The influence of surgical correction on white matter microstructural integrity in rabbits with familial coronal suture craniosynostosis.

OBJECT Craniosynostosis is a condition in which one or more of the calvarial sutures fuses prematurely. In addition to the cosmetic ramifications attributable to premature suture fusion, aberrations in neurophysiological parameters are seen, which may result in more significant damage. This work examines the microstructural integrity of white matter, using diffusion tensor imaging (DTI) in a homogeneous strain of rabbits with simple, familial coronal suture synostosis before and after surgical correction. METHODS After diagnosis, rabbits were assigned to different groups: wild-type (WT), rabbits with early-onset complete fusion of the coronal suture (BC), and rabbits that had undergone surgical correction with suturectomy (BC-SU) at 10 days of age. Fixed rabbit heads were imaged at 12, 25, or 42 days of life using a 4.7-T, 40-cm bore Avance scanner with a 7.2-cm radiofrequency coil. For DTI, a 3D spin echo sequence was used with a diffusion gradient (b = 2000 sec/mm(2)) applied in 6 directions. RESULTS As age increased from 12 to 42 days, the DTI differences between WT and BC groups became more pronounced (p < 0.05, 1-way ANOVA), especially in the corpus callosum, cingulum, and fimbriae. Suturectomy resulted in rabbits with no significant differences compared with WT animals, as assessed by DTI of white matter tracts. Also, it was possible to predict to which group an animal belonged (WT, BC, and BC-SU) with high accuracy based on imaging data alone using a linear support vector machine classifier. The ability to predict to which group the animal belonged improved as the age of the animal increased (71% accurate at 12 days and 100% accurate at 42 days). CONCLUSIONS Craniosynostosis results in characteristic changes of major white matter tracts, with differences becoming more apparent as the age of the rabbits increases. Early suturectomy (at 10 days of life) appears to mitigate these differences.

Traumatic brain injury alters long-term hippocampal neuron morphology in juvenile, but not immature, rats.

PURPOSE: Pediatric traumatic brain injury (TBI) represents a prominent yet understudied medical condition that can profoundly impact brain development. As the juvenile injured brain matures in the wake of neuropathological cascades during potentially critical periods, circuit alterations may explain neurological consequences, including cognitive deficits. We hypothesize that experimental brain injury in juvenile rats, with behavioral deficits that resolve, will lead to quantifiable structural changes in hippocampal neurons at chronic time points post-injury. METHODS: Controlled cortical impact (CCI), a model of focal TBI with contusion, was used to induce brain injury on post-natal day (PND) 17 juvenile rats. The histological consequence of TBI was quantified in regions of the hippocampus at post-injury day 28 (PID28) on sections stained using a variation of the Golgi-Cox staining method. Individual neuronal morphologies were digitized from the dentate gyrus (DG), CA3, and CA1 regions. RESULTS: Soma area in the ipsilateral injured DG and CA3 regions of the hippocampus increased significantly at PID28 in comparison to controls. In CA1, dendritic length and dendritic branching decreased significantly in comparison to controls and the contralateral hemisphere, without change in soma area. To extend the study, we examined neuronal morphology in rats with CCI at PND7. On PID28 after CCI on PND7 rats, CA1 neurons showed no injury-induced change in morphology, potentially indicating an age-dependent morphological response to injury. CONCLUSIONS: Long-lasting structural alterations in hippocampal neurons of brain-injured PND17 juvenile animals, but not PND7 immature animals, suggest differential plasticity depending on age-at-injury, with potential consequences for later function.

Morris water maze function and histologic characterization of two age-at-injury experimental models of controlled cortical impact in the immature rat.

