Differences in growth of Trypanoplasma borreli in carp serum is dependent on transferrin genotype.

Kinetoplastid parasites require transferrin (Tf), being the main source of iron, for growth and multiplication. This group of parasites developed a unique receptor-mediated system for acquiring host Tf which bears no structural homology with the host transferrin receptor. Trypanoplasma borreli, a blood parasite of common carp, probably uses a similar mechanism to sequester iron from host transferrin. In this study, we demonstrate a critical role of Tf for parasite growth. For in vitro studies we isolated and purified Tf from carp homozygous for the D or G allele of Tf. We obtained Tf-depleted serum using specific antibodies to carp Tf and studied gene expression in vivo during T. borreli infection with Real Time-quantitative PCR. We demonstrate that T. borreli cannot survive in medium supplemented with Tf-depleted serum while reconstitution with Tf restores normal growth. The critical role of Tf for parasite survival was shown in incomplete medium (medium without serum): addition of purified Tf significantly increased parasite survival. We also demonstrate that Tf polymorphism has a significant impact on T. borreli multiplication. Cultured parasites die more quickly in an environment containing D-typed Tf, as compared to medium with G-typed Tf. Gene expression during T. borreli infection in carp did not show an acute phase response. We could, however, observe an increased transcription of Tf in the head kidney, which may be associated with an immunological function of the Tf protein.

Serum Cyclophilin A Correlates with Increased Tissue MMP-9 in Patients with Ulcerative Colitis, but Not with Crohn's Disease.

BACKGROUND: Cyclophilin A (CyPA) is an immunomodulatory protein, high expression of which correlates with poor outcome of patients with inflammatory diseases. However, its role in inflammatory bowel disease (IBD) has not been studied. AIM: This study analyzes the correlation between cyclophilin A, matrix metalloproteinase (MMP)-9, and tissue inhibitor of MMP (TIMP)/MMP-9 complexes in the inflamed and non-inflamed colon mucosa of UC and CD patients. METHODS: Serum and biopsy specimens from inflamed and non-inflamed colonic mucosa of 38 patients with IBD (19 with UC and 19 with CD) and 16 controls were included in our study. We measured serum and tissue level of CyPA, and tissue level of TNF-α, MMP-9, TIMP-1/MMP-9, and TIMP-2/MMP-9 using ELISA method. RESULTS: Our results indicated that serum, but not tissue CyPA is increased in UC, rather than in CD patients, compared to the control. The increase correlated with higher tissue concentration of MMP-9 and TNF-α, especially in the UC group. Moreover, we observed significantly higher level of TIMP-1/MMP-9 in UC and CD group, which overlapped with the change in MMP-9. There was no change in TIMP-2/MMP-9 in the analyzed groups. CONCLUSION: The current study suggests that serum CyPA may be an independent additional marker of IBD, especially of UC. Higher CyPA level may be followed by increased MMP-9 in those patients. However, further studies are necessary to verify the role of CyPA in IBD development.

Polish Scientists in Fish Immunology: A Short History.

This review describes the role played by Polish scientists in the field of fish immunology and vaccination starting around 1900. In the early days, most publications were dealing with a description of relevant cells and organs in fish. Functional studies (phagocytosis, antibody response) came later starting in the late 1930s. Detailed papers on fish vaccination were published from 1970 onwards. Another important development was the unraveling of neuro-endocrine-immune interactions in the 1970s until today. Around 1980, it became more and more clear how important immunomodulation (stimulation or suppression by environmental factors, food components, drugs) was for fish health. The most recent findings are focusing on the discovery of genetic factors, signaling molecules, and receptors, which play a crucial role in the immune response. It can be concluded, that Polish scientists made considerable contributions to our present understanding of fish immunity and to applications in aquaculture worldwide.

Anti-inflammatory and antinociceptive action of the dimeric enkephalin peptide biphalin in the mouse model of colitis: new potential treatment of abdominal pain associated with inflammatory bowel diseases.

