Staying in the game: a pilot study examining the efficacy of protective headgear in an animal model of mild traumatic brain injury (mTBI).

PRIMARY OBJECTIVE: Rugby is one of the few contact sports that do not mandate protective headgear, possibly because studies have shown poor efficacy for protection related to concussion pathology with existing headguards. RESEARCH DESIGN: Following innovative material technology utilization to produce headgear believed to have protective capabilities, this study examined the effects of a soft-shell headgear constructed from a novel viscoelastic material, on both behaviour and serum biomarkers after high and average impact force mild traumatic brain injuries (mTBI). METHODS AND PROCEDURES: Seventy-five male Sprague Dawley rats were divided into five groups: control, average - 37G impact, with and without headgear, and high - 106G impact, with and without headgear. Rats were sacrificed at 3 or 48 hours and serum samples were analyzed for levels of TNF-α, NEF-L, and GFAP. Animals sacrificed at 48 hours also underwent testing for balance and motor coordination, and exploratory/locomotor behaviour. MAIN OUTCOMES AND RESULTS: The novel headgear offered significant protection against mTBI symptomology and biomarkers in the group that experienced an average impact force, but only moderated protection for the animals in the high impact group. CONCLUSIONS: This innovative headgear may prevent some of the negative sequel associated with concussion pathology.

Mesenchymal stem cells and a vitamin D receptor agonist additively suppress T helper 17 cells and the related inflammatory response in the kidney.

Mesenchymal stem cells (MSCs) suppress T helper (Th)17 cell differentiation and are being clinically pursued for conditions associated with aberrant Th17 responses. Whether such immunomodulatory effects are enhanced by coadministration of MSCs with other agents is not well known. In the present study, individual and combined effects of MSCs and the vitamin D receptor (VDR) agonist paricalcitol on Th17 induction were investigated in vitro and in a mouse model of sterile kidney inflammation (unilateral ureteral obstruction). In vitro, MSCs and paricalcitol additively suppressed Th17 differentiation, although only MSCs suppressed expression of Th17-associated transcriptions factors. Combined administration of MSCs and paricalcitol resulted in an early (day 3) reduction of intrarenal CD4(+) and CD8(+) T cells, CD11b(+)/lymphocyte antigen 6G(+) neutrophils, and inflammatory (lymphocyte antigen 6C(hi)) monocytes as well as reduced transcript for IL-17 compared with untreated animals. Later (day 8), obstructed kidneys of MSC/paricalcitol double-treated mice, but not mice treated with either intervention alone, had reduced tubular injury and interstitial fibrosis as well as lower numbers of neutrophils and inflammatory monocytes and an increase in the ratio between M2 (CD206(+)) and M1 (CD206(-)) macrophages compared with control mice. Adjunctive therapy with VDR agonists may enhance the immunosuppressive properties of MSCs in the setting of pathogenic Th17-type immune responses and related inflammatory responses.

Modelling iron mismanagement in neurodegenerative disease in vitro: paradigms, pitfalls, possibilities & practical considerations.

Although aberrant metabolism and deposition of iron has been associated with aging and neurodegeneration, the contribution of iron to neuropathology is unclear. Well-designed model systems that are suited to studying the putative pathological effect of iron are likely to be essential if such unresolved details are to be clarified. In this review, we have evaluated the utility and effectiveness of the reductionist in vitro platform to study the molecular mechanisms putatively underlying iron perturbations of neurodegenerative disease. The expression and function of iron metabolism proteins in glia and neurons and the extent to which this iron regulatory system is replicated in in vitro models has been comprehensively described, followed by an appraisal of the inherent suitability of different in vitro and ex vivo models that have been, or might be, used for iron loading. Next, we have identified and critiqued the relevant experimental parameters that have been used in in vitro iron loading experiments, including the choice of iron reagent, relevant iron loading concentrations and supplementation with serum or ascorbate, and propose optimal iron loading conditions. Finally, we have provided a synthesis of the differential iron accumulation and toxicity in glia and neurons from reported iron loading paradigms. In summary, this review has amalgamated the findings and paradigms of the published reports modelling iron loading in monocultures, discussed the limitations and discrepancies of such work to critically propose a robust, relevant and reliable model of iron loading to be used for future investigations.