PURPOSE: Controlled cortical impact (CCI) is commonly used in adult animals to study focal traumatic brain injury (TBI). Our study aims to further study injury mechanisms in children and variable models of pathology in the developing brain. METHODS: Develop a focal injury model of experimental TBI in the immature, postnatal days (PND) 7 and 17 rats that underwent a CCI at varying depths of deflection, 1.5-2.5 mm compared with sham and then tested using the Morris water maze (MWM) beginning on post-injury day (PID) 11. Histopathologic analysis was performed at PID 1 and 28. RESULTS: In PND 7, the 1.75- and 2.0-mm deflections (diameter (d) = 3 mm; velocity = 4 m/s; and duration = 500 ms) resulted in significant MWM deficits while the 1.5-mm injury did not produce MWM deficits vs. sham controls. In PND 17, all injury levels resulted in significant MWM deficits vs. sham controls with a graded response; the 1.5-mm deflection (d = 6 mm; velocity = 4 m/s; and duration = 500 ms) produced significantly less deficits as compared WITH the 2.0- and 2.5-mm injuries. Histologically, a graded injury response was also seen in both ages at injury with cortical and more severe injuries, hippocampal damage. Cortical contusion volume increased in most injury severities from PID 1 to 28 in both ages at injury while hippocampal volumes subsequently decreased. CONCLUSIONS: CCI in PND 7 and 17 rat results in significant MWM deficits and cortical histopathology providing two different and unique experimental models of TBI in immature rats that may be useful in further investigations into the mechanisms and treatments of pediatric TBI.

A neurodevelopmental epigenetic programme mediated by SMARCD3-DAB1-Reelin signalling is hijacked to promote medulloblastoma metastasis.

How abnormal neurodevelopment relates to the tumour aggressiveness of medulloblastoma (MB), the most common type of embryonal tumour, remains elusive. Here we uncover a neurodevelopmental epigenomic programme that is hijacked to induce MB metastatic dissemination. Unsupervised analyses of integrated publicly available datasets with our newly generated data reveal that SMARCD3 (also known as BAF60C) regulates Disabled 1 (DAB1)-mediated Reelin signalling in Purkinje cell migration and MB metastasis by orchestrating cis-regulatory elements at the DAB1 locus. We further identify that a core set of transcription factors, enhancer of zeste homologue 2 (EZH2) and nuclear factor I X (NFIX), coordinates with the cis-regulatory elements at the SMARCD3 locus to form a chromatin hub to control SMARCD3 expression in the developing cerebellum and in metastatic MB. Increased SMARCD3 expression activates Reelin-DAB1-mediated Src kinase signalling, which results in a MB response to Src inhibition. These data deepen our understanding of how neurodevelopmental programming influences disease progression and provide a potential therapeutic option for patients with MB.

Identification of ATP citrate lyase as a positive regulator of glycolytic function in glioblastomas.

Glioblastomas, the most malignant type of glioma, are more glycolytic than normal brain tissue. Robust migration of glioblastoma cells has been previously demonstrated under glycolytic conditions and their pseudopodia contain increased glycolytic and decreased mitochondrial enzymes. Glycolysis is suppressed by metabolic acids, including citric acid which is excluded from mitochondria during hypoxia. We postulated that glioma cells maintain glycolysis by regulating metabolic acids, especially in their pseudopodia. The enzyme that breaks down cytosolic citric acid is ATP citrate lyase (ACLY). Our identification of increased ACLY in pseudopodia of U87 glioblastoma cells on 1D gels and immunoblots prompted investigation of ACLY gene expression in gliomas for survival data and correlation with expression of ENO1, that encodes enolase 1. Queries of the NIH's REMBRANDT brain tumor database based on Affymetrix data indicated that decreased survival correlated with increased gene expression of ACLY in gliomas. Queries of gliomas and glioblastomas found an association of upregulated ACLY and ENO1 expression by chi square for all probe sets (reporters) combined and correlation for numbers of probe sets indicating shared upregulation of these genes. Real-time quantitative PCR confirmed correlation between ACLY and ENO1 in 21 glioblastomas (p < 0.001). Inhibition of ACLY with hydroxycitrate suppressed (p < 0.05) in vitro glioblastoma cell migration, clonogenicity and brain invasion under glycolytic conditions and enhanced the suppressive effects of a Met inhibitor on cell migration. In summary, gene expression data, proteomics and functional assays support ACLY as a positive regulator of glycolysis in glioblastomas.

Radiosurgery to the spinal dorsal root ganglion induces fibrosis and inhibits satellite glial cell activation while preserving axonal neurotransmission.

