Complement Activity in the Egg Cytosol of Zebrafish Danio rerio: Evidence for the Defense Role of Maternal Complement Components

Most fish embryos that develop externally are exposed to an environment full of microbes. How they survive microbial attacks are not understood to date. Here we demonstrated that the egg cytosol prepared from the newly fertilized eggs of zebrafish Danio rerio is capable of killing the Gram-negative bacterium Escherichia coli, via in vitro assay system of the complement activity established. All findings indicate that it is the complement system operating via the alternative pathway that is attributable to the bacteriolytic activity. This is the first report providing the evidence for the functional role of the maternal complement components in fish eggs, paving the way for study of maternal immunity in other organisms whose eggs are fertilized in vitro

The evolution and appearance of c3 duplications in fish originate an exclusive teleost c3 gene form with anti- inflammatory activity

12 páginas, 6 figuras, 3 tablas.-- This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are creditedThe complement system acts as a first line of defense and promotes organism homeostasis by modulating the fates of diverse physiological processes. Multiple copies of component genes have been previously identified in fish, suggesting a key role for this system in aquatic organisms. Herein, we confirm the presence of three different previously reported complement c3 genes (c3.1, c3.2, c3.3) and identify five additional c3 genes (c3.4, c3.5, c3.6, c3.7, c3.8) in the zebrafish genome. Additionally, we evaluate the mRNA expression levels of the different c3 genes during ontogeny and in different tissues under steady-state and inflammatory conditions. Furthermore, while reconciling the phylogenetic tree with the fish species tree, we uncovered an event of c3 duplication common to all teleost fishes that gave rise to an exclusive c3 paralog (c3.7 and c3.8). These paralogs showed a distinct ability to regulate neutrophil migration in response to injury compared with the other c3 genes and may play a role in maintaining the balance between inflammatory and homeostatic processes in zebrafishThis work has been funded by the project CSD2007-00002 “Aquagenomics” from the Spanish Ministerio de Ciencia e Innovación, the ITN 289209 “FISHFORPHARMA” (EU) and project 201230E057 from the Agencia Estatal Consejo Superior de Investigaciones Científicas (CSIC).Peer reviewe

Genomic view of the evolution of the complement system

The recent accumulation of genomic information of many representative animals has made it possible to trace the evolution of the complement system based on the presence or absence of each complement gene in the analyzed genomes. Genome information from a few mammals, chicken, clawed frog, a few bony fish, sea squirt, fruit fly, nematoda and sea anemone indicate that bony fish and higher vertebrates share practically the same set of complement genes. This suggests that most of the gene duplications that played an essential role in establishing the mammalian complement system had occurred by the time of the teleost/mammalian divergence around 500 million years ago (MYA). Members of most complement gene families are also present in ascidians, although they do not show a one-to-one correspondence to their counterparts in higher vertebrates, indicating that the gene duplications of each gene family occurred independently in vertebrates and ascidians. The C3 and factor B genes, but probably not the other complement genes, are present in the genome of the cnidaria and some protostomes, indicating that the origin of the central part of the complement system was established more than 1,000 MYA

Toll-like receptors (TLRs) and mannan-binding lectin (MBL): On constant alert in a hostile environment

In the beginning were neither B cells nor T cells nor antibodies, but innate immune defense alone. The primary functional theme of innate immunity is the distinction between self and non-self, which is maintained by a vast number of cellular and subcellular components. In this context, the immense importance of the Toll-like receptors (TLRs) is well established. Positive (Darwinian) selection seems to be acting on the ligand-binding domains of these molecules, suggesting a selection pattern similar to that previously observed in the MHC proteins. In sharp contrast to TLRs, the biological significance of mannan-binding lectin (MBL) is controversial, and, concerning humans, it has been suggested that low concentration of MBL in serum represents a selective advantage. In this mini-review, based on a doctoral thesis, evolutionary aspects of TLRs and MBL are discussed

Characterization of a gene coding for the complement system component FB from loxosceles laeta spider venom glands

