Immunostimulatory effects of natural human interferon-alpha (huIFN-alpha) on carps Cyprinus carpio L.

Human interferon-alpha (huIFN-alpha) is an important immunomodulatory substance used in the treatment and prevention of numerous infectious and immune-related diseases in animals. However, the immunostimulatory effects of huIFN-alpha in fish remain to be investigated. In the current study, the immune responses of the carp species Cyprinus carpio L. to treatment with huIFN-alpha were analyzed via measurement of superoxide anion production, phagocytic activity and the expression of cytokine genes including interleukin-1beta, tumor necrosis factor-alpha and interleukin 10. Low doses of huIFN-alpha were administered orally once a day for 3 days, and sampling was carried out at 1, 3 and 5 days post-treatment. Our results indicate that a low dose of huIFN-alpha significantly increased phagocytic activity and superoxide anion production in the carp kidney. The huIFN-alpha-treated fish also displayed a significant upregulation in cytokine gene expression. The current study demonstrates the stimulatory effects of huIFN-alpha on the carp immune system and highlights the immunomodulatory role of huIFN-alpha in fish.

The in vitro effects of CpG oligodeoxynucleotides on the expression of cytokine genes in the common carp (Cyprinus carpio L.) head kidney cells.

Synthetic oligodeoxynucleotides with CpG motifs (CpG-ODNs) have gained attention because it elicits strong innate immune responses. CpG-ODNs promote the production of T-helper 1(T(H)1) and pro-inflammatory cytokines. In fish, knowledge of the effects of CpG-ODNs on the expression of cytokine genes is scarce. In this study, we report that CpG-ODNs induce the expression of pro-inflammatory cytokines in common carp head kidney leucocytes in vitro. Evidence of a T(H)1 type immune stimulation was observed as the fish homologs of IP-10 (interferon gamma-inducible protein 10; CXCL10) and MCP (monocyte chemotactic protein a CC-chemokine) were induced by CpG. Interleukin 10, a cytokine inhibitory factor, failed to be induced by CpG. These results suggest a robust immune stimulation can be achieved by CpGs and has a potential as an immunostimulant in aquaculture.

Discovery of a novel immunoglobulin heavy chain gene chimera from common carp (Cyprinus carpio L.).

In fish, two types of immunoglobulin heavy chain (IGH) genes, namely, IgM and IgD, have been cloned and characterized. Recently, a new IGH isotype specific to teleosts had been identified from zebra fish, rainbow trout, and fugu. In zebra fish, the domains of this new gene are present upstream of the mu region along the IGH locus. During this study, a novel IGH chimera (IgM-IgZ) has been discovered from common carp. The cloned cDNA encodes a typical leader peptide, a variable region, two constant regions, and a secretory tail. The first constant region is made up of the C(H1) domain of carp IgM, while the second constant region shares a high similarity to the C(H4) domain of the IgZ from zebrafish. Southern hybridization studies of the mu and zeta domains, conducted separately, revealed the presence of at least three copies of the respective genes, and mu and zeta domains might be present on the same loci, although far apart. Expression studies of the IGH genes suggest that there is an increase in chimeric immunoglobulin gene transcription when stimulated with lipopolysaccharide.

Analysis of genes expressed in head kidney of common carp Cyprinus carpio L treated with cortisol.

We analyzed genes expressed from head kidney of common carp Cyprinus carpio L. treated with cortisol. The results of single-pass sequencing of expressed sequence tags (ESTs) from 188 clones (AU240288-AU240367, AU301120-AU301227) from kidney cDNA are presented. One-hundred-twenty-seven clones (67.6%) were completely unknown and are likely to represent newly described genes, whereas 61 clones (32.4%) were identified based on matches to sequences in the database. The putative genes contain several ribosomal proteins, cytochrome oxidase subunits. Immune related cDNA clones identified from kidney were immunoglobulin light chain (n=4), FK506/rapamycin-binding protein (FKBP), CXC chemokine receptor type 4, complement factor B/C2-A3, peptidylprolyl isomerase (cyclophilin; Cyp)-like1, cyclophilin S1, heat shock-70 kDa protein-4, stress-activated protein kinase-3 (n=2). FKBP and cyclophilin genes expressed in normal tissues (head kidney, spleen, liver, brain and heart). Expression of FKBP and cyclophilin genes were not detected in liver, brain and heart when treated with cortisol for 16 h.

In vitro modulation of common carp (Cyprinus carpio L.) phagocytic cells by Di-n-butyl phthalate and Di-2-ethylhexyl phthalate.

The modulation of common carp, Cyprinus carpio L. phagocytic cells by Di-n-butyl phthalate (DBP) and Di-2-ethylhexyl phthalate (DEHP) were analyzed in vitro. Carp head kidney leucocytes were cultured in a RPMI 1640 medium containing DBP or DEHP at concentration of 1-1000 nM and the production of superoxide anion was measured via the in vitro reduction of nitroblue tetrazolium (NBT). Macrophages incubated with 100 up to 1000 nM of DBP showed an increase in their production of superoxide anion in comparison with the control macrophages which, were cultured without DBP. DEHP also increased the production of superoxide anion in phagocytic cells from carp kidneys. Phagocytic cells treated with 10 and 100 ng of DEHP showed increased phagocytic activity. DBP and DEHP also significantly increased the production of nitric oxide in carp macrophages. These results showed that DBP and DEHP modulate the function of phagocytic cells in vitro.

Immunomodulatory effects of alpha melanocyte stimulating hormone on common carp (Cyprinus carpio L.).

The immunomodulatory effects of phagocytic cells in common carp Cyprinus carpio L., by alpha melanocyte stimulating hormone (alpha-MSH) was analyzed in vitro. Carp head kidney leucocytes were cultured in RPMI 1640 medium containing 0.05, 0.1, 1 and 10 nM alpha-MSH and the production of superoxide anion was measured via the reduction of nitroblue tetrazolium (NBT) in vitro. Macrophages incubated with alpha-MSH showed an increase in the production of superoxide anion in comparison to control macrophages cultured without hormone. Phagocytic cells treated with alpha-MSH also displayed increased phagocytosis. Furthermore, carp lymphocytes treated with alpha-MSH increased the mitogenic responses to phytohaemoagglutinin (PHA-P). These results show that alpha-MSH directly influences fish immune responses.

Suppression in function of phagocytic cells in common carp Cyprinus carpio L. injected with estradiol, progesterone or 11-ketotestosterone.

The modulation of phagocytic cells by beta-estradiol, 11-ketotestosterone, progesterone and cortisol was analysed in common carp (Cyprinus carpio L.). Carp were injected intraperitoneally (1 and 5 microg/kg fish) with each steroid and the activity of kidney macrophages was recorded 1 and 3 days post-injection. Bovine serum albumin-injected fish served as controls. Phagocytosis, superoxide anion and nitric oxide production were suppressed in all sex steroid treatments, and the suppression was in a dose-dependent manner. Cortisol treatment also inhibited the immune parameters to an extent similar to each of the steroid hormone. These results show that sex steroids can modulate the function of the phagocytic cells.