OBJECTIVE: Stereotactic radiosurgery (SRS) has been used to treat trigeminal neuralgia by targeting the cisternal segment of the trigeminal nerve, which in turn triggers changes in the gasserian ganglion. In the lumbar spine, the dorsal root ganglion (DRG) is responsible for transmitting pain sensitivity and is involved in the pathogenesis of peripheral neuropathic pain. Therefore, radiosurgery to the DRG might improve chronic peripheral pain. This study evaluated the clinical and histological effects of high-dose radiosurgery to the DRG in a rodent model. METHODS: Eight Sprague-Dawley rats received either 40- or 80-Gy SRS to the fifth and sixth lumbar DRGs using the Leksell Gamma Knife Icon. Animals were euthanized 3 months after treatment, and the lumbar spine was dissected and taken for analysis. Simple histology was used to assess collagen deposition and inflammatory response. GFAP, Neu-N, substance P, and internexin were used as a measure of peripheral glial activation, neurogenesis, pain-specific neurotransmission, and neurotransmission in general, respectively. The integrity of the spinothalamic tract was assessed by means of the von Frey test. RESULTS: The animals did not exhibit any signs of motor or sensory deficits during the experimentation period. Edema, fibrosis, and vascular sclerotic changes were present on the treated, but not the control, side. SRS reduced the expression of GFAP without affecting the expression of Neu-N, substance P, or internexin. The von Frey sensory perception elicited equivalent results for the control side and both radiosurgical doses. CONCLUSIONS: SRS did not alter sensory or motor function but reduced the activation of satellite glial cells, a pathway for DRG-mediated pain perpetuation. Radiosurgery provoked changes equivalent to the effects of focal radiation on the trigeminal ganglion after SRS for trigeminal neuralgia, suggesting that radiosurgery could be successful in relieving radiculopathic pain.

Therapeutic efficacy of a herpes simplex virus with radiation or temozolomide for intracranial glioblastoma after convection-enhanced delivery.

The herpes simplex virus-1 (HSV-1)-infected cell protein 0 (ICP0) is an E3 ubiquitin ligase implicated in cell cycle arrest and DNA repair inhibition. Convection-enhanced delivery (CED) of either the replication-defective, ICP0-producing HSV-1 mutant, d106, or the recombinant d109, devoid of all viral genome expression, was performed to determine the in vivo efficacy of ICP0 in combination with ionizing radiation (IR) or systemic temozolomide (TMZ) in the treatment of glioblastoma multiforme (GBM). Intracranial U87-MG xenografts were established in athymic nude mice. Animal survival was determined after mice underwent intracranial CED of either the replication-defective d106 or d109 viruses, or Hanks' balanced salt solution (HBSS), before a single session of whole-brain irradiation or TMZ treatment. Median survival for animals that underwent treatment with HBSS alone, d109 alone, d106 alone, HBSS + IR, HBSS + TMZ, d109 + IR, d106 + IR, and d106 + TMZ was 28, 35, 41, 39, 44, 39, 68 (P < 0.01), and 66 days (P < 0.01), respectively. Intracerebral d106 CED resulted in a significant increase in athymic nude mouse survival when combined with IR or TMZ. d106 CED allows for distribution of HSV-1 in human GBM xenografts and persistent viral infection.

Age-related peridural hyperemia in craniosynostotic rabbits.

OBJECTIVE: Craniosynostosis is the premature fusion of the calvarial sutures and is associated with aesthetic impairment and secondary damage to brain growth. Associated neurological injuries can result from increased intracranial pressure (ICP) and abnormal cerebral blood flow (CBF). Arterial spin-labeling (ASL) MRI was used to assess regional CBF in developing rabbits with early-onset coronal suture synostosis (EOCS) and age-matched wild-type controls (WT). METHODS: Rabbits were subjected to ASL MRI at or near 10, 25, or 42 days of age. Differences in regional CBF were assessed using one-way ANOVA. CONCLUSION: CBF was similar in WT and EOCS rabbits with the exception of the peridural surfaces in EOCS rabbits at 25 days of age. A twofold increase in peridural CBF at 25 days of age coincides with a transient increase in ICP. By 42 days of age, CBF in peridural surfaces had decreased.

