The development of cellular immune defence in marine medaka Oryzias melastigma.

Environmentally induced alterations of the immune system during sensitive developmental stages may manifest as abnormalities in immune organ configuration and/or immune cell differentiation. These not only render the early life stages more vulnerable to pathogens, but may also affect the adult immune competence. Knowledge of these sensitive periods in fish would provide an important prognostic/diagnostic tool for aquatic risk assessment of immunotoxicants. The marine medaka Oryzias melastigma is an emerging seawater fish model for immunotoxicology. Here, the presence and onset of four potentially sensitive periods during the development of innate and adaptive cellular immune defence were revealed in O. melastigma: 1.) initiation of phagocyte differentiation, 2.) migration and expansion of lymphoid progenitor cells, 3.) colonization of immune organs through lymphocyte progenitors and 4.) establishment of immune competence in the thymus. By using an established bacterial resistance assay for O. melastigma, larval immune competence (from newly hatched 1dph to 14dph) was found concomitantly increased with advanced thymus development and the presence of mature T-lymphocytes. A comparison between the marine O. melastigma and the freshwater counterpart Oryzias latipes disclosed a disparity in the T-lymphocyte maturation pattern, resulting in differences in the length of T-lymphocyte maturation. The results shed light on a potential difference between seawater and freshwater medaka in their sensitivity to environmental immunotoxicants. Further, medaka immune system development was compared and contrasted to economically important fish. The present study has provided a strong scientific basis for advanced investigation of critical windows for immune system development in fish.

Ancestral benzo[a]pyrene exposure affects bone integrity in F3 adult fish (Oryzias latipes).

Benzo[a]pyrene (BaP) at an environmentally relevant concentration (1µg/L) has previously been shown to affect bone development in a transgenerational manner in F3 medaka (Oryzias latipes) larvae (17dph). Here, we provide novel histomorphometric data demonstrating that the impaired bone formation at an early life stage is not recoverable and can result in a persistent transgenerational impairment of bone metabolism in F3 adult fish. A decrease in bone thickness and the occurrence of microcracks in ancestrally BaP-treated adult male fish (F3) were revealed by MicroCt measurement and histopathological analysis. The expression of twenty conserved bone miRNAs were screened in medaka and their relative expression (in the F3 ancestral BaP treatment vs the F3 control fish) were determined by quantitative real-time PCR. Attempt was made to link bone miRNA expression with the potential target bone mRNA expression in medaka. Five functional pairs of mRNA/miRNA were identified (Osx/miR-214, Col2a1b/miR-29b, Runx2/miR-204, Sox9b/miR-199a-3p, APC/miR-27b). Unique knowledge of bone-related miRNA expression in medaka in response to ancestral BaP-exposure in the F3 generation is presented. From the ecological risk assessment perspective, BaP needs to be regarded as a transgenerational skeletal toxicant which exerts a far-reaching impact on fish survival and fitness. Given that the underlying mechanisms of cartilage/bone formation are conserved between medaka and mammals, the results may also shed light on the potential transgenerational effect of BaP on skeletal disorders in mammals/humans.

Modulation of telomerase activity in fish muscle by biological and environmental factors.

Telomerase expression has long been linked to promotion of tumor growth and cell proliferation in mammals. Interestingly, telomerase activity (TA) has been detected in skeletal muscle for a variety of fish species. Despite this being a unique feature in fish, very few studies have investigated the potential role of TA in muscle. The present study was set to prove the concepts that muscle telomerase in fish is related to body growth, and more specifically, to muscle cell proliferation and apoptosis in vivo. Moreover, muscle TA can be influenced by biotic factors and modulated by environmental stress. Using three fish species, mangrove red snapper (Lutjanus argentimaculatus), orange-spotted grouper (Epinephelus coioides), and marine medaka (Oryzias melastigma), the present work reports for the first time that fish muscle TA was sensitive to the environmental stresses of starvation, foodborne exposure to benzo[a]pyrene, and hypoxia. In marine medaka, muscle TA was coupled with fish growth during early life stages. Upon sexual maturation, muscle TA was confounded by sex (female>male). Muscle TA was significantly correlated with telomerase reverse transcriptase (TERT) protein expression (Pearson correlation r=0.892; p≤0.05), which was coupled with proliferating cell nuclear antigen (PCNA) cell proliferation, but not associated with apoptosis (omBax/omBcl2 ratio) in muscle tissue. The results reported here have bridged the knowledge gap between the existence and function of telomerase in fish muscle. The underlying regulatory mechanisms of muscle TA in fish warrant further exploration for comparison with telomerase regulation in mammals.