Examination of several potential mechanisms for the negative outcome in a clinical stroke trial of enlimomab, a murine anti-human intercellular adhesion molecule-1 antibody: a bedside-to-bench study.

BACKGROUND AND PURPOSE: Enlimomab, a murine monoclonal anti-human intercellular adhesion molecule (ICAM)-1 antibody, had a negative outcome in a multicenter acute-stroke trial. We did a bedside-to-bench study in standardized rat stroke models to explore mechanisms for these untoward results. METHODS: After focal brain ischemia in Wistar rats and spontaneously hypertensive rats (SHR), we administered murine anti-rat ICAM-1 antibody (1A29), subclass-matched murine immunoglobulin (IgG1), or vehicle intravenously. To examine whether rat anti-mouse antibodies were generated against the mouse protein and whether these were deleterious, we sensitized Wistar rats with 1A29 or vehicle 7 days before surgery. Infarct volume, tissue myeloperoxidase activity, neutrophil CD11b expression, and microvascular E-selectin, P-selectin, and ICAM-1 expression were examined 48 hours after surgery. Complement activation was serially assessed for 2 hours after a single injection of either 1A29 or vehicle. RESULTS: 1A29 treatment did not significantly reduce infarct size in either strain. 1A29 sensitization augmented infarct size and generated rat anti-mouse antibodies. Although 1A29 inhibited neutrophil trafficking shown by reduction in brain myeloperoxidase activity, circulating neutrophils were activated and displayed CD11b upregulation. Complement was activated in 1A29-sensitized Wistar rats and 1A29-treated SHR. E-selectin (SHR), endothelial P-selectin (Wistar and SHR), and ICAM-1 (SHR) were upregulated in animals treated with 1A29. CONCLUSIONS: Administration to rats of a murine antibody preparation against ICAM-1, 1A29, elicits the production of host antibodies against the protein, activation of circulating neutrophils, complement activation, and sustained microvascular activation. These observations provide several possible mechanisms for central nervous system-related clinical deterioration that occurred when Enlimomab was given in acute ischemic stroke.

Neutrophil priming state predicts capillary leak after gut ischemia in rats.

BACKGROUND: Multiple organ failure after serious injury or illness is a major determinant of mortality. An initial insult is believed to "prime" circulating neutrophils and induce systemic inflammation. Thereafter, a second insult will precipitate distant organ injury. The aim of these studies was to evaluate circulating neutrophil function after mesenteric ischemia-reperfusion to determine the neutrophil "priming state," a quantitative and clinically useful predictor of multiple organ failure. MATERIALS AND METHODS: Anesthetized Sprague-Dawley rats underwent superior mesenteric artery occlusion for 30 min or sham operation and were euthanized after 2, 6, or 24 h of reperfusion. Control animals had blood taken without any intervention. To determine changes in lung capillary permeability, another group of rats received Evan's blue, a dye that binds albumin, 1 h before sacrifice. Flow cytometric analysis was performed on 5 million white blood cells after removal of red cells by lysis and centrifugation. Neutrophil number, oxidative burst, and CD18 expression were measured. RESULTS: The number of circulating neutrophils was elevated similarly in rats subjected to sham operation or ischemia-reperfusion. Oxidative burst potential was increased at 2 h, maximum at 6 h, and normal at 24 h after reperfusion, but not in sham rats. CD18 expression was similar in all groups. There was a significant temporal correlation between the "priming state" of the circulating neutrophil, defined as the product of the neutrophil number times oxidative burst, and lung leak. CONCLUSIONS: The neutrophil "priming state" may allow the clinician to better predict those patients at greatest risk for multiple organ failure.

The p65 subunit of NF-kappa B is redundant with p50 during B cell proliferative responses, and is required for germline CH transcription and class switching to IgG3.

B cells lacking individual NF-kappa B/Rel family members exhibit defects in activation programs. We generated small resting B cells lacking p65 or p50 alone, or lacking both p50 and p65, then evaluated the ability of these cells to proliferate, secrete Ig, and undergo Ig class switching. B cells lacking p65 proliferated well in response to all stimuli tested. However, these cells demonstrated an isolated defect in switching to IgG3, which was associated with a decrease in gamma 3 germline CH gene expression. Whereas, previously reported, B cells lacking p50 alone had a severe proliferative defect in response to LPS, a moderate defect in response to CD40 ligand (CD40L), and normal proliferation to Ag receptor cross-linking using dextran-conjugated anti-IgD Abs (alpha delta-dex), B cells lacking both p50 and p65 exhibited severely impaired proliferation in response to LPS, alpha delta-dex, and CD40L. This defect could be overcome by simultaneous administration of alpha delta-dex and CD40L. In response to this latter combination of stimuli, B cells lacking both p50 and p65 secreted Ig and underwent isotype switching to IgG1 as efficiently as B cells lacking p50 alone. These data demonstrate a role for the p65 subunit of NF-kappa B in germline CH gene expression as well as functional redundancy between p50 and p65 during proliferative responses.

Polyvalent antisera to Pseudomonas ribosomal vaccines: protection of mice against clinically isolated strains.

The preparation of polyvalent antisera to ribosomal vaccines from Pseudomonas aeruginosa is described. The ability of these antisera to protect mice by passive immunization against challenge with randomly chosen, clinically isolated strains of P. aeruginosa is reported. Significant protection was achieved against 34 of 40 strains tested (85%). Included among these strains against which protection was achieved were four mucoid strains. In addition, the degree of cross-protection attainable by the ribosomal vaccines was investigated. The results obtained indicated that these vaccines are generally serotype specific.