Evaluation of Clinical and Histologic Effects of High-Dose Radiosurgery on Rat Dorsal Root Ganglion.

BACKGROUND: Stereotactic radiosurgery (SRS) is an effective technique to create lesions of the trigeminal nerve to treat refractory trigeminal neuralgia. In the lumbar spine, the dorsal root ganglion (DRG) contains the body of the sensory neurons responsible for pain sensitivity. Neuromodulation of the DRG might therefore improve chronic peripheral pain. This study was performed to determine the feasibility, clinical, and histologic effects of delivering high-dose SRS targeted to the lumbar DRG in a rat model. METHODS: Four Sprague Dawley male rats underwent 80 Gy maximum-dose single-fraction SRS to the left L5 and L6 DRG using the Leksell Gamma Knife Icon (Elekta, Atlanta, Georgia, USA). The right L5 and L6 DRGs served as controls. The animals were evaluated for motor and sensory deficits every 2 weeks and were sacrificed at 3 and 6 months after SRS. Common histologic techniques were used to assess for fibrosis and demyelination at the target levels. RESULTS: No detectable motor or sensory deficits were seen in any animal. Histologic changes including fibrosis and loss of myelin were noted to the left L5 and L6 DRGs, but not the right side control DRGs. Fibrotic changes within the vertebral body were also evident on the treated sides of the vertebral bodies. CONCLUSIONS: We were able to detect a demyelinating response from SRS delivered to the DRG in rats. Because such changes mimic those seen after trigeminal SRS in experimental animals, we hypothesize that radiosurgery may be a potential option in chronic spinal radicular pain amenable to neuromodulation.

The influence of suturectomy on age-related changes in cerebral blood flow in rabbits with familial bicoronal suture craniosynostosis: A quantitative analysis.

BACKGROUND: Coronal suture synostosis is a condition which can have deleterious physical and cognitive sequelae in humans if not corrected. A well-established animal model has previously demonstrated disruptions in intracranial pressure and developmental abnormalities in rabbits with congenital craniosynostosis compared to wild type rabbits. OBJECTIVE: The current study aimed to measure the cerebral blood flow (CBF) in developing rabbits with craniosynostosis who underwent suturectomy compared to those with no intervention and compared to wild type rabbits. METHODS: Rabbits with early onset coronal suture synostosis were assigned to have suturectomy at 10 days of age (EOCS-SU, n = 15) or no intervention (EOCS, n = 18). A subset of each group was randomly selected for measurement at 10 days of age, 25 days of age, and 42 days of age. Wild type rabbits (WT, n = 18) were also randomly assigned to measurement at each time point as controls. Cerebral blood flow at the bilateral hemispheres, cortices, thalami, and superficial cortices was measured in each group using arterial spin-labeling MRI. RESULTS: At 25 days of age, CBF at the superficial cortex was significantly higher in EOCS rabbits (192.6 ± 10.1 mL/100 mg/min on the left and 195 ± 9.5 mL/100 mg/min on the right) compared to WT rabbits (99.2 ± 29.1 mL/100 mg/min on the left and 96.2 ± 21.4 mL/100 mg/min on the right), but there was no significant difference in CBF between EOCS-SU (97.6 ± 11.3 mL/100 mg/min on the left and 99 ± 7.4 mL/100 mg/min on the right) and WT rabbits. By 42 days of age the CBF in EOCS rabbits was not significantly different than that of WT rabbits. CONCLUSION: Suturectomy eliminated the abnormally increased CBF at the superficial cortex seen in EOCS rabbits at 25 days of age. This finding contributes to the evidence that suturectomy limits abnormalities of ICP and CBF associated with craniosynostosis.

Toward Improving Safety in Neurosurgery with an Active Handheld Instrument.

