Development of a sandwich ELISA for quantifying hepcidin in Rainbow trout

Santana, Paula A.; Álvarez, Claudio; Guzmán, Fanny; Mercado, Luís

doi:10.1016/j.fsi.2013.06.005

Cited by 32 publications

(16 citation statements)

References 39 publications

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“…The peptide changes to an alpha-helix conformation when in the reduced state, while it shows a β-sheet structure when in the oxidized state through the formation of M A N U S C R I P T A C C E P T E D ACCEPTED MANUSCRIPT disulfide bonds [14]. Furthermore, we have shown the presence of this peptide in trout liver and anterior kidney, and that it is upregulated in response to Escherichia coli or Aeromonas salmonicida lipopolysaccharide [15,16]. These results suggest that it participates in cellular defense responses of teleost fish against pathogen-associated molecular patterns (PAMPs), possibly being involved in post-invasion by the infectious agents [17].…”

Section: Introductionmentioning

confidence: 75%

Synthetic hepcidin from fish: Uptake and protection against Vibrio anguillarum in sea bass (Dicentrarchus labrax)

Álvarez

Acosta

Montero

et al. 2016

Fish & Shellfish Immunology

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Section: Introductionmentioning

confidence: 75%

Synthetic hepcidin from fish: Uptake and protection against Vibrio anguillarum in sea bass (Dicentrarchus labrax)

Álvarez

Acosta

Montero

et al. 2016

Fish & Shellfish Immunology

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“…Antiserum specificities were evaluated through an indirect enzyme-linked immunosorbent assay and immunoblotting by seeding each peptide (at 1, 0.5, 0.25, and 0.125 μg) onto a nitrocellulose membrane (0.45 μm; Thermo Scientific) (Fig C and D in S1 File ). The membranes were blocked and washed as previously reported [ 28 ], and then incubated with the anti-IL-10 antiserum and pre-immune antiserum for 1 h. After this, the membranes were washed and incubated with anti-mouse IgG-HRP (Thermo Scientific) at a 1:7000 dilution. The membranes were revealed with the Enhanced Chemiluminescence Western Blotting Substrate (Pierce, Thermo Scientific).…”

Section: Methodsmentioning

confidence: 99%

“…The membranes were revealed with the Enhanced Chemiluminescence Western Blotting Substrate (Pierce, Thermo Scientific). Evaluations of hepcidin were performed using a previously reported specific antiserum [ 27 , 28 ].…”

Section: Methodsmentioning

confidence: 99%

Piscirickettsia salmonis Imbalances the Innate Immune Response to Succeed in a Productive Infection in a Salmonid Cell Line Model

et al. 2016

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Piscirickettsia salmonis is a facultative intracellular bacterium that causes the disease called “salmon rickettsial syndrome”. Attempts to control this disease have been unsuccessful, because existing vaccines have not achieved the expected effectiveness and the antibiotics used fail to completely eradicate the pathogen. This is in part the product of lack of scientific information that still lacks on the mechanisms used by this bacterium to overcome infected–cell responses and survive to induce a productive infection in macrophages. For that, this work was focused in determining if P. salmonis is able to modify the expression and the imbalance of IL-12 and IL-10 using an in vitro model. Additionally, we also evaluated the role the antimicrobial peptide hepcidin had in the control of this pathogen in infected cells. Therefore, the expression of IL-10 and IL-12 was evaluated at earlier stages of infection in the RTS11 cell line derived from Oncorhynchus mykiss macrophages. Simultaneously, the hepcidin expression and location was analyzed in the macrophages infected with the pathogen. Our results suggest that IL-10 is clearly induced at early stages of infection with values peaking at 36 hours post infection. Furthermore, infective P. salmonis downregulates the expression of antimicrobial peptide hepcidin and vesicles containing this peptide were unable to merge with the infective bacteria. Our results suggest that P. salmonis is able to manipulate the behavior of host cytokines and likely might constitute a virulence mechanism that promotes intracellular bacterial replication in leukocytes cells lines of trout and salmon. This mechanism involves the generation of an optimum environment for the microorganism and the downregulation of antimicrobial effectors like hepcidin.

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“…To reduce non-specific binding, slides were incubated with 2% bovine serum albumin (BSA) in PBS for 10 min at RT. The sections were then incubated with mouse anti-human MICA/B polyclonal antibody generated as previously described ( 32 , 33 ) (1:50 dilution in 2% BSA/PBS) for 1 h at 37°C in a humid chamber. The specificity of the reaction was tested by omission of the primary antibody.…”

Section: Methodsmentioning

confidence: 99%

Clinical significance of tumor expression of major histocompatibility complex class I-related chains A and B (MICA/B) in gastric cancer patients

et al. 2015

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Gastric cancer (GC) is the third most common cause of cancer death worldwide. Natural killer cells play an important role in the immune defense against transformed cells. They express the activating receptor NKG2D, whose ligands belong to the MIC and ULBP/RAET family. Although it is well established that these ligands are generally expressed in tumors, the association between their expression in the tumor and gastric mucosa and clinical parameters and prognosis of GC remains to be addressed. In the present study, MICA and MICB expression was analyzed, by flow cytometry, in 23 and 20 pairs of gastric tumor and adjacent non-neoplasic gastric mucosa, respectively. Additionally, ligands expression in 13 tumors and 7 gastric mucosa samples from GC patients were evaluated by immunohistochemistry. The mRNA levels of MICA in 9 pairs of tumor and mucosa were determined by quantitative PCR. Data were associated with the clinicopathological characteristics and the patient outcome. MICA expression was observed in 57% of tumors (13/23) and 44% of mucosal samples (10/23), while MICB was detected in 50% of tumors (10/20) and 45% of mucosal tissues (9/20). At the protein level, ligand expression was significantly higher in the tumor than in the gastric mucosa. MICA mRNA levels were also increased in the tumor as compared to the mucosa. However, clinicopathological analysis indicated that, in patients with tumors >5 cm, the expression of MICA and MICB in the tumor did not differ from that of the mucosa, and tumors >5 cm showed significantly higher MICA and MICB expression than tumors ≤5 cm. Patients presenting tumors >5 cm that expressed MICA and MICB had substantially shorter survival than those with large tumors that did not express these ligands. Our results suggest that locally sustained expression of MICA and MICB in the tumor may contribute to the malignant progression of GC and that expression of these ligands predicts an unfavorable prognosis in GC patients presenting large tumors.

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Development of a sandwich ELISA for quantifying hepcidin in Rainbow trout

Cited by 32 publications

References 39 publications

Synthetic hepcidin from fish: Uptake and protection against Vibrio anguillarum in sea bass (Dicentrarchus labrax)

Synthetic hepcidin from fish: Uptake and protection against Vibrio anguillarum in sea bass (Dicentrarchus labrax)

Piscirickettsia salmonis Imbalances the Innate Immune Response to Succeed in a Productive Infection in a Salmonid Cell Line Model

Clinical significance of tumor expression of major histocompatibility complex class I-related chains A and B (MICA/B) in gastric cancer patients

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