Heechul Kim

MRI-detectable pH nanosensors incorporated into hydrogels for in vivo sensing of transplanted-cell viability

Biocompatible nanomaterials and hydrogels have become an important tool for improving cell-based therapies by promoting cell survival and protecting cell transplants from immune rejection. Although their potential benefit has been widely evaluated, it is currently not possible to determine, in vivo, if and how long cells remain viable following their administration without the use of a reporter gene. We here report a pH nanosensor-based magnetic resonance imaging (MRI) technique that can monitor cell death in vivo non-invasively. We demonstrate that specific MRI parameters that change upon cell death of microencapsulated hepatocytes are associated with the measured bioluminescence imaging (BLI) radiance. Moreover, the readout from this pH-sensitive nanosensor can be directly co-registered with high-resolution anatomical images. All the components of these nanosensors are clinical-grade and hence this approach should be a translatable and universal modification of hydrogels.

Multifunctional capsule-in-capsules for immunoprotection and trimodal imaging.

Type I diabetes mellitus (T1DM) is a T-cell-mediated autoimmune disease that results in destruction of insulin-producing β cells and subsequent hyperglycemia.[1,2] The current way of treating T1DM is insulin replacement therapy through repetitive injections of recombinant insulin. In more serious cases, a cadaveric pancreas[3] or purified pancreatic islets[4,5] can be transplanted to restore proper glucose regulation. However, the risks of surgery and the accompanying life-long immunosuppression outweigh the disadvantages of continued administration of insulin. The immunoisolation of islets by alginate microencapsulation is an emerging and promising solution to circumvent immune rejection and so overcome this limitation.[6] While the semipermeable alginate membrane blocks penetration of immune cells and antibodies, it allows the unhindered passage of nutrients, metabolites, and insulin that are produced by encapsulated islet cells.[7] Intraperitoneal administration of microencapsulated islets in monkeys and humans has showed considerable promise for the treatment of T1DM.[8,9]

Optogenetic-guided cortical plasticity after nerve injury

Peripheral nerve injury causes sensory dysfunctions that are thought to be attributable to changes in neuronal activity occurring in somatosensory cortices both contralateral and ipsilateral to the injury. Recent studies suggest that distorted functional response observed in deprived primary somatosensory cortex (S1) may be the result of an increase in inhibitory interneuron activity and is mediated by the transcallosal pathway. The goal of this study was to develop a strategy to manipulate and control the transcallosal activity to facilitate appropriate plasticity by guiding the cortical reorganization in a rat model of sensory deprivation. Since transcallosal fibers originate mainly from excitatory pyramidal neurons somata situated in laminae III and V, the excitatory neurons in rat S1 were engineered to express halorhodopsin, a light-sensitive chloride pump that triggers neuronal hyperpolarization. Results from electrophysiology, optical imaging, and functional MRI measurements are concordant with that within the deprived S1, activity in response to intact forepaw electrical stimulation was significantly increased by concurrent illumination of halorhodopsin over the healthy S1. Optogenetic manipulations effectively decreased the adverse inhibition of deprived cortex and revealed the major contribution of the transcallosal projections, showing interhemispheric neuroplasticity and thus, setting a foundation to develop improved rehabilitation strategies to restore cortical functions.

Human Glial-Restricted Progenitors Survive, Proliferate, and Preserve Electrophysiological Function in Rats with Focal Inflammatory Spinal Cord Demyelination

Transplantation of glial progenitor cells results in transplant‐derived myelination and improved function in rodents with genetic dysmyelination or chemical demyelination. However, glial cell transplantation in adult CNS inflammatory demyelinating models has not been well studied. Here we transplanted human glial‐restricted progenitor (hGRP) cells into the spinal cord of adult rats with inflammatory demyelination, and monitored cell fate in chemically immunosuppressed animals. We found that hGRPs migrate extensively, expand within inflammatory spinal cord lesions, do not form tumors, and adopt a mature glial phenotype, albeit at a low rate. Human GRP‐transplanted rats, but not controls, exhibited preserved electrophysiological conduction across the spinal cord, though no differences in behavioral improvement were noted between the two groups. Although these hGRPs myelinated extensively after implantation into neonatal shiverer mouse brain, only marginal remyelination was observed in the inflammatory spinal cord demyelination model. The low rate of transplant‐derived myelination in adult rat spinal cord may reflect host age, species, transplant environment/location, and/or immune suppression regime differences. We conclude that hGRPs have the capacity to myelinate dysmyelinated neonatal rodent brain and preserve conduction in the inflammatory demyelinated adult rodent spinal cord. The latter benefit is likely dependent on trophic support and suggests further exploration of potential of glial progenitors in animal models of chronic inflammatory demyelination.

