Marsha A. Widmayer

A prospective, randomized, controlled cervical fusion study using recombinant human bone morphogenetic protein-2 with the CORNERSTONE-SR™ allograft ring and the ATLANTIS™ anterior cervical plate

Study Design. A prospective, randomized, pilot clinical trial compared recombinant human bone morphogenetic protein-2 (rhBMP-2) with iliac crest autograft bone for the treatment of human cervical disc disease. Objective. To examine the safety and effectiveness of using INFUSE™ Bone Graft (rhBMP-2 applied to an absorbable collagen sponge), as compared with an autogenous iliac crest bone graft placed inside the CORNERSTONE-SR™ fibular allograft, in anterior cervical discectomy and interbody fusion. Summary of Background Data. Recombinant human bone morphogenetic protein-2 is an osteoinductive protein that induces a reliable fusion in the lumbar spine, but it has not been studied in patients with degenerative cervical disc disease. Methods. For this study, 33 patients with degenerative cervical disc disease were randomly assigned to investigational or control groups. The investigational group received a fibular allograft (CORNERSTONE-SR™ Allograft Ring) with an rhBMP-2–laden collagen carrier inside the graft along with an ATLANTIS™ anterior cervical plate. The control group received a fibular allograft with cancellous iliac crest autograft placed inside it, along with an ATLANTIS anterior cervical plate. The patients underwent plain radiographs at 6 weeks, then at 3, 6, 12, and 24 months, and CT scans at 3 and 6 months after surgery. They also completed general health profiles and self-evaluation scales. Adverse events were evaluated for severity, duration, association with the implant, and the need for a second surgical procedure. Results. All the patients evaluated had solid fusions 6, 12, and 24 months after surgery. There were no device-related adverse events. At 24 months, the investigational group had mean improvement superior to that of the control group in neck disability and arm pain scores (P < 0.03 each). Conclusions. This pilot study demonstrates the feasibility of using rhBMP-2 safely and effectively in the cervical spine.

Evaluation of delayed treatment of focal cerebral ischemia with three selective kappa-opioid agonists in cats.

The purpose of this study was to determine the therapeutic efficacy of three K-opioid agonists used for delayed treatment of experimental focal cerebral ischemia. Methods Forty halothane-anesthetized cats underwent permanent occlusion of the right intracranial internal carotid, middle cerebral, and anterior cerebral arteries via a transorbital, microsurgical approach. Six hours after occlusion, animals received a blinded bolus injection, and a subcutaneous osmotic pump was implanted to provide continuous release for 7 days. The injection and pump contained either saline or one of three K-agonists: dynorphin (1-13), U-50,488, or DuP E3800. Survival, neurological function, tissue damage, and brain weight were assessed. Results As a group, K-agonist-treated animals had higher survival (P<.02), less tissue damage (P<.02), and lower brain weight (P<.05) than saline controls. U-50,488 more effectively improved survival (P<.03) than dynorphin (P<.07) or E3800 (P<.07). Each of the three K compounds improved tissue damage (dynorphin, P<.02; U-50,488, P<.05; E3800, P<.05). Greater improvement in neurological function was seen after treatment with dynorphin (P<.05) than with U-50,488 (P<.6) or E3800 (P<.1). The only significant reduction in brain weight was seen after dynorphin treatment (P<.01). Conclusions Compounds that act at the K subclass of opiate receptors are effective in increasing survival, improving neurological function, and decreasing tissue damage and edema in a cat model of focal cerebral ischemia. The current study provides support for the benefits of treatment of acute cerebrovascular ischemia with K-opioid agonists. The agents may prove to be of superior clinical utility because of efficacy even when administered 6 hours after the onset of stroke.