The human complement system is composed of more than 30 proteins and many of these have conserved domains that allow tracing the phylogenetic evolution. The complement system seems to be initiated with the appearance of C3 and factor B (FB), the only components found in some protostomes and cnidarians, suggesting that the alternative pathway is the most ancient. Here, we present the characterization of an arachnid homologue of the human complement component FB from the spider Loxosceles laeta. This homologue, named Lox-FB, was identified from a total RNA L. laeta spider venom gland library and was amplified using RACE-PCR techniques and specific primers. Analysis of the deduced amino acid sequence and the domain structure showed significant similarity to the vertebrate and invertebrate FB/C2 family proteins. Lox-FB has a classical domain organization composed of a control complement protein domain (CCP), a von Willebrand Factor domain (vWFA), and a serine protease domain (SP). The amino acids involved in Mg2+ metal ion dependent adhesion site (MIDAS) found in the vWFA domain in the vertebrate C2/FB proteins are well conserved; however, the classic catalytic triad present in the serine protease domain is not conserved in Lox-FB. Similarity and phylogenetic analyses indicated that Lox-FB shares a major identity (43%) and has a close evolutionary relationship with the third isoform of FB-like protein (FB-3) from the jumping spider Hasarius adansoni belonging to the Family Salcitidae

Granulocyte-macrophage colony-stimulating factor improves immunological parameters in patients with refractory solid tumours receiving second-line chemotherapy: Correlation with clinical responses

In this report, we studied the immunorestorative properties of subcutaneously administered granulocyte-macrophage colony-stimulating factor (GM-CSF) in patients with refractory solid tumours receiving second-line chemotherapy. Such patients exhibit abnormal immune responses in vivo and in vitro and, therefore, it was of interest to examine the effect of GM-CSF-induced immunomodulation on clinical response. We examined patients with primary malignant carcinomas (head and neck, n = 10; urogenital tract, n = 17; penis n = 6; colorectal, n = 8) who were treated with carboplatin (JM8), 300 ng/m(2) on days 1 and 22, leucovorin (LV), 200 mg/m(2) plus 5-fluoracil (5-FU), 500 mg/m(2) on days 8, 15 and 29 and four cycles of daily injections with placebo or GM-CSF, 300 mu g/day on days 3-6, 10-13, 17-20 and 24-27. Peripheral blood was collected from the patients one day after the end of each of the four-cycle injections with placebo or GM-CSF, namely on days 7, 14, 21 and 28. Peripheral blood mononuclear cells (PBMC) were tested in the autologous mixed lymphocyte reaction (AMLR) and for natural killer (NK) or lymphokine-activated killer (LAK) cell activity. Cytokine levels in serum were measured by immunoenzymatic (ELISA) assay. A total of 21 patients received a four-cycle regimen with GM-CSF (Group 1) and 20 were similarly treated with placebo (Group 2). All received standard chemotherapy as outlined above. Before GM-CSF treatment, all patients exhibited increased serum levels of interleukin-1 (IL-1 beta), tumour necrosis factor-alpha (TNF-alpha), IL-6 and prostaglandin E-2 (PGE(2)) and decreased serum levels of IL-2. Cellular immune responses (AMLR, NK- and LAK-cytotoxicity) were also low in all patients. Five patients from Group 1 had a PR (partial response), 2 patients had CR (complete response), and 14 patients had stable disease. Seven patients from Group 2 showed progressive disease, 3 had a PR and 10 had stable disease. All immune parameters were significantly improved during treatment in Group 1 but remained unchanged or even deteriorated in Group 2. Administration of GM-CSF during treatment of cancer patients with conventional chemotherapeutic drugs results in a marked potentiation of deficient cellular immune responses in vitro and a change towards normalisation of cytokine serum levels. The results reported herein support the use of GM-CSF as immunopotentiator during chemotherapy, but more patients must be studied before definite conclusions can be drawn. (C) 1997 Published by Elsevier Science Ltd