Co-amplification of phosphoinositide 3-kinase enhancer A and cyclin-dependent kinase 4 triggers glioblastoma progression.

Glioblastoma (GBM) is the most common primary brain tumor and has a dismal prognosis. Amplification of chromosome 12q13-q15 (Cyclin-dependent kinase 4 (CDK4) amplicon) is frequently observed in numerous human cancers including GBM. Phosphoinositide 3-kinase enhancer (PIKE) is a group of GTP-binding proteins that belong to the subgroup of centaurin GTPase family, encoded by CENTG1 located in CDK4 amplicon. However, the pathological significance of CDK4 amplicon in GBM formation remains incompletely understood. In the current study, we show that co-expression of PIKE-A and CDK4 in TP53/PTEN double knockout GBM mouse model additively shortens the latency of glioma onset and survival compared to overexpression of these genes alone. Consequently, p-mTOR, p-Akt and p-ERK pathways are highly upregulated in the brain tumors, in alignment with their oncogenic activities by CDK4 and PIKE-A stably transfected in GBM cell lines. Hence, our findings support that PIKE amplification or overexpression coordinately acts with CDK4 to drive GBM tumorigenesis.

Fyn-phosphorylated PIKE-A binds and inhibits AMPK signaling, blocking its tumor suppressive activity.

The AMP-activated protein kinase, a key regulator of energy homeostasis, has a critical role in metabolic disorders and cancers. AMPK is mainly regulated by cellular AMP and phosphorylation by upstream kinases. Here, we show that PIKE-A binds to AMPK and blocks its tumor suppressive actions, which are mediated by tyrosine kinase Fyn. PIKE-A directly interacts with AMPK catalytic alpha subunit and impairs T172 phosphorylation, leading to repression of its kinase activity on the downstream targets. Mutation of Fyn phosphorylation sites on PIKE-A, depletion of Fyn, or pharmacological inhibition of Fyn blunts the association between PIKE-A and AMPK, resulting in loss of its inhibitory effect on AMPK. Cell proliferation and oncogenic assays demonstrate that PIKE-A antagonizes tumor suppressive actions of AMPK. In human glioblastoma samples, PIKE-A expression inversely correlates with the p-AMPK levels, supporting that PIKE-A negatively regulates AMPK activity in cancers. Thus, our findings provide additional layer of molecular regulation of the AMPK signaling pathway in cancer progression.

Merlin sumoylation is required for its tumor suppressor activity.

Merlin, encoded by the Neurofibromatosis 2 (NF2) gene, is a multifunctional tumor suppressor that integrates and regulates extracellular cues and intracellular signaling pathways, both at the plasma membrane and in the nucleus, to control cell proliferation, migration and invasion. Molecular mechanisms regulating merlin's tumor-suppressive activity have not been clearly defined. Here we report that merlin can be sumoylated on Lysine residue (K76) in vitro and in vivo. Sumoylation mediates merlin's intramolecular and intermolecular binding activities and regulates its cytoplasm/nucleus trafficking. Interestingly, sumoylation of merlin is regulated by its phosphorylation via Akt and PAK2 kinases. Mutation of K76 into arginine (R) abolishes its sumoylation, disrupts merlin cortical cytoskeleton residency and attenuates its stability. Using a K76R mutant merlin in a subcutaneous U87MG xenograft model, we demonstrate that merlin sumoylation is required for tumor-suppressive activity. Taken together, our findings indicate that merlin is sumoylated and that this post-translational modification is essential for tumor suppression.

Extraventricular neurocytomas: pathologic features and clinical outcome.