Development of a model for Parkinson's disease in sheep using unilateral intracarotid injection of MPTP via slow continuous infusion

The effects of unilateral intracarotid administration of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) in sheep were studied with the goal of producing a non-primate, large animal model of Parkinsons Disease. Adult female sheep were given an acute (over 30 min) or chronic (over 1 week) injection of MPTP (0.4-5.0 mg/kg) via the common carotid artery. Both methods produced parkinsonian-like behavior. Turning contralateral to the side of injection was induced by apomorphine (APO) in both groups. However, amphetamine (AMP) induced ipsilateral turning only in the chronic treatment group. Acute and chronic MPTP treatment resulted in a loss of substantia nigra tyrosine hydroxylase immunoreactive (THIR) neurons with a significantly greater loss ipsilateral to the injection in each treatment group (acute p < 0.05; chronic p < 0.01). Caudate dopamine (DA) was depleted in both treatment groups, although the difference between ipsilateral and contralateral DA content was significant only in the chronic treatment group (p < 0.05). The best results were seen in those animals with chronic infusion with the occipital artery occluded to prevent entry of drug into the posterior circulation with subsequent bilateral distribution. Use of slow and continuous intracarotid administration of MPTP with the ipsilateral occipital artery occluded can prevent some of the bilateral effects of acute treatment, and results in statistically significant ipsilateral reduction of THIR neurons in the substantia nigra and reduction of tissue levels of DA in the caudate nucleus. Such treatment produces appropriate turning responses to both AMP and APO challenge not seen in the acute treatment group, and appears to be an effective method of producing parkinsonian-like behavior in a large animal.

Suppression of post-ischemic-induced fos protein expression by an antisense oligonucleotide to c-fos mRNA leads to increased tissue damage.

Activation of c-fos, an immediate early gene, and the subsequent upregulation of Fos protein expression occur following neural injury, including focal cerebral ischemia (fci). Fos and Jun form a heterodimer known as activator protein 1, which regulates the expression of many late effector genes. To study the downstream effects of c-fos expression following ischemia, we suppressed the translation of c-fos by administering an antisense oligonucleotide (AO) to c-fos mRNA. Eighteen hours prior to fci, male, Long Evans (LE) rats received intraventricular injections of AO, mismatched AO (MS) or artificial cerebrospinal fluid (aCSF). Fci was induced by permanent right middle cerebral artery occlusion. At 24-h post-occlusion, neurological function was assessed, and the animals were sacrificed. The brains were removed and stained with triphenyltetrazolium chloride for infarct volume determination. Fos immunohistochemistry was performed in separate animals to determine the effects of treatment on Fos expression number of Fos positive cells. AO administration reduced the number of cells with fci-induced Fos expression by approximately 75%. No differences in neurological scores existed between any of the groups. AO-treated LE developed larger infarcts (40.1+/-1.0%, mean+/-S.D., p<0.001) than MS- or aCSF-treated controls (34.3+/-1.0%, 34.6+/-1.0%, respectively). These results suggest that c-fos activation and subsequent Fos protein expression exerts a neuroprotective effect, which is likely via upregulation of neurotrophins, following focal cerebral ischemia. This response, among others, may contribute to brain adaptation to injury that underlies functional recovery after stroke.

Antibody-targeted nanovectors for the treatment of brain cancers

Introduced here is the hydrophilic carbon clusters (HCCs) antibody drug enhancement system (HADES), a methodology for cell-specific drug delivery. Antigen-targeted, drug-delivering nanovectors are manufactured by combining specific antibodies with drug-loaded poly(ethylene glycol)-HCCs (PEG-HCCs). We show that HADES is highly modular, as both the drug and antibody component can be varied for selective killing of a range of cultured human primary glioblastoma multiforme. Using three different chemotherapeutics and three different antibodies, without the need for covalent bonding to the nanovector, we demonstrate extreme lethality toward glioma, but minimal toxicity toward human astrocytes and neurons.

