Advances in nanotechnology have opened up a new era of diagnosis, prevention and treatment of diseases and traumatic injuries. Nanomaterials, including those with potential for clinical applications, possess novel physicochemical properties that have an impact on their physiological interactions, from the molecular level to the systemic level. There is a lack of standardized methodologies or regulatory protocols for detection or characterization of nanomaterials. This review summarizes the techniques that are commonly used to study the size, shape, surface properties, composition, purity and stability of nanomaterials, along with their advantages and disadvantages. At present there are no FDA guidelines that have been developed specifically for nanomaterial based formulations for diagnostic or therapeutic use. There is an urgent need for standardized protocols and procedures for the characterization of nanoparticles, especially those that are intended for use as theranostics.

Yeast deprived of nutrients exhibit a marked life span extension that requires the activity of the NAD ؉ -dependent histone deacetylase, Sir2p. Here we show that increased dosage of NPT1, encoding a nicotinate phosphoribosyltransferase critical for the NAD ؉ salvage pathway, increases Sir2-dependent silencing, stabilizes the rDNA locus, and extends yeast replicative life span by up to 60%. Both NPT1 and SIR2 provide resistance against heat shock, demonstrating that these genes act in a more general manner to promote cell survival. We show that Npt1 and a previously uncharacterized salvage pathway enzyme, Nma2, are both concentrated in the nucleus, indicating that a significant amount of NAD ؉ is regenerated in this organelle. Additional copies of the salvage pathway genes, PNC1, NMA1, and NMA2, increase telomeric and rDNA silencing, implying that multiple steps affect the rate of the pathway. Although SIR2-dependent processes are enhanced by additional NPT1, steady-state NAD ؉ levels and NAD ؉ /NADH ratios remain unaltered. This finding suggests that yeast life span extension may be facilitated by an increase in the availability of NAD ؉ to Sir2, although not through a simple increase in steady-state levels. We propose a model in which increased flux through the NAD ؉ salvage pathway is responsible for the Sir2-dependent extension of life span.

A relationship between life span and cellular glucose metabolism has been inferred from genetic manipulations and caloric restriction of model organisms. In this report, we have used the Snf1p glucose-sensing pathway of Saccharomyces cerevisiae to explore the genetic and biochemical linkages between glucose metabolism and aging. Snf1p is a serine/threonine kinase that regulates cellular responses to glucose deprivation. Loss of Snf4p, an activator of Snf1p, extends generational life span whereas loss of Sip2p, a presumed repressor of the kinase, causes an accelerated aging phenotype. An annotated data base of global age-associated changes in gene expression in isogenic wild-type, sip2⌬, and snf4⌬ strains was generated from DNA microarray studies. The transcriptional responses suggested that gluconeogenesis and glucose storage increase as wild-type cells age, that this metabolic evolution is exaggerated in rapidly aging sip2⌬ cells, and that it is attenuated in longerlived snf4⌬ cells. To test this hypothesis directly, we applied microanalytic biochemical methods to generation-matched cells from each strain and measured the activities of enzymes and concentrations of metabolites in the gluconeogenic, glycolytic, and glyoxylate pathways, as well as glycogen, ATP, and NAD ؉ . The sensitivity of the assays allowed comprehensive biochemical profiling to be performed using aliquots of the same cell populations employed for the transcriptional profiling. The results provided additional evidence that aging in S. cerevisiae is associated with a shift away from glycolysis and toward gluconeogenesis and energy storage. They also disclosed that this shift is forestalled by two manipulations that extend life span, caloric restriction and genetic attenuation of the normal age-associated increase in Snf1p activity. Together, these findings indicate that Snf1p activation is not only a marker of aging but also a candidate mediator, because a shift toward energy storage over expenditure could impact myriad aspects of cellular maintenance and repair. Genetic studies in model organisms imply that changes in glucose and energy metabolism can alter life span (1-4), although there has been little direct biochemical analysis of this hypothesis in aging cells (5). Saccharomyces cerevisiae is an attractive model for studying how glucose and energy metabolism are linked to aging. Age-associated alterations in energy metabolism can be analyzed more readily in a unicellular eukaryote than in a multicellular organism with diverse

Use of clinical-grade human induced pluripotent stem cell (iPSC) lines as a starting material for the generation of cellular therapeutics requires demonstration of comparability of lines derived from different individuals and in different facilities. This requires agreement on the critical quality attributes of such lines and the assays that should be used. Working from established recommendations and guidance from the International Stem Cell Banking Initiative for human embryonic stem cell banking, and concentrating on those issues more relevant to iPSCs, a series of consensus workshops has made initial recommendations on the minimum dataset required to consider an iPSC line of clinical grade, which are outlined in this report. Continued evolution of this field will likely lead to revision of these guidelines on a regular basis.

Saccharomyces cerevisiae has evolved a number of mechanisms for sensing glucose. In the present study we examine the mechanism by which one of these pathways, involving Snf1, regulates cellular aging. Snf1 is a heterotrimer composed of a catalytic ␣ subunit (Snf1p) that phosphorylates target proteins at Ser/Thr residues, an activating ␥ subunit (Snf4p), and a ␤ subunit (Sip1p, Sip2p, or Gal83). We previously showed that forced expression of Snf1p or loss of Sip2p, but not the other ␤ subunits, causes accelerated aging, while removal of Snf4p extends life span (Ashrafi, K., Lin, S. S., Manchester, J. K., and Gordon, J. I. (2000) Genes Dev. 14, 1872-1885). We now demonstrate that in wild type cells, there is an age-associated shift in Sip2p from the plasma membrane to the cytoplasm, a prominent redistribution of Snf4p from the plasma membrane to the nucleus, a modest increase in nuclear Snf1p, and a concomitant increase in cellular Snf1 histone H3 kinase activity. Covalent attachment of myristate to the N-terminal Gly of Sip2p is essential for normal cellular life span. When plasma membrane association of Sip2p is abolished by a mutation that blocks its N-myristoylation, Snf4p is shifted to the nucleus. Rapidly aging sip2⌬ cells have higher levels of histone H3 kinase activity than their generation-matched isogenic wild type counterparts. Increased Snf1 activity is associated with augmented recombination at rDNA loci, plus desilencing at sites affected by Snf1-catalyzed Ser 10 phosphorylation of histone H3 (the INO1 promoter plus targets of the transcription factor Adr1p). The rapidaging phenotype of sip2⌬ cells is fully rescued by blocking recombination at rDNA loci with a fob1⌬ allele; rescue is not accompanied by amelioration of an age-associated shift toward gluconeogenesis and glucose storage. Together, these findings suggest that Sip2p acts as a negative regulator of nuclear Snf1 activity in young cells by sequestering its activating ␥ subunit at the plasma membrane and that loss of Sip2p from the plasma membrane to the cytoplasm in aging cells facilities Snf4p entry into the nucleus so that Snf1 can modify chromatin structure.Saccharomyces cerevisiae is a unicellular model for studying the molecular mechanisms of aging. Mother yeast cells undergo replicative senescence, with different strains having characteristic mean and maximum life spans (1). Senescence is associated with a number of cellular and molecular phenotypes. Progressive sterility arises from the loss of silencing at HM loci (2). This loss of silencing is accompanied by a redistribution of Sir3p, a component of the Sir-silencing complex, from HM loci and telomeres to the nucleolus (3). Homologous recombination at rDNA loci liberates an extrachromosomal rDNA circle (ERC) 1 that contains an autonomously replicating sequence (4). With each round of cell division, replicated ERCs segregate to the mother rather than to daughter, leading to an exponential increase in cellular ERC concentrations over successive generations.ERC formation is thought to ...