Neurocytic neoplasms usually arise within the lateral ventricles, generally as circumscribed, slowly growing masses curable by total resection. Both subtotal resection and histologic atypia are associated with an increased risk of recurrence. In contrast, neurocytic neoplasms situated within brain parenchyma, so-called "extraventricular neurocytomas" (EVNs), are not as well characterized. The relationships between histologic features and extent of resection versus clinical behavior have not been defined. We evaluated pathologic features, clinical data, and neuroimaging of 35 examples. The tumors occurred in 18 males and 17 females, age 5-76 years (median 34 years). All tumors involved the cerebrum. On imaging, EVNs were solitary, variably contrast-enhancing, and often (57%) cystic. Tumor cells were arranged in sheets, clusters, ribbons, or rosettes, in association with fine neuropil dispersed in broad zones that separated cell aggregates. Ganglion cell differentiation was seen in 66%. All tumors showed strong synaptophysin immunoreactivity. Despite the lack of apparent astrocytes in hematoxylin and eosin-stained sections, focal glial fibrillary acidic protein reactivity was seen in 46%. Eleven EVNs were designated "atypical" based on the presence of necrosis, vascular proliferation, or elevated mitotic activity (> or = 3 mitoses/10 high power fields). Nineteen tumors were subtotally resected or biopsied, whereas 14 were totally resected grossly. Seventeen patients underwent radiotherapy (mean 55 Gy). In 30 cases with follow-up, 10 tumors recurred, 3 causing death at 6, 14, and 43 months. All 10 recurrences followed subtotal resection. No totally resected tumors recurred. Thus, the majority of EVNs are well differentiated and appear unlikely to recur after gross total resection. Subtotal resection, atypical histologic features, and high cell proliferation rates correlate with recurrence.

Tau-associated neuropathology in ganglion cell tumours increases with patient age but appears unrelated to ApoE genotype.

Ganglion cell tumours, including gangliogliomas and gangliocytomas, are low grade neoplasms with a mature neuronal component. Ganglion cells within these lesions occasionally exhibit neurodegenerative changes including neurofibrillary tangles (NFT) similar to those in Alzheimer's disease. The frequency and spectrum of degenerative pathology in these lesions have not been defined, nor has their relation to patient age or factors such as apolipoprotein E (ApoE) genotype that predispose to Alzheimer's disease. We studied 72 ganglion cell tumours (61 gangliogliomas, 11 gangliocytomas) from patients 7 months to 72-years-old. Haematoxylin and eosin (H&E), silver stains (Hirano method) and immunohistochemistry for tau, alpha-synuclein and beta-amyloid were performed on formalin-fixed, paraffin-embedded tissue from surgical specimens. Tau-and silver-positive NFT and neuropil threads (NPT) were present in four of 26 ganglion cell tumours from patients over 30-years-old (ages 31, 38, 50, and 58 years). Neuronal granulovacuolar degeneration (GVD) was noted in five of 26 tumours from patients over 30-years-old (mean, 48 years). NFT, NPT, and GVD were not seen in ganglion cell tumours from patients under 30-years-old[0/46]. Cytoplasmic argentophilic bodies distinct from NFT were present in five of 26 tumours from patients over 30-years-old and in two of 46 under 30 years. Neither alpha-synuclein positive neuronal inclusions nor beta-amyloid immunoreactivity was noted in ganglion cell tumours from any age group. The distribution of ApoE genotypes was similar among those tumours that contained tau-associated neuropathology and those that did not. Neurodegenerative changes are uncommon in ganglion cell tumours, but increase in frequency with patient age. GVD, tau-positive NFT and NPT, and argentophilic bodies occur more often in ganglion cell tumours from patients over 30-yrs-old, but do not appear to be associated with a specific ApoE genotype.

Controlled local delivery of interleukin-2 by biodegradable polymers protects animals from experimental brain tumors and liver tumors.

PURPOSE: The purpose of our study was to develop an injectable polymeric system for the long-term localized delivery of bioactive interleukin-2 for antitumor immunotherapy. METHODS: IL-2 was encapsulated into gelatin and chondroitin-6-sulfate using an aqueous-based complex coacervation. CTLL-2 cells were used to measure the bioactivity of released IL-2 and radiolabeled IL-2 was used for release studies in the rat brain and mouse liver. Antitumor efficacy studies were carried out in primary (9L gliosarcoma) and metastatic (B16-F10 melanoma) brain tumor models in rats and mice, respectively, as well as a murine liver tumor model (CT26 carcinoma). Survivors of the metastatic brain tumor challenge were rechallenged with tumor in the opposite lobe of the brain to confirm that antitumor immunologic memory had developed. RESULTS: Bioactive IL-2 was released for over 2 weeks in vitro and in vivo IL-2 release showed significant IL-2 levels for up to 21 days. Polymeric IL-2 microspheres injected intratumorally were statistically more effective in protecting animals challenged with fatal tumor doses in the brain and the liver than placebo or autologous tumor cells genetically engineered to secrete IL-2. Immunologic memory was induced following IL-2 microsphere therapy in the B16-F10 brain tumor model that was capable of protecting 42% of animals from a subsequent intracranial tumor challenge, suggesting that tumor destruction was mediated by the immune system. CONCLUSIONS: Local IL-2 therapy using novel polymeric carriers. aimed at stimulating long-lasting antitumor immunity, may provide an improved method of treating a variety of cancers.

