Comprehensive Proteomic Analysis of the Differential Expression of 83 Proteins Following Intracortical Microelectrode Implantation.

Intracortical microelectrodes (IMEs) are devices designed to be implanted into the cerebral cortex for various neuroscience and neuro-engineering applications. A critical feature of these devices is their ability to detect neural activity from individual neurons. Currently, IMEs are limited by chronic failure, largely considered to be caused by the prolonged neuroinflammatory response to the implanted devices. Over the decades, characterization of the neuroinflammatory response has grown in sophistication, with the most recent advances including advanced genomics and spatially resolved transcriptomics. While gene expression studies increase our broad understanding of the relationship between IMEs and cortical tissue, advanced proteomic techniques have not been reported. Proteomic evaluation is necessary to describe the diverse changes in protein expression specific to neuroinflammation, neurodegeneration, or tissue and cellular viability, which could lead to the development of more targeted intervention strategies designed to improve IME function. In this study, we have characterized the expression of 83 proteins within 180 µm of the IME implant site at 4-, 8-, and 16-weeks post-implantation. We identified potential targets for immunotherapies, as well as key pathways and functions that contribute to neuronal dieback around the IME implant.

Depletion of complement factor 3 delays the neuroinflammatory response to intracortical microelectrodes.

The neuroinflammatory response to intracortical microelectrodes (IMEs) used with brain-machine interfacing (BMI) applications is regarded as the primary contributor to poor chronic performance. Recent developments in high-plex gene expression technologies have allowed for an evolution in the investigation of individual proteins or genes to be able to identify specific pathways of upregulated genes that may contribute to the neuroinflammatory response. Several key pathways that are upregulated following IME implantation are involved with the complement system. The complement system is part of the innate immune system involved in recognizing and eliminating pathogens - a significant contributor to the foreign body response against biomaterials. Specifically, we have identified Complement 3 (C3) as a gene of interest because it is the intersection of several key complement pathways. In this study, we investigated the role of C3 in the IME inflammatory response by comparing the neuroinflammatory gene expression at the microelectrode implant site between C3 knockout (C3-/-) and wild-type (WT) mice. We have found that, like in WT mice, implantation of intracortical microelectrodes in C3-/- mice yields a dramatic increase in the neuroinflammatory gene expression at all post-surgery time points investigated. However, compared to WT mice, C3 depletion showed reduced expression of many neuroinflammatory genes pre-surgery and 4 weeks post-surgery. Conversely, depletion of C3 increased the expression of many neuroinflammatory genes at 8 weeks and 16 weeks post-surgery, compared to WT mice. Our results suggest that C3 depletion may be a promising therapeutic target for acute, but not chronic, relief of the neuroinflammatory response to IME implantation. Additional compensatory targets may also be required for comprehensive long-term reduction of the neuroinflammatory response for improved intracortical microelectrode performance.

Differential expression of genes involved in the chronic response to intracortical microelectrodes.

Brain-Machine Interface systems (BMIs) are clinically valuable devices that can provide functional restoration for patients with spinal cord injury or improved integration for patients requiring prostheses. Intracortical microelectrodes can record neuronal action potentials at a resolution necessary for precisely controlling BMIs. However, intracortical microelectrodes have a demonstrated history of progressive decline in the recording performance with time, inhibiting their usefulness. One major contributor to decreased performance is the neuroinflammatory response to the implanted microelectrodes. The neuroinflammatory response can lead to neurodegeneration and the formation of a glial scar at the implant site. Historically, histological imaging of relatively few known cellular and protein markers has characterized the neuroinflammatory response to implanted microelectrode arrays. However, neuroinflammation requires many molecular players to coordinate the response - meaning traditional methods could result in an incomplete understanding. Taking advantage of recent advancements in tools to characterize the relative or absolute DNA/RNA expression levels, a few groups have begun to explore gene expression at the microelectrode-tissue interface. We have utilized a custom panel of ∼813 neuroinflammatory-specific genes developed with NanoString for bulk tissue analysis at the microelectrode-tissue interface. Our previous studies characterized the acute innate immune response to intracortical microelectrodes. Here we investigated the gene expression at the microelectrode-tissue interface in wild-type (WT) mice chronically implanted with nonfunctioning probes. We found 28 differentially expressed genes at chronic time points (4WK, 8WK, and 16WK), many in the complement and extracellular matrix system. Further, the expression levels were relatively stable over time. Genes identified here represent chronic molecular players at the microelectrode implant sites and potential therapeutic targets for the long-term integration of microelectrodes. STATEMENT OF SIGNIFICANCE: Intracortical microelectrodes can record neuronal action potentials at a resolution necessary for the precise control of Brain-Machine Interface systems (BMIs). However, intracortical microelectrodes have a demonstrated history of progressive declines in the recording performance with time, inhibiting their usefulness. One major contributor to the decline in these devices is the neuroinflammatory response against the implanted microelectrodes. Historically, neuroinflammation to implanted microelectrode arrays has been characterized by histological imaging of relatively few known cellular and protein markers. Few studies have begun to develop a more in-depth understanding of the molecular pathways facilitating device-mediated neuroinflammation. Here, we are among the first to identify genetic pathways that could represent targets to improve the host response to intracortical microelectrodes, and ultimately device performance.

