Mesenchymal stem cell conditioned medium increases glial reactivity and decreases neuronal survival in spinal cord slice cultures.

Ex vivo spinal cord slice cultures (SCSC) allow study of spinal cord circuitry, maintaining stimuli responses comparable to live animals. Previously, we have shown that mesenchymal stem/stromal cell (MSC) transplantation in vivo reduced inflammation and increased nerve regeneration but MSC survival was short-lived, highlighting that beneficial action may derive from the secretome. Previous in vitro studies of MSC conditioned medium (CM) have also shown increased neuronal growth. In this study, murine SCSC were cultured in canine MSC CM (harvested from the adipose tissue of excised inguinal fat) and cell phenotypes analysed via immunohistochemistry and confocal microscopy. SCSC in MSC CM displayed enhanced viability after propidium iodide staining. GFAP immunoreactivity was significantly increased in SCSC in MSC CM compared to controls, but with no change in proteoglycan (NG2) immunoreactivity. In contrast, culture in MSC CM significantly decreased the prevalence of ßIII-tubulin immunoreactive neurites, whilst Ca2+ transients per cell were significantly increased. These ex vivo results contradict previous in vitro and in vivo reports of how MSC and their secretome may affect the microenvironment of the spinal cord after injury and highlight the importance of a careful comparison of the different experimental conditions used to assess the potential of cell therapies for the treatment of spinal cord injury.

An In Vitro Comparison of the Neurotrophic and Angiogenic Activity of Human and Canine Adipose-Derived Mesenchymal Stem Cells (MSCs): Translating MSC-Based Therapies for Spinal Cord Injury.

The majority of research into the effects of mesenchymal stem cell (MSC) transplants on spinal cord injury (SCI) is performed in rodent models, which may help inform on mechanisms of action, but does not represent the scale and wound heterogeneity seen in human SCI. In contrast, SCI in dogs occurs naturally, is more akin to human SCI, and can be used to help address important aspects of the development of human MSC-based therapies. To enable translation to the clinic and comparison across species, we have examined the paracrine, regenerative capacity of human and canine adipose-derived MSCs in vitro. MSCs were initially phenotyped according to tissue culture plastic adherence, cluster of differentiation (CD) immunoprofiling and tri-lineage differentiation potential. Conditioned medium (CM) from MSC cultures was then assessed for its neurotrophic and angiogenic activity using established cell-based assays. MSC CM significantly increased neuronal cell proliferation, neurite outgrowth, and ßIII tubulin immunopositivity. In addition, MSC CM significantly increased endothelial cell migration, cell proliferation and the formation of tubule-like structures in Matrigel assays. There were no marked or significant differences in the capacity of human or canine MSC CM to stimulate neuronal cell or endothelial cell activity. Hence, this study supports the use of MSC transplants for canine SCI; furthermore, it increases understanding of how this may subsequently provide useful information and translate to MSC transplants for human SCI.

Exposing mesenchymal stem cells to chondroitin sulphated proteoglycans reduces their angiogenic and neuro-adhesive paracrine activity.

The multifactorial complexity of spinal cord injuries includes the formation of a glial scar, of which chondroitin sulphated proteoglycans (CSPG) are an integral component. Previous studies have shown CSPG to have inhibitory effects on endothelial and neuronal cell growth, highlighting the difficulty of spinal cord regeneration. Mesenchymal stem/stromal cells (MSC) are widely used as a cell therapy, and there is mounting evidence for their angiogenic and neurotrophic paracrine properties. However, in vivo studies have observed poor engraftment and survival of MSC when injected into SCI. Currently, it is not known whether increasing CSPG concentrations seen after SCI may affect MSC; therefore we have investigated the effects of CSPG exposure to MSC in vitro. CSPG-mediated inhibition of MSC adhesion was observed when MSC were cultured on substrates of increasing CSPG concentration, however MSC viability was not affected even up to five days of culture. Culture conditioned medium harvested from these cultures (primed MSC CM) was used as both culture substrata and soluble medium for EA.hy926 endothelial cells and SH-SY5Y neuronal cells. MSC CM was angiogenic, promoting endothelial cell adhesion, proliferation and tubule formation. However, exposing MSC to CSPG reduced the effects of CSPG-primed MSC CM on endothelial cell adhesion and proliferation, but did not reduce MSC-induced endothelial tubule formation. Primed MSC CM also promoted neuronal cell adhesion, which was reduced following exposure to CSPG. There were no marked differences in neurite outgrowth in MSC CM from CSPG primed MSC cultures versus control conditions, although non-primed MSC CM from the same donors was found to significantly enhance neurite outgrowth. Taken together, these studies demonstrate that MSC are resilient to CSPG exposure, but that there is a marked effect of CSPG on their paracrine regenerative activity. The findings increase our understanding of how the wound microenvironment after SCI can mitigate the beneficial effects of MSC transplantation.

Application of gamma irradiation knowledge in tissue sterilisation: inactivation of malaria parasite.

Malaria is one of the exclusion criteria used in selecting tissue donors and the absence of this information can lead to rejection of tissues for transplant. The studies on the malaria parasite have been confined to low dose attenuation of parasites in blood for transfusion purposes. There is no published information relating to the inactivation of malaria parasites with irradiation for the sterilisation of tissues. A dose-surviving parasite population following radiation was replotted using D0 value from a published paper whereby D10 value of 41 Gy was obtained. Calculation of sterilisation dose for achieving SAL 10-6 of malaria parasites demonstrated the effectiveness of the sterilisation dose of 25 kGy being used in tissue banking.

An international survey of tissue banking: a preliminary report.

International Atomic Energy Agency is currently carrying out a survey on the tissue banking activities from the tissue banking organisations worldwide. The purpose of the survey is to establish regional and global overview of the current tissue banking activities and practices in order to assist the International Atomic Energy Agency to provide further guidance about the use of radiation technology to sterilise tissues as well as to promote better collaboration between the regional tissue banking associations. The survey is an on-going exercise and the preliminary data is presented here. This exercise is not completed and the authors urge the remaining tissue banks to participate in the survey.