Microphthalmia transcription factor immunohistochemistry: a useful diagnostic marker in the diagnosis and detection of cutaneous melanoma, sentinel lymph node metastases, and extracutaneous melanocytic neoplasms.

BACKGROUND: Melanoma is the most lethal form of skin cancer. Diagnosis of amelanotic melanoma and detection of micrometastases in sentinel lymph nodes pose diagnostic and therapeutic dilemmas for the dermatopathologist and clinician. OBJECTIVE: The purpose of this article is to determine the utility of immunohistochemistry using antibodies specific for microphthalmia in the identification of melanocytic lesions in the skin, eye, central nervous system, and sentinel lymph nodes. METHODS: Paraffin-embedded, formalin-fixed specimens of cutaneous melanoma, including amelanotic melanoma and lentigo maligna melanoma, were stained with antibodies specific for microphthalmia. In addition, paraffin sections of extracutaneous lesions, including sentinel lymph nodes, uveal melanoma, and central nervous system melanocytomas, were stained with the specific microphthalmia antibody. RESULTS: All cutaneous melanomas stained positively with microphthalmia, as did uveal melanomas and central nervous system melanocytomas. These findings confirm the melanocytic origin of melanocytomas and uveal melanomas and demonstrate that microphthalmia staining can be used to establish melanocytic origin of neoplasms. In addition, micrometastases were easily detected in sentinel lymph nodes. CONCLUSION: Microphthalmia transcription factor immunohistochemistry is a valuable tool in the identification of melanocytic lesions in numerous sites. Use of this stain may facilitate detection of micrometastases in sentinel lymph nodes.

Novel patterns of gene expression in pituitary adenomas identified by complementary deoxyribonucleic acid microarrays and quantitative reverse transcription-polymerase chain reaction.

Pituitary adenomas account for approximately 10% of intracranial tumors, but little is known of the oncogenesis of these tumors. The identification of tumor-specific genes may further elucidate the pathways of tumor formation. We used complementary DNA microarrays to examine gene expression profiles in nonfunctioning, PRL, GH, and ACTH secreting adenomas, compared with normal pituitary. Microarray analysis showed that 128 of 7075 genes examined were differentially expressed. We then analyzed three genes with unique expression patterns and oncogenic importance by RT-real time quantitative PCR in 37 pituitaries. Folate receptor gene was significantly overexpressed in nonfunctioning adenomas but was significantly underexpressed in PRL and GH adenomas, compared with controls and to other tumors. The ornithine decarboxylase gene was significantly overexpressed in GH adenomas, compared with other tumor subtypes but was significantly underexpressed in ACTH adenomas. C-mer proto-oncogene tyrosine kinase gene was significantly overexpressed in ACTH adenomas but was significantly underexpressed in PRL adenomas. We have shown that at least three genes involved in carcinogenesis in other tissues are also aberrantly regulated in the major types of pituitary tumors. The evaluation of candidate genes that emerge from these experiments provides a rational approach to investigate those genes significant in tumorigenesis.

Pancreatic intraepithelial neoplasia and infiltrating adenocarcinoma: analysis of progression and recurrence by DPC4 immunohistochemical labeling.