Microsurgical procedures, such as petroclival meningioma resection, require careful surgical actions in order to remove tumor tissue, while avoiding brain and vessel damaging. Such procedures are currently performed under microscope magnification. Robotic tools are emerging in order to filter surgeons' unintended movements and prevent tools from entering forbidden regions such as vascular structures. The present work investigates the use of a handheld robotic tool (Micron) to automate vessel avoidance in microsurgery. In particular, we focused on vessel segmentation, implementing a deep-learning-based segmentation strategy in microscopy images, and its integration with a feature-based passive 3D reconstruction algorithm to obtain accurate and robust vessel position. We then implemented a virtual-fixture-based strategy to control the handheld robotic tool and perform vessel avoidance. Clay vascular phantoms, lying on a background obtained from microscopy images recorded during petroclival meningioma surgery, were used for testing the segmentation and control algorithms. When testing the segmentation algorithm on 100 different phantom images, a median Dice similarity coefficient equal to 0.96 was achieved. A set of 25 Micron trials of 80 s in duration, each involving the interaction of Micron with a different vascular phantom, were recorded, with a safety distance equal to 2 mm, which was comparable to the median vessel diameter. Micron's tip entered the forbidden region 24% of the time when the control algorithm was active. However, the median penetration depth was 16.9 µm, which was two orders of magnitude lower than median vessel diameter. Results suggest the system can assist surgeons in performing safe vessel avoidance during neurosurgical procedures.

Toll like receptor-3 ligand poly-ICLC promotes the efficacy of peripheral vaccinations with tumor antigen-derived peptide epitopes in murine CNS tumor models.

BACKGROUND: Toll-like receptor (TLR)3 ligands serve as natural inducers of pro-inflammatory cytokines capable of promoting Type-1 adaptive immunity, and TLR3 is abundantly expressed by cells within the central nervous system (CNS). To improve the efficacy of vaccine strategies directed against CNS tumors, we evaluated whether administration of a TLR3 ligand, polyinosinic-polycytidylic (poly-IC) stabilized with poly-lysine and carboxymethylcellulose (poly-ICLC) would enhance the anti-CNS tumor effectiveness of tumor peptide-based vaccinations. METHODS: C57BL/6 mice bearing syngeneic CNS GL261 glioma or M05 melanoma received subcutaneous (s.c.) vaccinations with synthetic peptides encoding CTL epitopes--mEphA2 (671-679), hgp100 (25-33) and mTRP-2 (180-188) for GL261, or ovalbumin (OVA: 257-264) for M05. The mice also received intramuscular (i.m.) injections with poly-ICLC. RESULTS: The combination of subcutaneous (s.c.) peptide-based vaccination and i.m. poly-ICLC administration promoted systemic induction of antigen (Ag)-specific Type-1 CTLs expressing very late activation antigen (VLA)-4, which confers efficient CNS-tumor homing of vaccine-induced CTLs based on experiments with monoclonal antibody (mAb)-mediated blockade of VLA-4. In addition, the combination treatment allowed expression of IFN-gamma by CNS tumor-infiltrating CTLs, and improved the survival of tumor bearing mice in the absence of detectable autoimmunity. CONCLUSION: These data suggest that poly-ICLC, which has been previously evaluated in clinical trials, can be effectively combined with tumor Ag-specific vaccine strategies, thereby providing a greater index of therapeutic efficacy.

Tip Design for Safety of Steerable Needles for Robot-Controlled Brain Insertion.

BACKGROUND: Current practice in neurosurgical needle insertion is limited by the straight trajectories inherent with rigid probes. One technique allowing curvilinear trajectories involves flexible bevel-tipped needles, which bend during insertion due to their asymmetry. In the brain, safety will require avoidance of the sharp tips often used in laboratory studies, in favor of a more rounded profile. Steering performance, on the other hand, requires maximal asymmetry. Design of safe bevel-tipped brain needles thus involves management of this tradeoff by adjusting needle gauge, bevel angle, and fillet (or tip) radius to arrive at a design that is suitably asymmetrical while producing strain, strain rate, and stress below the levels that would damage brain tissue. METHODS: Designs with a variety of values of needle radius, bevel angle, and fillet radius were evaluated in finite-element simulations of simultaneous insertion and rotation. Brain tissue was modeled as a hyperelastic, linear viscoelastic material. Based on the literature available, safety thresholds of 0.19 strain, 10 s-1 strain rate, and 120 kPa stress were used. Safe values of needle radius, bevel angle, and fillet radius were selected, along with an appropriate velocity envelope for safe operation. The resulting needle was fabricated and compared with a Sedan side-cutting brain biopsy needle in a study in the porcine model in vivo (N=3). RESULTS: The prototype needle selected was 1.66 mm in diameter, with bevel angle of 10° and fillet radius of 0.25 mm. Upon examination of postoperative CT and histological images, no differences in tissue trauma or hemorrhage were noted between the prototype needle and the Sedan needle. CONCLUSIONS: The study indicates a general design technique for safe bevel-tipped brain needles based on comparison with relevant damage thresholds for strain, strain rate, and stress. The full potential of the technique awaits the determination of more exact safety thresholds.

