USO DE LA LECTINA DE OLNEYA TESOTA (PF2) PARA LA PURIFICACIÓN Y DETECCIÓN HISTOQUÍMICA DE UNA PROTEÍNA DE CHOQUE TÉRMICO (GP14) DE BAZO/ USE OF THE Olneya tesota LECTIN (PF2) FOR PURIFICATION AND HISTOCHEMICAL DETECTION OF A SPLEEN HEAT SHOCK PROTEIN

PF2 es una lectina de las semillas de Olneya tesota (Palo fierro) con afinidad alta hacia carbohidratos complejos triantenarios tetrasialilados. El objetivo de este estudio fue localizar glicoconjugados que interactúen con PF2 en secciones de tejido de bazo y purificar glicoproteínas usando cromatografía de afinidad con PF2. Se incubaron secciones finas de tejido de bazo con PF2 biotinilada y se observaron interacciones con linfocitos en los nódulos y estructuras internas de tipo reticular en la pulpa blanca. Se prepararon extractos de bazo para purificar la glicoproteína detectada, usando dos etapas cromatográficas, intercambio iónico (CM-celulosa) y afinidad (PF2-agarosa). Por electroforesis nativa y con SDS se mostró una glicoproteína monomérica de 14 kDa (GP14). Una vez digerida con una proteasa, se identificaron seis secuencias peptídicas en la GP14. Cada péptido mostró 100% de homología a secuencias internas de diferentes proteínas de choque térmico de la familia de 70 kDa (HSP70). Esta similitud con las HSP fue confirmada por electrotransferencia e inmunodetección usando anticuerpos específicos para HSP70. En conclusión, la lectina PF2 tiene especificidad hacia un glicoconjugado denominado GP14 que pudiera ser una glicoproteína de estrés del bazo humano, identificada por primera vez. ABSTRACT PF2 is a lectin from Olneya tesota (Palo fierro) seeds with high affinity towards triantennary tetrasialylated complex carbohydrates. The goal of this study was to localize PF2-interacting glycoconjugates in spleen tissue thin sections, and to purify glycoproteins using PF2 affinity chromatography. Biotinylated PF2 was incubated with thin sections of fixed spleen tissues and interactions with lymphocytes in nodules of splenic white pulp were observed. Two chromatographic steps, using ion exchange chromatography (CM-cellulose) and affinity chromatography (PF2-agarose) made possible the purification of a glycoprotein from spleen extracts. Both SDS-PAGE and PAGE showed a 14-kDa monomeric glycoprotein (GP14). GP14 was protease digested and the sequences of six peptides were identified. Each of the GP14 peptides showed 100% homology to different internal sequences of the 70-kDa family of heat shock proteins (HSP70). The similarity with HSP was confirmed by Western blot using an antibody specific to HSP70. In conclusion, PF2 lectin has specificity toward a glycoconjugate named GP14 from spleen extract. GP14 could be a novel stress glycoprotein from human spleen

Antibacterial Activity of Bovine and Porcine Lactoferrins Against Escherichia coli K88+

Se comparó el efecto antibacteriano de las lactoferrinas (Lfs) bovina y porcina sobre Escherichia coli K88+ (E. coli K88+), uno de los principales agentes etiológicos de las diarreas en lechones en el hemisferio norte. Las Lfs se purificaron por cromatografía de intercambio iónico, confirmando su pureza por electroforesis en condiciones desnaturalizantes y reductoras (SDS-PAGE) en geles al 8% y por inmuno-detección con anticuerpos anti- lactoferrina. La actividad bacteriostática se probó utilizando concentraciones de 0,5 y 1,0 mg/mL de Holo (saturada de hierro) y Apo-Lf (libre de hierro). En todos los casos la actividad bacteriostática de las Holo-Lfs fue insignificante, mientras que en ambas concentraciones, la Apo-Lf bovina mostró un mayor efecto en la inhibición del crecimiento de la E. coli K88+ que la Apo-Lf porcina. La actividad bactericida se ensayó utilizando concentraciones de 2,0; 4,0; 6,0 y 8,0 mg/mL de Lfs bovina o porcina. La Lf bovina mostró efecto bactericida a una concentración de 8 mg/mL, mientras que la Lf porcina no presentó este efecto. Los resultados indican que la fuente bovina puede ser útil en la prevención de diarreas en lechones.473 - [email protected]@ciad.mxBimestralThe antibacterial effect of bovine lactoferrin (Lf) towards Escherichia coli K88+ (E. coli K88+) was compared with that of porcine Lf. E. coli K88+ is one of the main etiological agents of piglet diarrhea in the northern hemisphere. Lactoferrins (Lfs) were purified by ion exchange chromatography and further analyzed by electrophoresis using 8% sodium dodecyl sulfate polyacrylamide gel (SDS-PAGE) and immuno detection with anti-lactoferrin antibodies. Bacteriostatic effect was assayed using 0.5 and 1.0 mg/mL of Holo-Lf (iron saturated) or Apo-Lf (iron free). In all test the holo-LFs showed negligible bacteriostatic activity while for Apo-Lfs the bovine protein had the highest activity. Bactericide activity was assayed using bovine or porcine Lf concentrations of 2.0, 4.0, 6.0 y 8.0 mg/mL. Bovine Lf had bactericide activity at 8.0 mg/mL while no growth inhibition was observed with porcine Lf. These results suggest that bovine Lf may serve in the prophylaxis of piglets’ diarrhea

