We found adult human stem cells that can generate, from a single cell, cells with the characteristics of the three germ layers. The cells are stress-tolerant and can be isolated from cultured skin fibroblasts or bone marrow stromal cells, or directly from bone marrow aspirates. These cells can self-renew; form characteristic cell clusters in suspension culture that express a set of genes associated with pluripotency; and can differentiate into endodermal, ectodermal, and mesodermal cells both in vitro and in vivo. When transplanted into immunodeficient mice by local or i.v. injection, the cells integrated into damaged skin, muscle, or liver and differentiated into cytokeratin 14-, dystrophin-, or albumin-positive cells in the respective tissues. Furthermore, they can be efficiently isolated as SSEA-3(+) cells. Unlike authentic ES cells, their proliferation activity is not very high and they do not form teratomas in immunodeficient mouse testes. Thus, nontumorigenic stem cells with the ability to generate the multiple cell types of the three germ layers can be obtained through easily accessible adult human mesenchymal cells without introducing exogenous genes. These unique cells will be beneficial for cell-based therapy and biomedical research.
The stochastic and elite models have been proposed for the mechanism of induced pluripotent stem (iPS) cell generation. In this study we report a system that supports the elite model. We previously identified multilineage-differentiating stress-enduring (Muse) cells in human dermal fibroblasts that are characterized by stress tolerance, expression of pluripotency markers, selfrenewal, and the ability to differentiate into endodermal-, mesodermal-, and ectodermal-lineage cells from a single cell. They can be isolated as stage-specific embryonic antigen-3/CD105 doublepositive cells. When human fibroblasts were separated into Muse and non-Muse cells and transduced with Oct3/4, Sox2, Klf4, and c-Myc, iPS cells were generated exclusively from Muse cells but not from non-Muse cells. Although some colonies were formed from non-Muse cells, they were unlike iPS cells. Furthermore, epigenetic alterations were not seen, and some of the major pluripotency markers were not expressed for the entire period during iPS cell generation. These findings were confirmed further using cells transduced with a single polycistronic virus vector encoding all four factors. The results demonstrate that in adult human fibroblasts a subset of preexisting adult stem cells whose properties are similar in some respects to those of iPS cells selectively become iPS cells, but the remaining cells make no contribution to the generation of iPS cells. Therefore this system seems to fit the elite model rather than the stochastic model.
Summary On the basis of discussions emerging from a workshop and discussions at the 7th meeting of the European Society for Pigment Cell Research in Geneva in 2012, this manuscript outlines useful criteria for defining the bona fide pigment cells as a functional entity of the vertebrate body plan and differentiating them from ‘pigmented’ cells in general. It also proposes a nomenclature for various types of pigment cells of vertebrates.
Measurements of spectral reflectance from the sky-blue portion of skin from the common surgeonfish, Paracanthurus hepatus, showed a relatively steep peak at around 490 nm. We consider that a multilayer thin-film interference phenomenon of the non-ideal type, which occurs in stacks of very thin light-reflecting platelets in iridophores of that region, is primarily responsible for the revelation of that hue. The structural organization of the iridophore closely resembles that of bluish damselfish species, although one difference is the presence of iridophores in a monolayer in the damselfish compared to the double layer of iridophores in the uppermost part of the dermis of surgeonfish. If compared with the vivid cobalt blue tone of the damselfish, the purity of the blue hue of the surgeonfish is rather low. This may be ascribable mainly to the double layer of iridophores in the latter since incident lightrays are complicatedly reflected and scattered in the strata. The dark-blue hue of the characteristic scissors-shaped pattern on the trunk of surgeonfish is mainly due to the dense population of melanophores, because iridophores are only present there in a scattered fashion. Photographic and spectral reflectance studies in vivo, as well as photomicrographic, photo-electric, and spectrometric examinations of the state of chromatophores in skin specimens in vitro, indicate that both melanophores and iridophores are motile. Physiological analyses disclosed that melanophores are under the control of the sympathetic nervous and the endocrine systems, while iridophores are regulated mainly by nerves. The body color of surgeonfish shows circadian changes, and becomes paler at night; this effect may be mediated by the pineal hormone, melatonin, which aggregates pigment in melanophores.
We report here the striking anisotropy of fluorescence exhibited by crystals of native green fluorescence protein (GFP). The crystals were generated by water dialysis of highly purified GFP obtained from the jellyfish Aequorea. We find that the fluorescence becomes six times brighter when the excitation, or emission, beam is polarized parallel (compared with perpendicular) to the crystal long axis. Thus, the major dipoles of the fluorophores must be oriented very nearly parallel to the crystal long axis. Observed in a polarizing microscope between parallel polars instead of either a polarizer or analyzer alone, the fluorescence polarization ratio rises to an unexpectedly high value of about 30:1, nearly the product of the fluorescence excitation and emission ratios, suggesting a sensitive method for measuring fluorophore orientations, even of a single fluorophore molecule. We have derived equations that accurately describe the relative fluorescence intensities of crystals oriented in various directions, with the polarizer and analyzer arranged in different configurations. The equations yield relative absorption and fluorescence coefficients for the four transition dipoles involved. Finally, we propose a model in which the elongated crystal is made of GFP molecules that are tilted 60°t o align the fluorophores parallel to the crystal long axis. The unit layer in the model may well correspond to the arrangement of functional GFP molecules, to which resonant energy is efficiently transmitted from Ca 2؉ -activated aequorin, in the jellyfish photophores. Green fluorescence protein (GFP), initially extracted and purified from the luminescent jellyfish, Aequorea sp, converts the blue light (that would otherwise be emitted by the Ca 2ϩ -sensitive protein aequorin) into a brilliant green fluorescence, the ''luminescence'' emitted by this jellyfish (1-3). Both of these proteins are localized at high concentration in the photophores of Aequorea. Today, GFP and its genetically encoded variants are widely used as noninvasive fluorescent biosensors for protein expression, protein dynamics, and protein-protein interactions in living cells (4,5).According to x-ray crystallographic analyses (6, 7), GFP is a barrel-shaped molecule (28 kDa, made of 238-aa residues), about 24 Å in diameter and 42 Å in length. The outer cylinder of the barrel (the '' can'') is composed of 11 antiparallel  sheets capped with ␣-helical stretches of the molecule, part of which extends to the interior of the can and forms the fluorescent chromophore. The chromophore is situated near the center of the  can and lies at Ϸ60°to the long axis of the can. Interaction between the (posttranslationally modified) tripeptide chromophore and those of neighboring residues is shown to determine the exact fluorescence property of GFP and its related constructs (6-8); the chromophore is no longer fluorescent when isolated from the  can.In this paper, we report on the anisotropic fluorescent properties of GFP as measured in crystals of native GFP (see Materials and ...
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