A neural tract-inspired conduit for facile, on-demand biopsy of glioblastoma.

Background: A major hurdle to effectively treating glioblastoma (GBM) patients is the lack of longitudinal information about tumor progression, evolution, and treatment response. Methods: In this study, we report the use of a neural tract-inspired conduit containing aligned polymeric nanofibers (i.e., an aligned nanofiber device) to enable on-demand access to GBM tumors in 2 rodent models. Depending on the experiment, a humanized U87MG xenograft and/or F98-GFP+ syngeneic rat tumor model was chosen to test the safety and functionality of the device in providing continuous sampling access to the tumor and its microenvironment. Results: The aligned nanofiber device was safe and provided a high quantity of quality genomic materials suitable for omics analyses and yielded a sufficient number of live cells for in vitro expansion and screening. Transcriptomic and genomic analyses demonstrated continuity between material extracted from the device and that of the primary, intracortical tumor (in the in vivo model). Conclusions: The results establish the potential of this neural tract-inspired, aligned nanofiber device as an on-demand, safe, and minimally invasive access point, thus enabling rapid, high-throughput, longitudinal assessment of tumor and its microenvironment, ultimately leading to more informed clinical treatment strategies.

Enriching neural stem cell and anti-inflammatory glial phenotypes with electrical stimulation after traumatic brain injury in male rats.

Traumatic brain injury (TBI) by an external physical impact results in compromised brain function via undesired neuronal death. Following the injury, resident and peripheral immune cells, astrocytes, and neural stem cells (NSCs) cooperatively contribute to the recovery of the neuronal function after TBI. However, excessive pro-inflammatory responses of immune cells, and the disappearance of endogenous NSCs at the injury site during the acute phase of TBI, can exacerbate TBI progression leading to incomplete healing. Therefore, positive outcomes may depend on early interventions to control the injury-associated cellular milieu in the early phase of injury. Here, we explore electrical stimulation (ES) of the injury site in a rodent model (male Sprague-Dawley rats) to investigate its overall effect on the constituent brain cell phenotype and composition during the acute phase of TBI. Our data showed that a brief ES for 1 hr on day 2 of TBI promoted anti-inflammatory phenotypes of microglia as assessed by CD206 expression and increased the population of NSCs and Nestin+ astrocytes at 7 days post-TBI. Also, ES effectively increased the number of viable neurons when compared to the unstimulated control group. Given the salience of microglia and neural stem cells for healing after TBI, our results strongly support the potential benefit of the therapeutic use of ES during the acute phase of TBI to regulate neuroinflammation and to enhance neuroregeneration.

Electrotaxis of Glioblastoma and Medulloblastoma Spheroidal Aggregates.

Treatment of neuroepithelial cancers remains a daunting clinical challenge, particularly due to an inability to address rampant invasion deep into eloquent regions of the brain. Given the lack of access, and the dispersed nature of brain tumor cells, we explore the possibility of electric fields inducing directed tumor cell migration. In this study we investigate the properties of populations of brain cancer undergoing electrotaxis, a phenomenon whereby cells are directed to migrate under control of an electrical field. We investigate two cell lines for glioblastoma and medulloblastoma (U87mg & DAOY, respectively), plated as spheroidal aggregates in Matrigel-filled electrotaxis channels, and report opposing electrotactic responses. To further understand electrotactic migration of tumor cells, we performed RNA-sequencing for pathway discovery to identify signaling that is differentially affected by the exposure of direct-current electrical fields. Further, using selective pharmacological inhibition assays, focused on the PI3K/mTOR/AKT signaling axis, we validate whether there is a causal relationship to electrotaxis and these mechanisms of action. We find that U87 mg electrotaxis is abolished under pharmacological inhibition of PI3Kγ, mTOR, AKT and ErbB2 signaling, whereas DAOY cell electrotaxis was not attenuated by these or other pathways evaluated.

Immunoengineering nerve repair.

Injuries to the peripheral nervous system are major sources of disability and often result in painful neuropathies or the impairment of muscle movement and/or normal sensations. For gaps smaller than 10 mm in rodents, nearly normal functional recovery can be achieved; for longer gaps, however, there are challenges that have remained insurmountable. The current clinical gold standard used to bridge long, nonhealing nerve gaps, the autologous nerve graft (autograft), has several drawbacks. Despite best efforts, engineering an alternative "nerve bridge" for peripheral nerve repair remains elusive; hence, there is a compelling need to design new approaches that match or exceed the performance of autografts across critically sized nerve gaps. Here an immunomodulatory approach to stimulating nerve repair in a nerve-guidance scaffold was used to explore the regenerative effect of reparative monocyte recruitment. Early modulation of the immune environment at the injury site via fractalkine delivery resulted in a dramatic increase in regeneration as evident from histological and electrophysiological analyses. This study suggests that biasing the infiltrating inflammatory/immune cellular milieu after injury toward a proregenerative population creates a permissive environment for repair. This approach is a shift from the current modes of clinical and laboratory methods for nerve repair, which potentially opens an alternative paradigm to stimulate endogenous peripheral nerve repair.

