipophosphoglycan (LPG) plus Mycobacterium bovis Bacille Calmette-Guérin (BCG) as a vaccine candidate against visceral leishmaniosis in susceptible BALB/c mouse and Syrian golden hamster (Mesocricetus auratus) models was investigated. Following a triple vaccination with a total dose of 150 µl BCG plus 60 µg or 30 µg of LPG for hamsters and BALB/c mice respectively, there were no noticeable side effects both locally and systemically; implying that the molecule was safe at this dosage level. Vaccinated animals demonstrated an activation of both the humoral as well as cell-mediated responses to LPG, which correlated with resistance against the disease. Protection by LPG plus BCG, was however, poor as the remaining immunized animals showed disease progression leading to severity of the disease as illustrated by emaciation, mass loss and heavy splenic parasitaemia in hamsters. These data nevertheless suggest that it may be rewarding to further evaluate the potential of LPG as a vaccine candidate in leishmaniosis using other adjuvants, which may enhance its immunogenicity

Leishmania donovani-derived lipophosphoglycan plus BCG induces a Th1 type immune response but does not protect Syrian golden hamsters (Mesocricetus auratus) and BALB/c mice against Leishmania donovani

The efficacy of Leishmania donovani-derived lipophosphoglycan (LPG) plus Mycobacterium bovis Bacille Calmette-Guerin (BCG) as a vaccine candidate against visceral leishmaniosis in susceptible BALB/c mouse and Syrian golden hamster (Mesocricetus auratus) models was investigated. Following a triple vaccination with a total dose of 150 µI BCG plus 60 µg or 30 µg of LPG for hamsters and BALB/c mice respectively, there were no noticeable side effects both locally and systemically; implying that the molecule was safe at this dosage level. Vaccinated animals demonstrated an activation of both the humoral as well as cell-mediated responses to LPG, which correlated with resistance against the disease. Protection by LPG plus BCG, was however, poor as the remaining immunized animals showed disease progression leading to severity of the disease as illustrated by emaciation, mass loss and heavy splenic parasitaemia in hamsters. These data nevertheless suggest that it may be rewarding to further evaluate the potential of LPG as a vaccine candidate in leishmaniosis using other adjuvants, which may enhance its immunogenicity.The articles have been scanned with a HP Scanjet 8300; 600dpi, saved in TIFF format. Adobe Acrobat v.9 was used to OCR the text and also for the merging and conversion to the final presentation PDF-format.Intemational Society for Infectious diseases (lSID). Intemational Atomic Agency (IAEA). Kenya Medical Research Institute.mn201

TRANSMISSION BLOCKING VACCINE STUDIES IN LEISHMANIASIS: 11. EFFECT OF IMMUNISATION USING LEISHMANIA MAJOR DERIVED 63 KILODALTON GLYCOPROTEIN, LIPOPHOSPHOGLYCAN AND WHOLE PARASITE ANTIGENS ON THE COURSE OF L. MAJOR INFECTION IN BALBIC MICE

Background: Safe, effective and inexpensive vaccines may be the most practical tool for controlof any form of leishmaniasis. Leishmaniasis produces a state of pre-immunition which is theunderlying mechanism for prolonged immunity to re-infection. Low doses of parasites has beenshown to beable to induce protection in mice. It is not known, however, how immunesel-a froma susceptible host imrnunised with Leishmania-derived antigens when taken in by the sandflyaffects the development and the subsequent transmission of the parasite to naive hosts.Objective: To monitor the course of disease in BALBlc mice following challenge using L.rnajor infected P. duboscqi which had previously fed on immunised mice.Methods: BALBIc mice were immunised adequately withLeishrnania major-derived antigensnamely, crude whole parasite (WPA), recombinant 63 kilodalton glycoprotein (rgp63),lipophosphoglycan (LPG;) ancl a cocktail composed of rgp63 plus LPG antigens. Laboratoryreared Phlebotornus duboscqi sandflies, the natural vector for L. major were later allwwed tofeed on immunised animals, interrupted and allowed to continue feeding on infected animalsfor an equal amount of time until they became fully engorged. The sandflies were maintainedon apples as a carbohydrate source in an insectary maintained at a temperature of 2S°C and80% relative humidity. On the seventh day these sandflies were used to infect naivc B:ALB/c mice and the course of infection followed for a period of at least three months.Results: Mice infected usingsandflies which had previously fed on WPA or rgp63-immunizedmice showed disease exacerbation as the infection progressed, whereas those infected usingsandflies which had previously fed on LPG-immunised mice had the least lesion sizescompared to control mice infected using sandflies which had fed on saline immunised mice(p&lt;0.05).Conclusions: Results from this study indicate that the course of L. major infection in BALBIc mice was dependent on the infective dose of parasites transmitted by the sandflies. R~:sultsfrom this study suggests that sub-infective doses of the parasite from sandflies previously fedon animals immunised with Leishmania-derived antigens needs to be evaluated for theirpotential in vaccine development against Leishmania infections

TRANSMISSION BLOCKING VACCINE STUDIES IN LElISHMANIASIS: I . LIPOPHOSPHOGLYCAN IS A PROMISING TRANSMISSION BLOCKING VACCINE MOLECULE AGAINST CUTANEOUS LEISHMANIASIS

