Gene Duplication of Androgen Receptor As An Evolutionary Driving Force Underlying the Diversity of Sexual Characteristics in Teleost Fishes.

Sexual dimorphism allows species to meet their fitness optima based on the physiological availability of each sex. Although intralocus sexual conflict appears to be a genetic constraint for the evolution of sex-specific traits, sex-linked genes and the regulation of sex steroid hormones contribute to resolving this conflict by allowing sex-specific developments. Androgens and their receptor, androgen receptor (Ar), regulate male-biased phenotypes. In teleost fish, ar ohnologs have emerged as a result of teleost-specific whole genome duplication (TSGD). Recent studies have highlighted the evolutionary differentiation of ar ohnologs responsible for the development of sexual characteristics, which sheds light on the need for comparative studies on androgen regulation among different species. In this review, we discuss the importance of ar signaling as a regulator of male-specific traits in teleost species because teleost species are suitable experimental models for comparative studies owing to their great diversity in male-biased morphological and physiological traits. To date, both in vivo and in vitro studies on teleost ar ohnologs have shown a substantial influence of ars as a regulator of male-specific reproductive traits such as fin elongation, courtship behavior, and nuptial coloration. In addition to these sexual characteristics, ar substantially influences immunity, inducing a sex-biased immune response. This review aims to provide a comprehensive understanding of the current state of teleost ar studies and emphasizes the potential of teleost fishes, given their availability, to find molecular evidence about what gives rise to the spectacular diversity among fish species.

Non-specific cytotoxic cell receptor protein-1 (NCCRP-1) is involved in anti-parasite innate CD8+ T cell-mediated cytotoxicity in ginbuna crucian carp.

CD8+ cytotoxic T cells (CTLs) are a main cellular component of adaptive immunity. Our previous research has shown that CD8+ cells demonstrate spontaneous cytotoxic activity against the parasite Ichthyophthirius multifiliis in ginbuna crucian carp, suggesting that CD8+ cells play an important role in innate immunity. Herein, we investigated the molecules and cellular signal pathways involved in the cytotoxic response of ginbuna crucian carp. We considered non-specific cytotoxic receptor protein-1 (NCCRP-1) as candidate molecule for parasite recognition. We detected NCCRP-1 protein in CD8+ cells and the thymus as well as in other cells and tissues. CD8+ cells expressed mRNA for NCCRP-1, Jak2, and T cell-related molecules. In addition, treatment with a peptide containing the presumed antigen recognition site of ginbuna NCCRP-1 significantly inhibited the cytotoxic activity of CD8+ cells against the parasites. The cytotoxic activity of CD8+ cells was significantly inhibited by treatment with the JAK1/2 inhibitor baricitinib. These results suggest that teleost CTLs recognize I. multifiliis through NCCRP-1 and are activated by JAK/STAT signaling.

Local immune responses to two stages of Ichthyophthirius multifiliis in ginbuna crucian carp.

Ichthyophthirius multifiliis is a ciliated protozoan parasite and is known to infect many freshwater teleosts. Characterizing the immune system in epithelial tissues, where the parasites penetrate and settle, is key to understanding host-parasite interactions. This study examined local immune responses in vivo to the infective stage (theront and trophont) of the parasites using intra-fin administration, which has been developed to analyze in vivo immune responses using fish fin. CD8α+ and CD4+ T-cell compositions were increased significantly in the fin cavity injected with theront or trophont antigens. The expression of GATA-3 and T-bet mRNA, which regulate differentiation of helper T-cells, was upregulated significantly in leukocytes from the trophont antigen-injected site. In contrast, the percentages of macrophages and neutrophils, which are innate immunity components, were decreased significantly in the injection sites. These results suggest that I. multifiliis antigens inhibit the migration of macrophages and neutrophils, and T-cells are the first responders to I. multifiliis. Thus, to better understand the interaction of host immunity and I. multifiliis, further studies should focus on exploring the inhibitory factors from I. multifiliis or examining innate functions of teleost T-cells.

Evaluation of probiotic Bacillus aerius B81e isolated from healthy hybrid catfish on growth, disease resistance and innate immunity of Pla-mong Pangasius bocourti.