Delivery of interferon-alpha transfected dendritic cells into central nervous system tumors enhances the antitumor efficacy of peripheral peptide-based vaccines.

We evaluated the effects, on immunity and survival, of injection of interferon (IFN)-alpha-transfected dendritic cells (DC-IFN-alpha) into intracranial tumors in mice immunized previously with syngeneic dendritic cells (DCs) pulsed either with ovalbumin-derived CTL or T helper epitopes. These immunizations protected animals from s.c. challenge with ovalbumin-expressing M05 melanoma (class I+ and class II-negative). Notably, antiovalbumin CTL responses were observed in animals vaccinated with an ovalbumin-derived T helper epitope but only after the mice were challenged with M05 cells. This cross-priming of CTL was dependent on both CD4+ and CD8+ T cells. Because we observed that s.c., but not intracranial, tumors were infiltrated with CD11c+ DCs, and because IFN-alpha promotes the activation and survival of both DCs and T cells, we evaluated the combinational antitumor effects of injecting adenoviral (Ad)-IFN-alpha-engineered DCs into intracranial M05 tumors in preimmunized mice. Delivery of DC-IFN-alpha prolonged survival. This was most notable for animals prevaccinated with both the CTL and T helper ovalbumin epitopes, with 60% (6 of 10) of mice (versus 0 of 10 of control animals) surviving for > 80 days after tumor challenge. DC-IFN-alpha appeared to persist longer than mock-transfected DCs within the intracranial tumor microenvironment, and DC-IFN-alpha-treated mice exhibited enhanced levels of ovalbumin-specific CTL in draining cervical lymph nodes. On the basis of these results, we believe that local expression of IFN-alpha by DCs within the intracranial tumor site may enhance the clinical efficacy of peripheral vaccine approaches for brain tumors.

Human Connectome-Based Tractographic Atlas of the Brainstem Connections and Surgical Approaches.

BACKGROUND: The brainstem is one of the most challenging areas for the neurosurgeon because of the limited space between gray matter nuclei and white matter pathways. Diffusion tensor imaging-based tractography has been used to study the brainstem structure, but the angular and spatial resolution could be improved further with advanced diffusion magnetic resonance imaging (MRI). OBJECTIVE: To construct a high-angular/spatial resolution, wide-population-based, comprehensive tractography atlas that presents an anatomical review of the surgical approaches to the brainstem. METHODS: We applied advanced diffusion MRI fiber tractography to a population-based atlas constructed with data from a total of 488 subjects from the Human Connectome Project-488. Five formalin-fixed brains were studied for surgical landmarks. Luxol Fast Blue-stained histological sections were used to validate the results of tractography. RESULTS: We acquired the tractography of the major brainstem pathways and validated them with histological analysis. The pathways included the cerebellar peduncles, corticospinal tract, corticopontine tracts, medial lemniscus, lateral lemniscus, spinothalamic tract, rubrospinal tract, central tegmental tract, medial longitudinal fasciculus, and dorsal longitudinal fasciculus. Then, the reconstructed 3-dimensional brainstem structure was sectioned at the level of classic surgical approaches, namely supracollicular, infracollicular, lateral mesencephalic, perioculomotor, peritrigeminal, anterolateral (to the medulla), and retro-olivary approaches. CONCLUSION: The advanced diffusion MRI fiber tracking is a powerful tool to explore the brainstem neuroanatomy and to achieve a better understanding of surgical approaches. ABBREVIATIONS: CN, cranial nerveCPT, corticopontine tractCST, corticospinal tractCTT, central tegmental tractDLF, dorsal longitudinal fasciculusHCP, Human Connectome ProjectML, medial lemniscusMLF, medial longitudinal fasciculusRST, rubrospinal tractSTT, spinothalamic tract.