IMMUNOMODULATION BY TRANSPLANTED HUMAN EMBRYONIC STEM CELL-DERIVED OLIGODENDROGLIAL PROGENITORS IN EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS

Transplantation of embryonic stem cells and their neural derivatives can lead to amelioration of the disease symptoms of experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis (MS). Oligodendroglial progenitors (OPs), derived from human embryonic stem cells (hESC, HES‐1), were labeled with superparamagnetic iron oxide and transduced with luciferase. At 7 days following induction of EAE in C57/BL6 mice, 1 × 106 cells were transplanted in the ventricles of C57/BL6 mice and noninvasively monitored by magnetic resonance and bioluminescence imaging. Cells were found to remain within the cerebroventricular system and did not survive for more than 10 days. However, EAE mice that received hESC‐OPs showed a significant improvement in neurological disability scores (0.9 ± 0.2; n = 12) compared to that of control animals (3.3 ± 0.4; n = 12) at day 15 post‐transplantation. Histopathologically, transplanted hESC‐OPs generated TREM2‐positive CD45 cells, increased TIMP‐1 expression, confined inflammatory cells within the subarachnoid space, and gave rise to higher numbers of Foxp3‐positive regulatory T cells in the spinal cord and spleen. Our results suggest that transplantation of hESC‐OPs can alter the pathogenesis of EAE through immunomodulation, potentially providing new avenues for stem cell‐based treatment of MS. STEM CELLS 2012;30:2820–2829

Neural Precursors Exhibit Distinctly Different Patterns of Cell Migration Upon Transplantation During Either the Acute or Chronic Phase of EAE: A Serial MR Imaging Study

As the complex pathogenesis of multiple sclerosis contributes to spatiotemporal variations in the trophic micromilieu of the central nervous system, the optimal intervention period for cell‐replacement therapy must be systematically defined. We applied serial, 3D high‐resolution magnetic resonance imaging to transplanted neural precursor cells (NPCs) labeled with superparamagnetic iron oxide nanoparticles and 5‐bromo‐2‐deoxyuridine, and compared the migration pattern of NPCs in acute inflamed (n = 10) versus chronic demyelinated (n = 9) brains of mice induced with experimental allergic encephalomyelitis (EAE). Serial in vivo and ex‐vivo 3D magnetic resonance imaging revealed that NPCs migrated 2.5 ± 1.3 mm along the corpus callosum in acute EAE. In chronic EAE, cell migration was slightly reduced (2.3 ± 1.3 mm) and only occurred in the lateral side of transplantation. Surprisingly, in 6/10 acute EAE brains, NPCs were found to migrate in a radial pattern along RECA‐1+ cortical blood vessels, in a pattern hitherto only reported for migrating glioblastoma cells. This striking radial biodistribution pattern was not detected in either chronic EAE or disease‐free control brains. In both acute and chronic EAE brain, Iba1+ microglia/macrophage number was significantly higher in central nervous system regions containing migrating NPCs. The existence of differential NPC migration patterns is an important consideration for implementing future translational studies in multiple sclerosis patients with variable disease. Magn Reson Med, 2011.

ICV-transplanted human glial precursor cells are short-lived yet exert immunomodulatory effects in mice with EAE.

Human glial precursor cells (hGPs) have potential for remyelinating lesions and are an attractive cell source for cell therapy of multiple sclerosis (MS). To investigate whether transplanted hGPs can affect the pathogenesis of experimental autoimmune encephalomyelitis (EAE), an animal model of MS, we evaluated the therapeutic effects of transplanted hGPs together with the in vivo fate of these cells using magnetic resonance imaging (MRI) and bioluminescence imaging (BLI). At 14 days post‐EAE induction, mice (n = 19) were intracerebroventricularly (ICV) injected with 5 × 105 hGPs that were magnetically labeled with superparamagnetic iron oxide (SPIO) particles as MR contrast agent and transduced with firefly luciferase for BLI of cell survival. Control mice (n = 18) received phosphate buffered saline (PBS) vehicle only. The severity of EAE clinical disability in the hGP‐transplanted group was significantly suppressed (P < 0.05) with concomitant inhibition of ConA and MOG‐specific T cell proliferation in the spleen. Astrogliosis was reduced and a lower activity of macrophages and/or microglia was observed in the spinal cord (P < 0.05). On MRI, SPIO signal was detected within the lateral ventricle from 1 day post‐transplantation and remained there for up to 34 days. BLI indicated that most cells did not survive beyond 5–10 days, consistent with the lack of detectable migration into the brain parenchyma and the histological presence of an abundance of apoptotic cells. Transplanted hGPs could not be detected in the spleen. We conclude that ICV transplantation of short‐lived hGPs can have a remote therapeutic effect through immunomodulation from within the ventricle, without cells directly participating in remyelination.

Immunohistochemical studies on disabled-2 protein in the spinal cords of rats with experimental autoimmune encephalomyelitis.

Disabled-2 (Dab-2), an adaptor protein of transforming growth factor beta (TGF-β) signaling, was studied in the spinal cords of rats with experimental autoimmune encephalomyelitis (EAE) to evaluate the possible involvement of Dab-2 in the pathogenesis of EAE using Western blot and immunohistochemical analyses. Western blot analysis showed that two isoforms (p96 kDa and p67 kDa) of Dab-2 were detected in the spinal cords of rats used as controls. Both isoforms of Dab-2 were significantly elevated in the EAE spinal cord at the peak stage of EAE (P<0.05) and declined at the recovery stage. However, only the p96 kDa isoform was markedly phosphorylated in the EAE spinal cord. Immunohistochemistry showed that Dab-2 and p-Dab-2 were detected in some vascular endothelial cells, glial cells, and some neurons in the rat spinal cords of normal and immunized CFA-alone controls. In EAE lesions, Dab-2 and p-Dab-2 were immunodetected in some inflammatory cells (mainly in ED1-positive macrophages and R73-positive T cells), while the enhanced immunoreactivity of Dab-2 in spinal cord cells suggested constitutive expression. Additionally, TGF-β1 immunoreactivity showed a similar expression pattern of Dab-2 in EAE lesions. These findings suggest that Dab-2 is transiently upregulated and phosphorylated (particularly the p96 kDa isoform) in EAE, a CNS autoimmune disease, and may be involved in TGF-β signaling.