Transcriptional and proteomic analysis reveal recombinant galectins of Haemonchus contortus down-regulated functions of goat PBMC and modulation of several signaling cascades in vitro (original) (raw)
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Biomolecules
Galectins are glycan-binding proteins that are widely expressed and distributed in mammalian tissues as well as cells of innate and adaptive immune responses. CD4+ T-helper cells differentiate into effector subsets in response to cytokines. T helper 9 cells are one of the recently described subsets of effector T cells that are relatively new and less studied. In this study, galectin domain containing protein from Haemonchus contortus (Hc-GDC) was cloned, expressed in pET32a, and immunoblotting was performed. Localization of recombinant (r)Hc-GDC on outer and inner surface of H. contortus worm and binding with goat Peripheral Blood Mononuclear cells (PBMCs) were performed using immunofluorescence assay. Moreover, effects of rHc-GDC on proliferation, apoptosis, cell migration, and the nitric oxide production in goat PBMCs were evaluated. Furthermore, modulatory effects of rHc-GDC on production of Th1, Th2, and Th9 cells were evaluated by flowcytometry and on interferon gamma, interleu...
Parasites & vectors, 2015
Hco-gal-m and -f were two isoforms of galectin cloned from male and female Haemonchus contortus, respectively, and it was demonstrated that recombinant Hco-gal-m and -f could act as immune suppressors. However, little is known about the receptors or binding partners of these galectins in the host. The research of the molecular mechanisms that govern the interactions between these galectins and host molecules will fill a gap in our understanding how parasite galectins interact with host cells. A yeast two-hybrid system was used to identify the binding partners of Hco-gal-m and -f in this research. The interaction between rHco-gal-m and candidate binding protein was validated by co-immunoprecipitation. The localization of transmembrane protein 63A (TMEM63A) in peripheral blood mononuclear cells (PBMCs) was detected by immunofluorescence. The distribution of TMEM63A in T cells, B cells and monocytes in PBMCs was detected by flow cytometry. The immunomodulatory effects of Hco-gal-m and ...
PeerJ, 2018
is the most pathogenic nematode of small ruminants. Infection in sheep and goats results in anaemia that decreases animal productivity and can ultimately cause death. The involvement of ruminant-specific galectin-11 (LGALS-11) and galectin-14 (LGALS-14) has been postulated to play important roles in protective immune responses against parasitic infection; however, their ligands are unknown. In the current study, LGALS-11 and LGALS-14 ligands in were identified from larval (L4) and adult parasitic stages extracts using immobilised LGALS-11 and LGALS-14 affinity column chromatography and mass spectrometry. Both LGALS-11 and LGALS-14 bound more putative protein targets in the adult stage of (43 proteins) when compared to the larval stage (two proteins). Of the 43 proteins identified in the adult stage, 34 and 35 proteins were bound by LGALS-11 and LGALS-14, respectively, with 26 proteins binding to both galectins. Interestingly, hematophagous stage-specific sperm-coating protein and zi...
Galectins and collectinis expression are increased in Haemonchus contortus-infected corriedale sheep
Revista Brasileira de Parasitologia Veterinária, 2015
Galectins and collectins are proteins classified in the lectin family that have the ability to recognize molecular patterns associated with pathogens. Studies on cattle have demonstrated high expression of these proteins during infection with gastrointestinal nematodes. The aim of this study was to investigate whether the level of Haemonchus contortus infection would alter the expression of galectins (Gal11 and Gal14) and collectins (SPA and CGN) in sheep. Twelve Corriedale sheep exposed to natural infection with nematodes were divided into two groups: group 1 (G1, n = 7) and group 2 (G2, n = 5), with low and high parasite burdens, respectively, based on fecal egg counts and abomasal parasite counts. The fecal egg counts and abomasal parasite counts were significantly different (p < 0.05) between the groups. Galectin and collectin gene expression was observed in all sheep abomasal samples. However, animals with lower infection levels showed lower expression of the genes Gal14, SP...
Journal of Animal Science and Technology
Pathogen-associated Molecular Patterns (PAMPs) are highly conserved structural motifs that are recognized by Pathogen Recognition receptors (PRRs) to initiate immune responses. Infection by these pathogens and the immune response to PAMPS such as lipopolysaccharide (LPS), Peptidoglycan (PGN), bacterial oligodeoxynucleotides [CpG oligodeoxynucleotides 2006 (CpG ODN2006) and CpG oligodeoxynucleotides 2216 (CpG ODN2216)], and viral RNA Polyinosinic-Polycytidylic Acid (Poly I:C), are associated with infectious and metabolic diseases in animals impacting health and production. It is established that PAMPs mediate the production of cytokines by binding to PRRs such as Toll-like receptors (TLR) on immune cells. Galectins (Gal) are carbohydrate-binding proteins that when expressed play essential roles in the resolution of infectious and metabolic diseases. Thus it is important to determine if the expression of galectin gene (LGALS) and Gal secretion in blood are affected by exposure to LPS and PGN, PolyI:C and bacterial CpG ODNs. LPS increased transcription of LGALS4 and 12 (2.5 and 2.02 folds respectively) and decreased secretion of Gal 4 (p < 0.05). PGN increased transcrip
Toll-Like Receptor Agonists Modulate Galectin Gene Expression in Goat Blood
Journal of Applied Biotechnology
Pathogenesis and lack of effective therapeutics pose a threat to animal and public health. Recognition of pathogen-associated molecular patterns (PAMPS) by Toll-like receptors is associated with gene activation. Galectins are sugar-binding proteins that play a role in pathogen-recognition and immune modulation. The objective of this study was to investigate the effect of different TLR agonists on galectin gene expression and secretion in goat blood. Blood was collected from the jugular vein of BoerXSpanish goats (n=3). Blood was treated with 10 µg/ml Lipopolysaccharide, Peptidoglycan, 12.5 µg/ml of Polyinosinic-polycytidylic acid, 10 µg/ml CpG ODN (2216) and 10 µg/ml CpG ODN (2006) or PBS control. Secretion of galectin-1, -3 and -9 was measured by ELISA. Expression of LGALS-1, -2, -3, -4, -7, -8, -9, -11, -12, -14, -15, -16 was assessed using real-time PCR. Beta-actin and GAPDH served as controls. Fold change in gene expression was calculated using the 2−ΔΔCt method. Data were analy...
