Use of ICAM-1 antibodies and antisense oligonucleotides to inhibit transmigration of neutrophils

Objective: The increase in adhesion molecule expression during the initial phase of inflammation leads to transmigration of neutrophils. Inhibition of this transmigration could decrease inflammatory response and tissue damage.Materials and methods: Uptake of fluorescein-labelled oligonucleotides, expression of ICAM-1 and neutrophil migration were studied using the bilayer of epithelial (H292) and endothelial (ECV-304) cell lines and neutrophils from peripheral blood of healthy volunteers.Results: The inhibition of ICAM-1 expression was time dependent for both cell lines, with inhibition of more than 50% after 48 h. Antisense oligos inhibited transmigration already after 4 h of incubation (6.6 ± 2.5% versus 18.6 ± 2.5% inhibition, p < 0.01). Antisense was more effective in preventing fMLP-induced neutrophil migration than ICAM-1 antibodies (88 ± 3.8% versus 67 ± 7% inhibition, p = 0.02).Conclusions: Antisense oligos cause a decrease in ICAM-1 expression and inhibit transmigration of neutrophils. However the effectiveness of antisense is higher than monoclonal antibodies, neither of them is able to block the migration completely. This suggests the existence of ICAM-1 independent mechanisms that are responsible for migration.Received 12 June 2003; returned for revision 9 Ocotober 2003; accepted by I. Ahnfelt-Rønne 14 November 2003     

Osteoprotegerin and Osteoprotegerin-Ligand Balance: a New Paradigm in Bone Metabolism Providing New Therapeutic Targets

Osteoprotegerin-ligand, also called Osteoclast Differentiation Factor or RANK-ligand, its receptor RANK and its decoy receptor Osteoprotegerin are key molecules regulating osteoclast differentiation and activation. In this view we discuss structure and expression of these molecules, the genetic models addressing their function and their role in in vivo models of osteoclast differentiation and activation. The new paradigm that has evolved from these studies, is not only important in normal bone homeostasis but also appears to play a role in different diseases that affect the skeleton, such as osteoporosis, inflammatory joint disease and cancer. It has opened a new era in bone research by increasing our molecular knowledge and providing new therapeutic targets in bone disease. 5 November 2000 / Accepted: 15 December 2000     

Histomorphometry and micro-computed tomography of bone augmentation under a titanium membrane

OBJECTIVES: Bone augmentation underneath an occlusive titanium membrane is evaluated in most cases by means of serial histological sections and histomorphometry. Micro-computed tomography (micro-CT) is a less invasive and dynamic technique to measure bone volume in animals of a size that fits into the gantry. The aim of the present study was to evaluate whether the latter approach could match histomorphometry to assess bone augmentation under a titanium membrane. MATERIAL AND METHODS: Pre-formed titanium cups were placed on the skull of 16 rabbits. Bone formation underneath the cups was allowed to occur for 12 weeks. The amount of bone volume assessed by micro-CT was expressed as a numerical unit. One unit volume corresponds to 0.043 mm3. The measurements reveal the volume of bone-like tissue under the membrane, with the same density as that of the original rabbit skull bone. Histological sections were cut along the same plane as the one used for the micro-CT images. The total bone surface was assessed by a digital image system in double-stained undecalcified histological sections and related to the maximum available surface of the titanium cups, which was on average 1366 mm2. RESULTS: The amount of total bone surface found under the titanium membrane varied between 40 and 163 mm2. Measured by micro-CT, the bone detected ranged from 3.7 to 396 numerical units. A highly significant (P<0.001) correlation was found between the total bone volume measured in conventional serial histological sections and by the micro-CT technique (r2=0.72). CONCLUSIONS: The total bone volume measured underneath a membrane using the micro-CT when compared with histological sections remained within a 16% error. This is because of the scattering effect of the metallic membrane and the impossibility to distinguish newly formed bone from the original skull bone on the micro-CT images.     

Presence of substance P and neurokinin 1 receptors in human sputum macrophages and U-937 cells.

Tachykinins such as substance P (SP) may be involved in the pathogenesis of inflammatory airway diseases such as asthma. This study investigated the presence of SP and its receptor in the differentiated macrophage-like U-937 cell line and in macrophages from sputum induced in healthy subjects (n=8). In situ hybridization with digoxigenin-labelled sense and antisense complementary ribonucleic acid (cRNA) probes was used to determine the expression of SP and its receptor (neurokinin (NK)1 receptor). SP-immunoreactive material was detected using a rabbit anti-SP antiserum and the alkaline phosphatase anti-alkaline phosphatase technique. Beta-preprotachykinin (PPT)-I messenger ribonucleic acid (mRNA) encoding SP, was detected using in situ hybridization in differentiated U-937 cells as well as in CD45+ human leukocyte antigen (HLA) DR+ sputum macrophages. The expression of the beta-PPT-I mRNA was increased in lipopolysaccharide (LPS)-stimulated U-937 cells. SP-immunoreactive material was found in differentiated U-937 cells and in CD68+ sputum macrophages. NK1 receptor mRNA was detected in differentiated U-937 cells and sputum macrophages. Incubation of U-937 cells with SP considerably increased the expression of NK1 receptor mRNA. This study demonstrates that human monocytes/macrophages express substance P and that this expression is upregulated by lipopolysacharide. Human monocytes/macrophages also express neurokinin1 receptor messenger ribonucleic acid, suggesting an autocrine effect of substance P on these cells.     

Regional differences of brain glucose metabolic compensation after unilateral labyrinthectomy in rats: a [C]2-deoxyglucose study

A unilateral labyrinthectomy was performed on anesthetized adult albino rats. Brain [¹⁴C]2-deoxyglucose (2DG) uptake was measured autoradiographically 3.5 h to 20 days later and compared to sham-operated controls. In the vestibular nuclei (nn.) of labyrinthectomized subjects, large left-right differences of 2DG uptake occurred, which decreased over time. The equalization of vestibular nuclear 2DG uptake paralleled behavioral compensation of body, neck and head postural abnormalities, and known equalization of vestibular nuclear cell firing rates during compensation. There was a small difference of 2DG uptake in medial and lateral vestibular nn. 20 days after lesions when animals had a residual head tilt and tonic eye deviation. In the oculomotor nn., trochlear nn. and interstitial n. of Cajal, large left-right differences of 2DG uptake occurred, which did not change over time. The higher 2DG uptake in these nn. occurred ipsilateral to the labyrinthine lesion and did not correlate with the onset and cessation of nystagmus. The persistent asymmetry did appear to correlate with ipsilateral downward and contralateral upward eye deviation which continued for long periods after the lesion. We hypothesize that the non-compensating metabolic asymmetry in the oculomotor and trochlear nn. could be due to lesioned otolithic input to the vestibular nn. which relays to trochlear and oculomotor nn.