Infectious diseases have been found to be a major cause of mortality in fish hatcheries. Probiotics have been introduced to replace antibiotics commonly used for treatment of bacterial infection in aquaculture. This study was conducted to isolate, screen, and evaluate the probiotic Bacillus spp. for potential use as a feed supplement to enhance fish growth, disease resistance and innate immunity of Pla-mong Pangasius bocourti. Bacillus aerius strain B81e was selectively isolated from the intestine of healthy catfish and chosen based on its probiotic properties both in vitro and in vivo. This bacterium produced a bacteriocin-like substance and exhibited a broad-spectrum antibacterial activity inhibiting both Gram-positive and Gram-negative bacteria especially the fish pathogens Aeromonas hydrophila and Streptococcus agalactiae. The susceptibility to all 8 antibiotics tested implies that it is unlikely to be an antibiotic-resistant bacterium. B. aerius strain B81e possessed interesting adhesion properties as shown by its high percentages of hydrophobicity, auto-aggregation, co-aggregation with fish pathogens A. hydrophila FW52 and S. agalactiae F3S and mucin binding. The strain B81e survived simulated gastrointestinal conditions, producing protease and lipase but not ß-haemolysin. The study also evaluated the effects of dietary supplementation with strain B81e on growth performance, innate immunity, and the disease resistance of P. bocourti against A. hydrophila infection. Fish with a mean body weight of 69 g were fed strain B81e at 0 (control) and 107 CFU g-1 feed (test) for 60 days. Various growth and immune parameters were examined at 30 and 60 days post-feeding. Fish were challenged with A. hydrophila 60 days post-feeding and mortalities were recorded over 14 days post-infection. Results showed that the administration of strain B81e for 60 days had significant effects (p < 0.05) on weight gain, specific growth rate and feed utilization efficiency of P. bocourti. Dietary administration of strain B81e increased the serum lysozyme and bactericidal activities of P. bocourti significantly throughout the experimental period whereas the alternative complement, phagocytic and respiratory burst activities were significantly (p < 0.05) higher in the test fish compared to the control fish after 60 days of feeding. In addition, the fish fed a strain B81e supplemented diet had a significantly higher (p < 0.05) post-challenge survival rate than the control fish. The results in this study indicate that B. aerius B81e has beneficial effects on growth performance, innate immunity and disease resistance of P. bocourti. This is the first report on the probiotic roles of B. aerius in aquaculture.

A CD46-like molecule functional in teleost fish represents an ancestral form of membrane-bound regulators of complement activation.

In the complement system, the regulators of complement activation (RCA) play crucial roles in controlling excessive complement activation and in protecting host cell from misdirected attack of complement. Several members of RCA family have been cloned from cyclostome and bony fish species and classified into soluble and membrane-bound type as in mammalian RCA factors. Complement-regulatory functions have been described only for soluble RCA of lamprey and barred sand bass; however, little is known on the biological function of the membrane-bound RCA proteins in the lower vertebrates. In this study, a membrane-bound RCA protein, designated teleost complement-regulatory membrane protein (Tecrem), was cloned and characterized for its complement-regulatory roles. Carp Tecrem, an ortholog of a zebrafish type 2 RCA, ZCR1, consists of four short consensus repeat modules, a serine/threonine/proline-rich domain, a transmembrane region, and a cytoplasmic domain, from the N terminus, as does mammalian CD46. Tecrem showed a ubiquitous mRNA expression in carp tissues, agreeing well with the putative regulatory role in complement activation. A recombinant Chinese hamster ovary cell line bearing carp Tecrem showed a significantly higher tolerance against lytic activity of carp complement and less deposition of C3-S, the major C3 isotypes acting on the target cell, than control Chinese hamster ovary (mock transfectant). Anti-Tecrem mAb enhanced the depositions of carp C3 and two C4 isotypes on autologous erythrocytes. Thus, the present findings provide the evidence of complement regulation by a membrane-bound group 2 RCA in bony fish, implying the host-cell protection is an evolutionarily conserved mechanism in regulation of the complement system.

Polymorphic Sirpa is the genetic determinant for NOD-based mouse lines to achieve efficient human cell engraftment.

Current mouse lines efficient for human cell xenotransplantation are backcrossed into NOD mice to introduce its multiple immunodeficient phenotypes. Our positional genetic study has located the NOD-specific polymorphic Sirpa as a molecule responsible for its high xenograft efficiency: it recognizes human CD47 and the resultant signaling may cause NOD macrophages not to engulf human grafts. In the present study, we established C57BL/6.Rag2(nullIl2rgnull) mice harboring NOD-Sirpa (BRGS). BRGS mice engrafted human hematopoiesis with an efficiency that was equal to or even better than that of the NOD.Rag1(nullIl2rgnull) strain, one of the best xenograft models. Consequently, BRGS mice are free from other NOD-related abnormalities; for example, they have normalized C5 function that enables the evaluation of complement-dependent cytotoxicity of antibodies against human grafts in the humanized mouse model. Our data show that efficient human cell engraftment found in NOD-based models is mounted solely by their polymorphic Sirpa. The simplified BRGS line should be very useful in future studies of human stem cell biology.

