Isolation and Cryopreservation of Animal Mesenchymal Stromal Cells

Scientific progress in cellular and molecular biotechnology has led to the development of advanced therapies, such as gene therapy, cell therapy, and tissue engineering. The application of stem cells as therapeutic agents has been investigated for several years in human medicine and, more recently, the same approach has been considered in the veterinary field as a novel opportunity for the treatment of animal diseases. Mesenchymal stem cell (MSC)-based therapies seem to contribute to the healing process by several mechanisms due to their peculiar biological features. It has been shown that MSCs could effectively differentiate into the required cell type to replace the damaged tissue. Furthermore, due to their autocrine and paracrine secretory activities, these cells are a powerful source of trophic mediators, growth factors, cytokines, and extracellular matrix components. The clinical application of MSCs needs great amounts of cells designed for in vivo implantation that can be obtained following their in vitro isolation, serial subcultivations, cryopreservation, and thawing. These procedures could determine their feature changes which could interfere with the therapeutic outcome. For these reasons, to preserve MSCs after in vitro manipulation for future applications, standardized quality controls and a reliable long-term cryopreservation method are required

Transformation and tumorigenicity testing of simian cell lines and evaluation of poliovirus replication

The key role of cell cultures in different scientific fields is worldwide recognized, both as in vitro research models alternative to laboratory animals and substrates for biological production. However, many safety concerns rise from the use of animal/human cell lines that may be tumorigenic, leading to potential adverse contaminations in cell-derived biologicals. In order to evaluate the suitability of 13 different cell lines for Poliovirus vaccine production, safety and quality, in vitro/in vivo tumorigenicity and Poliovirus propagation properties were evaluated. Our results revealed that non-human primate cell lines CYNOM-K1, FRhK-4, 4MBr-5 and 4647 are free of tumorigenic features and represent highly susceptible substrates for attenuated Sabin Poliovirus strains. In particular, FRhK-4 and 4647 cell lines are characterized by a higher in vitro replication, resulting indicated for the use in large-scale production field

Intracellular retention of ABL kinase inhibitors determines commitment to apoptosis in CML cells

Clinical development of imatinib in CML established continuous target inhibition as a paradigm for successful tyrosine kinase inhibitor (TKI) therapy. However, recent reports suggested that transient potent target inhibition of BCR-ABL by highdose TKI (HD-TKI) pulse-exposure is sufficient to irreversibly commit cells to apoptosis. Here, we report a novel mechanism of prolonged intracellular TKI activity upon HD-TKI pulse-exposure (imatinib, dasatinib) in BCR-ABL-positive cells. Comprehensive mechanistic exploration revealed dramatic intracellular accumulation of TKIs which closely correlated with induction of apoptosis. Cells were rescued from apoptosis upon HD-TKI pulse either by repetitive drug wash-out or by overexpression of ABC-family drug transporters. Inhibition of ABCB1 restored sensitivity to HD-TKI pulse-exposure. Thus, our data provide evidence that intracellular drug retention crucially determines biological activity of imatinib and dasatinib. These studies may refine our current thinking on critical requirements of TKI dose and duration of target inhibition for biological activity of TKIs.Daniel B. Lipka, Marie-Christine Wagner, Marek Dziadosz, Tina Schnöder, Florian Heidel, Mirle Schemionek, Junia V. Melo, Thomas Kindler, Carsten Müller-Tidow, Steffen Koschmieder and Thomas Fische

Protein mutations following adaptation of avian influenza viruses in different biological systems

Traditionally, embryonated chicken eggs (ECE) are considered the gold standard for Influenza virus isolation and vaccine production. Nowadays, different biological systems have been improved and performed, in order to evaluate a feasible alternative to ECE. In fact, in a previous study, mammalian and avian cell cultures were successfully used for avian influenza viruses primary isolation from target tissues and virus propagation. This research is focused on the investigation of adaptive mutations that occur after influenza A virus amplification in ECE and cell cultures. The results of the study shows that avian influenza viruses after multiple passages in different biological systems undergo mutations, in particular, the largest number of amino acid substitutions occurred in all biological substrates in the hemagglutinin

Cell line quality and tumorigenicity parameters.

<p>Cell line quality and tumorigenicity parameters.</p

Cell lines used in the study.

<p>Cell lines used in the study.</p

Soft agar colony assay results.

<p>Representative captures of <i>in vitro</i> transformation assay results are reported for each investigated cell line.</p

Tumorigenic evaluation and histology.

<p>Representative captures of the absence (A) and presence (B) of a nodular lesion, localized at the site of injection of cells, in treated nude mice. In Panel C histological sections derived from cell-treated mice are reported.</p

The problem with reproductive freedom:Procreation beyond procreators’ interests

Reproductive freedom plays a pivotal role in debates on the ethics of procreation. This moral principle protects people’s interests in procreative matters and allows them discretion over whether to have children, the number of children they have and, to a certain extent, the type of children they have. Reproductive freedom’s theoretical and political emphasis on people’s autonomy and well-being is grounded in an individual-centred framework for discussing the ethics of procreation. It protects procreators’ interests and significantly reduces the permissible grounds for interference by third parties. In this article I show that procreative decisions have far-reaching effects on the composition and size of the population. The upshot of considering these effects allows for the appreciation of the inadequacy of a framework that solely considers individual (i.e. procreators’) interests to discuss the ethics of procreation. To address such inadequacy, I assess costs and benefits of past and present proposals to reflect on procreation in such a way as to consider its far-reaching effects. I conclude by arguing that reproductive freedom should be defended as an imperfect but instrumentally necessary tool. This framing would enable those participating in debates on the ethics of procreative decisions to work towards an ethical framework that accounts for the cumulative effects of these decisions