International Journal for Parasitology, 2011
Gastrointestinal nematode parasites undergo several developmental stages within their mammalian host, each presenting different antigenic challenges to the immune system. To examine the expression of different immune mediators over time, biopsy samples were collected from the cannulated abomasum (true stomach) of immune sheep at several times after a challenge infection with Haemonchus contortus L3s. IL-5 and IL-13 mRNA expression levels were significantly increased above saline-challenged control levels at 5 and 7 days post challenge, while IL-4 showed an earlier peak at day 2 post challenge. IL-5, IL-13 and IL-4, as well as IFN-c mRNA levels, peaked at 7 days before decreasing to non-significant levels at 28 days post challenge. TNF-a followed a similar profile while there was a slight increase in TGF-b in both control and challenged sheep. There was a significant increase in galectin-14 mRNA in the L3 challenged compared with the saline challenged group at 7 days while both galectin-11 protein and mRNA levels increased significantly by day 3 post challenge, peaking at 5-7 days post challenge. Distinct correlations were observed between these immune parameters at different times after L3 challenge. The galectin-14 protein level at day 2 post challenge was the only measured mediator significantly negatively correlated with worm burden. These studies highlight the dynamic nature of the immune response during parasite infection and the need to consider the different life cycle stages involved.
1 Systemic expression of galectin genes in 2 periparturient goats 3
2018
Galectins constitute an evolutionarily conserved family of β-galactoside-binding proteins. 14 They regulate innate and adaptive immunity and homeostasis. Expression of Galectins may regulate 15 periparturient immune suppression. Galectin gene expression was studied in goat blood during the 16 periparturient period. Body weight, body condition and FAMACHA scores, and fecal and blood 17 samples were collected from Five BoerXSpanish goats at 14 days and 7 days after parturition. Fecal 18 samples were used to assess parasite load. Total RNA was isolated from blood using Trizol and 19 converted to cDNA for real-time PCR using specific primers for goat LGALs-1, -2, -3, -4, -7, -8, -9, 20 11, -12, -14, -15, -16, and ligand Gal3bp, T-cell immunoglobulin domain, and mucin domain 3(TIM21 3). Beta-actin and GAPDH housekeeping genes were used as internal controls. Fold changes in 22 transcript abundance were compared to non-pregnant goats and calculated using the Livak method. 23 Secretion of G...
Systemic Expression of Galectin Genes in Periparturient Goats
Galectins constitute an evolutionarily conserved family of β-galactoside-binding proteins. They regulate innate and adaptive immunity and homeostasis. Expression of Galectins may regulate periparturient immune suppression. Galectin gene expression was studied in goat blood during the periparturient period. Body weight, body condition and FAMACHA scores, and fecal and blood samples were collected from Five BoerXSpanish goats at 14 days and 7 days after parturition. Fecal samples were used to assess parasite load. Total RNA was isolated from blood using Trizol and converted to cDNA for real-time PCR using specific primers for goat LGALs-1, -2, -3, -4, -7, -8, -9, -11, -12, -14, -15, -16, and ligand Gal3bp, T-cell immunoglobulin domain, and mucin domain 3(TIM-3). Beta-actin and GAPDH housekeeping genes were used as internal controls. Fold changes in transcript abundance were compared to non-pregnant goats and calculated using the Livak method. Secretion of GALS-1, -3 and -9 in pla...
International journal of biological macromolecules, 2018
Galectins are mammalian lectins characterized by affinity for β-galactosides and the presence of a conserved carbohydrate recognition domain (CRD). Galectins play crucial role in the causation and progression of deadly human diseases like cancer, neurodegenerative disorders and cardiovascular disorders. Available literature reports relevant roles of galectins in innate as well as adaptive immune responses, along with the modulation of acute inflammatory response. In the current study, we purified the goat heart galectin-1 (GHG-1) and carried out its extensive immunological studies. Immunodiffusion studies revealed that anti-GHG-1 antibodies recognize the GHG-1 more readily as compared to the other galectins, suggesting its preferred utilization in various recognition studies. Antigenic cross-reactivity between galectins isolated from different tissues and species suggest their evolutionary preserved fundamental biological roles. A gradual increase in the lysozyme release was evident...