Autologous bone marrow mononuclear cells to treat severe traumatic brain injury in children.

Autologous bone marrow mononuclear cells (BMMNCs) infused after severe traumatic brain injury have shown promise for treating the injury. We evaluated their impact in children, particularly their hypothesized ability to preserve the blood-brain barrier and diminish neuroinflammation, leading to structural CNS preservation with improved outcomes. We performed a randomized, double-blind, placebo-sham-controlled Bayesian dose-escalation clinical trial at two children's hospitals in Houston, TX and Phoenix, AZ, USA (NCT01851083). Patients 5-17 years of age with severe traumatic brain injury (Glasgow Coma Scale score ≤ 8) were randomized to BMMNC or placebo (3:2). Bone marrow harvest, cell isolation and infusion were completed by 48 h post-injury. A Bayesian continuous reassessment method was used with cohorts of size 3 in the BMMNC group to choose the safest between two doses. Primary end points were quantitative brain volumes using MRI and microstructural integrity of the corpus callosum (diffusivity and oedema measurements) at 6 months and 12 months. Long-term functional outcomes and ventilator days, intracranial pressure monitoring days, intensive care unit days and therapeutic intensity measures were compared between groups. Forty-seven patients were randomized, with 37 completing 1-year follow-up (23 BMMNC, 14 placebo). BMMNC treatment was associated with an almost 3-day (23%) reduction in ventilator days, 1-day (16%) reduction in intracranial pressure monitoring days and 3-day (14%) reduction in intensive care unit (ICU) days. White matter volume at 1 year in the BMMNC group was significantly preserved compared to placebo [decrease of 19 891 versus 40 491, respectively; mean difference of -20 600, 95% confidence interval (CI): -35 868 to -5332; P = 0.01], and the number of corpus callosum streamlines was reduced more in placebo than BMMNC, supporting evidence of preserved corpus callosum connectivity in the treated groups (-431 streamlines placebo versus -37 streamlines BMMNC; mean difference of -394, 95% CI: -803 to 15; P = 0.055), but this did not reach statistical significance due to high variability. We conclude that autologous BMMNC infusion in children within 48 h after severe traumatic brain injury is safe and feasible. Our data show that BMMNC infusion led to: (i) shorter intensive care duration and decreased ICU intensity; (ii) white matter structural preservation; and (iii) enhanced corpus callosum connectivity and improved microstructural metrics.

Autologous cellular therapy for cerebral palsy: a randomized, crossover trial.

We examined an autologous mononuclear-cell-therapy-based approach to treat cerebral palsy using autologous umbilical cord blood or bone-marrow-derived mononuclear cells. The primary objective was to determine if autologous cells are safe to administer in children with cerebral palsy. The secondary objectives were to determine if there was improvement in motor function of patients 12 months after infusion using the Gross Motor Function Measure and to evaluate impact of treatment on corticospinal tract microstructure as determined by radial diffusivity measurement. This Phase 1/2a trial was a randomized, blinded, placebo-controlled, crossover study in children aged 2-10 years of age with cerebral palsy enrolled between November 2013 and November 2016. Participants were randomized to 2:1 treatment:placebo. Treatment was either autologous bone-marrow-derived mononuclear cells or autologous umbilical cord blood. All participants who enrolled and completed their baseline visit planned to return for follow-up visits at 6 months, 12 months and 24 months after the baseline visit. At the 12-month post-treatment visit, participants who originally received the placebo received either bone-marrow-derived mononuclear cell or umbilical cord blood treatment. Twenty participants were included; 7 initially randomized to placebo, and 13 randomized to treatment. Five participants randomized to placebo received bone-marrow-derived mononuclear cells, and 2 received umbilical cord blood at the 12-month visit. None of the participants experienced adverse events related to the stem cell infusion. Cell infusion at the doses used in our study did not dramatically alter motor function. We observed concordant bilateral changes in radial diffusivity in 10 of 15 cases where each corticospinal tract could be reconstructed in each hemisphere. In 60% of these cases (6/10), concordant decreases in bilateral corticospinal tract radial diffusivity occurred post-treatment. In addition, 100% of unilateral corticospinal tract cases (3/3) exhibited decreased corticospinal tract radial diffusivity post-treatment. In our discordant cases (n = 5), directionality of changes in corticospinal tract radial diffusivity appeared to coincide with handedness. There was a significant improvement in corticospinal tract radial diffusivity that appears related to handedness. Connectivity strength increased in either or both pathways (corticio-striatal and thalamo-cortical) in each participant at 12 months post-treatment. These data suggest that both stem cell infusions are safe. There may be an improvement in myelination in some groups of patients that correlate with small improvements in the Gross Motor Function Measure scales. A larger autologous cord blood trial is impractical at current rates of blood banking. Either increased private banking or matched units would be required to perform a larger-scale trial.