Differential activation of ER stress pathways in myelinating cerebellar tracts.

Myelin production during brain development requires an increase in membrane protein and lipid production in oligodendrocytes and this primarily occurs in the endoplasmic reticulum (ER), an organelle which initiates the Unfolded Protein Response (UPR) when under stress. We hypothesise that the UPR is activated in white matter tracts during myelination in order to expand the ER capacity of oligodendrocytes. Using early and late stage markers, critical myelination time points were identified by immunohistochemistry in developing rat cerebellum. These were correlated to peaks in ER stress signalling by staining for activated UPR transducers (pIRE1, ATF6 and pPERK) and associated downstream molecules (peIF2α, PDI, GRP78, GRP94, CHOP and calreticulin) in cerebellar tracts III and IV. Gene expression in developing cerebellum was assessed by qPCR. Actively myelinating tracts were shown to have differential expression of pIRE1, PERK and ATF6 as well as UPR targets GRP94, GRP78 and PDI. Activated pIRE1-positive cells were widespread at P14 and P17 and at significantly higher numbers during myelination than at other stages. Nuclear-localised ATF6 (indicative of the active transcription factor) peaked at P10, concurrent with the initial phase of myelination. The percentage of cells positive for pPERK was less than 1% at postnatal ages but increased significantly in adult tissue. The downstream targets GRP78, GRP94 and PDI were significantly up-regulated at P17 compared to P7 and remained significantly elevated in adults. The majority of cells positive for these markers and ATF6 were oligodendrocytes as confirmed by dual-labelling. Although gene expression in the cerebellum for GRP78, GRP94 and PDI did not change significantly over time, ATF6 and XBP1s both showed significant fold changes between early and late timepoints. This data helps promote understanding of events occurring during developmental myelination and may have implications for the development of reparative treatments in diseases such as multiple sclerosis.

Comparison of viral and nonviral vectors for gene transfer to human endothelial progenitor cells.

BACKGROUND/AIMS: The ability of endothelial progenitor cells (EPCs) to home to sites of neoangiogenesis makes them attractive candidates for use in the field of gene therapy. The efficacy of this approach depends on the efficiency of the vector used for transgene delivery. METHODS/RESULTS: In this study, we have compared the efficiency of adenovirus, five serotypes of AAV2, VSVG-pseudotyped lentivirus, and nonviral plasmid/liposome DNA vectors to deliver the green fluorescence protein reporter gene to human early EPCs to determine efficacy and vector-related cell toxicity. Adenovirus proved most effective with efficiencies of up to 80% with low levels of cell death. Lower levels of expression were seen with other vectors. Electroporation proved unsuitable at the parameters tested. We have also identified at least two distinct subpopulations that exist in the heterogeneous parent EPC culture, one of which is amenable to transduction with adenovirus and one that is not. In addition, adenoviral transduction did not disrupt the ability of the cells to incorporate into endothelial structures in vitro. CONCLUSION: We have found adenovirus to be the most efficient of the vector systems tested for gene delivery to EPCs, an effect that is mediated almost entirely by one of two identified subpopulations.

A non-apoptotic role for caspase-9 in muscle differentiation.

Caspases, a family of cysteine proteases most often investigated for their roles in apoptosis, have also been demonstrated to have functions that are vital for the efficient execution of cell differentiation. One such role that has been described is the requirement of caspase-3 for the differentiation of skeletal myoblasts into myotubes but, as yet, the mechanism leading to caspase-3 activation in this case remains elusive. Here, we demonstrate that caspase-9, an initiator caspase in the mitochondrial death pathway, is responsible for the activation of caspase-3 in differentiating C2C12 cells. Reduction of caspase-9 levels, using an shRNA construct, prevented caspase-3 activation and inhibited myoblast fusion. Myosin-heavy-chain expression, which accompanies myoblastic differentiation, was not caspase-dependent. Overexpression of Bcl-xL, a protein that inhibits caspase-9 activation, had the same effect on muscle differentiation as knockdown of caspase-9. These data suggest that the mitochondrial pathway is required for differentiation; however, the release of cytochrome c or Smac (Diablo) could not be detected, raising the possibility of a novel mechanism of caspase-9 activation during muscle differentiation.