Pancreatic intraepithelial neoplasia (PanIN) is thought to be a precursor lesion of infiltrating pancreatic ductal adenocarcinoma (IPA). DPC4 is a tumor-suppressor gene on chromosome 18q21.1 and is inactivated in approximately 55% of IPAs. Recently, immunohistochemical labeling using a monoclonal antibody to the Dpc4 protein has been shown to mirror DPC4 genetic status in invasive adenocarcinomas of the pancreas. In the present study, we examined the role of Dpc4 loss in neoplastic progression and recurrence. Two cases in which a PanIN clinically progressed to an invasive adenocarcinoma and a third of a patient with IPA of the head of the pancreas who later developed invasive adenocarcinoma in the tail of the pancreas were studied using Dpc4 immunolabeling. The first patient underwent pancreatic resection, which revealed PanIN-3 that lacked Dpc4 expression, and the patient developed an invasive pancreatic ductal carcinoma 10 years later that shared this loss of expression. The second patient had a pancreaticoduodenectomy for recurrent pancreatitis, and the resected pancreas contained PanIN-3 with intact Dpc4 expression. Seventeen months later, the patient developed an invasive adenocarcinoma of the distal pancreas that also had intact Dpc4 expression. In the third case, the patient underwent pancreaticoduodenectomy for an invasive ductal adenocarcinoma with negative margins. This carcinoma lacked Dpc4 expression. Three years later, resection of the pancreatic tail showed a second invasive adenocarcinoma. The cancer in the tail of the gland showed intact Dpc4 expression, suggesting it represented a second primary tumor, not a recurrence. We conclude that Dpc4 expression in PanIN can be predictive of Dpc4 expression in the subsequent invasive ductal adenocarcinoma. Additionally, Dpc4 expression can be used to differentiate recurrent or persistent adenocarcinoma from a second primary adenocarcinoma.

Intrasellar pituicytoma in a patient with other endocrine neoplasms.

Considered a neoplasm of pituicytes, pituicytoma is a rare and distinct type of glioma that arises in the suprasellar space and within the sella turcica. Only 12 previously reported cases of pituicytoma are documented in the literature. We report an intrasellar pituicytoma in a 66-year-old man presenting with symptoms and radiologic appearance indistinguishable from a nonfunctional pituitary adenoma. The patient also had a medical history significant for parathyroid adenomas and follicular carcinoma of the thyroid. The intrasellar tumor had morphologic features of a pituicytoma, with interlacing fascicles and a storiform pattern much like a benign fibrous histiocytoma. Immunoreactivity for S100 was strong, but the tumor lacked intercellular collagen type IV. The differential diagnosis of a low-grade spindle cell lesion of the sellar space is discussed, and the literature is reviewed. A summary of the clinical and pathologic features of this case, as well as the 12 previously reported cases, is presented.

Chordoid glioma: a neoplasm unique to the hypothalamus and anterior third ventricle.

BACKGROUND AND PURPOSE: Chordoid glioma is a new clinicopathologic entity that occurs in the region of the hypothalamus/anterior third ventricle. The aims of this study were to describe the characteristic radiographic features of chordoid glioma, identify specific imaging features that may enable differentiation of chordoid glioma from other suprasellar tumors, and increase neuroradiologists' awareness of this newly described tumor, facilitating prospective diagnosis. METHODS: CT scans and/or MR images of six patients with chordoid glioma were reviewed retrospectively to determine whether any characteristic radiographic features would emerge. Reports of the clinical presentation, pathologic findings, and radiographic findings of another six patients were reviewed and included, for a total patient population of 12 (mean age +/- SD, 46 +/- 13 years). RESULTS: Imaging features were strikingly similar for all tumors. In each case, the mass was ovoid, was well circumscribed, was located in the region of the hypothalamus/anterior third ventricle, and enhanced uniformly and intensely. Tumors were hyperdense to gray matter on CT scans and were isointense on T1-weighted MR images and slightly hyperintense on long-TR MR images. In two patients, vasogenic edema extended into the optic tracts, and in three, there was hydrocephalus. CONCLUSION: Chordoid glioma is a recently described unique histopathologic entity that has been added to the World Health Organization glioma classification scheme and must be included in the differential diagnosis of a suprasellar mass. Distinctive imaging features are its location, ovoid shape, hyperdensity on CT scans, and uniform intense contrast enhancement.

Chromosomal abnormalities subdivide ependymal tumors into clinically relevant groups.

Ependymoma occurs most frequently within the central nervous system of children and young adults. We determined relative chromosomal copy-number aberrations in 44 ependymomas using comparative genomic hybridization. The study included 24 intracranial and 20 spinal cord tumors from pediatric and adult patients. Frequent chromosomal aberrations in intracranial tumors were gain of 1q and losses on 6q, 9, and 13. Gain of 1q and loss on 9 were preferentially associated with histological grade 3 tumors. On the other hand, gain on chromosome 7 was recognized almost exclusively in spinal cord tumors, and was associated with various other chromosomal aberrations including frequent loss of 22q. We conclude that cytogenetic analysis of ependymomas may help to classify these tumors and provide leads concerning their initiation and progression. The relationship of these aberrations to patient outcome needs to be addressed.

