Angiopoietin/Tie2 Axis Regulates the Age-at-Injury Cerebrovascular Response to Traumatic Brain Injury.

Although age-at-injury influences chronic recovery from traumatic brain injury (TBI), the differential effects of age on early outcome remain understudied. Using a male murine model of moderate contusion injury, we investigated the underlying mechanism(s) regulating the distinct response between juvenile and adult TBI. We demonstrate similar biomechanical and physical properties of naive juvenile and adult brains. However, following controlled cortical impact (CCI), juvenile mice displayed reduced cortical lesion formation, cell death, and behavioral deficits at 4 and 14 d. Analysis of high-resolution laser Doppler imaging showed a similar loss of cerebral blood flow (CBF) in the ipsilateral cortex at 3 and 24 h post-CCI, whereas juvenile mice showed enhanced subsequent restoration at 2-4 d compared with adults. These findings correlated with reduced blood-brain barrier (BBB) disruption and increased perilesional vessel density. To address whether an age-dependent endothelial cell (EC) response affects vessel stability and tissue outcome, we magnetically isolated CD31+ ECs from sham and injured cortices and evaluated mRNA expression. Interestingly, we found increased transcripts for BBB stability-related genes and reduced expression of BBB-disrupting genes in juveniles compared with adults. These differences were concomitant with significant changes in miRNA-21-5p and miR-148a levels. Accompanying these findings was robust GFAP immunoreactivity, which was not resolved by day 35. Importantly, pharmacological inhibition of EC-specific Tie2 signaling abolished the juvenile protective effects. These findings shed new mechanistic light on the divergent effects that age plays on acute TBI outcome that are both spatial and temporal dependent.SIGNIFICANCE STATEMENT Although a clear "window of susceptibility" exists in the developing brain that could deter typical developmental trajectories if exposed to trauma, a number of preclinical models have demonstrated evidence of early recovery in younger patients. Our findings further demonstrate acute neuroprotection and improved restoration of cerebral blood flow in juvenile mice subjected to cortical contusion injury compared with adults. We also demonstrate a novel role for endothelial cell-specific Tie2 signaling in this age-related response, which is known to promote barrier stability, is heightened in the injured juvenile vasculature, and may be exploited for therapeutic interventions across the age spectrum following traumatic brain injury.

A comparison of non-integrating reprogramming methods.

Human induced pluripotent stem cells (hiPSCs) are useful in disease modeling and drug discovery, and they promise to provide a new generation of cell-based therapeutics. To date there has been no systematic evaluation of the most widely used techniques for generating integration-free hiPSCs. Here we compare Sendai-viral (SeV), episomal (Epi) and mRNA transfection mRNA methods using a number of criteria. All methods generated high-quality hiPSCs, but significant differences existed in aneuploidy rates, reprogramming efficiency, reliability and workload. We discuss the advantages and shortcomings of each approach, and present and review the results of a survey of a large number of human reprogramming laboratories on their independent experiences and preferences. Our analysis provides a valuable resource to inform the use of specific reprogramming methods for different laboratories and different applications, including clinical translation.