Luigi Vitelli

Eicosapentaenoic acid stimulates the expression of myelin proteins in rat brain.

We have previously demonstrated that, in C6 glioma cells, eicosapentaenoic acid (EPA) stimulates the expression of proteolipid protein (PLP) via cAMP‐mediated pathways. In this study, we investigated whether n‐3 polyunsaturated fatty acids can affect myelinogenesis in vivo. A single dose of either EPA or docosahexaenoic acid (DHA) was injected intracerebroventricularly into 2‐day‐old rats, which were then killed after 3 days post‐injection (p.i.). Total RNA was isolated from the medulla, cerebellum, and cortex, and the expression of myelin‐specific mRNAs was analyzed by real‐time PCR. The levels of PLP, myelin basic protein, and myelin oligodendrocyte protein mRNAs increased in nearly all brain regions of DHA‐ and EPA‐treated animals, but the effect was more pronounced in EPA‐treated rats. The enhancement in PLP transcript levels was followed by an increase in PLP translation in EPA‐treated rats. A further indicator of accelerated myelination was the increase in 2′‐3′‐cyclic nucleotide 3′‐phosphodiesterase (CNPase) protein levels. In EPA‐treated rats, the increased expression of myelin genes coincided with a decrease of cAMP‐response element‐binding protein (CREB)‐DNA binding in the cerebellum and cortex (1 hr p.i.). After 16 hr, this effect was still present in the same cerebral regions even though the decrease in EPA‐treated rats was less pronounced than in controls. The down‐regulation of CREB activity was due to a decrease in the levels of CREB phosphorylation. In conclusion, our data suggest that EPA stimulates the expression of specific myelin proteins through decreased CREB phosphorylation. These results corroborate the clinical studies of the n‐3 PUFA beneficial effects on several demyelinating diseases.

Chromatin immunoselection defines a TAL-1 target gene

Despite the major functions of the basic helix–loop–helix transcription factor TAL‐1 in hematopoiesis and T‐cell leukemogenesis, no TAL‐1 target gene has been identified. Using immunoprecipitation of genomic fragments bound to TAL‐1 in the chromatin of murine erythro‐leukemia (MEL) cells, we found that 10% of the immunoselected fragments contained a CAGATG or a CAGGTG E‐box, followed by a GATA site. We studied one of these fragments containing two E‐boxes, CAGATG and CAGGTC, followed by a GATA motif, and showed that TAL‐1 binds to the CAGGTG E‐box with an affinity modulated by the CAGATG or the GATA site, and that the CAGGTG–GATA motif exhibits positive transcriptional activity in MEL but not in HeLa cells. This immunoselected sequence is located within an intron of a new gene co‐expressed with TAL‐1 in endothelial and erythroid cells, but not expressed in fibroblasts or adult liver where no TAL‐1 mRNA was detected. Finally, in vitro differentiation of embryonic stem cells towards the erythro/megakaryocytic pathways showed that the TAL‐1 target gene expression followed TAL‐1 and GATA‐1 expression. These results establish that TAL‐1 is likely to activate its target genes through a complex that binds an E‐box–GATA motif and define the first gene regulated by TAL‐1.

Pressure overload causes cardiac hypertrophy in β1-adrenergic and β2-adrenergic receptor double knockout mice

Objective Cardiac hypertrophy arises as an adaptive response to increased afterload. Studies in knockout mice have shown that catecholamines, but not α1-adrenergic receptors, are necessary for such an adaptation to occur. However, whether β-adrenergic receptors are critical for the development of cardiac hypertrophy in response to pressure overload is not known at this time. Methods and results Pressure overload was induced by transverse aortic banding in β1-adrenergic and β2-adrenergic receptor double knockout (DβKO) mice, in which the predominant cardiac β-adrenergic receptor subtypes are lacking. Chronic pressure overload for 4 weeks induced cardiac hypertrophy in both DβKO and wild-type mice. There were no significant differences between banded mice in left ventricular weight to body weight ratio, in the left ventricular wall thickness, in the cardiomyocyte size or in the expression levels of the load-sensitive cardiac genes such as ANF and β-MHC. Additionally, the left ventricular systolic pressure, an index of afterload, and cardiac contractility, evaluated as dp/dtmax, the maximal slope of systolic pressure increment, and Ees, end-systolic elastance, were increased at a similar level in both wild-type and DβKO banded mice, and were significantly greater than in sham controls. Conclusion Despite chronic activation of the cardiac β-adrenergic system being sufficient to induce a pathological hypertrophy, we show that β1-adrenergic and β2-adrenergic receptors are not an obligatory component of the signaling pathway that links the increased afterload to the development of cardiac hypertrophy.

Cell Dynamics in Early Embryogenesis and Pluripotent Embryonic Cell Lines: From Sea Urchin to Mammals

From oogenesis through fertilization and gastrulation, embryos use various mechanisms to regulate cell expansion, keeping a strict balance between cell proliferation, cell differentiation and cell death. While rapid divisions are necessary at the initial stage to ensure early embryo survival, further developmental transitions are marked by changes in cell cycle and transcriptional regulation. Pluripotency and capability of self-renewal are maintained in a low percentage of cells, the embryonic stem cells (ESCs), which will be later used as a cellular source for tissue replacement. Clearly, some essential characteristics of cell cycle and transcriptional regulation of the early embryo will be conserved in ESC lines. We addressed here the peculiarities of these developmental programs in early embryos and pluripotent embryonic cell lines, considering examples from marine invertebrates to mammals. Finally, we discussed the importance of transcription regulation and chromatin remodelling and their peculiar features in embryonic cells from these species.