Suppression of glioma growth in vitro and in vivo by glia maturation factor.

Glia maturation factor (GMF), a 14,000 Mr acidic protein of the brain, is capable of promoting differentiation of cultured astroblasts. In this study we report the effect of GMF on two glioma cell lines: the C6 line, of rodent origin, and the HG-1 line, of human origin. When tested in culture, GMF promotes the initial growth of the two cell lines when the cells are sparse but limits proliferation by restoring contact inhibition when the cells are confluent. Cell cycle analysis confirms the arrest of the cells at the G0/G1 phase when the tumor cells are contact inhibited by GMF. When C6 cells are inoculated into the athymic (nude) mice at a s.c. site, a single solid tumor grows out with a 100% take. Intraperitoneal injection of GMF leads to the slowing down of tumor growth. That the in vivo effect of GMF is not due to cytotoxicity is evidenced by the lack of necrosis and by the appearance of more mature astrocytic cells in the tumors. The results lend support to the concept of GMF as a cellular regulator and suggest the therapeutic potential of GMF for brain tumors.

Initial clinical experience with frameless stereotactic radiosurgery: analysis of accuracy and feasibility.

PURPOSE: To report on preliminary clinical experience with a novel image-guided frameless stereotactic radiosurgery system. METHODS AND MATERIALS: Fifteen patients ranging in age from 14 to 81 received radiosurgery using a commercially available frameless stereotactic radiosurgery system. Pathologic diagnoses included metastases (12), recurrent primary intracranial sarcoma (1), recurrent central nervous system (CNS) lymphoma (1), and medulloblastoma with supratentorial seeding (1). Treatment accuracy was assessed from image localization of the stereotactic reference array and reproducibility of biteplate reseating. We chose 0.3 mm vector translation error and 0.3 degree rotation about each axis as the maximum tolerated misalignment before treating each arc. RESULTS: The biteplates were found on average to reseat with a reproducibility of 0.24 mm. The mean registration error from CT localization was found to be 0.5 mm, which predicts that the average error at isocenter was 0.82 mm. No patient treatment was delivered beyond the maximum tolerated misalignment. The radiosurgery treatment was delivered in approximately 25 min per patient. CONCLUSION: Our initial clinical experience with stereotactic radiotherapy using the infrared camera guidance system was promising, demonstrating clinical feasibility and accuracy comparable to many frame-based systems.

Endogenous immunoreactive glia maturation factor-like molecule in astrocytes and glioma cells.

Using the monoclonal antibody G2-09 raised against bovine glia maturation factor (GMF), we conducted a survey of GMF-like immunoreactivity in various cell types. Of all the normal and neoplastic cells tested, only extracts from astroblasts, gliomas, Schwann cells and schwannomas, but not their conditioned media, possessed endogenous GMF-like immunoreactivity. The presence of immunoreactive GMF correlated well with GMF bioactivity. Using the same monoclonal antibody, the GMF-like factor in astroblasts and C6 glioma cells was characterized by immunofluorescence, immunoadsorption and immunoblotting. Immunofluorescence confirmed the intracellular location of GMF. Immunoadsorption completely eliminated the GMF-like bioactivity from the cell extracts. Immunoblotting identified a protein band having a mol. wt. of 14,000 Da. Thus, the evidence strongly supports the argument that the GMF-like factor in astroblasts and C6 cells is identical with GMF from the bovine brain. In order to explain the fact that astroblasts and C6 cells are both the source and targets of GMF, we propose the hypothesis that GMF functions as an injury signal, being released from the injured glia and serving as a stimulant for gliosis in the neighboring intact glia.

Endogenous immunoreactive glia maturation factor-like molecule in cultured rat Schwann cells.

Using the monoclonal antibody G2-09 raised against bovine glia maturation factor (GMF), we demonstrated that cultured rat Schwann cells and Schwannoma cells, but not their conditioned media, possessed endogenous GMF-like immunoreactivity. The presence of immunoreactive GMF correlated well with GMF bioactivity. The GMF-like factor in Schwann cells was characterized by immunodotting, immunofluorescence, immunoadsorption and immunoblotting. Immunofluorescence confirmed the intracellular location of GMF. Immunoadsorption completely eliminated the GMF-like bioactivity from the cell extracts. Immunoblotting identified a protein band with a molecular weight of 14,000. Thus, the evidence strongly supports the argument that the GMF-like factor in rat Schwann cells is identical with GMF from the bovine brain. The GMF-like molecule in Schwannoma cells showed properties similar to those in Schwann cells, but for unknown reasons was not detectable by immunofluorescence. The presence of GMF in cultured rat Schwann cells suggests that the factor may play a role in the peripheral nervous system.