PET Imaging of Peripheral Benzodiazepine Receptor Standard Uptake Value Increases After Controlled Cortical Impact, a Rodent Model of Traumatic Brain Injury.

Traumatic brain injury (TBI) is a chronic, life threatening injury for which few effective interventions are available. Evidence in animal models suggests un-checked immune activation may contribute to the pathophysiology. Changes in regional density of active brain microglia can be quantified in vivo with positron emission topography (PET) with the relatively selective radiotracer, peripheral benzodiazepine receptor 28 (11 C-PBR28). Phenotypic assessment (activated vs resting) can subsequently be assessed (ex vivo) using morphological techniques. To elucidate the mechanistic contribution of immune cells in due to TBI, we employed a hybrid approach involving both in vivo (11 C-PBR28 PET) and ex vivo (morphology) to elucidate the role of immune cells in a controlled cortical impact (CCI), a rodent model for TBI. Density of activated brain microglia/macrophages was quantified 120 hours after injury using the standardized uptake value (SUV) approach. Ex vivo morphological analysis from specific brain regions using IBA-1 antibodies differentiated ramified (resting) from amoeboid (activated) immune cells. Additional immunostaining of PBRs facilitated co-localization of PBRs with IBA-1 staining to further validate PET data. Injured animals displayed greater PBR28suv when compared to sham animals. Immunohistochemistry demonstrated elevated density of amoeboid microglia/macrophages in the ipsilateral dentate gyrus, corpus callosum, thalami and injury penumbra of injured animals compared to sham animals. PBR co-stained with amoeboid microglia/macrophages in the injury penumbra and not with astrocytes. These data suggest the technologies evaluated may serve as bio-signatures of neuroinflammation following severe brain injury in small animals, potentially enabling in vivo tracking of neuroinflammation following TBI and cellular-based therapies.

Lack of Racial, Ethnic, and Sex Disparities in Ischemic Stroke Care Metrics within a Tele-Stroke Network.

INTRODUCTION: Differences in access to stroke care and compliance with standard of care stroke management among patients of varying racial and ethnic backgrounds and sex are well-characterized. However, little is known on the impact of telestroke in addressing disparities in acute ischemic stroke care. METHODS: We conducted a retrospective review of acute ischemic stroke patients evaluated over our 17-hospital telestroke network in Texas from 2015-2018. Patients were described as Non-Hispanic White (NHW) male or female, Non-Hispanic Black (NHB) male or female, or Hispanic (HIS) male or female. We compared frequency of tPA and mechanical thrombectomy (MT) utilization, door-to-consultation times, door-to-tPA times, and time-to-transfer for patients who went on to MT evaluation at the hub after having been screened for suspected large vessel occlusion at the spoke. RESULTS: Among 3873 patients (including 1146 NHW male (30%) and 1134 NHW female (29%), 405 NHB male (10%) and 491 NHB female (13%), and 358 HIS male (9%) and 339 HIS female (9%) patients), we did not find any differences in door-to consultation time, door-to-tPA time, time-to-transfer, frequency of tPA administration, or incidence of MT utilization. CONCLUSION: We did not find racial, ethnic, and sex disparities in ischemic stroke care metrics within our telestroke network. In order to fully understand how telestroke alleviates disparities in stroke care, collaboration among networks is needed to formulate a multicenter telestroke database similar to the Get-With-The Guidelines.

Call to Action: SARS-CoV-2 and CerebrovAscular DisordErs (CASCADE).

