Evaluating serum proteinogram methodologies for the diagnosis of inflammation in fish: Acute and chronic patterns in gilthead seabream (Sparus aurata) injected with λ-carrageenan.

Abstract

Proteinogram is a semiquantitative method specially used in clinic to separate the serum proteins from patients for use in the diagnosis of diseases. However, this methodology has only been applied very recently with this approach in farmed fish. Thus, the aim of this study was to explore the changes in the serum proteinogram of gilthead seabream (Sparus aurata), after triggering an acute or chronic sterile inflammation. For this, two experiments were carried out: i) Acute inflammation experiment: seabream specimens were injected intramuscularly with 50 μL of λ-carrageenan (0.5 mg fish-1) or buffer (control) and blood samples were collected at 3, 6 and 24 h post-injection; ii) Chronic inflammation experiment: specimens were injected at 0, 7 and 14 days with 500, 250 and 250 μL of λ-carrageenan, respectively (20 mg fish-1) or buffer, and blood samples were collected at 15 days post-injection. In both cases, serum was obtained and processed by electropherograms and HPLC-mass spectrometry. Results of electropherograms of control fish revealed four major proteins of 19.5, 76.3, 104.4, and 156.7 kDa in the serum. These four proteins were correlated with apolipoprotein A-II (II (the counterpart of mammalian albumin, Apo fraction), serotransferrin (β fraction), inter-α-trypsin inhibitor heavy chain H3-like (α1 fraction) and α-2-macroglobulin-like (α2 fraction) according to the results obtained with HPLC-mass spectrometry. In a statistical view (p<0.05), no variations were detected in the four major serum protein bands between the control and the acutely inflamed groups. However, in chronically inflamed fish, the Apo fraction decreased statistically compared to the control group. In contrast, the α1 and α2 fractions were statistically increased in the serum of fish sampled 15 days after λ-carrageenan injection, compared to those found in the control fish. α1 and α2 protein fractions are recognized indicators of inflammation in mammals. Consequently, our study presents a novel method for assessing both acute and chronic λ-carrageenan-induced sterile inflammation in gilthead seabream, which could be applicable to other marine species for diagnostic purposes.