Induction of cytoskeletal alterations in C6 glioma by glia maturation factor.

C6 rat glioma cells respond to glia maturation factor (GMF) with characteristic morphological alterations. Observed under phase-contrast microscopy, the cells changed from a rounded morphology in random formation to a spindle-shaped appearance in parallel arrays. Observed under scanning electron microscopy, GMF led to a decrease in the number of microvilli and cell surface knobs. Transmission electron microscopy demonstrated the appearance of numerous microtubules aligned with the long axis of the cells after GMF stimulation. The change in cell shape and histotypic pattern was inhibited by vinblastin, further implicating the involvement of microtubules. Immunofluorescence using anti-alpha-tubulin revealed a well-defined cytoskeletal system in GMF-stimulated cells but not in the control cells. Finally, an increase in tubulin was confirmed with enzyme-linked immunosorbent assay (ELISA) on extracts from these cultures. The findings indicate that morphological alterations induced by GMF are associated with changes in the quantity and arrangement of microtubules.

Cytotoxicity of cis-parinaric acid in cultured malignant gliomas.

The cytotoxic effects of cis-parinaric acid, a plant-derived 18-carbon polyunsaturated fatty acid, were assessed in vitro on normal and neoplastic glia. After being incubated for 24 hours in the presence of 12 mumol/L cis-parinaric acid, 36B10 glioma cultures demonstrated nearly 90% toxicity (unpaired Student's t test, P < 0.001). Similar results were obtained after the exposure of C6 rat glioma cultures, A172 human glioma cultures, and U-937 human monocytic leukemia cultures to cis-parinaric acid. In contrast, fetal rat astrocytes incubated with 12 mumol/L cis-parinaric acid demonstrated no significant toxicity (3% reduction, P = 0.12); fetal rat astrocytes showed only 20% toxicity after exposure to 40 mumol/L cis-parinaric acid (P = 0.001). The cytotoxic effects of cis-parinaric acid were antagonized with the addition of equimolar concentrations of alpha-tocopherol. Enzyme immunoassay of treated 36B10 glioma supernatant fluid for 8-isoprostane (a known oxidative metabolite) demonstrated a 10-fold increase of 8-isoprostane over 24 hours (123.0 +/- 10.3 versus 10.0 +/- 0.7 pg/ml for control, P < 0.001). These studies indicate that cis-parinaric acid may be significantly cytotoxic to malignant glioma cells in concentrations that spare normal astrocytes and that the mechanism of cytotoxicity is related to an oxidative process. The selective cytotoxic effect of cis-parinaric acid we describe represents the first step in the development of new chemotherapeutic agents for gliomas; these new agents act by preferentially enhancing lipid peroxidation in neoplastic cells.

Tumor Fas (APO-1/CD95) up-regulation results in increased apoptosis and survival times for rats with intracranial malignant gliomas.

OBJECTIVE: The cellular "death" receptor Fas has been proposed to be a potential specific target for anti-glioma therapy. However, little is known regarding the effects of Fas expression on glioma viability in vivo. The goal of this study was to clarify the relationships among Fas expression, apoptosis, and survival rates for high-grade astrocytomas. METHODS: Fas expression was measured in several human glioblastoma multiforme cell lines and a malignant rat glioma cell line (36B10), before and after Fas up-regulation by gene transfer. Expression was correlated with the degree of Fas-mediated cytotoxicity and apoptosis induced after Fas activation. Subsequently, rats underwent intracranial implantation of either wild-type or genetically altered 36B10 cell lines, for study of the effects of Fas up-regulation on survival rates. RESULTS: Low levels of cell surface Fas expression in glioblastomas multiforme were correlated with their limited susceptibility to Fas-mediated cytotoxicity. Through Fas receptor up-regulation, relationships among increased Fas expression, Fas-mediated cytotoxicity, and apoptosis were demonstrated. The percentage of cells undergoing apoptosis after exposure to a Fas ligand-producing cell line increased from 4% in the sham-transfected line (36B10-) to 27% in the Fas-transfected line (36B10-Fas). After intracranial implantation of these tumors into rats, the median survival time increased significantly from 14 days (36B10 and 36B10-) to 24.5 days (36B10-Fas), which represents a 75% increase in the survival time for the greater Fas-expressing group (P = 0.0005). CONCLUSION: It seems that the overall low rate of apoptosis in high-grade astrocytomas is related to low levels of cell surface Fas expression. With increases in cellular Fas expression, rates of Fas-mediated apoptosis and survival rates were increased.

