Urokinase-Type Plasminogen Activator Enhances the Neuroprotective Activity of Brain-Derived Neurotrophic Factor in a Model of Intracerebral Hemorrhage.

Brain-derived neurotrophic factor (BDNF) is a classic neuroprotective and pro-regenerative factor in peripheral and central nervous tissue. Its ability to stimulate the restoration of damaged nerve and brain tissue after ischemic stroke and intraventricular hemorrhage has been demonstrated. However, the current concept of regeneration allows us to assert that one factor, even if essential, cannot be the sole contributor to this complex biological process. We have previously shown that urokinase-type plasminogen activator (uPA) complements BDNF activity and stimulates restoration of nervous tissue. Using a model of intracerebral hemorrhage in rats, we investigated the neurotrophic and neuroprotective effect of BDNF combined with uPA. The local simultaneous administration of BDNF and uPA provided effective neuroprotection of brain tissue after intracerebral hemorrhage, promoted survival of experimental animals and their neurological recovery, and decreased lesion volume. The study of cellular mechanisms of the observed neurotrophic effect of BDNF and uPA combination revealed both known mechanisms (neuronal survival and neurite growth) and new ones (microglial activation) that had not been shown for BDNF and uPA. Our findings support the concept of using combinations of biological factors with diverse but complementary mechanisms of action as a promising regenerative approach.

MSC Secretome as a Promising Tool for Neuroprotection and Neuroregeneration in a Model of Intracerebral Hemorrhage.

Multipotent mesenchymal stromal cells (MSCs) are considered to be critical contributors to injured tissue repair and regeneration, and MSC-based therapeutic approaches have been applied to many peripheral and central neurologic disorders. It has been demonstrated that the beneficial effects of MSC are mainly mediated by the components of their secretome. In the current study, we have explored the neuroprotective potential of the MSC secretome in a rat model of intracerebral hemorrhage and shown that a 10-fold concentrated secretome of human MSC and its combination with the brain-derived neurotrophic factor (BDNF) provided a better survival and neurological outcome of rats within 14 days of intracerebral hemorrhage compared to the negative (non-treated) and positive (BDNF) control groups. We found that it was due to the ability of MSC secretome to stimulate neuron survival under conditions of glutamate-induced neurotoxicity. However, the lesion volume did not shrink in these rats, and this also correlated with prominent microglia activation. We hypothesize that this could be caused by the species-specificity of the used MSC secretome and provide evidence to confirm this. Thus, we have found that allogenic rat MSC secretome was more effective than xenogenic human MSC secretome in the rat intracerebral hemorrhage model: it reduced the volume of the lesion and promoted excellent survival and neurological outcome of the treated rats.

Optimization of CRISPR/Cas9 Technology to Knock Out Genes of Interest in Aneuploid Cell Lines.

IMPACT STATEMENT: Cell lines represent convenient models to elucidate specific causes of multigenetic and pluricausal diseases, to test breakthrough regenerative technologies. Most commonly used cell lines surpass diploid cells in their accessibility for delivery of large DNA molecules and genome editing, but the main obstacles for obtaining cell models with knockout-targeted protein from aneuploid cells are multiple allele copies and karyotype/phenotype heterogeneity. In the study, we report an original approach to CRISPR-/Cas9-mediated genome modification of aneuploid cell cultures to create functional cell models, achieving highly efficient targeted protein knockout and avoiding "clonal effect" (for the first time to our knowledge).