Epidermal growth factor receptor-transfected bone marrow stromal cells exhibit enhanced migratory response and therapeutic potential against murine brain tumors.

We have created a novel cellular vehicle for gene therapy of malignant gliomas by transfection of murine bone marrow stroma cells (MSCs) with a cDNA encoding epidermal growth factor receptor (EGFR). These cells (EGFR-MSCs) demonstrate enhanced migratory responses toward glioma-conditioned media in comparison to primary MSCs in vitro. Enhanced migration of EGFR-MSC was at least partially dependent on EGF-EGFR, PI3-, MAP kinase kinase, and MAP kinases, protein kinase C, and actin polymerization. Unlike primary MSCs, EGFR-MSCs were resistant to FasL-mediated cytotoxicity and were capable of stimulating allogeneic mixed lymphocyte reaction, suggesting EGFR-MSCs possess suitable characteristics as vehicles for brain tumor immuno-gene therapy. Following injection at various sites, including the contralateral hemisphere in the brain of syngeneic mice, EGFR-MSCs were able to migrate toward GL261 gliomas or B16 melanoma in vivo. Finally, intratumoral injection with EGFR-MSC adenovirally engineered to secrete interferon-alpha to intracranial GL261 resulted in significantly prolonged survival in comparison to controls. These data indicate that EGFR-MSCs may serve as attractive vehicles for infiltrating brain malignancies such as malignant gliomas.

Hippocampal neurotransplantation evaluated in the rat kainic acid epilepsy model.

OBJECTIVE: Neurotransplantation has focused on disorders that involve subcortical brain targets. We evaluated the concepts of epileptic focus repair and changes in animal behavior through replacement of lost hippocampal neurons. The safety of hippocampal neurotransplantation was assessed in the rat kainic acid (KA) epilepsy model. METHODS: Sixty-three rats were studied and classified into six groups: KA plus 40,000 LBS-Neurons (Layton BioScience, Sunnyvale, CA; n = 13); KA plus 80,000 cells (n = 12); KA plus media (n = 9); no-KA plus 40,000 cells (n = 12); no-KA plus 80,000 cells (n = 12); and no-KA plus media (n = 5). Clinical observation (2 h daily) and electroencephalogram recording (3 h every other week) were performed to check for seizures until Week 11 after KA injection. On Week 12, the Morris water maze test was performed to assess spatial learning and memory. RESULTS: Four rats were excluded because of intracranial hematoma or abscess. In the clinical observation of seizures, the no-KA plus media group had significantly fewer seizures than rats that received KA followed by injection of 40,000 cells, 80,000 cells, or media (P = 0.001, 0.0004, and 0.004, respectively). On electroencephalographic analysis, there was no significant difference between any of the groups. Transplanted rats with KA-induced epilepsy did not have an increased number of seizures. In the Morris water maze test, the hidden platform task showed that the KA plus 80,000 cell group had significantly longer swim latencies than groups with no-KA plus 40,000 cells (P = 0.035) or no-KA plus 80,000 cells (P = 0.015), demonstrating the behavioral deficits caused by KA injection. The probe trial showed no significant difference for the percentage of time in the target quadrant between any of the groups. Histological studies showed that 26 (59%) of 44 transplanted rats had evidence of graft survival. CONCLUSION: The safety of cortical neurotransplantation was demonstrated, even in an animal model predisposed to epilepsy. We did not find evidence for cessation of seizures or improvement in behavior using this model.

Intracranial pressure changes in craniosynostotic rabbits.