Effects of halothane, α-chloralose, and pCO2 on injury volume and CSF β-endorphin levels in focal cerebral ischemia

Anesthetic agent, arterial pCO2 level, and opioid peptides have all been implicated in the pathophysiology of experimental stroke models. The effects of halothane, α-chloralose, and differing concentrations of arterial pCO2 on injury volume and CSF β-endorphin levels were studied in a feline model of experimental focal cerebral ischemia. The type of anesthetic agent used had no effect on injury volume following 6 h of focal cerebral ischemia. Over a 6-h period, β-endorphin levels significantly increased from 10.1±5.0 fmol/mL at zero time to 14.4±7.2 fmol/mL at 6 h under halothane anesthesia (p<0.05), whereas they did not significantly change (10.1±6.7 to 7.8 ±4.7 fmol/mL) under α-chloralose anesthesia. In contrast, hypercapnia had no effect on β-endorphin levels, but significantly increased injury volume from 30.6±5.7% of the ipsilateral hemisphere under normocapnic conditions to 37.1±5.9% under hypercapnic conditions (p<0.05). These results suggest that hypercapnia increases injury volume in a feline model of focal cerebral ischemia, and pCO2 should be controlled in experimental focal cerebral ischemia models.

U50488 reduces the severity of tissue damage in a rabbit model of focal cerebral ischemia.

Many pharmacotherapies for stroke that have been successful in the laboratory have proven to be ineffective in the clinical setting, often because patients do not arrive for treatment until hours after the onset of the ischemic insult. Kappa opioid treatment of cerebral ischemia has been successful in the cat and mouse with treatment delays of up to 6 h. The purpose of the present study was to develop a model of delayed kappa opioid treatment of cerebral ischemia in the rabbit. Fourteen rabbits underwent permanent, unilateral occlusion of the internal carotid, middle cerebral, and anterior cerebral arteries via a transorbital, microsurgical approach. At 6 h postocclusion, animals received a blinded bolus injection and continuous infusion of either saline or the kappa agonist, U50488. Survival was not improved after U50488 treatment. U50488 treatment did, however, reduce areas of severe tissue damage and increase areas of modest tissue damage. This suggests U50488 arrested the progression of damage from noninfarcted to fully infarcted tissue. The present results show beneficial effects of delayed treatment with kappa agonists in a species similar in vasculature to humans, and much less costly than primates or cats.

Penetration of dynorphin 1-13 across the blood-brain barrier.

Previous studies have demonstrated neuroprotective effects of the opioid peptide dynorphin (dyn) 1-13 in focal cerebral ischemia. The passage of dyn 1-13 across the blood-brain barrier (BBB) was studied by a modification of the Oldendorf technique in the normal rat and cat, as well as in a feline model of experimentally induced focal cerebral ischemia. In the rat, dyn 1-13 penetration of the BBB could not be detected by this technique, even in the presence of peptidase inhibitors. In contrast, dyn 1-13 did cross the BBB into the normal cat hippocampus, cortex and cerebellum. The passage of dyn 1-13 across the BBB was greater in cats with experimentally induced focal cerebral ischemia. Some of the tritium-labeled material which crossed the BBB was confirmed by high performance liquid chromatography to be dyn 1-13. These studies support the hypothesis that the therapeutic effects observed after the peripheral administration of dyn 1-13 to cats with focal cerebral ischemia can be produced by a central mechanism of action.

Increased intracranial pressure is associated with elevated cerebrospinal fluid ADH levels in closed-head injury

Abstract Objectives: Head injury frequently results in increased intracranial pressure and brain edema. Investigators have demonstrated that ischemic injury causes an increase in cerebrospinal fluid (CSF) levels of antidiuretic hormone (ADH); increased CSF ADH levels exacerbate cerebral edema, and inhibition of the ADH system with specific ADH antagonists reduces cerebral edema. The current study was designed to test the hypothesis that elevated levels of ADH are present in the CSF of subjects with head injury. Methods: Ventricular CSF and blood samples were taken from 11 subjects with head injury and 12 subjects with no known head trauma or injury. ADH levels were analyzed using radioimmunoassay. Severity of increased intracranial pressure (ICP) was rated in head-injured subjects using a four-point ordinal scale, based on which treatments were necessary to reduce ICP. Results: Subjects with head injury had higher CSF (3.2 versus 1.2 pg/ml; P<0.02) and plasma (4.1 versus 1.4 pg/ml; P<0.02) levels of ADH than did control subjects. In head-injured subjects, CSF ADH levels positively correlated with severity of ICP. Discussion: The results of this study suggest that ADH plays a role in brain edema associated with closed head injury.