Characterization of a novel cell line from the caudal fin of koi carp Cyprinus carpio

A continuous cell line (KF‐101) derived from the caudal fin of the koi carp Cyprinus carpio was established and characterized. The KF‐101 cell line multiplied abundantly in Leibovitz's L‐15 medium containing 10% foetal bovine serum at 25° C, and was subcultured for >90 passages over a period of 3 years. Immunocytochemistry revealed that the KF‐101 cells contain keratin, junction proteins connexin‐43 and occludin, and ectodermal stem‐cell marker Pax‐6, but not vimentin. Furthermore, the KF‐101 cells reacted with anti‐human DARPP‐32 and anti‐human GATA‐4 antibodies, and the labelling was regulated according to the cell cycle. The labels of the DARPP‐32 and GATA‐4 antibodies in the KF‐101 cells were the suggested phosphatase‐1 inhibitor‐1 and GATA‐3, respectively. In addition, the KF‐101 cells were susceptible to koi herpesvirus but were resistant to eel herpesvirus, iridovirus, grouper nodavirus and chum salmon (Oncorhynchus keta) virus. The results indicate that the KF‐101 cells are suitable materials for investigating biological and virological development.     

Histopathological and ultrastructural features of Koi herpesvirus (KHV)-infected carp Cyprinus carpio, and the morphology and morphogenesis of KHV

Epizootics of Koi herpesvirus (KHV) cause mass mortalities in koi carp and common carp worldwide. We used a newly developed 'per-gill infection' procedure with live KHV, and then conducted detailed histopathological and ultrastructural studies of KHV-infected cells including an examination of the morphology and morphogenesis of KHV. The primary target of KHV was respiratory epithelial cells of the gill lamellae, and release of virions from infected epithelial cells resulted in a systemic infection affecting the kidney, spleen, heart, brain and liver. The pathognomonic feature of infected cells was the formation of intranuclear inclusion bodies with marginal hyperchromatosis in the nucleus. Within the nucleus, assembly of capsids and nucleocapsids and an increase in filamentous nucleoproteins were evident. Enveloped nucleocapsids budded from the inner nuclear membrane into the perinuclear space. De-enveloped nucleocapsids were translocated in the cytoplasm to be embedded within inclusion bodies where tegumentation of the nucleocapsid occurred. Enveloped virions that had budded into intracytoplasmic vesicles and virions located extracellularly were composed of an electron-dense core, surrounded in turn by the capsid, the tegument and finally an envelope with projections. The morphology and morphogenesis of KHV were the same as those of viruses within the family Herpesviridae.     

生长激素促进斑马鱼尾鳍的再生

目的建立生长激素过表达的转基因斑马鱼,研究生长激素在斑马鱼尾鳍再生过程中的作用。方法利用Gateway技术构建表达质粒"pDestTol2CG2; ubi:GH-polyA",在一细胞期显微注射表达质粒和转座酶mRNA后,通过荧光显微镜和qPCR技术筛选鉴定GH过表达的转基因斑马鱼。将斑马鱼分为对照组(野生型)和生长激素过表达组,尾鳍横切后,记录分析斑马鱼尾鳍再生过程。结果转基因斑马鱼中心脏被绿色荧光蛋白标记。荧光定量PCR检测结果显示GH表达水平显著高于对照组(P<0.05)。斑马鱼尾鳍横断后,生长激素过表达组的再生速度显著提高(P<0.05)。结论建立稳定生长激素过表达的转基因斑马鱼品系,过表达生长激素能够提高斑马鱼尾鳍再生速度。     

The effect of primary and secondary immunization on the lymphoid tissues of the carp, Cyprinus carpio L

Mirror carp immunized with human gamma globulin (HGG) in Freund's complete adjuvant (FCA) show a proliferative response involving cells whose cytoplasm stains deep red with methyl green-pyronin (pyroninophilic cells). This response occurs particularly in the haemopoietic parenchyma of the pronephros and mesonephros. It peaks at week 3, with the formation of clusters of pyroninophilic cells in the pronephros. Immunization with Aeromonas salmonicida elicited a less intense pyroninophilic response but caused a larger increase in pigment-containing cells. After a secondary immunization with HGG in FCA, a distinct response was observed in the spleen: Pyroninophilic cells collected within the ellipsoid sheaths in large numbers and formed nodules. The reticulum of such nodules acquired spherical proportions and resembled the white pulp reticulum of the tetrapod spleen. The roles of such pyroninophilic cells and the possibility that aggregations of them may be functionally analogous to homoiotherm germinal centres are discussed.     