Dysembryoplastic neuroepithelial tumor-like neoplasm of the septum pellucidum: a lesion often misdiagnosed as glioma: report of 10 cases.

The authors report a series of 10 low-grade neoplasms arising in the midline anteriorly in the region of the septum pellucidum with many of the histologic features of dysembryoplastic neuroepithelial tumor (DNT). The patients (five female, five male) ranged in age from 6 to 35 years (mean age, 21.5 years). The most common presenting symptoms were headache, nausea and vomiting, and visual disturbances. Radiographically, the tumors extended into the lateral ventricles from the septal region and obstructed the foramen of Monro. Varying degrees of hydrocephalus were present. The lesions were lobular, well-delineated, hypointense to brain on T1-weighted magnetic resonance imaging, and hyperintense on T2-weighted images. They were uniformly nonenhancing or showed only minimal peripheral enhancement. The tumors, in aggregate, had the histologic features of DNT. These included a mucin-rich background, oligodendrocyte-like cells, "floating neurons," and a "specific glioneuronal element." Seven patients underwent gross total resection and two underwent subtotal resection. No patients received adjuvant chemotherapy or radiotherapy. On follow-up (n = 6; median, 14 months), all tumors had either not recurred or were radiologically stable. On the basis of both neuroimaging and histopathology, DNT-like lesions should be considered in the differential diagnosis of midline intraventricular tumors in children and young adults. Distinction from more aggressive neoplasms is essential because these tumors appear to behave in a benign fashion.

Ectopic cerebellum presenting as a suprasellar mass in infancy: implications for cerebellar development.

A 4-month-old infant with a history of nasopharyngeal teratoma developed progressive optic neuropathy. Neuroimaging studies demonstrated a solid, isointense, suprasellar mass impinging on optic nerves and chiasm superiorly. The mass was subtotally resected. No attachment of the mass to brain stem or cerebellar structures was noted. Histological examination identified the tissue as developing cerebellum. The cytoarchitecture and cellular constituents of the cerebellar tissue were only slightly distorted. All cerebellar cortical constituents were arranged anatomically, and an external granular cell layer was present superficially. The latter was actively proliferating and appropriately cellular for the infant's age. The clinical presentation of ectopic cerebellum as a suprasellar mass in an infant is highly unusual. Moreover, this example illustrates the ability of cerebellar tissue to mature appropriately in a site distant from the posterior fossa, removed from ascending and descending afferent projections. Intrinsic signaling mechanisms appear sufficient to direct histogenesis in developing cerebellar cortex.

Differential neuronal localizations and dynamics of phosphorylated and unphosphorylated type 1 inositol 1,4,5-trisphosphate receptors.

Type 1 inositol 1,4,5-trisphosphate receptors are phosphorylated by cyclic-AMP-dependent protein kinase A at serines 1589 and 1755, with serine 1755 phosphorylation greatly predominating in the brain. Inositol 1,4,5-trisphosphate receptor protein kinase A phosphorylation augments Ca(2+) release. To assess type 1 protein kinase A phosphorylation dynamics in the intact organism, we developed antibodies selective for either serine 1755 phosphorylated or unphosphorylated species. Immunohistochemical studies reveal marked variation in localization. For example, in the hippocampus the phosphorylated type 1 inositol 1,4,5-trisphosphate receptor is restricted to CA1, while the unphosphorylated receptor occurs ubiquitously in CA1-CA3 and dentate gyrus granule cells. Throughout the brain the phosphorylated type 1 inositol 1,4,5-trisphosphate receptor is selectively enriched in dendrites, while the unphosphorylated receptor predominates in cell bodies. Focal cerebral ischemia in rats and humans is associated with dephosphorylation of type 1 inositol 1,4,5-trisphosphate receptors, and glutamatergic excitation of cerebellar Purkinje cells mediated by ibogaine elicits dephosphorylation of type 1 inositol 1,4,5-trisphosphate receptors that precedes evidence of excitotoxic neuronal degeneration. We have demonstrated striking variations in regional and subcellular distribution of inositol 1,4,5-trisphosphate receptor phosphorylation that may influence normal physiological intracellular Ca(2+) signaling in rat and human brain. We have further shown that the subcellular distribution of inositol 1,4,5-trisphosphate receptor phosphorylation in neurons is regulated by excitatory neurotransmission, as well as excitotoxic insult and neuronal ischemia-reperfusion. Phosphorylation dynamics of type 1 inositol 1,4,5-trisphosphate receptors may modulate intracellular Ca(2+) release and influence the cellular response to neurotoxic insults.

