Mesenchymal stem/stromal cells (MSCs) can regenerate tissues by direct differentiation or indirectly by stimulating angiogenesis, limiting inflammation, and recruiting tissue-specific progenitor cells. MSCs emerge and multiply in long-term cultures of total cells from the bone marrow or multiple other organs. Such a derivation in vitro is simple and convenient, hence popular, but has long precluded understanding of the native identity, tissue distribution, frequency, and natural role of MSCs, which have been defined and validated exclusively in terms of surface marker expression and developmental potential in culture into bone, cartilage, and fat. Such simple, widely accepted criteria uniformly typify MSCs, even though some differences in potential exist, depending on tissue sources. Combined immunohistochemistry, flow cytometry, and cell culture have allowed tracking the artifactual cultured mesenchymal stem/stromal cells back to perivascular anatomical regions. Presently, both pericytes enveloping microvessels and adventitial cells surrounding larger arteries and veins have been described as possible MSC forerunners. While such a vascular association would explain why MSCs have been isolated from virtually all tissues tested, the origin of the MSCs grown from umbilical cord blood remains unknown. In fact, most aspects of the biology of perivascular MSCs are still obscure, from the emergence of these cells in the embryo to the molecular control of their activity in adult tissues. Such dark areas have not compromised intents to use these cells in clinical settings though, in which purified perivascular cells already exhibit decisive advantages over conventional MSCs, including purity, thorough characterization and, principally, total independence from in vitro culture. A growing body of experimental data is currently paving the way to the medical usage of autologous sorted perivascular cells for indications in which MSCs have been previously contemplated or actually used, such as bone regeneration and cardiovascular tissue repair.

The non‐structural proteins of Sindbis virus, nsP1, 2, 3 and 4, are produced upon cleavage of polyproteins P123 and P1234 by a proteinase residing in nsP2. We used cell free translation of SP6 transcripts to study the proteolytic activity of nsP2 and of nsP2‐containing polyproteins. To generate polyprotein enzymes, a set of plasmids was made in which cleavage sites were eliminated and new initiation and termination codons introduced by in vitro mutagenesis. As a substrate, we used a polyprotein in which the nsP2 proteinase had been inactivated by a single amino acid substitution. All nsP2‐containing polyproteins cleaved the nsP1/2 site in trans. However, proteinases containing nsP1 were unable to cleave the nsP2/3 site. Furthermore, only proteinases containing nsP3 could cleave the nsP3/4 site. These differences in cleavage site specificity result in a temporal regulation of processing in vivo. At 1.7 h post infection P123 and nsP4 accumulated and only small amounts of P34 were found. However, at 4 h post infection P123 was processed rapidly and P34 was produced rather than nsP4. Since nsP4 is thought to be the viral RNA polymerase, the temporal regulation of the nsP4/P34 ratio may be responsible for the temporal regulation of RNA synthesis.

Electrochemistry of surface-modified cytochrome c (cyt c) bound electrostatically to carboxylate-terminated alkanethiol self-assembled monolayers (SAM) reveals highly anisotropic electronic coupling across the protein/ monolayer interface. Substitution of a lysine residue with alanine at position 13 in recombinant rat cyt c (RC9-K13A) lowers the interfacial electron transfer (ET) rate more than 5 orders of magnitude, whereas ET is only slightly affected by replacement of lysine-72 or lysine-79 with alanine. The results clearly show that lysine-13 is directly involved in coupling the protein to the SAM carboxylate terminus. Interfacial ET rates for both yeast iso-1 cyt c and the mutant RC9-K13R indicate that arginine-13 couples the protein to the carboxylate interface less well than lysine-13.