Early corticogenesis was studied in human embryos and early fetuses from Carnegie stages 16 to 22 (5-8 gestational weeks) by using immunohistochemistry for Reelin (Reln), calretinin (CR), and glutamic acid decarboxylase (GAD). A first population of Reln-positive cells appears in the neocortical anlage at stage 16 and increases in number at stages 17-18. At stages 19-20, a monolayer of horizontal CR- and GAD-positive, Reln-negative neurons forms in the preplate, whereas Reln-positive cells shift into a subpial position. Another cell class, the pioneer projection neuron, is CR-positive but GAD- and Reln-negative; pioneer cells contribute early corticofugal axons. Pioneer cells first appear below the monolayer at stage 20 and form a pioneer plate at stage 21. The cortical plate (CP) proper emerges at stage 21 and inserts itself within the pioneer plate, which is thus split into a minor superficial component and a larger deep component that presumably corresponds to the subplate. Initial CP neurons are radially organized and mostly CR-negative. Reln-positive cells remain consistently segregated from the pioneer cells and are thus not directly involved in preplate partition. Our data indicate that the neuronal composition of the human neocortical preplate is more complex than generally described and that various neurons participate in a sequence of events that precede the emergence of the CP.
A family of new uridine phosphocholine amphiphiles that were prepared using a convenient four-step synthetic route is described. Physicochemical studies (differential scanning calorimetry, small-angle X-ray scattering, UV-vis and circular dichroism spectroscopies, light microscopy, transmission electronic microscopy, and scanning electron microscopy) show that these amphiphiles spontaneously assemble into supramolecular structures including vesicles, fibers, hydrogels, and organogels. In aqueous solution, the amphiphiles possessing saturated alkyl chains self-assemble into DNA-like helical fibers in the crystalline state below T(m) and compact bilayers above the melting temperature (T(m)). The transition from bilayers to fibers is thermally reversible. Above a threshold concentration (>6% w/w), a hydrogel is formed due to an entangled network of the fibers. A therapeutic agent such as DNA can be entrapped within the hydrogel structure. In addition to forming bilayer vesicles and hydrogels in aqueous solution, these nucleoside amphiphiles also form organogels in cyclohexane above T(m). Scanning electron microscopy shows a continuous multilamellar phase in the organogels.
A novel uridine-based nucleo-lipid, DOTAU (N-[5'-(2',3'-dioleoyl)uridine]-N',N',N'-trimethylammonium tosylate) was prepared by using a convenient four-step synthetic pathway. From the preliminary physicochemical studies (quasielastic light scattering and light microscopy), this amphiphilic structure forms supramolecular organizations in aqueous solution. In addition, in the presence of nucleic acids, transmission electronic microscopy experiments (TEM) and small angle X-ray scattering (SAXS) reveal the formation of multilamellar structures similar to lipoplexes (cationic liposome-DNA complexes) with cationic lipids. The formation of a complex was confirmed by fluorescence spectroscopic assays involving ethidium bromide. Transfection assays of mammalian cell lines (HeLa and MCF-7) indicate that DOTAU can transfect efficiently an expression vector (pEGFP) encoding GFP. Proliferation assays realized on these cell lines show that DOTAU does not inhibit cell proliferation and is less toxic than the commercial Lipofectamine 2000.
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