Characterization of CD83 homologs differently expressed during monocytes differentiation in ginbuna crucian carp, Carassius auratus langsdorfii.

CD83 is a costimulatory molecule of antigen-presenting cells (APCs) that plays an important role in eliciting adaptive responses. It is also a well-known surface protein on mature dendritic cells (DCs). Furthermore, monocytes have been reported to differentiate into macrophages and monocyte-derived dendritic cells, which play an important role in innate immunity. CD83 expression affects the activation and maturation of DCs and stimulates cell-mediated immune responses. This study aims to reveal the CD83 expression during monocyte differentiation in teleosts, and the CD83 homologs evolutionary relationship. This study found two distinct CD83 homologs (GbCD83 and GbCD83-L) in ginbuna crucian carp (Gb) and investigated the evolutionary relationship among GbCD83 homologs and other vertebrates and the gene and protein expression levels of the homologs during 4 days of monocyte culture. The phylogenetic tree showed that the two GbCD83 homologs are classified into two distinct branches. Interestingly, only ostariophysians (Gb, common carp, rohu, fathead minnow and channel catfish), but not neoteleosts, mammals, and others, have two CD83 homologs. Morphological observation and colony-stimulating factor-1 receptor (CSF-1R), CD83, CD80/86, and CCR7 gene expressions illustrated that there is a differentiation of monocytes isolated from peripheral blood leukocytes after 4 days. Specifically, gene expression and immunocytochemistry revealed that GbCD83 is mainly expressed on monocytes at the early stage of cell culture, whereas GbCD83-L is expressed in the latter stage. These findings provided the first evidence of differential expression of CD83 homologs during monocytes differentiation in teleost.

Molecular characterization and expression analysis of three membrane-bound complement regulatory protein isoforms in the ginbuna crucian carp Carassius auratus langsdorfii.

Regulators of complement activation (RCA) play a role in protecting cells from excessive complement activation in humans. cDNA corresponding to three isoforms of teleost membrane-bound RCA protein (gTecrem) have been identified in the ginbuna crucian carp. gTecrem-1 consists of seven short consensus repeats (SCRs), whereas gTecrem-2 and gTecrem-3 have four SCRs. While gTecrem-1 possesses a tyrosine phosphorylation site in its cytoplasmic region, gTecrem-2 and gTecrem-3 lack the site. Tissue distribution analysis showed that gTecrem-1 and gTecrem-2 mRNAs were expressed in almost all tissues examined, whereas gTecrem-2 expression was not significantly detected in gill, liver, or intestine. Furthermore, analysis showed that gTecrem-1 was expressed in both peripheral blood leukocytes (PBLs) and erythrocytes and was also expressed in T cell subsets such as CD4(+), CD8(+) T cells, and IgM(+) B cells. gTecrem-2 expression was not detected in either PBLs or erythrocytes, whereas gTecrem-3 was expressed only in erythrocytes. These results suggested that gTecrem isoforms may serve different functional roles; gTecrem-1, which is expressed in T cells and possesses a tyrosine phosphorylation site, may act as a complement regulator and a cellular receptor in adaptive immunity.

Guideline for hereditary angioedema (HAE) 2010 by the Japanese Association for Complement Research - secondary publication.

This guideline was provided by the Japanese Association for Complement Research targeting clinicians for making an accurate diagnosis of hereditary angioedema (HAE), and for prompt treatment of the HAE patient in Japan. This is a 2010 year version and will be updated according to any pertinent medical advancements.

Development of a filter device for the prevention of aquatic bacterial disease using a single-chain variable fragment (scFv)-conjugated affinity silk.

Infectious disease is one of the most serious problems in the aquaculture industry for ornamental or edible fish. This study attempted to develop a new device for preventing an aquatic bacterial disease, ulcer disease, caused by Aeromonas salmonicida (As), using "affinity silk". Affinity silk is a silk protein-containing fibroin L-chain (FibL) fused to the single-chain variable fragment (scFv). It can be easily processed into different formats such as fibers, gels, sponges, or films. A transgenic silkworm that could express a cDNA construct containing FibL fused to an scFv derived from a monoclonal antibody (MAb) against As was successfully generated. An enzyme-linked immunosorbent assay was used to detect As by employing 96-well plates coated with scFv-conjugated affinity silk. As could be captured efficiently by glass wool coated with affinity silk in the column. Furthermore, the air-lift water filter equipped with the affinity silk-coated wool could considerably reduce the concentration of As in water and was estimated to have sufficient ability to trap a lethal dose of As. These findings show that the "affinity silk filter" is a potential device for the prophylaxis of aquatic animal diseases.