The 7th amendment to the EU Cosmetics Directive prohibits to put animal-tested cosmetics on the market in Europe after 2013. In that context, the European Commission invited stakeholder bodies (industry, non-governmental organisations, EU Member States, and the Commission's Scientific Committee on Consumer Safety) to identify scientific experts in five toxicological areas, i.e. toxicokinetics, repeated dose toxicity, carcinogenicity, skin sensitisation, and reproductive toxicity for which the Directive foresees that the 2013 deadline could be further extended in case alternative and validated methods would not be available in time. The selected experts were asked to analyse the status and prospects of alternative methods and to provide a scientifically sound estimate of the time necessary to achieve full replacement of animal testing. In summary, the experts confirmed that it will take at least another 7-9 years for the replacement of the current in vivo animal tests used for the safety assessment of cosmetic ingredients for skin sensitisation. However, the experts were also of the opinion that alternative methods may be able to give hazard information, i.e. to differentiate between sensitisers and non-sensitisers, ahead of 2017. This would, however, not provide the complete picture of what is a safe exposure because the relative potency of a sensitiser would not be known. For toxicokinetics, the timeframe was 5-7 years to develop the models still lacking to predict lung absorption and renal/biliary excretion, and even longer to integrate the methods to fully replace the animal toxicokinetic models. For the systemic toxicological endpoints of repeated dose toxicity, carcinogenicity and reproductive toxicity, the time horizon for full replacement could not be estimated.

Somatic stem cell (SSC) dysfunction is typical for different progeroid phenotypes in mice with genomic DNA repair defects. MtDNA mutagenesis in mice with defective Polg exonuclease activity also leads to progeroid symptoms, by an unknown mechanism. We found that Polg-Mutator mice had neural (NSC) and hematopoietic progenitor (HPC) dysfunction already from embryogenesis. NSC self-renewal was decreased in vitro, and quiescent NSC amounts were reduced in vivo. HPCs showed abnormal lineage differentiation leading to anemia and lymphopenia. N-acetyl-L-cysteine treatment rescued both NSC and HPC abnormalities, suggesting that subtle ROS/redox changes, induced by mtDNA mutagenesis, modulate SSC function. Our results show that mtDNA mutagenesis affected SSC function early but manifested as respiratory chain deficiency in nondividing tissues in old age. Deletor mice, having mtDNA deletions in postmitotic cells and no progeria, had normal SSCs. We propose that SSC compartment is sensitive to mtDNA mutagenesis, and that mitochondrial dysfunction in SSCs can underlie progeroid manifestations.

In this article, we show that, in transfected COS-1 cells, protein tyrosine phosphatase (PTP)-PEST translocates to the membrane periphery following stimulation by the extracellular matrix protein fibronectin. When plated on fibronectin, PTP-PEST (−/−) fibroblasts display a strong defect in motility. 3 h after plating on fibronectin, the number and size of vinculin containing focal adhesions were greatly increased in the homozygous PTP-PEST mutant cells as compared with heterozygous cells. This phenomenon appears to be due in part to a constitutive increase in tyrosine phosphorylation of p130CAS, a known PTP-PEST substrate, paxillin, which associates with PTP-PEST in vitro, and focal adhesion kinase (FAK). Another effect of this constitutive hyperphosphorylation, consistent with the focal adhesion regulation defect, is that (−/−) cells spread faster than the control cell line when plated on fibronectin. In the PTP-PEST (−/−) cells, an increase in affinity for the SH2 domains of Src and Crk towards p130CAS was also observed. In (−/−) cells, we found a significant increase in the level of tyrosine phosphorylation of PSTPIP, a cleavage furrow–associated protein that interacts physically with all PEST family members. An effect of PSTPIP hyperphosphorylation appears to be that some cells remain attached at the site of the cleavage furrow for an extended period of time. In conclusion, our data suggest PTP-PEST plays a dual role in cell cytoskeleton organization, by promoting the turnover of focal adhesions required for cell migration, and by directly or indirectly regulating the proline, serine, threonine phosphatase interacting protein (PSTPIP) tyrosine phosphorylation level which may be involved in regulating cleavage furrow formation or disassembly during normal cell division.