No abstract

We used zebrafish as a whole-organism model to screen new compounds for sun protection activity. First of all, we designed a series of UVB exposure experiments and recorded the phenotypic changes of zebrafish embryos. Results showed that 100 mJ/cm(2) of UVB given six times separated by 30 min intervals is the best condition. Fin malformation (reduced and/or absent fin) phenotypes are the most evident consequences after exposure to UVB. Each fin was affected by UVB, including pelvic, ventral, caudal, and dorsal fin, but pelvic fin seemed to be the most sensitive target after UVB exposure. We furthermore carried out "prevention" and "treatment" experiments using green tea extract and/or (-)-epigallocatechin (EGCG) to test this whole-organism model by observing the morphological changes of all fins (especially pelvic fin) after UVB exposure. Effects of UVB, green tea extract and EGCG on fin development were assessed using the Kaplan-Meier analysis, log-rank test and Cox proportional hazards regression. Results showed that a zebrafish pelvic fin in the UVB + green tea (treatment) group is 5.51 (range from 2.39 to 14.90) times, one in the UVB + green tea (prevention) group is 7.04 (range from 3.11 to 18.92) times, and one in the 25 ppm of EGCG (prevention) group is 22.19 (range from 9.40 to 61.50) times more likely to return to normal fin than one in the UVB only group. On the basis of these observations, we believe this model is effective for screening the higher stability and lower toxicity of new compounds, such as small chemicals which are derivative from EGCG or other dietary agents for sun protection.

We generated a transgenic line Tg(k18:shh:RFP) with overexpression of Sonic hedgehog in the skin epidermis. By 5 day-post-fertilization (dpf), many epidermal lesions were clearly observed, including a swollen yolk sac, epidermis growth malformation around the eyes and at the basement of the pectoral fins. Skin histology revealed embryos derived from Tg(k18:shh:RFP) displayed an elevated Nuclear/Cytoplasmic ratio and pleomorphic nuclei compared to their wild type littermates, suggesting the abnormal growth pattern on the epidermis of Tg(k18:shh:RFP) embryos were dysplasia. Later (by 7 dpf), Tg(k18:shh:RFP) embryos displayed broader pectoral fins which are similar to the polydactyly phenotypes of Nevoid basal cell carcinoma syndrome (NBCCS)/Gorlin patients and polydactylous mice. In addition, treatment with cyclopamine is able to enhance and prolong the survival rates and survival durations of Tg(k18:shh:RFP) embryos. In conclusion, this unique Tg(k18:shh:RFP) fish line, should be an excellent experimental animal for screening for a lower toxicity level of the new Hh-inhibitor and can even be used as a new anti-cancer drug-screening platform.

No abstract

The objective of the current study was to investigate the effects of Ca(2+) levels on myofibril alignment during zebrafish embryogenesis. To investigate how altered cytoplasmic Ca(2+) levels affect myofibril alignment, we exposed zebrafish embryos to 2-aminothoxyldiphenyl borate (2-APB; an inositol 1,4,5-trisphosphate receptor inhibitor that reduces cytosolic Ca(2+) levels) and caffeine (a ryanodine receptor activator that enhances cytosolic Ca(2+) levels). The results demonstrated that the most evident changes in zebrafish embryos treated with 2-APB were shorter body length, curved trunk and malformed somite boundary. In contrast, such malformed phenotypes were evident neither in untreated controls nor in caffeine-treated embryos. Subtle morphological changes, including changes in muscle fibers, F-actin and ultrastructures were easily observed by staining with specific monoclonal antibodies (F59 and α-laminin), fluorescent probes (phalloidin) and by transmission electron microscopy. Our data suggested that: (1) the exposure to 2-APB and/or caffeine led to myofibril misalignment; (2) 2-APB-treated embryos displayed split and short myofibril phenotypes, whereas muscle fibers from caffeine-treated embryos were twisted and wavy; and (3) zebrafish embryos co-exposed to 2-APB and caffeine resulted in normal myofibril alignment. In conclusion, we proposed that cytosolic Ca(2+) is important for myogenesis, particularly for myofibril alignment.