BACKGROUND AND PURPOSE: The novel severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2), now named coronavirus disease 2019 (COVID-19), may change the risk of stroke through an enhanced systemic inflammatory response, hypercoagulable state, and endothelial damage in the cerebrovascular system. Moreover, due to the current pandemic, some countries have prioritized health resources towards COVID-19 management, making it more challenging to appropriately care for other potentially disabling and fatal diseases such as stroke. The aim of this study is to identify and describe changes in stroke epidemiological trends before, during, and after the COVID-19 pandemic. METHODS: This is an international, multicenter, hospital-based study on stroke incidence and outcomes during the COVID-19 pandemic. We will describe patterns in stroke management, stroke hospitalization rate, and stroke severity, subtype (ischemic/hemorrhagic), and outcomes (including in-hospital mortality) in 2020 during COVID-19 pandemic, comparing them with the corresponding data from 2018 and 2019, and subsequently 2021. We will also use an interrupted time series (ITS) analysis to assess the change in stroke hospitalization rates before, during, and after COVID-19, in each participating center. CONCLUSION: The proposed study will potentially enable us to better understand the changes in stroke care protocols, differential hospitalization rate, and severity of stroke, as it pertains to the COVID-19 pandemic. Ultimately, this will help guide clinical-based policies surrounding COVID-19 and other similar global pandemics to ensure that management of cerebrovascular comorbidity is appropriately prioritized during the global crisis. It will also guide public health guidelines for at-risk populations to reduce risks of complications from such comorbidities.

Clinical parameters affecting multipotent adult progenitor cells in vitro.

BACKGROUND: Human multipotent adult progenitor cells (MAPC®) are an emerging therapy for traumatic brain injury (TBI); however, clinically translating a therapy involves overcoming many factors in vivo which are not present in pre-clinical testing. In this study we examined clinical parameters in vitro that may impact cell therapy efficacy. METHODS: MAPC were infused through varying gauged needles and catheters with and without chlorhexidine, and their viability tested with trypan blue exclusion. MAPC were co-cultured with phenytoin and celecoxib at relevant clinical concentrations for 1 h and 24 h. Anti-inflammatory potency was tested using a stimulated rat splenocyte co-culture and ELISA for TNF-α production. MAPC were cultured under different osmolar concentrations and stained with propidium iodide for viability. Anti-inflammatory potency was tested by co-culture of MAPC with naïve lymphocytes activated by CD3/CD28 beads, and Click-iT® Plus EdU was used to quantify proliferation by flow cytometry. RESULTS: The mean viability of the MAPC infused via needles was 95 ± 1%; no difference was seen with varying flow rate, but viability was notably reduced by chlorhexidine. MAPC function was not impaired by co-culture with phenytoin, celecoxib, or combination with both. Co-culture with phenytoin showed a decrease in TNF-α production as compared to the MAPC control. MAPC cultured at varying osmolar concentrations all had viabilities greater than 90% with no statistical difference between them. Co-culture of MAPC with CD3/CD28 activated PBMCs showed a significant reduction in proliferation as measured by EdU uptake. DISCUSSION: Needle diameter, phenytoin, celecoxib, and a relevant range of osmolarities do not impair MAPC viability or anti-inflammatory potency in vitro.

Pre-injury monocyte/macrophage depletion results in increased blood-brain barrier permeability after traumatic brain injury.

