Regulation of Growth and Differentiation in Early Mammalian Embryo by FGF and NGF Growth Factors in Organ Culture in VitroReportar como inadecuado

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Acta clinica Croatica, Vol.44 No.2 June 2005. -

The aim was to analyze the regulation of growth and tissue differentiation in a unique in vitro4culture model of gastrulating mammalian embryo by fibroblast growth factor FGF and nerve growth factor NGF during two weeks. They both play a crucial role during embryogenesis and the purpose of this study was to test their possible synergistic influence in the period when mammalian embryos are extremely sensitive to external factors. We cultivated 9.5-day-old rat embryos on metal grid supported lens paper, at the air-liquid interface in culture medium Eagle-s minimal essential medium MEM with 50% of homologous serum with the addition of FGF, NGF and a combination of FGF-NGF in a time frame of 9 days. Other three groups of embryos were for 24 hours pretreated with 5-azacytidine 5azaC, an agent that can activate repressed genes. A parallel group of nontreated control embryos were cultivated with each experimental group. During 14 days of culture embryos grown in teratoma-like explants and growth rate were evaluated by measuring average size of explants using an eyepiece micrometer on days 5, 7, 11 and 14 after the addition of growth factors. Differences between respective groups were estimated by Student-s t-test. Differentiated tissues were estimated on serial histologic sections. c2-test or Fisher exact test were used to compare the proportion of tissues between respective groups. In embryo-derived teratomas NGF or FGF- NGF combination used within the 9 day time frame did not stimulate differentiation of any kind of tissues; moreover, FGF-NGF inhibited maturation of epidermis, while FGF stimulated differentiation of neural tissue, hemopoiesis and myotubes. We did not observe any kind of stimulative cooperative action of FGF and NGF in differentiation processes. So it seems that NGF hinders the stimulating effect of FGF. NGF alone impaired growth of explants, but in combination with FGF acted synergistically, thus improving the growth rate of cultivated embryos. Additional activation of genes with 5azaC had no the effect on possible NGF influence on neural tissue differentiation, but resulted in improved myotube differentiation. The activation of genes with 5azaC-FGF signal and 5azaC-FGF-NGF combination improved the proportion of neural tissue and myotubes as well as hemopoiesis. Obviously, these results supported the role of FGF as neural inducer and mesoderm inducer. Anyway, FGF or NGF induced differentiation at least partially depends on the status of gene methylation.

Embryo culture; Fibroblast growth factor; Nerve growth factor; Embryonic teratomas

Autor: Vesna Crnek-Kunstelj - Jagoda Stipić - Tomislav Zeljko -



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