Cervical spine immobilization before admission to the hospital.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: All trauma patients with a cervical spinal column injury or with a mechanism of injury having the potential to cause cervical spine injury should be immobilized at the scene and during transport by using one of several available methods. A combination of a rigid cervical collar and supportive blocks on a backboard with straps is effective in limiting motion of the cervical spine and is recommended. The long-standing practice of attempted cervical spine immobilization using sandbags and tape alone is not recommended.

Transportation of patients with acute traumatic cervical spine injuries.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Expeditious and careful transport of patients with acute cervical spine or spinal cord injuries is recommended, from the site of injury by the most appropriate mode of transportation available to the nearest capable definitive care medical facility.

Clinical assessment after acute cervical spinal cord injury.

UNLABELLED: NEUROLOGICAL EXAMINATION: STANDARDS: There is insufficient evidence to support neurological examination standards. GUIDELINES: There is insufficient evidence to support neurological examination guidelines. OPTIONS: The American Spinal Injury Association international standards for neurological and functional classification of spinal cord injury are recommended as the preferred neurological examination tool for clinicians involved in the assessment and care of patients with acute spinal cord injuries. FUNCTIONAL OUTCOME ASSESSMENT: STANDARDS: There is insufficient evidence to support functional outcome assessment standards. GUIDELINES: The Functional Independence Measure is recommended as the functional outcome assessment tool for clinicians involved in the assessment and care of patients with acute spinal cord injuries. OPTIONS: The modified Barthel index is recommended as a functional outcome assessment tool for clinicians involved in the assessment and care of patients with acute spinal cord injuries.

Radiographic assessment of the cervical spine in asymptomatic trauma patients.

STANDARDS: Radiographic assessment of the cervical spine is not recommended in trauma patients who are awake, alert, and not intoxicated, who are without neck pain or tenderness, and who do not have significant associated injuries that detract from their general evaluation.

Radiographic assessment of the cervical spine in symptomatic trauma patients.

STANDARDS: A three-view cervical spine series (anteroposterior, lateral, and odontoid views) is recommended for radiographic evaluation of the cervical spine in patients who are symptomatic after traumatic injury. This should be supplemented with computed tomography (CT) to further define areas that are suspicious or not well visualized on the plain cervical x-rays. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: It is recommended that cervical spine immobilization in awake patients with neck pain or tenderness and normal cervical spine x-rays (including supplemental CT as necessary) be discontinued after either a) normal and adequate dynamic flexion/extension radiographs, or b) a normal magnetic resonance imaging study is obtained within 48 hours of injury. Cervical spine immobilization in obtunded patients with normal cervical spine x-rays (including supplemental CT as necessary) may be discontinued a) after dynamic flexion/extension studies performed under fluoroscopic guidance, or b) after a normal magnetic resonance imaging study is obtained within 48 hours of injury, or c) at the discretion of the treating physician.

Initial closed reduction of cervical spine fracture-dislocation injuries.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. Early closed reduction of cervical spine fracture-dislocation injuries with craniocervical traction is recommended to restore anatomic alignment of the cervical spine in awake patients. Closed reduction in patients with an additional rostral injury is not recommended. Patients with cervical spine fracture-dislocation injuries who cannot be examined during attempted closed reduction, or before open posterior reduction, should undergo magnetic resonance imaging (MRI) before attempted reduction. The presence of a significant disc herniation in this setting is a relative indication for a ventral decompression before reduction. MRI study of patients who fail attempts at closed reduction is recommended. Prereduction MRI performed in patients with cervical fracture dislocation injury will demonstrate disrupted or herniated intervertebral discs in one-third to one-half of patients with facet subluxation. These findings do not seem to significantly influence outcome after closed reduction in awake patients; therefore, the usefulness of prereduction MRI in this circumstance is uncertain.

Pharmacological therapy after acute cervical spinal cord injury.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Treatment with methylprednisolone for either 24 or 48 hours is recommended as an option in the treatment of patients with acute spinal cord injuries that should be undertaken only with the knowledge that the evidence suggesting harmful side effects is more consistent than any suggestion of clinical benefit. GM-1 GANGLIOSIDE: STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Treatment of patients with acute spinal cord injuries with GM-1 ganglioside is recommended as an option without demonstrated clinical benefit.

Blood pressure management after acute spinal cord injury.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Hypotension (systolic blood pressure <90 mmHg) should be avoided if possible or corrected as soon as possible after acute spinal cord injury. Maintenance of mean arterial blood pressure at 85 to 90 mmHg for the first 7 days after acute spinal cord injury to improve spinal cord perfusion is recommended.

Deep venous thrombosis and thromboembolism in patients with cervical spinal cord injuries.