Pediatric chordoid glioma with chondroid metaplasia.

Chordoid gliomas are uncommon primary brain tumors that arise in the region of the third ventricle. Reports of this entity to date have been limited to adults. We present a case of a chordoid glioma arising in the hypothalamic/third ventricle region of a 12-year-old male who presented with visual symptoms. The neoplasm consisted of cords and clusters of well-differentiated, spindled-to-rounded cells containing abundant eosinophilic cytoplasm within a prominent mucinous matrix. Unlike other chordoid gliomas, this lesion contained islands and sheets showing cartilaginous differentiation intermixed with the glial component. A graded transition between neoplastic glial and chondroid regions was evident, and cells in both regions were strongly immunoreactive for GFAP and S-100. Cartilaginous metaplasia is infrequent in gliomas, but occurs most often in pediatric neoplasms of the midline such as this chordoid glioma. Thus, chondroid metaplasia represents an unusual histopathologic feature of chordoid glioma-in this case, presenting in a child.

Extraneural metastatic glioblastoma after interstitial brachytherapy.

PURPOSE: This is a report of 3 cases of extraneural metastasis of glioblastoma after interstitial radiation and assessment of pertinent literature addressing concern over an increased risk of these events with this therapy. METHODS AND MATERIALS: In a series of 82 patients treated with (125)I brachytherapy for primary malignant brain tumors over a 7-year interval, 3 cases of extraneural glioblastoma were identified. The multicatheter technique for delivery of (125)I sources was utilized in all. Extraneural metastases were documented by imaging studies or biopsy. Over the same period, 310 patients with primary malignant brain tumors were treated without brachytherapy. RESULTS: Biopsy-proven scalp and skull metastases occurred in 2 patients, at 3 and 8 months following brachytherapy. Each developed radiographic evidence of systemic metastases at 7 and 14 months postbrachytherapy, respectively. The third patient developed biopsy-proven cervical node involvement 4 months after brachytherapy. No patients with malignant gliomas undergoing craniotomy or stereotactic biopsy, but not brachytherapy, during the same time period developed extraneural metastases. Incidence in previously reported series commenting on this otherwise rare process range from 0% to 4.3%. The incidence of extraneural metastases in this series is 3.7% (3/82) and is comparable to those reports. CONCLUSIONS: Percutaneous catheter-delivered brachytherapy may be associated with an increased incidence of extraneural metastatic glioma.

Intracranial paracrine interleukin-2 therapy stimulates prolonged antitumor immunity that extends outside the central nervous system.

To explore the potential efficacy of local cytokine delivery against tumors in the central nervous system (CNS), C57BL6 mice were simultaneously given intracranial injections of tumor challenge and of irradiated B16F10 melanoma cells transduced to secrete interleukin-2 (IL-2). Intracranial IL-2 therapy generated antitumor responses capable of extending the survival of animals that received simultaneous intracranial tumor challenge either locally or at distant sites in the brain. Nontransduced melanoma cells had little effect. Animals that survived intracranial IL-2 therapy and tumor challenge showed prolonged survival compared with controls when challenged with a second tumor dose 70 days after initial treatment. In addition, animals that rejected intracranial tumors were also protected from tumor growth upon rechallenge at sites outside the CNS (i.e., subcutaneous tumor challenge). Conversely, identical or 10-fold larger doses of IL-2-transduced cells administered by subcutaneous injection failed to generate protection against intracranial tumor challenges. Elimination of T-cell and natural killer (NK) subsets using gene knockout mice and antibody-depletion techniques demonstrated that NK cells were most important for the initial antitumor response, whereas CD4+ T-cells were not necessary. These studies demonstrate that local IL-2 therapy in the brain not only generates an immediate local antitumor immune response, but also establishes long-term immunologic memory capable of eliminating subsequent tumor challenges within and outside of the CNS. Furthermore, the antitumor response to paracrine IL-2 in the brain differed significantly from that in the flank, suggesting that the intrinsic CNS cells involved in initiating immunity within the brain have different cytokine requirements from their peripheral counterparts.