Cytotoxicity assays provide an in vitro evaluation of the lytic activity of NK and T cells against tumors or transformed cells. However, none of these methods allow the recovery of cells or supernatants after the assay. We standardized a microcytotoxicity test using calcein-acetoxymethyl (calcein-AM) dye that requires very small quantities of cells while maintaining the same sensitivity as the traditional⁵¹Cr assay. The assay is applicable to resting as well as activated human effector cells and uses different targets such as human cell lines that are adherent or growing in suspension and resistant or sensitive. The most important feature of the method is the possibility of recovering cells and supernatants for additional analyses such as phenotyping and evaluation of soluble factors.

Mesenchymal stem/stromal cells (MSC) show widespread application for a variety of clinical conditions; therefore, their use necessitates continuous monitoring of their safety. The risk assessment of mesenchymal stem cell-based therapies cannot be separated from an accurate and deep knowledge of their biological properties and in vitro and in vivo behavior. One of the most relevant safety issues is represented by the genetic stability of MSCs, that can be altered during in vitro manipulation, frequently required before clinical application. MSC genetic stability has the potential to influence the transformation and the therapeutic effect of these cells. At present, karyotype evaluation represents the definitely prevailing assessment of MSC stability, but DNA alterations of smaller size should not be underestimated. This review will focus on current scientific knowledge about the genetic stability of mesenchymal stem cells. The techniques used and possible improvements together with regulatory aspects will also be discussed.

Mesenchymal stem/stromal cells (MSCs) are a reservoir for tissue homeostasis and repair that age during organismal aging. Beside the fundamental in vivo role of MSCs, they have also emerged in the last years as extremely promising therapeutic agents for a wide variety of clinical conditions. MSC use frequently requires in vitro expansion, thus exposing cells to replicative senescence. Aging of MSCs (both in vivo and in vitro) can affect not only their replicative potential, but also their properties, like immunomodulation and secretory profile, thus possibly compromising their therapeutic effect. It is therefore of critical importance to unveil the underlying mechanisms of MSC senescence and to define shared methods to assess MSC aging status. The present review will focus on current scientific knowledge about MSC aging mechanisms, control and effects, including possible anti-aging treatments.

In this study we investigated whether purified NK cells, derived from a group of nonagenarian healthy subjects, were able to produce the chemokines MIP‐1α, RANTES and IL‐8, and also characterized the effect of IL‐12 or IL‐2 immunomodulatory cytokines (that are among the most effective inducers of NK lytic activity and soluble factor secretion) on the induction, in vitro, of these chemokines and on the modulation of the corresponding receptors. This study provides evidence that human NK cells from healthy subjects over 90 years old retain the ability to synthesize MIP‐1α, Rantes and IL‐8 chemotactic cytokines, that NK cells isolated from these subjects can be activated to significantly up‐regulate the production of these chemokines in response to stimulation by IL‐12 or IL‐2 cytokines (even though production remains lower than that observed in young subjects), and that NK cells express the corresponding chemokine receptors.