The presence of skeletal anomalies in farmed teleost fish is currently a major problem in aquaculture, entailing economical, biological and ethical issues. The common occurrence of skeletal abnormalities in farmed fish and the absence of effective solutions for avoiding their onset or definitely culling out the affected individuals as early as possible from the productive cycle, highlight the need to improve our knowledge on the basic processes regulating fish skeletogenesis and skeletal tissues differentiation, modelling and remodelling. Severe skeletal anomalies may actually occur throughout the entire life cycle of fish, but their development often begins with slight aberrations of the internal elements. Comprehensive investigation efforts conducted on reared larvae and juveniles could provide a great contribution in filling the gap in knowledge, as skeletogenesis and skeletal tissue differentiation occur during these early life stages. The aim of this review is to provide a synthetic but comprehensive picture of the actual knowledge on the ontogeny, typologies and occurrence of skeletal anomalies, and on the proposed causative factors for their onset in larvae and juveniles of European farmed fish. The state-of-art of knowledge of these issues is analysed critically intending to individualize the main gaps of knowledge that require to be filled, in order to optimize the morphological quality of farmed juveniles.

Skeletal anomalies in reared European fish larvae and juveniles, part 2 : main typologies, occurrences and causative factors

This critical review summarizes the knowledge about fish skeletal tissues and inherent normal and anomalous development. Particular emphasis is given to existing literature on reared European fishes. The aim was to identify the main gaps of knowledge that require to be filled, in order to precociously identify anomalous developmental patterns that lead to skeletal anomalies in reared finfish larvae and juveniles. The review also aims to extend our knowledge about the factors that are possibly involved in the onset of skeletal anomalies. The final goal is the optimization of the morphological quality of farmed juvenile fish.

Skeletal Anomalies in Reared European Fish Larvae and Juveniles. Part 1: Normal and Anomalous Skeletogenic Processes

Fish as a model to assess chemical toxicity in bone

http://www.unitedfisheries.co.nz/sites/default/files/REF-2-Fish-a-suitable-system-to-model-human-bone.pdf

Fish: a suitable system to model human bone disorders and discover drugs with osteogenic or osteotoxic activities

Synthesis, biological and environmental applications of hydroxyapatite and its composites with organic and inorganic coatings

Vitamin K (VK) acts as a cofactor driving the biological activation of VK-dependent proteins and conferring calcium-binding properties to them. As a result, VK is converted into VK epoxide, which must be recycled by VK epoxide reductases (Vkors) before it can be reused. Although VK has been shown to play a central role in fish development, particularly during skeletogenesis, pathways underlying VK actions are poorly understood, while good and reliable molecular markers for VK cycle/homeostasis are still lacking in fish. In the present work, expression of 2 zebrafish vkor genes was characterized along larval development and in adult tissues through qPCR analysis. Zebrafish cell line ZFB1 was used to evaluate in vitro regulation of vkors and other VK cycle-related genes during mineralization and upon 24 h exposure to 0.16 and 0.8 µM phylloquinone (VK1), 0.032 µM warfarin, or a combination of both molecules. Results showed that zebrafish vkors are differentially expressed during larval development, in adult tissues, and during cell differentiation/mineralization processes. Further, several VK cycle intermediates were differentially expressed in ZFB1 cells exposed to VK1 and/or warfarin. Present work provides data identifying different developmental stages and adult tissues where VK recycling is probably highly required, and shows how genes involved in VK cycle respond to VK nutritional status in skeletal cells. Expression of vkor genes can represent a reliable indicator to infer VK nutritional status in fish, while ZFB1 cells could represent a suitable in vitro tool to get insights into the mechanisms underlying VK action on fish bone.

Zebrafish vitamin K epoxide reductases: expression in vivo, along extracellular matrix mineralization and under phylloquinone and warfarin in vitro exposure

Mechanisms of bone formation and skeletal development have been successfully investigated in zebrafish using a variety of in vivo approaches, but in vitro studies have been hindered due to a lack of homologous cell lines capable of producing an extracellular matrix (ECM) suitable for mineral deposition. Here we describe the development and characterization of a new cell line termed ZFB1, derived from zebrafish calcified tissues. ZFB1 cells have an epithelium-like phenotype, grow at 28°C in a regular L-15 medium supplemented with 15% of fetal bovine serum, and are maintained and manipulated using standard methods (e.g., trypsinization, cryopreservation, and transfection). They can therefore be propagated and maintained easily in most cell culture facilities. ZFB1 cells show aneuploidy with 2n=78 chromosomes, indicative of cell transformation. Furthermore, because DNA can be efficiently delivered into their intracellular space by nucleofection, ZFB1 cells are suitable for gene targeting approaches ...

/pdf/development-of-an-in-vitro-cell-system-from-zebrafish-25c1kb6acc.pdf

Development of an in vitro cell system from zebrafish suitable to study bone cell differentiation and extracellular matrix mineralization.

Development and characterization of a skin cell line (SGA) from the mosquitofish Gambusia affinis and its susceptibility to fish Betanodavirus

Over the past decade, hydroxyapatite nanoparticles (HAp Nps) have been utilized effectively in many biomedical applications. The present study focuses on the preparation and characterization of HAp Nps through various analyses such as TEM, FTIR and XRD. The TEM results showed that the particles were rod-shaped with size ranging from 20 to 100 nm. The FTIR spectrum showed the presence of O–H group, amine group and also the calcium and phosphate groups. The XRD spectrum showed characteristic peaks at 24.4 A and 35.4 A, which indicated the crystalline nature of the particles. The HAp Nps did not show cytotoxicity in a fish vertebra derived BGA cell line. The AO/EtBr staining showed the presence of apoptotic bodies in cells treated with a higher concentration (300 μg mL-1). These results suggest that HAp Nps could be useful in investigating bone cell differentiation and mineralization in fish models.

Synthesis and characterization of hydroxyapatite nanoparticles and their cytotoxic effect on a fish vertebra derived cell line

Sulfated polysaccharides (SP) from seaweeds exhibit numerous biological activities and are less toxic than conventional antiviral drugs. However, little is known about their mode of action. This study aims at extracting the polysaccharides from brown seaweed Sargassum ilicifolium and then evaluating its antiviral activity against fish Betanodavirus. The extracted SP was characterized through FT-IR, 1H NMR, and thermal gravimetric analysis (TGA). The antiviral activity of SP was tested on SGA cell line by syncytia reduction and the polysaccharide represented a prominent antiviral activity to Betanodavirus by reducing the cytopathic effect (CPE) formation in vitro. The study revealed that SP effectively blocked viral attachment and entry. Moreover, during the in vivo challenge experiment in Gambusia affinis, SP treatment showed a significant decrease in the viral titers. Hence, these results indicate that the polysaccharides obtained from S. ilicifolium exhibited the high antiviral potential to Betanodavirus infection in fishes both in vitro and in vivo.

Parameswaran Vijayakumar

Papers

Zebrafish vitamin K epoxide reductases: expression in vivo, along extracellular matrix mineralization and under phylloquinone and warfarin in vitro exposure

Development of an in vitro cell system from zebrafish suitable to study bone cell differentiation and extracellular matrix mineralization.

Development and characterization of a skin cell line (SGA) from the mosquitofish Gambusia affinis and its susceptibility to fish Betanodavirus

Synthesis and characterization of hydroxyapatite nanoparticles and their cytotoxic effect on a fish vertebra derived cell line

Antiviral activity of sulfated polysaccharides from Sargassum ilicifolium against fish Betanodavirus infection