Core-shell nanoparticles from fucoidan neoglycans: Synthesis, characterization with capability of glycomimetic ligands for Campylobacter jejuni

Campylobacteriosis produced by Campylobacter jejuni is the main cause of bacterial gastroenteritis worldwide and the binding of this pathogen to fucosylated glycoconjugates expressed on host cells is a determining factor for the infection. The aim of this work was to synthesize and characterize core-shell nanoparticles from fucoidan neoglycans as potential glycomimetic ligands for Campylobacter jejuni. Fucoidan oligosaccharides (OFuc) were obtained by mild hydrolysis of fucoidan. OFuc were separated by ultrafiltration, characterized by dynamic light scattering (DLS), infrared spectroscopy (FTIR), Fluorophore-assisted carbohydrate electrophoresis and lectin recognition. The OFuc1 were then conjugated with bovine serum albumin (BSA) by controlled glycation. The formation of fucosylated BSA (BSA-OFuc1) was confirmed by FTIR, increased surface charge, polyacrylamide gel electrophoresis, mass spectrometry and Ulex europaeus I lectin recognition. Afterwards core-shell BSA/BSA-OFuc1 nanoparticles were obtained by crosslinking. These nanoparticles presented hemispherical form with size and charge distribution of 312.1 ± 3.2 nm and -34.4 ± 0.3 mV respectively and were selectively recognized by C. jejuni by an ELISA-like lectin assay (ELLA). These results revel the potential of core-shell nanoparticles from fucoidan neoglycans as glycomimetic ligands for Campylobacter jejuni and open further opportunities to explore potential applications in the pharmaceutic industry

Lactosylated Albumin Nanoparticles: Potential Drug Nanovehicles with Selective Targeting Toward an In Vitro Model of Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) ranks fifth in occurrence and second in mortality of all cancers. The development of effective therapies for HCC is urgently needed. Anticancer drugs targeted to the liver-specific asialoglycoprotein receptors (ASGPRs) are viewed as a promising potential treatment for HCC. ASGPRs facilitate the recognition and endocytosis of molecules, and possibly vehicles with galactose end groups, by the liver. In this study, bovine serum albumin (BSA) was conjugated with lactose using a thermal treatment. The formation of lactosylated BSA (BSA-Lac) was confirmed by a change of the chemical structure, increased molecular mass, and Ricinus communis lectin recognition. Subsequently, the low-crosslinking BSA-Lac nanoparticles (LC BSA-Lac NPs) and high-crosslinking BSA-Lac nanoparticles (HC BSA-Lac NPs) were synthesized. These nanoparticles presented spherical shapes with a size distribution of 560 ± 18.0 nm and 539 ± 9.0 nm, as well as an estimated surface charge of −26 ± 0.15 mV and −24 ± 0.45 mV, respectively. Both BSA-Lac NPs were selectively recognized by ASGPRs as shown by biorecognition, competition, and inhibition assays using an in vitro model of HCC. This justifies pursuing the strategy of using BSA-Lac NPs as potential drug nanovehicles with selective direction toward hepatocellular carcinoma

Purification, Biochemical Characterization, and Bioactive Properties of a Lectin Purified from the Seeds of White Tepary Bean (Phaseolus Acutifolius Variety Latifolius)

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Global attitudes in the management of acute appendicitis during COVID-19 pandemic: ACIE Appy Study

Background: Surgical strategies are being adapted to face the COVID-19 pandemic. Recommendations on the management of acute appendicitis have been based on expert opinion, but very little evidence is available. This study addressed that dearth with a snapshot of worldwide approaches to appendicitis. Methods: The Association of Italian Surgeons in Europe designed an online survey to assess the current attitude of surgeons globally regarding the management of patients with acute appendicitis during the pandemic. Questions were divided into baseline information, hospital organization and screening, personal protective equipment, management and surgical approach, and patient presentation before versus during the pandemic. Results: Of 744 answers, 709 (from 66 countries) were complete and were included in the analysis. Most hospitals were treating both patients with and those without COVID. There was variation in screening indications and modality used, with chest X-ray plus molecular testing (PCR) being the commonest (19\ub78 per cent). Conservative management of complicated and uncomplicated appendicitis was used by 6\ub76 and 2\ub74 per cent respectively before, but 23\ub77 and 5\ub73 per cent, during the pandemic (both P < 0\ub7001). One-third changed their approach from laparoscopic to open surgery owing to the popular (but evidence-lacking) advice from expert groups during the initial phase of the pandemic. No agreement on how to filter surgical smoke plume during laparoscopy was identified. There was an overall reduction in the number of patients admitted with appendicitis and one-third felt that patients who did present had more severe appendicitis than they usually observe. Conclusion: Conservative management of mild appendicitis has been possible during the pandemic. The fact that some surgeons switched to open appendicectomy may reflect the poor guidelines that emanated in the early phase of SARS-CoV-2