The effect of intestinal ischemia and reperfusion injury on ICAM-1 expression,endothelial barrier function,neutrophil tissue influx,and protease inhibitor levels in rats

Multiple organ dysfunction syndrome (MODS) is mediated by complex mechanisms in which interactions between activated leukocytes and endothelial cells play a central role. ICAM-1 (intercellular adhesion molecule-1) mediates firm adhesion and transendothelial migration of activated leukocytes from postcapillary venules into the tissue. The present study evaluated the ICAM-1 expression in various organs after 40 min of intestinal ischemia and 1, 3, 6, 12 h of reperfusion (I/R) in the rat, using a dual monoclonal antibody technique (n = 36). Endothelial barrier permeability, using the vascular leakage of radiolabeled human serum albumin was also assessed (n = 12). Neutrophil sequestration in the lungs was quantitated by myeloperoxidase activity and plasma protease inhibitor levels were measured with electroimmunoassay. Significant regional differences were found in ICAM-1 expression between organs, both constitutively and after I/R-injury. The highest constitutive levels were observed in the liver and lungs, followed by the kidneys. The constitutive ICAM-1 expression in the intestines and in the heart was about 1/20 compared with that found in the liver and lungs. The brain and muscle had levels of about 1/150 of that in the liver and lungs. After intestinal I/R, significant increases (17-45%) were found in the lungs, intestines, brain, heart, and muscle. Albumin leakage index (ALI) in all examined organs and myeloperoxidase activity in the lungs increased after I/R-injury. Serum levels of albumin and most protease inhibitors decreased significantly after I/R challenge. Intestinal I/R results in an increase of systemic ICAM-1 expression with marked organ variability. The upregulation of ICAM-1 could represent a crucial step in the adherence- and migration process of activated leukocytes and potentially in the development of tissue injury.     

DNA arrays as diagnostic tools in human healthcare

The sequencing of the entire human genome was completed in June 2000. The sequence, however, is only a starting point and gene-function is now of major interest. All this information shows that gene-based diagnostics can be helpful for treatment targeting and patient surveillance. High volume gene expression assays can optimize pharmaceutical therapies by targeting genome-based treatments to specific patient populations and providing methods to study genes involved with cancer growth patterns and tumor suppression. Molecular biology, so far, has elucidated many of the genetic mechanisms underlying heritable metabolic diseases, so that appropriate diagnostic assays will revolutionize molecular diagnostics in medicine and pharmaceuticals. One of the most promising new technologies designed to analyze large amounts of genomic information rapidly is the DNA chip.     

Foreign body reaction to calcium alginate fibre mimicking recurrent tumour of the submandibular salivary gland

A 50-year-old woman was referred after the discovery of adenoid cystic carcinoma in an excised left submandibular gland. Treatment involved clearance of the left submandibular fossa, and bilateral levels II and III selective neck dissections. A left-sided submandibular haematoma developed during the immediate postoperative period. After removal of the clot, there was a persistent, low volume capillary ooze from the left submandibular fossa and a calcium alginate fibre pack (Kaltostat) was left in place to control the bleeding. After an extended period of time the pack excited a foreign body reaction which, on a computed tomogram, mimicked a recurrence of the tumour. We review the role of Kaltostat in this setting and its potential for foreign body reaction, which may mimic serious disease.     

Long-term administration of L-carnitine to humans: effect on skeletal muscle carnitine content and physical performance

BACKGROUND: Long-term administration of high oral doses of L-carnitine on the skeletal muscle composition and the physical performance has not been studied in humans. METHODS: Eight healthy male adults were treated with 2 x 2 g of L-carnitine per day for 3 months. Muscle biopsies and exercise tests were performed before, immediately after, and 2 months after the treatment. Exercise tests were performed using a bicycle ergometer for 10 min at 20%, 40%, and 60% of the individual maximal workload (P(max)), respectively, until exhaustion. RESULTS: There were no significant differences between V(O(2)max), RER(max), and P(max) between the three time points investigated. At submaximal intensities, the only difference to the pretreatment values was a 5% increase in V(O(2)) at 20% and 40% of P(max) 2 months after the cessation of the treatment. The total carnitine content in the skeletal muscle was 4.10 +/- 0.82 micromol/g before, 4.79 +/- 1.19 micromol/g immediately after, and 4.19 +/- 0.61 micromol/g wet weight 2 months after the treatment (no significant difference). Activities of the two mitochondrial enzymes citrate synthase and cytochrome oxidase, as well as the skeletal muscle fiber composition also remained unaffected by the administration of L-carnitine. CONCLUSIONS: Long-term oral treatment of healthy adults with L-carnitine is not associated with a significant increase in the muscle carnitine content, mitochondrial proliferation, or physical performance. Beneficial effects of the long-term treatment with L-carnitine on the physical performance of healthy adults cannot be explained by an increase in the carnitine muscle stores.     

