In Vivo Imaging to Characterize Dynamic Tissue Responses after Neural Electrode Implantation

Implantable neural electrodes are promising technologies to restore motor, sensory, and cognitive function in many neural pathologies through brain-computer interfacing (BCI). Many BCI applications require electrode implantation within neural tissue to resolve and/or modulate the physiological activity of individual neurons via electrical recording and stimulation. This invasive implantation leads to acute and long-term deterioration of both the electrode device as well as the neurons surrounding the device. Ultimately, damage to the electrode and neural tissue results in electrode recording failure within the first years after implantation. Many strategies to improve BCI longevity focus on mitigating tissue damage through improving neuronal survival or reducing inflammatory activity around implants. Despite incremental improvements, electrode failure persists as an obstacle to wide-spread clinical deployment of BCIs. This can be partly attributed to an incomplete understanding of the biological correlates of recording performance. These correlates have largely been identified through post-mortem histological staining, which cannot capture dynamic changes in cellular physiology and morphology. In the following dissertation, we use longitudinal two-photon in vivo imaging to quantify how neurons, microglia, and meningeal immune cells are affected by an intracortical electrode during and after implantation in mouse cortex. We go beyond conventional histological techniques to show the time-course of neuronal injury and microglial recruitment after implantation. Neuronal injury occurs instantaneously, with prolonged, high calcium levels evident in neurons within 100 µm of implants. Microglial activation occurs within minutes of implantation and subsequent microglial encapsulation of electrodes can be modulated by bioactive surface coatings. Within the first day post-implant, there is high trafficking of peripheral immune cells through venules at the surface of the brain as well as along the electrode’s shank at the surface of the brain. Over the next month, calcium activity in neurons increases while the collagenous meningeal tissues at the surface of the brain thicken. We further show that meningeal thickening can have profound implications for devices implanted into non-human primates as well. In sum, these results define new potential therapeutic targets and windows that could improve the longevity of implantable neural electrodes

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

ROS responsive resveratrol delivery from LDLR peptide conjugated PLA-coated mesoporous silica nanoparticles across the blood–brain barrier

Abstract Background Oxidative stress acts as a trigger in the course of neurodegenerative diseases and neural injuries. An antioxidant-based therapy can be effective to ameliorate the deleterious effects of oxidative stress. Resveratrol (RSV) has been shown to be effective at removing excess reactive oxygen species (ROS) or reactive nitrogen species generation in the central nervous system (CNS), but the delivery of RSV into the brain through systemic administration is inefficient. Here, we have developed a RSV delivery vehicle based on polylactic acid (PLA)-coated mesoporous silica nanoparticles (MSNPs), conjugated with a ligand peptide of low-density lipoprotein receptor (LDLR) to enhance their transcytosis across the blood–brain barrier (BBB). Results Resveratrol was loaded into MSNPs (average diameter 200 nm, pore size 4 nm) at 16 μg/mg (w/w). As a gatekeeper, the PLA coating prevented the RSV burst release, while ROS was shown to trigger the drug release by accelerating PLA degradation. An in vitro BBB model with a co-culture of rat brain microvascular endothelial cells (RBECs) and microglia cells using Transwell chambers was established to assess the RSV delivery across BBB. The conjugation of LDLR ligand peptides markedly enhanced the migration of MSNPs across the RBECs monolayer. RSV could be released and effectively reduce the activation of the microglia cells stimulated by phorbol-myristate-acetate or lipopolysaccharide. Conclusions These ROS responsive LDLR peptides conjugated PLA-coated MSNPs have great potential for oxidative stress therapy in CNS

MOESM1 of ROS responsive resveratrol delivery from LDLR peptide conjugated PLA-coated mesoporous silica nanoparticles across the blood–brain barrier

Additional file 1: Figure S1. Characterization of eluted PLA from MSNPs by NMR. The NMR spectra of PLA (A) and coated PLA with 1:0.25 (B), 1:0.5 (C) and 1:1 (D) mass ratio on MSNPs. PLA coated MNSPs at different ratios were re-dissolved in CDCl3 solvent and the solutions were analyzed by NMR. Figure S2. RSV adsorption profile onto MSNPs from 30 μg/mL RSV solutions. The RSV was dissolved in 50% ethanol/PBS solution and then mixed with MSNPs at room temperature. The mixture was centrifuged at specified time point, and the supernatant was analyzed with spectroscopy at 304 nm to determine the amount of free RSV remaining in solution and the amount of RSV loaded in the MSNPs was determined by subtracting the remaining RSV from the total amount originally present. n = 3. Figure S3. Dynamic Light Scattering measurement displaying the hydrodynamic diameters for uncoated mesoporous silica nanoparticles and PMSNPs in water over 1 h. DLS measurements were performed by Malvern ZS90 Zetasizer. 1:0.25 PMSNPs were suspended at 0.5 mg/ml in water then pipetted into a disposable polystyrene cuvette for analysis. Measurements were taken every 15 min. Table S1. Sample identification codes and preparation conditions