Engineering challenges for brain tumor immunotherapy.

Malignant brain tumors represent one of the most devastating forms of cancer with abject survival rates that have not changed in the past 60years. This is partly because the brain is a critical organ, and poses unique anatomical, physiological, and immunological barriers. The unique interplay of these barriers also provides an opportunity for creative engineering solutions. Cancer immunotherapy, a means of harnessing the host immune system for anti-tumor efficacy, is becoming a standard approach for treating many cancers. However, its use in brain tumors is not widespread. This review discusses the current approaches, and hurdles to these approaches in treating brain tumors, with a focus on immunotherapies. We identify critical barriers to immunoengineering brain tumor therapies and discuss possible solutions to these challenges.

Bacterial Carriers for Glioblastoma Therapy.

Treatment of aggressive glioblastoma brain tumors is challenging, largely due to diffusion barriers preventing efficient drug dosing to tumors. To overcome these barriers, bacterial carriers that are actively motile and programmed to migrate and localize to tumor zones were designed. These carriers can induce apoptosis via hypoxia-controlled expression of a tumor suppressor protein p53 and a pro-apoptotic drug, Azurin. In a xenograft model of human glioblastoma in rats, bacterial carrier therapy conferred a significant survival benefit with 19% overall long-term survival of >100 days in treated animals relative to a median survival of 26 days in control untreated animals. Histological and proteomic analyses were performed to elucidate the safety and efficacy of these carriers, showing an absence of systemic toxicity and a restored neural environment in treated responders. In the treated non-responders, proteomic analysis revealed competing mechanisms of pro-apoptotic and drug-resistant activity. This bacterial carrier opens a versatile avenue to overcome diffusion barriers in glioblastoma by virtue of its active motility in extracellular space and can lead to tailored therapies via tumor-specific expression of tumoricidal proteins.

Enhanced therapeutic neovascularization by CD31-expressing cells and embryonic stem cell-derived endothelial cells engineered with chitosan hydrogel containing VEGF-releasing microtubes.

Various stem cells and their progeny have been used therapeutically for vascular regeneration. One of the major hurdles for cell-based therapy is low cell retention in vivo, and to improve cell survival several biomaterials have been used to encapsulate cells before transplantation. Vascular regeneration involves new blood vessel formation which consists of two processes, vasculogenesis and angiogenesis. While embryonic stem cell (ESC)-derived endothelial cells (ESC-ECs) have clearer vasculogenic potency, adult cells exert their effects mainly through paracrine angiogenic activities. While these two cells have seemingly complementary advantages, there have not been any studies to date combining these two cell types for vascular regeneration. We have developed a novel chitosan-based hydrogel construct that encapsulates both CD31-expressing BM-mononuclear cells (BM-CD31(+) cells) and ESC-ECs, and is loaded with VEGF-releasing microtubes. This cell construct showed high cell survival and minimal cytotoxicity in vitro. When implanted into a mouse model of hindlimb ischemia, it induced robust cell retention, neovascularization through vasculogenesis and angiogenesis, and efficiently induced recovery of blood flow in ischemic hindlimbs. This chitosan-based hydrogel encapsulating mixed adult and embryonic cell derivatives and containing VEGF can serve as a novel platform for treating various cardiovascular diseases.

Noninvasive imaging of peripheral nerves.

Recent developments in the field of peripheral nerve imaging extend the capabilities of imaging modalities to assist in the diagnosis and treatment of patients with peripheral nerve maladies. Methods such as magnetic resonance imaging (MRI) and its derivative diffusion tensor imaging (DTI), ultrasound (US) and positron emission tomography (PET) are capable of assessing nerve structure and function following injury and relating the state of the nerve to electrophysiological and histological analysis. Of the imaging methods surveyed here, each offered unique and interesting advantages related to the field. MRI offered the opportunity to visualize immune activity on the injured nerve throughout the course of the regeneration process, and DTI offered numerical characterization of the injury and the ability to develop statistical bases for diagnosing injury. US extends imaging to the treatment phase by enabling more precise analgesic applications following surgery, and PET represents a novel method of assessing nerve injury through analysis of relative metabolism rates in injured and healthy tissue. Exciting new possibilities to enhance and extend the abilities of imaging methods are also discussed, including innovative contrast agents, some of which enable multimodal imaging approaches and present opportunities for treatment application.

