Seawater acidification induced immune function changes of haemocytes in Mytilus edulis: a comparative study of CO2 and HCl enrichment

Abstract

The present study was performed to evaluate the effects of CO₂- or HCl-induced seawater acidification (pH 7.7 or 7.1; control: pH 8.1) on haemocytes of Mytilus edulis, and the changes in the structure and immune function were investigated during a 21-day experiment. The results demonstrated that seawater acidification had little effect on the cellular mortality and granulocyte proportion but damaged the granulocyte ultrastructure. Phagocytosis of haemocytes was also significantly inhibited in a clearly concentration-dependent manner, demonstrating that the immune function was affected. Moreover, ROS production was significantly induced in both CO₂ and HCl treatments, and four antioxidant components, GSH, GST, GR and GPx, had active responses to the acidification stress. Comparatively, CO₂ had more severe destructive effects on haemocytes than HCl at the same pH level, indicating that CO₂ stressed cells in other ways beyond the increasing H⁺ concentration. One possible explanation was that seawater acidification induced ROS overproduction, which damaged the ultrastructure of haemocytes and decreased phagocytosis.

Figure 1. Representative TEM micrographs of observed histopathological alterations in the granulocyte of M. edulis.

(a) Normal structure of a granulocyte of a mussel, with dense and boundary clear lysosomes (ly), complete and smooth cytomembranes (cy) and karyothecas (ka), and chromatin evenly distributed in the nucleus (n). (b) Cytoplasmic vacuolation (cv), with many lysosomes having lost their contents. (c) Swollen cytomembrane (cys) and swollen karyotheca (kas), with obvious membrane separations. Severe swelling causes breakages. (d) A seriously injured granulocyte, with a fuzzy boundary of lysosomes (lysosomes dissolved, lyd), chromatin condensation (cc) and extensively swollen cytomembrane (cys).

Figure 2. Structural damaging effects of seawater acidification induced structural changes in haemocytes exposed to seawater acidification (mean ± SEM, n = 5).

Note: Different lower case letters indicated the significant difference between the treated groups with the control group at the P < 0.05 level. (a) Acidification caused membrane damage in haemocytes detected by LDH release assay; (b) Acidification caused lysosomal membrane stability damage in haemocytes detected by NRRT assay.

Figure 3. The functional effects of seawater acidification in haemocytes exposed to HG or CG induced seawater acidification at different pH values compared to the control group (mean ± SEM, n = 5).

Different lower case letters indicate significant differences between haemocyte treatments (P < 0.05, ANOVA). (a) Phagocytosis levels of haemocytes; (b) Percent differences in production of ROS in haemocytes; (c) The concentration of GSH in haemocytes; (d) GST activity changes in haemocytes; (e) GR activity changes in haemocytes; (f) GPx activity changes in haemocytes.

Figure 4. Changes of antioxidant system (IBR consolidation) and ROS concentration in each pH level on 21 d in the HCl adjustment and the CO₂ enrichment groups

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Figure 5. The conjectured pathway of how seawater acidification acts on the structure and immune function of haemocytes of M. edulis

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