Neuroinflammatory Gene Expression Analysis Reveals Pathways of Interest as Potential Targets to Improve the Recording Performance of Intracortical Microelectrodes.

Intracortical microelectrodes are a critical component of brain-machine interface (BMI) systems. The recording performance of intracortical microelectrodes used for both basic neuroscience research and clinical applications of BMIs decreases over time, limiting the utility of the devices. The neuroinflammatory response to the microelectrode has been identified as a significant contributing factor to its performance. Traditionally, pathological assessment has been limited to a dozen or so known neuroinflammatory proteins, and only a few groups have begun to explore changes in gene expression following microelectrode implantation. Our initial characterization of gene expression profiles of the neuroinflammatory response to mice implanted with non-functional intracortical probes revealed many upregulated genes that could inform future therapeutic targets. Emphasis was placed on the most significant gene expression changes and genes involved in multiple innate immune sets, including Cd14, C3, Itgam, and Irak4. In previous studies, inhibition of Cluster of Differentiation 14 (Cd14) improved microelectrode performance for up to two weeks after electrode implantation, suggesting CD14 can be explored as a potential therapeutic target. However, all measures of improvements in signal quality and electrode performance lost statistical significance after two weeks. Therefore, the current study investigated the expression of genes in the neuroinflammatory pathway at the tissue-microelectrode interface in Cd14-/- mice to understand better how Cd14 inhibition was connected to temporary improvements in recording quality over the initial 2-weeks post-surgery, allowing for the identification of potential co-therapeutic targets that may work synergistically with or after CD14 inhibition to improve microelectrode performance.

Understanding the Effects of Both CD14-Mediated Innate Immunity and Device/Tissue Mechanical Mismatch in the Neuroinflammatory Response to Intracortical Microelectrodes.

Intracortical microelectrodes record neuronal activity of individual neurons within the brain, which can be used to bridge communication between the biological system and computer hardware for both research and rehabilitation purposes. However, long-term consistent neural recordings are difficult to achieve, in large part due to the neuroinflammatory tissue response to the microelectrodes. Prior studies have identified many factors that may contribute to the neuroinflammatory response to intracortical microelectrodes. Unfortunately, each proposed mechanism for the prolonged neuroinflammatory response has been investigated independently, while it is clear that mechanisms can overlap and be difficult to isolate. Therefore, we aimed to determine whether the dual targeting of the innate immune response by inhibiting innate immunity pathways associated with cluster of differentiation 14 (CD14), and the mechanical mismatch could improve the neuroinflammatory response to intracortical microelectrodes. A thiol-ene probe that softens on contact with the physiological environment was used to reduce mechanical mismatch. The thiol-ene probe was both softer and larger in size than the uncoated silicon control probe. Cd14-/- mice were used to completely inhibit contribution of CD14 to the neuroinflammatory response. Contrary to the initial hypothesis, dual targeting worsened the neuroinflammatory response to intracortical probes. Therefore, probe material and CD14 deficiency were independently assessed for their effect on inflammation and neuronal density by implanting each microelectrode type in both wild-type control and Cd14-/- mice. Histology results show that 2 weeks after implantation, targeting CD14 results in higher neuronal density and decreased glial scar around the probe, whereas the thiol-ene probe results in more microglia/macrophage activation and greater blood-brain barrier (BBB) disruption around the probe. Chronic histology demonstrate no differences in the inflammatory response at 16 weeks. Over acute time points, results also suggest immunomodulatory approaches such as targeting CD14 can be utilized to decrease inflammation to intracortical microelectrodes. The results obtained in the current study highlight the importance of not only probe material, but probe size, in regard to neuroinflammation.

Alternative splicing of CHEK2 and codeletion with NF2 promote chromosomal instability in meningioma.

Mutations of the NF2 gene on chromosome 22q are thought to initiate tumorigenesis in nearly 50% of meningiomas, and 22q deletion is the earliest and most frequent large-scale chromosomal abnormality observed in these tumors. In aggressive meningiomas, 22q deletions are generally accompanied by the presence of large-scale segmental abnormalities involving other chromosomes, but the reasons for this association are unknown. We find that large-scale chromosomal alterations accumulate during meningioma progression primarily in tumors harboring 22q deletions, suggesting 22q-associated chromosomal instability. Here we show frequent codeletion of the DNA repair and tumor suppressor gene, CHEK2, in combination with NF2 on chromosome 22q in a majority of aggressive meningiomas. In addition, tumor-specific splicing of CHEK2 in meningioma leads to decreased functional Chk2 protein expression. We show that enforced Chk2 knockdown in meningioma cells decreases DNA repair. Furthermore, Chk2 depletion increases centrosome amplification, thereby promoting chromosomal instability. Taken together, these data indicate that alternative splicing and frequent codeletion of CHEK2 and NF2 contribute to the genomic instability and associated development of aggressive biologic behavior in meningiomas.