Cranial vault and brain deformities in individuals with craniosynostosis are thought to result, in part, from changes in intracranial pressure, but clinical findings are still inconclusive. The present study describes intracranial pressure changes in a rabbit model with naturally occurring, uncorrected coronal suture synostosis. Longitudinal and cross-sectional intracranial pressure data were collected from 241 New Zealand White rabbits, divided into four groups: normal controls (n = 81); rabbits with delayed-onset coronal suture synostosis (n = 78); rabbits with early-onset unilateral coronal suture synostosis (n = 32); and rabbits with early-onset bilateral coronal suture synostosis (n = 50). Epidural intracranial pressure measurements were obtained at 10, 25, 42, and 84 days of age using a NeuroMonitor microsensor transducer. Normal rabbits and rabbits with delayed-onset coronal suture and early-onset unilateral coronal suture synostosis showed a similar oscillating pattern of age-related changes in normal and head-down intracranial pressure from 10 to 84 days of age. In contrast, rabbits with early-onset bilateral coronal suture synostosis showed markedly elevated normal and head-down intracranial pressure levels from 10 to 25 days and showed a different pattern through 84 days. Results from one-way analysis of variance revealed significant (p < 0.01) group differences only at 25 days of age. Rabbits with early-onset bilateral coronal suture synostosis had significantly (p < 0.05) greater normal and head-down intracranial pressure (by 42 percent) than the other three groups. These results showed differing intracranial pressure compensations in rabbits with uncorrected multiple-suture synostosis compared with normal rabbits or rabbits with uncorrected single-suture synostosis, possibly through progressive cerebral atrophy and decreased intracranial volume, abnormal intracranial vascular patterns and blood volume, and/or differing cranial vault compensatory changes.

Premetastatic soil and prevention of breast cancer brain metastasis.

BACKGROUND: As therapies for systemic cancer improve and patients survive longer, the risk for brain metastases increases. We evaluated whether immune mechanisms are involved in the development of brain metastasis. METHODS: We conducted our studies using BALB/c mice bearing syngeneic 4T1 mammary adenocarcinoma cells in the mammary gland. RESULTS: The brains of mice bearing 4T1 tumors at day 14 had no detectable metastatic tumor cells but presented with marked accumulation of bone marrow-derived CD11b(+)Gr1(+) myeloid cells, which express high levels of inflammatory chemokines S100A8 and S100A9. In vitro, S100A9 attracts 4T1 cells through Toll-like receptor 4 and CD11b(+)Gr1(+) myeloid cells through Toll-like receptor 4 and the receptor for advanced glycation end-products. Systemic treatment of 4T1-bearing mice with anti-Gr1 (RB6-8C5) monoclonal antibody reduces accumulation of CD11b(+)Gr1(+) myeloid cells in the day-14 premetastatic brain as well as subsequent brain metastasis of 4T1 cells detected on day 30. Furthermore, treatment of 4T1 tumor-bearing mice with the cyclooxygenase-2 inhibitor celecoxib or genetic disruption of cyclooxygenase-2 in 4T1 cells inhibits the inflammatory chemokines and infiltration of CD11b(+)Gr1(+) myeloid cells in the premetastatic brain and subsequent formation of brain metastasis. CONCLUSIONS: Our results suggest that the primary tumor induces accumulation of CD11b(+)Gr1(+) myeloid cells in the brain to form "premetastatic soil" and inflammation mediators, such as S100A9, that attract additional myeloid cells as well as metastatic tumor cells. Celecoxib and anti-Gr1 treatment may be useful for blockade of these processes, thereby preventing brain metastasis in patients with breast cancer.

Injection parameters affect cell viability and implant volumes in automated cell delivery for the brain.

The technique of central nervous system cell implantation can affect the outcome of preclinical or clinical studies. Our goal was to evaluate the impact of various injection parameters that may be of consequence during the delivery of solute-suspended cells. These parameters included (1) the type and concentration of cells used for implantation, (2) the rate at which cells are injected (flow rate), (3) the acceleration of the delivery device, (4) the period of time between cell loading and injection into the CNS (delay), and (5) the length and gauge of the needle used to deliver the cells. Neural progenitor cells (NPCs) and bone marrow stromal cells (BMSCs) were injected an automated device. These parameters were assessed in relation to their effect on the volume of cells injected and cell viability. Longer and thinner cannulae and higher cell concentrations were detrimental for cell delivery. Devices and techniques that optimize these parameters should be of benefit.