STANDARDS: Prophylactic treatment of thromboembolism in patients with severe motor deficits due to spinal cord injury is recommended. The use of low-molecular-weight heparins, rotating beds, adjusted dose heparin, or a combination of modalities is recommended as a prophylactic treatment strategy. Low-dose heparin in combination with pneumatic compression stockings or electrical stimulation is recommended as a prophylactic treatment strategy. GUIDELINES: Low-dose heparin therapy alone is not recommended as a prophylactic treatment strategy. Oral anticoagulation alone is not recommended as a prophylactic treatment strategy. OPTIONS: Duplex Doppler ultrasound, impedance plethysmography, and venography are recommended for use as diagnostic tests for deep venous thrombosis in the spinal cord-injured patient population. A 3-month duration of prophylactic treatment for deep venous thrombosis and pulmonary embolism is recommended. Vena cava filters are recommended for patients who do not respond to anticoagulation or who are not candidates for anticoagulation therapy and/or mechanical devices.

Nutritional support after spinal cord injury.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Nutritional support of patients with spinal cord injuries is recommended. Energy expenditure is best determined by indirect calorimetry in these patients because equation estimates of energy expenditure and subsequent caloric need tend to be inaccurate.

Management of pediatric cervical spine and spinal cord injuries.

STANDARDS: There is insufficient evidence to support diagnostic standards. GUIDELINES: In children who have experienced trauma and are alert, conversant, have no neurological deficit, no midline cervical tenderness, and no painful distracting injury, and are not intoxicated, cervical spine x-rays are not necessary to exclude cervical spine injury and are not recommended. In children who have experienced trauma and who are either not alert, nonconversant, or have neurological deficit, midline cervical tenderness, or painful distracting injury, or are intoxicated, it is recommended that anteroposterior and lateral cervical spine x-rays be obtained. OPTIONS: In children younger than age 9 years who have experienced trauma, and who are nonconversant or have an altered mental status, a neurological deficit, neck pain, or a painful distracting injury, are intoxicated, or have unexplained hypotension, it is recommended that anteroposterior and lateral cervical spine x-rays be obtained. In children age 9 years or older who have experienced trauma, and who are nonconversant or have an altered mental status, a neurological deficit, neck pain, or a painful distracting injury, are intoxicated, or have unexplained hypotension, it is recommended that anteroposterior, lateral, and open-mouth cervical spine x-rays be obtained. Computed tomographic scanning with attention to the suspected level of neurological injury to exclude occult fractures or to evaluate regions not seen adequately on plain x-rays is recommended. Flexion/extension cervical x-rays or fluoroscopy may be considered to exclude gross ligamentous instability when there remains a suspicion of cervical spine instability after static x-rays are obtained. Magnetic resonance imaging of the cervical spine may be considered to exclude cord or nerve root compression, evaluate ligamentous integrity, or provide information regarding neurological prognosis. STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Thoracic elevation or an occipital recess to prevent flexion of the head and neck when restrained supine on an otherwise flat backboard may allow for better neutral alignment and immobilization of the cervical spine in children younger than 8 years because of the relatively large head in these younger children and is recommended. Closed reduction and halo immobilization for injuries of the C2 synchondrosis between the body and odontoid is recommended in children younger than 7 years. Consideration of primary operative therapy is recommended for isolated ligamentous injuries of the cervical spine with associated deformity.

Diagnosis and management of traumatic atlanto-occipital dislocation injuries.

STANDARDS: There is insufficient evidence to support diagnostic standards. GUIDELINES: There is insufficient evidence to support diagnostic guidelines. OPTIONS: A lateral cervical x-ray is recommended for the diagnosis of atlanto-occipital dislocation. If a radiological method for measurement is used, the basion-axial interval-basion-dental interval method is recommended. The presence of upper cervical prevertebral soft tissue swelling on an otherwise nondiagnostic plain x-ray should prompt additional imaging. If there is clinical suspicion of atlanto-occipital dislocation, and plain x-rays are nondiagnostic, computed tomography or magnetic resonance imaging is recommended, particularly for the diagnosis of non-Type II dislocations. STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Treatment with internal fixation and arthrodesis using one of a variety of methods is recommended. Traction may be used in the management of patients with atlanto-occipital dislocation, but it is associated with a 10% risk of neurological deterioration.

Isolated fractures of the atlas in adults.

STANDARDS: There is insufficient evidence to support treatment standards. GUIDELINES: There is insufficient evidence to support treatment guidelines. OPTIONS: Treatment options in the management of isolated fractures of the atlas are based on the specific atlas fracture type. It is recommended that isolated fractures of the atlas with an intact transverse atlantal ligament be treated with cervical immobilization alone. It is recommended that isolated fractures of the atlas with disruption of the transverse atlantal ligament be treated with either cervical immobilization alone or surgical fixation and fusion.