Retracted: Evolution and development of the homocercal caudal fin in teleosts

The vertebrate caudal skeleton is one of the most innovative structures in vertebrate evolution and has been regarded as an excellent model for functional morphology, a discipline that relates a structure to its function. Teleosts have an internally‐asymmetrical caudal fin, called the homocercal caudal fin, formed by the upward bending of the caudal‐most portion of the body axis, the ural region. This homocercal type of the caudal fin ensures powerful and complex locomotion and is thought to be one of the most important evolutionary innovations for teleosts during adaptive radiation in an aquatic environment. In this review, we summarize the past and present research of fish caudal skeletons, especially focusing on the homocercal caudal fin seen in teleosts. A series of studies with a medaka spontaneous mutant have provided important insight into the evolution and development of the homocercal caudal skeleton. By comparing developmental processes in various vertebrates, we propose a scenario for acquisition and morphogenesis of the homocercal caudal skeleton during vertebrate evolution.     

Saltatory control of isometric growth in the zebrafish caudal fin is disrupted in long fin and rapunzel mutants

Zebrafish fins grow by sequentially adding new segments of bone to the distal end of each fin ray. In wild type zebrafish, segment addition is regulated such that an isometric relationship is maintained between fin length and body length over the lifespan of the growing fish. Using a novel, surrogate marker for fin growth in conjunction with cell proliferation assays, we demonstrate here that segment addition is not continuous, but rather proceeds by saltation. Saltation is a fundamental growth mechanism shared by disparate vertebrates, including humans. We further demonstrate that segment addition proceeds in conjunction with cyclic bursts of cell proliferation in the distal fin ray mesenchyme. In contrast, cells in the distal fin epidermis proliferate at a constant rate throughout the fin ray growth cycle. Finally, we show that two separate fin overgrowth mutants, long fin and rapunzel, bypass the stasis phase of the fin ray growth cycle to develop asymmetrical and symmetrical fin overgrowth, respectively.     

The regenerative capacity of the zebrafish caudal fin is not affected by repeated amputations

Background

The zebrafish has the capacity to regenerate many tissues and organs. The caudal fin is one of the most convenient tissues to approach experimentally due to its accessibility, simple structure and fast regeneration. In this work we investigate how the regenerative capacity is affected by recurrent fin amputations and by experimental manipulations that block regeneration.

Methodology/Principal Findings

We show that consecutive repeated amputations of zebrafish caudal fin do not reduce its regeneration capacity and do not compromise any of the successive regeneration steps: wound healing, blastema formation and regenerative outgrowth. Interfering with Wnt/ß-catenin signalling using heat-shock-mediated overexpression of Dickkopf1 completely blocks fin regeneration. Notably, if these fins were re-amputated at the non-inhibitory temperature, the regenerated caudal fin reached the original length, even after several rounds of consecutive Wnt/ß-catenin signalling inhibition and re-amputation.

Conclusions/Significance

We show that the caudal fin has an almost unlimited capacity to regenerate. Even after inhibition of regeneration caused by the loss of Wnt/ß-catenin signalling, a new amputation resets the regeneration capacity within the caudal fin, suggesting that blastema formation does not depend on a pool of stem/progenitor cells that require Wnt/ß-catenin signalling for their survival.     

The effects of carotenoid and food intake on caudal fin regeneration in male guppies

The trade‐offs involved in allocating carotenoid pigments and food to healing and regrowing damaged caudal fin tissue v . other functions were examined in guppies Poecilia reticulata , a species in which females prefer males that display larger amounts of carotenoids in their skin. The guppies were derived from four natural populations in Trinidad that differed in resource availability but not predation intensity. Carotenoids, food and site of origin did not affect either absolute or relative fin regrowth, which suggested that fin regeneration in guppies was not constrained by carotenoid availability. It is possible that carotenoid intake influences fin regeneration in the presence of natural stressors such as predators. There was a significant negative interaction between food level in the laboratory and resource availability in the field: males from low‐resource‐availability sites regrew more fin tissue when raised on the high food level, and males from high‐resource‐availability sites regrew more fin tissue when raised on the low food level. The direction of this interaction runs counter to theoretical expectations.     