Human adrenal cells in culture produce both ouabain-like and dihydroouabain-like factors

BACKGROUND: Ouabain-like factor (OLF) and its newly discovered reduced species, dihydroouabain-like factor (Dh-OLF), are mammalian cardenolides whose structural and functional characteristics are similar to the plant-derived compounds ouabain and dihydroouabain. These endogenous compounds are believed to be produced by the adrenals and to constitute part of an hormonal axis that may regulate the catalytic activity of the alpha-subunit of Na(+),K(+)-ATPase. We developed antibodies sufficiently specific to distinguish between OLF and Dh-OLF, and in this study demonstrate the selective secretion of OLF and Dh-OLF from human H295R-1 adrenocortical cells in culture. METHODS: We used reversed-phase HPLC, inhibition of Na(+),K(+)-ATPase catalytic activity, and two enzyme immunoassays developed with antibodies specific to ouabain and dihydroouabain to purify and characterize the secretion of these two compounds by human adrenal cells in culture. Purified antisera had high titers (1 x 10(6) for ouabain and 8 x10(5) for dihydroouabain) and were specific to their corresponding antigens. RESULTS: Human H295R-1 cells grown in serum-free medium secreted 0.18 +/- 0.03 pmol of OLF and 0.39 +/- 0.04 pmol of Dh-OLF per 10(6) cells in 24 h. Both OLF and Dh-OLF inhibited the ouabain-sensitive catalytic activity of the sodium pump (0.03 micro mol/L OLF inhibited 29% of the catalytic activity; 0.07 micro mol/L Dh-OLF inhibited 17%). Stimulation of the cell culture by dibutryl cAMP increased the secretion of Dh-OLF 50% over control (unstimulated), whereas the secretion of OLF did not increase significantly. CONCLUSIONS: OLF and Dh-OLF are secreted by human adrenal cells, and antibodies specific to these two compounds can be developed, using the plant-derived counterparts as antigens. The secretion of Dh-OLF is responsive to a cAMP-dependent stimulation mechanism, whereas OLF is not. Our data suggest that either the secretory or biosynthetic pathways for production of these two compounds by human adrenal cells may have different control mechanisms or that they may be linked via a precursor-product relationship.     

Impact of microscope numerical aperture on microspectrophotometric measurements of hemoglobin in microvessels

In the microspectrophotometric method to measure hemoglobin concentration and oxygen saturation in microvessels it is recommended that a low numerical aperture (NA) condenser be employed to ensure that the recorded image is a true projection of the object. However, this tenet has never been rigorously justified. In this study, the microspectrophotometric method is evaluated using the theory of three-dimensional image formation by a light microscope for a wide range of NA. The results of the calculations show that for measurements for hemoglobin concentration, the recorded image is close to the true projection only when the size of the microvessel is large compared to the degree of smearing ( proportional, variant 1/NA) but small compared to the degree of defocus ( proportional, variant NA(2)). These opposing tendencies lead to an optimum NA for which the errors are minimum. This optimum NA is a function of the size of the microvessel and the manner in which the hemoglobin concentration is distributed within the lumen. For measurements of oxygen saturation, the recorded image is the true projection as long as the measurements are made in regions near the microvessel centerline. For measurements made in regions away from the centerline, good agreement was obtained only when the distribution of oxygen saturation was uniform. Reconstruction of the axisymmetric profiles from the recorded projections showed that the errors in the projections cause the recovered profiles to deviate from the true profiles. These deviations are directly related to the extent by which the recorded projections deviate from the true projection.