Traumatic brain injury (TBI) effects both the brain and the immune system. Circulating monocytes/macrophages (Mo /Ma ) after a TBI may play an important role in preserving the blood-brain barrier (BBB), reducing brain edema, and interacting with resident microglia. To elucidate the role of circulating Mo /Ma , we utilized a monocyte/macrophage depletion model in response to TBI in male rats. Clodronate liposomes (CL) were used to deplete circulating Mo /Ma . A controlled cortical impact (CCI) injury model was used to create a TBI. All animals received either CL or PBS liposomes (PL), 48 and 24 hr prior to the procedure, and were sacrificed 72 hr post-injury for analysis of BBB permeability, brain edema, whole blood (Mo /Ma and granulocytes), and/or microglial analysis. Animals undergoing Mo /Ma depletion with CL prior to CCI (CCI-CL) were found to have increased BBB permeability when compared to non-depleted CCI (CCI-PL) animals. At 72 hr following injury, Sham-CL maintained on average an 82% reduction in the whole blood monocytes when compared to Sham-PL (p < 0.001). Monocytes in the whole blood remained significantly lower in CCI-CL animals when compared to CCI-PL (p < 0.001). The number of granulocytes in the whole blood of CCI-CL animals was higher at 3 days when compared to CCI-PL (p < 0.022). Surprisingly, the depletion of Mo /Ma did not affect brain edema. However, the depletion of Mo /Ma did result in a significant decrease in microglia (CCI-CL vs. CCI-PL, p < 0.012). In conclusion, an intact Mo /Ma population is required to repair BBB integrity and microglial response following injury.

Therapeutic time window of multipotent adult progenitor therapy after traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) is a major cause of death and disability. TBI results in a prolonged secondary central neuro-inflammatory response. Previously, we have demonstrated that multiple doses (2 and 24 h after TBI) of multipotent adult progenitor cells (MAPC) delivered intravenously preserve the blood-brain barrier (BBB), improve spatial learning, and decrease activated microglia/macrophages in the dentate gyrus of the hippocampus. In order to determine if there is an optimum treatment window to preserve the BBB, improve cognitive behavior, and attenuate the activated microglia/macrophages, we administered MAPC at various clinically relevant intervals. METHODS: We administered two injections intravenously of MAPC treatment at hours 2 and 24 (2/24), 6 and 24 (6/24), 12 and 36 (12/36), or 36 and 72 (36/72) post cortical contusion injury (CCI) at a concentration of 10 million/kg. For BBB experiments, animals that received MAPC at 2/24, 6/24, and 12/36 were euthanized 72 h post injury. The 36/72 treated group was harvested at 96 h post injury. RESULTS: Administration of MAPC resulted in a significant decrease in BBB permeability when administered at 2/24 h after TBI only. For behavior experiments, animals were harvested post behavior paradigm. There was a significant improvement in spatial learning (120 days post injury) when compared to cortical contusion injury (CCI) in groups when MAPC was administered at or before 24 h. In addition, there was a significant decrease in activated microglia/macrophages in the dentate gyrus of hippocampus of the treated group (2/24) only when compared to CCI. CONCLUSIONS: Intravenous injections of MAPC at or before 24 h after CCI resulted in improvement of the BBB, improved cognitive behavior, and attenuated activated microglia/macrophages in the dentate gyrus.

Biomechanical Forces Promote Immune Regulatory Function of Bone Marrow Mesenchymal Stromal Cells.

Mesenchymal stromal cells (MSCs) are believed to mobilize from the bone marrow in response to inflammation and injury, yet the effects of egress into the vasculature on MSC function are largely unknown. Here we show that wall shear stress (WSS) typical of fluid frictional forces present on the vascular lumen stimulates antioxidant and anti-inflammatory mediators, as well as chemokines capable of immune cell recruitment. WSS specifically promotes signaling through NFκB-COX2-prostaglandin E2 (PGE2 ) to suppress tumor necrosis factor-α (TNF-α) production by activated immune cells. Ex vivo conditioning of MSCs by WSS improved therapeutic efficacy in a rat model of traumatic brain injury, as evidenced by decreased apoptotic and M1-type activated microglia in the hippocampus. These results demonstrate that force provides critical cues to MSCs residing at the vascular interface which influence immunomodulatory and paracrine activity, and suggest the potential therapeutic use of force for MSC functional enhancement. Stem Cells 2017;35:1259-1272.

Treatment of Severe Adult Traumatic Brain Injury Using Bone Marrow Mononuclear Cells.