A perspective on immunomodulation and tissue repair.

An immune response involves the action of all types of macrophages, classically activated subtype (M1) in the early inflammatory phase and regulatory and wound-healing subtypes (M2) in the resolution phase. The remarkable plasticity of macrophages makes them an interesting target in the context of immunomodulation. Here, we reviewed the current state of understanding regarding the role that different phenotypes of macrophages and monocytes play following injury and during the course of remodeling in different tissue types. Moreover, we explored recent designs of macrophage modulatory biomaterials for tissue engineering and regenerative medicine applications.

A microfluidic approach to synthesizing high-performance microfibers with tunable anhydrous proton conductivity.

Here, we demonstrate a new approach for the synthesis of ion exchange microfibers with finely tuned anhydrous conductivity. This work presents microfluidics as a system to control the size and phosphoric acid (PA) doping level of the polybenzimidazole (PBI) microfibers. It has been shown that the PA doping level can be controlled by varying the flow ratios in the microfluidic channel. The diameter of the microfibers increased with extending mixing time, whereas the doping level decreased with increasing flow ratio. The highest doping level, 16, was achieved at the flow ratio of 0.175. The anhydrous proton conductivity of the microfibers was found to be adjustable between 0.01 and 0.1 S cm(-1) at 160 °C, which is considerably higher than for conventionally doped PBI cast membranes (0.004 S cm(-1)). Furthermore, molecular dynamic simulation of proton conduction through the microfibers at different doping levels was in good agreement with the experimental results. These results demonstrate the potential of the microfluidic technique to precisely tune the physicochemical properties of PBI microfibers for various electrochemical applications such as hydrogen sensors, fuel cells as well as artificial muscles.

Effect of modulating macrophage phenotype on peripheral nerve repair.

Peripheral nerve repair across long gaps remains clinically challenging despite progress made with autograft transplantation. While scaffolds that present trophic factors and extracellular matrix molecules have been designed, matching the performance of autograft-induced repair has been challenging. In this study, we explored the effect of cytokine mediated 'biasing' of macrophage phenotypes on Schwann cell (SC) migration and axonal regeneration in vitro and in vivo. Macrophage phenotype was successfully modulated by local delivery of either Interferon-gamma (IFN-γ) or Interleukin-4 (IL-4) within polymeric nerve guidance channels, polarizing them toward pro-inflammatory (M1) or pro-healing (M2a and M2c) phenotypes, respectively. The initial polarization of macrophages to M2a and M2c phenotype results in enhanced SC infiltration and substantially faster axonal growth in a critically-sized rat sciatic nerve gap model (15 mm). The ratio of pro-healing to pro-inflammatory population of macrophages (CD206+/CCR7+), defined as regenerative bias, demonstrates a linear relationship with the number of axons at the distal end of the nerve scaffolds. The present results clearly suggest that rather than the extent of macrophage presence, their specific phenotype at the site of injury regulates the regenerative outcomes.

Overcoming endogenous constraints on neuronal regeneration.

One of the grand challenges in neuroengineering is to stimulate regeneration after central nervous system (CNS) or peripheral nervous system (PNS) injury to restore function. The state of the art today is that PNS injuries heal to a limited extent, whereas CNS injuries are largely intractable to regeneration. In this context, we examine the underlying biochemical and cellular constraints on endogenous healing of neural tissues. Identification and characterization of endogenous "rate-limiting" processes that constrain regeneration would allow one to craft solutions to overcome critical impediments for accelerated healing. It is increasingly evident that biochemical pathways triggered by the nature and duration of injury-triggered inflammatory response may determine the endogenous constraints and subsequently determine regenerative fate. In this paper, critical endogenous constraints of PNS and CNS regeneration are identified, and the effects of modulating the phenotypes of immune cells on neuronal regeneration are discussed.

Novel high-performance nanohybrid polyelectrolyte membranes based on bio-functionalized montmorillonite for fuel cell applications.

This study is concerned with electrochemical investigation of novel high-performance proton exchange membranes based on bio-functionalized montmorillonite and Nafion. It was found that the incorporation of 2 wt% BMMT into Nafion polyelectrolyte matrix results in significantly improved methanol-air fuel cell efficiency of 30% compared to 14% for Nafion(R)117, and about 23-times higher membrane selectivity.