No abstract

Olfactomedin 1 (Olfm1) is a secreted glycoprotein belonging to a family of olfactomedin domain-containing proteins. It is involved in the regulation of neural crest production in chicken and promotes neuronal differentiation in Xenopus. Here, we investigate the functions of Olfm1 in zebrafish eye development. Overexpression of full-length Olfm1, and especially its BMY form lacking the olfactomedin domain, increased the thickness of the optic nerve and produced a more extended projection field in the optic tectum compared with control embryos. In contrast, injection of olfm1-morpholino oligonucleotide (Olfm1-MO) reduced the eye size, inhibited optic nerve extension, and increased the number of apoptotic cells in the retinal ganglion cell and inner nuclear layers. Overexpression of full-length Olfm1 increased the lateral separation of the expression domains of eye-field markers, rx3 and six3. The Olfm1-MO had the opposite effect. These data suggest that zebrafish Olfm1 may play roles in the early eye determination, differentiation, optic nerve extension, and branching of the retinal ganglion cell axon terminals, with the N-terminal region of Olfm1 being critical for these effects. Injection of RNA encoding WIF-1, a secreted inhibitor of Wnt signaling, caused changes in the expression pattern of rx3 similar to those observed after Olfm1-MO injection. Simultaneous overexpression of WIF-1 and Olfm1 abolished the WIF-1 effect. Physical interaction of WIF-1 and Olfm1 was demonstrated by coimmunoprecipitation experiments. We concluded that Olfm1 serves as a modulator of Wnt signaling.

The unlimited proliferative and differentiative capacities of embryonic stem cells (ESCs) are tightly regulated by their microenvironment. Local concentrations of soluble factors, cell-cell interactions and extracellular matrix signaling are just a few variables that influence ESC fate. A common method employed to induce ESC differentiation involves the formation of cell aggregates called embryoid bodies (EBs), which recapitulate early stages of embryonic development. EBs are normally formed in suspension cultures, producing heterogeneously shaped and sized aggregates. The present study demonstrates the usage of a microfluidic traps system which supports prolonged EB culturing. The traps are uniquely designed to facilitate cell capture and aggregation while offering efficient gas/nutrients exchange. A finite element simulation is presented with emphasis on several aspects critical to appropriate design of such bioreactors for ESC culture. Finally, human ESC, mouse Nestin-GFP ESC and OCT4-EGFP ESCs were cultured using this technique and demonstrated extended viability for more than 5 days. In addition, EBs developed and maintained a polarized differentiation pattern, possibly as a result of the nutrient gradients imposed by the traps bioreactor. The novel microbioreactor presented here can enhance future embryogenesis research by offering tight control of culturing conditions.

No abstract

Under aquaculture conditions, European eels (Anguilla anguilla) produce a high percentage of males (809 5%) that normally stop growing at 100^200 g. Females continue to grow to 500^750 g and obtain higher market value. Therefore, increasing the percentage of females in a population would be bene¢cial to the culture of eels. The present study was carried out in order to examine the e¡ect of oestradiol and phytooestrogens on sex di¡erentiation and growth rate of eels. Juvenile European eels with undi¡erentiated gonads were fed pellets containing oestradiol-17b (E2) or phytooestrogens for 100^150 days. Feeding E2 resulted in 50^61% increase in body weight compared with the control. Oestradiol-17b and phytooestrogens both elevated signi¢cantly the percentage of females in the population. Feeding E2 at 20 mg kg À1 feed resulted in 70% females, while lower concentration of E2 (2 mg kg À1 ) resulted in only 30% after 100 days (Experiment 1). The same dose given for 150 days (Experiment 2) resulted in 88% females, indicating that both, the concentration and duration of E2 treatments had a signi¢cant e¡ect on sex di¡erentiation. Fish fed genistein at 2 mg kg À1 for 100 days, resulted in 55% of females, but at a higher dose of 20 mg kg À1 there were only15% females. These results demonstrate that phytooestrogens can be used as alternatives to gonadal steroids for sex manipulation in eels, but the optimal concentrations and duration are still to be determined.