Preclinical studies using bone marrow derived cells to treat traumatic brain injury have demonstrated efficacy in terms of blood-brain barrier preservation, neurogenesis, and functional outcomes. Phase 1 clinical trials using bone marrow mononuclear cells infused intravenously in children with severe traumatic brain injury demonstrated safety and potentially a central nervous system structural preservation treatment effect. This study sought to confirm the safety, logistic feasibility, and potential treatment effect size of structural preservation/inflammatory biomarker mitigation in adults to guide Phase 2 clinical trial design. Adults with severe traumatic brain injury (Glasgow Coma Scale 5-8) and without signs of irreversible brain injury were evaluated for entry into the trial. A dose escalation format was performed in 25 patients: 5 controls, followed 5 patients in each dosing cohort (6, 9, 12 ×106 cells/kg body weight), then 5 more controls. Bone marrow harvest, cell processing to isolate the mononuclear fraction, and re-infusion occurred within 48 hours after injury. Patients were monitored for harvest-related hemodynamic changes, infusional toxicity, and adverse events. Outcome measures included magnetic resonance imaging-based measurements of supratentorial and corpus callosal volumes as well as diffusion tensor imaging-based measurements of fractional anisotropy and mean diffusivity of the corpus callosum and the corticospinal tract at the level of the brainstem at 1 month and 6 months postinjury. Functional and neurocognitive outcomes were measured and correlated with imaging data. Inflammatory cytokine arrays were measured in the plasma pretreatment, posttreatment, and at 1 and 6 month follow-up. There were no serious adverse events. There was a mild pulmonary toxicity of the highest dose that was not clinically significant. Despite the treatment group having greater injury severity, there was structural preservation of critical regions of interest that correlated with functional outcomes. Key inflammatory cytokines were downregulated. Treatment of severe, adult traumatic brain injury using an intravenously delivered autologous bone marrow mononuclear cell infusion is safe and logistically feasible. There appears to be a treatment signal as evidenced by central nervous system structural preservation, consistent with previous pediatric trial data. Inflammatory biomarkers are downregulated after cell infusion. Stem Cells 2016 Video Highlight: https://youtu.be/UiCCPIe-IaQ Stem Cells 2017;35:1065-1079.

Strategies for CNS repair following TBI.

Traumatic brain injury (TBI) imparts a significant health burden in the United States, leaving many patients with chronic deficits. Improvement in clinical outcome following TBI has been hindered by a lack of treatments that have proven successful during phase III trials. Research remains active into a variety of non-pharmacologic, small molecule, endocrine and cell based therapies. Of particular focus in this review are the recent therapeutic avenues that have undergone clinical investigation and the mechanisms by which cell therapies may mediate recovery in severe TBI. Preclinical data show cell therapies to provide benefit when administered systemically or with transplantation to the site of injury. Increasingly, studies have shown that these cells are able to attenuate the inflammatory response to injury and stimulate production of neurotrophic factors. In animal models, beneficial effects on blood-brain barrier permeability, neuroprotection and neural repair through enhanced axonal remodeling have been observed. Clinical investigation with cell therapies for TBI remains ongoing.

Pharmacological postconditioning with lactic acid and hydrogen rich saline alleviates myocardial reperfusion injury in rats.

This study investigated whether pharmacological postconditioning with lactic acid and hydrogen rich saline can provide benefits similar to that of mechanical postconditioning. To our knowledge, this is the first therapeutic study to investigate the co-administration of lactic acid and hydrogen. SD rats were randomly divided into 6 groups: Sham, R/I, M-Post, Lac, Hyd, and Lac + Hyd. The left coronary artery was occluded for 45 min. Blood was withdrawn from the right atrium to measure pH. The rats were sacrificed at different time points to measure mitochondrial absorbance, infarct size, serum markers and apoptotic index. Rats in Lac + Hyd group had similar blood pH and ROS levels when compared to the M-Post group. Additionally, the infarct area was reduced to the same extent in Lac + Hyd and M-Post groups with a similar trends observed for serum markers of myocardial injury and apoptotic index. Although the level of P-ERK in Lac + Hyd group was lower, P-p38/JNK, TNFα, Caspase-8, mitochondrial absorbance and Cyt-c were all similar in Lac + Hyd and M-Post groups. The Lac and Hyd groups were able to partially mimic this protective role. These data suggested that pharmacological postconditioning with lactic acid and hydrogen rich saline nearly replicates the benefits of mechanical postconditioning.