Concentrations of a Koi herpesvirus (KHV) in tissues of experimentally infected Cyprinus carpio koi as assessed by real-time TaqMan PCR

The Koi herpesvirus (KHV) is a herpes-like virus now recognized as a worldwide cause of mortality among populations of koi Cyprinus carpio koi and common carp Cyprinus carpio carpio. Temperature is a key factor influencing virus replication both in cell culture and in the tissues of experimentally infected fish. Genomic DNA sequences were used to optimize a rapid real-time TaqMan PCR assay to detect and quantify KHV DNA as found in the tissues of virus-exposed fish. The assay allowed analytical enumeration of target KHV genome copies ranging from 10(1) to 10(7) molecules as present in infected cell lines or fish tissues. The new assay was specific for KHV and did not detect DNA from 3 related herpes-like viruses found in fish, the Cyprinid herpesvirus 1 (CyHV-1), Cyprinid herpesvirus 2 (CyHV-2), Ictalurid herpesvirus 1 (IcHV-1) or the KF-1 cell line used for virus growth. Concentrations of KHV DNA were evaluated in 7 different tissues of replicate groups of virus-exposed koi held at water temperatures of 13, 18, 23 and 28 degrees C. Viral DNA was detected among virus-exposed koi at all 4 water temperatures but mortality was only observed among fish at 18, 23, and 28 degrees C. Time and temperature and the interactions between them affected concentrations of viral DNA detected in tissues of koi exposed to KHV. Although there were no recognized patterns to viral DNA concentrations as found in different tissues over time, KHV genome copies for all tissues increased with time post virus exposure and with water temperature. The remarkably rapid and systemic spread of the virus was demonstrated by the presence of viral DNA in multiple tissues 1 d post virus exposure. The greatest DNA concentrations found were in the gill, kidney and spleen, with virus genome equivalents consistently from 10(8) to 10(9) per 10(6) host cells. High levels of KHV DNA were also found in the mucus, liver, gut, and brain. Koi surviving infection at 62 to 64 d post virus exposure contained lower KHV genome copies (up to 1.99 x 10(2) per 10(6) host cells) as present in gill, kidney or brain tissues.     

The musculoskeletal system of the caudal fin in basal Actinopterygii: heterocercy,diphycercy, homocercy

The caudal fin represents the posteriormost region of the vertebrate axis and is one location where forces are exerted to the surrounding medium. The evolutionary changes of its skeleton have been well analyzed in gnathostomes and revealed transitions from heterocercal to diphycercal and homocercal tails. In contrast, we only know little about the evolutionary transformations of the muscular system of the caudalis and about possible ways of force transmission from anterior myomeres to the caudal fin. The goals of this study are to gain insight into evolutionary transformations of the musculoskeletal system in the four basal actinopterygian groups (Cladistia, Chondrostei, Ginglymodi, and Halecomorphi) and to identify likely pathways of force transmission to the tail. In this context, the connective tissue of the myosepta is considered to be an essential part of the musculoskeletal system. For the first time, this system is analyzed for the whole postanal region. The use of microdissection techniques and polarized light microscopy revealed the collagen fiber architecture and the insertions of all postanal myosepta from cleared and stained specimens. The collagen fiber architecture is similar in all investigated specimens and thus represents the primary actinopterygian condition. All parts of postanal myosepta are dominated by longitudinally arranged myoseptal tendons (lateral and myorhabdoid tendons) that span several vertebral segments. This architecture supports the view that posterior myosepta are well designed to transfer muscular forces that are generated in anterior myomeres. In contrast to the uniform myoseptal architecture, the musculoskeletal system differs between the four basal actinopterygian groups. Among them, chondrosteans have retained the plesiomorphic condition of actinopterygian tails. For the remaining taxa several evolutionary novelties in the musculoskeletal system of the tail are revealed. Most of these have evolved independently in the cladistian and neopterygian stem lineage. In these groups extensions of all epaxial and hypaxial parts of myosepta are present that insert on caudal fin rays. This remarkable contribution of epaxial muscle masses to the caudal fin organization is in contrast to the skeletal organization, that largely derives from hypaxial material only. In contrast to former studies the hypochordal longitudinalis muscle is shown to be a synapomorphy of Halecostomi (Halecomorphi + Teleostei). The morphological framework presented here allows to generate new hypotheses on the function of caudal fins that can be tested experimentally.     

The functional role of the caudal fin in the feeding ecology of the common thresher shark Alopias vulpinus

This study tests the hypothesis that the common thresher shark Alopias vulpinus uses its elongate caudal fin to both produce thrust and immobilize prey during feeding. Underwater video recorded in southern California from 2007 to 2009 revealed 34 feeding events, all of which were initiated with the upper lobe of the caudal fin.