Biphalin, a mixed MOP/DOP agonist, displays a potent antinociceptive activity in numerous animal models of pain. The aim of the study was to characterize the anti-inflammatory and antinociceptive action of biphalin in the mouse models of colitis. The anti-inflammatory effect of biphalin (5mg/kg, twice daily, i.c. and i.p.) was characterized in a semi-chronic mouse model of colitis, induced by i.c. injection of trinitrobenzenesulfonic acid (TNBS). The antinociceptive action of biphalin (5mg/kg, i.p. and i.c.) in inflamed mice was assessed in mustard oil-induced model of visceral pain and in the hot plate test. In the semi-chronic mouse model of colitis, biphalin i.c. (5mg/kg), but not i.p. improved colitis macroscopic score (2.88±0.19 and 4.99±0.80 units for biphalin and vehicle treated animals, respectively). Biphalin injected i.p. and i.c. (5mg/kg) displayed a potent antinociceptive action in the mustard oil-induced pain test. In the hot plate test, biphalin (5mg/kg, i.p.) produced a potent antinociceptive activity in inflamed mice, suggesting central site of action. Our data suggest that biphalin may become a novel opioid-based analgesic agent in IBD therapy and warrant further investigation of its pharmacological profile.

Genetic resistance of carp (Cyprinus carpio L.) to Trypanoplasma borreli: influence of transferrin polymorphisms.

In serum most of the iron molecules are bound to transferrin (Tf), which is a highly polymorphic protein in fish. Tf is an essential growth factor for mammalian trypanosomes. We performed a series of experiments with Trypanoplasma borreli to detect putative correlations between different Tf genotypes of common carp (Cyprinus carpio L.) and susceptibility to this blood parasite. Five genetically different, commercially exploited carp lines (Israelian 'D', Polish 'R2' and 'K', Ukrainian 'Ur', Hungarian 'R0') and a reference laboratory cross ('R3xR8') were challenged with T. borreli and parasitaemia measured to determine susceptibility to the parasite. Among the commercial carp lines, Israelian 'D' carp were identified as most and Polish 'R2' carp as least susceptible, and used to produce a next generation and reciprocal crosses. These progenies were challenged with T. borreli and parasitaemia measured. We demonstrated significant effects of genetic background of the carp lines on susceptibility to T. borreli. This genetic effect was preserved in a next generation. We also observed a significant male effect on susceptibility to T. borreli in the reciprocal crosses. Serum samples from a representative number of fish from two infection experiments were used for Tf genotyping by polyacrylamide gel electrophoresis (PAGE), identifying DD, DG and DF as most frequent Tf genotypes. We could detect a significant association of the homozygous DD genotype with low parasitaemia in the least susceptible 'R2' (and 'K') carp lines and the lack of a such an association in the most susceptible 'D' carp line. Upon examination of parasite growth in vitro in culture media supplemented with 3% serum taken from fish with different Tf genotypes, we could show a faster decrease in number of parasites in culture media with serum from DD-typed animals.

Differential transcription of multiple forms of alpha-2-macroglobulin in carp (Cyprinus carpio) infected with parasites.

Alpha-2-macroglobulin (a2M) is a non-specific protease inhibitor involved in host defense mechanisms, inhibiting both endogenous and exogenous proteases. It is unique among the plasma anti-proteases with respect to the diversity of proteases that it can inactivate. Carp a2M consists of an alpha and beta chain of which the first includes the bioactive regions. Previously, three a2M alpha chain sequences were reported for East-Asian common carp. We studied a2M alpha chain variability in European common carp and report the cloning of a fourth a2M alpha chain with distinct sequence diversity in the bait region. The role of a2M in the immune response to parasites was studied in the liver of carp infected with Trypanoplasma borreli or with Ichthyophthirius multifiliis. Quantitative gene transcription analysis showed a differential regulation of the four isoforms, most clearly seen in infections with I. multifiliis. A2M3 was the only a2M isoform with a highly upregulated transcription during infection, suggesting that this particular isoform is of foremost biological importance.

Genetic differences in natural antibody levels in common carp (Cyprinus carpio L.).

In mammals, natural antibodies (Nabs) are mostly of the IgM isotype and can bind to a particular antigen or pathogen even if the host has never been exposed. Despite their early detection and abundance, the exact role and genetic control of Nabs remain unclear. We have used an indirect ELISA with three different antigens (keyhole limpet haemocyanin, chicken ovalbumin and bovine serum albumin) to demonstrate the ubiquitous presence of Nabs in common carp. Serum levels of Nabs increased with age, i.e. 10-month-old fish showed higher levels than 4-month-old fish. Also, fish grown in earth ponds showed higher levels of Nabs than fish grown in a clean environment of UV-treated water. Furthermore, we show that Nabs are present in different levels in the serum of carp lines with a different genetic background, suggestive of a genetic control. These genetic differences were independent of antigen, age and environment. Genetic differences in levels of Nabs could not unequivocally be related to differences in survival under farmed conditions. The possibilities for using levels of Nabs as marker criterion for selection for genetic disease resistance are discussed.