Background: New strategies for control of leishmaniasis is needed as chemotherapy usingantimonial drugs is prolonged, expensive, associated with side effects and relapses. Vectorcontrol has limitations and a vaccine which may be the best approach is not available.Objectives: To assess the level of inhibition of promastigote development and gut morphologyin infected Phlebotomus dubotcqi sandflies fed on different groups of BALBlc mice immunisedwith rgp63, lipophosglycan (LPG) or their cocktail and whole parasite antigens preparedfrom L. major culture-derived promastigotes.Methods: BALBlc mice were immunised adequately with Leishmania major-derived antigensnamely, crude whole para5ite ( WPA), recombinant 63 kilodalton glycoprotein (rgp63), LPG anda cocktail composed of rgp63 plus LPG antigens . Laboratory reared Phlebotomus duboscqisandflies, the natural vector for I, major were later allowed to feed on immunised animals,interrupted and allowed to continue feeding on infected animals for an equal amount of tirne untilthey became fully engorged. The sandflies were maintained on apples as a carbohydrate sourcein an insectary maintained at a temperature of 2S°C and 80% relative humidity. Some of thesandflies were dissected on days 2,4 and 6 after feeding and observed using the light :and thetransmission electron microscopy for any changes in their gut morphology. The remainingsandflies werealldissectdon thesixthday post-feeding and examinedfor procyclics,necton~onads,haptomonads and metacyclic promastigote forms of Leishmania.Results: Sandflies which had previously fed on WPA, LPG plus rgp63 cocktail ant1 LPGimmunisedmice showed the lowest infection rates compared to control sandflies fed on salineimmunised mice (p&lt;0.05). A significant number of procyclic promastigotes, th~e firstdevelopmental form of the parasite in culture as well as in the sandfly was observed insandflies which fed on LPG-immunised mice (pe0.05). The dominant parasite form insandflies which fed on rgp63 or LPG-immunised mice was the nectomonad form but very fewof the infective metacyclic forms (pe0.05). Control sandflies fed on saline immunised orinfected mice alone displayed a normal pattern of parasite development up to the metacyclicstage. Studies showed that two possible mechanisms through which immune sera fromimmunised mice may came inhibition of parasite development is by exflagellation ofnectomonad forms and degeneration of the sandfly midgut epithelium as revealed by lightand electron microscopy studies respectively.Conclusions: This studj has shown that immune-mediated transmission blocking rnay beapplied to Leishmania infections. Based on observation of the procyclic promastigoles, thedominance of the nectomonad forms, low infectivity rates in sandflies fed on LPG-immunisedmice, we concluded that LPG stands out to be a promising transmission blocking vaccinecandidate in leishmaniasis

Fast wave direct electron heating in advanced inductive and ITER baseline scenario discharges in DIII-D

Fast Wave (FW) heating and electron cyclotron heating (ECH) are used in the DIII-D tokamak to study plasmas with low applied torque and dominant electron heating characteristic of burning plasmas. FW heating via direct electron damping has reached the 2.5 MW level in high performance ELMy H-mode plasmas. In Advanced Inductive (AI) plasmas, core FW heating was found to be comparable to that of ECH, consistent with the excellent first-pass absorption of FWs predicted by ray-tracing models at high electron beta. FW heating at the ∼2 MW level to ELMy H-mode discharges in the ITER Baseline Scenario (IBS) showed unexpectedly strong absorption of FW power by injected neutral beam (NB) ions, indicated by significant enhancement of the D-D neutron rate, while the intended absorption on core electrons appeared rather weak. The AI and IBS discharges are compared in an effort to identify the causes of the different response to FW

Emerald zoyzia grass development regarding photosynthetically active radiation in different slopes Desenvolvimento da grama-esmeralda em relação à radiação fotossinteticamente ativa em diferentes declividades

With this study, the objective was to estimate the photosynthetically active radiation (PAR) and to correlate it with the dry matter (MMSPA) of the emerald zoysia (Zoysia japonica Steud.) on surfaces with different expositions and slopes. The research was conducted at the Experimental Watershed of the Agricultural Engineering Department, School of Agriculture and Veterinary Sciences of São Paulo State University (FCAV/UNESP), Brazil, where the surfaces (H, 10 N, 30 N, 50 N, 10 S, 30 S, 50 S, 10 L, 30 L, 50 L, 10 O, 30 O and 50 O) were used. To obtain the global solar radiation, it was installed an automated weather station where the PAR (dependent variable) was obtained by the equation y = a + bx, and the global radiation was independent. To compare means of MMSPA, it was used the Tukey test at 5% probability, and to assess the relation PAR/MMSPA, the simple linear correlation coefficient. The result showed that the accumulation of these effects in the PAR increases with North exposure and decreases with the South, and exposure to 50N is most suitable for slopes, not having correlation between the PAR and the MMSPA for the surfaces evaluated for the study period.<br>Com este trabalho, o objetivo foi estimar a radiação fotossinteticamente ativa (PAR) e correlacioná-la com a massa de matéria seca (MMSPA) da grama-esmeralda (Zoysia japonica Steud.), em superfícies com diferentes exposições e declividades. A pesquisa foi desenvolvida na Bacia Hidrográfica Experimental do Departamento de Engenharia Rural, FCAV/UNESP, Brasil, onde foram utilizadas as superfícies (H; 10 N; 30 N; 50 N; 10 S; 30 S; 50 S; 10 L; 30 L; 50 L; 10 O; 30 O e 50 O). Para a obtenção da radiação solar global, foi instalada uma estação meteorológica automatizada, onde a PAR (variável dependente) foi obtida por meio da equação y = a + bx, e a radiação global foi a independente. Para comparação de médias da MMSPA, utilizou-se o teste de Tukey, a 5% de probabilidade, e para verificar a relação existente PAR/MMSPA, o coeficiente de correlação linear simples. O resultado mostrou que o acúmulo desses efeitos na PAR aumenta com a exposição norte e decresce com a sul, sendo a exposição 50 N a mais indicada para taludes, não havendo correlação entre a PAR e a MMSPA para as superfícies avaliadas para o período estudado