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C L Klein

Publications and source records attributed to C L Klein.

At least 37 records · Page 2Linked to original sources

The role of the microcirculation in multiple organ dysfunction syndrome (MODS): a review and perspective.

Major advances in intensive care medicine during the past two decades have altered the spectrum of disease encountered by intensive care physicians, anaesthesiologists, traumatologists and pathologists. One of the most important manifestations of severe trauma or infections is the multiple organ dysfunction syndrome (MODS), a life-threatening condition that often ends in multiple organ failure (MOF) and death. Evidence gathered from clinical and morphological observations in humans, taken together with experimental animal studies and a vast accumulation of in vitro data, clearly indicate that the microcirculation lies at the centre of this complex process, which results in peripheral vascular insufficiency, inadequate oxygen delivery to vital organs, and hence, severe organ dysfunction. The multifunctional nature of the endothelium makes it a prime candidate for study of the pathomechanisms of MODS. This paper reviews the evidence for the hypothesis that the microcirculation, and in particular its endothelial component, has a central role in the pathogenesis of MODS. The evidence is reviewed principally from the standpoints of classical morbid anatomy and cell pathobiology.

Animals↗

Biomaterial-induced alterations of human neutrophils under fluid shear stress: scanning electron microscopical study in vitro.

Morphological changes of human polymorphonuclear neutrophils (PMN) adhering to hydrophilic (glass) and hydrophobic (FEP-Teflon, polyethylene, polypropylene) surfaces were studied in a parallel-plate flow chamber at the light and scanning electron microscopical levels. The PMN were exposed to a shear stress of 0.19 Pa (1.9 dynes cm-2) or were allowed to adhere without the stress component (static control) during 30 min for all four biomaterials. Observation by light microscopy was performed in situ in the flow chamber at 1, 5, 10, 15, 20, 25 and 30 min. The total number of adherent cells as a function of time and the activation status of the population on the basis of morphological criteria were determined. On the hydrophilic material adhesion of activated PMN was significantly higher (P < 0.05) than on the more hydrophobic surfaces. This effect was most pronounced for the adhesion of neutrophils to glass and polypropylene (PP). Polyethylene (PE) showed only minor adhesion rates. Scanning electron microscopy revealed details of cell shape changes and permitted a more precise classification of populations of neutrophils based on distinctive shapes. As PMN were exposed to shear stress on glass, the majority of cells exhibited surface veils, ridges and ruffles, suggesting a high level of cell migration. In this case, on polymeric surfaces the presence of filopodial networks (FEP-Teflon) and ameoboid cell shapes (PP and PE) was noted. The results suggest that a low shear stress, as well as various chemical and physical properties of biomaterial surfaces, are together responsible for differentiation of PMN populations on solid substrata.

Biocompatible Materials↗

Local and systemic inflammatory mediator release in patients with acute and chronic posttraumatic osteomyelitis.

The local and systemic release of thromboxane A2, prostaglandin I2, leukotriene B4 (LTB4), tumor necrosis factor alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and interleukin-8 (IL-8) were studied before and after operation in 29 patients with acute and 22 with chronic posttraumatic osteomyelitis. Twenty patients without osteomyelitis, who underwent operations for fractures of the lower extremities, served as controls. Blood and tissue samples from the osteomyelitic and control groups were collected under defined conditions and mediators were determined by radioimmunoassay (thromboxane B2, 6-keto-prostaglandin F1 alpha, LTB4) or by enzyme-linked immunosorbent assay (TNF-alpha, IL-1 beta, and IL-8). In addition, common parameters (leukocyte count, C-reactive protein, temperature) were measured. The best correlation with acute disease activity was given by TNF-alpha, IL-6, IL-8, and LTB4. Plasma levels of these mediators in acute osteomyelitis were significantly increased compared to chronic osteomyelitis and to controls, respectively. Tissue samples from osteomyelitic focus showed significantly increased levels for IL-8, IL-6, TNF-alpha, IL-1 beta, and LTB4 in acute osteomyelitis, whereas the values for TxB2 and 6-keto-prostaglandin F1 alpha were only slightly increased compared to the chronic osteomyelitis group. This study describes the local and systemic liberation of various mediators in acute and chronic posttraumatic osteomyelitis in detail for the first time and provides data for pre- and postoperative monitoring of disease activity and demonstrates new pathogenetic and therapeutic aspects of bone modulation in osteomyelitis.

Acute Disease↗

PECAM-1 expression in human mesothelial cells: an in vitro study.

Mesothelial cells are actively involved in inflammatory processes by expressing a set of cell adhesion molecules (CAMs). Transmigration of leukocytes into inflamed tissues requires a chemotactic stimulus and engagement of platelet-endothelial cell adhesion molecule-1 (PECAM-1). To investigate the kinetics involved in peritonitis, pure cultures of mesothelial cells are necessary. In previous studies, we have found that human mesothelial cells (HOMES) show a weak constitutive expression of PECAM-1, which cannot be further stimulated by cytokines. It is known that all serous cavities and body fluids contain numerous macrophages which strongly express this adhesion molecule. To identify the cells responsible for the expression of PECAM-1, mesothelial cells freshly obtained from omental tissue were isolated using PECAM-1-conjugated magnetic beads by cell sorting. For these studies, the negative as well as the positive fraction of isolated cells were used. As a control, freshly isolated monocytes were studied. Cell cultures were characterized by light and electron microscopy, as well as immunocytochemistry. The negative cell fraction was cultivated and stimulated for different times with tumor necrosis factor-alpha (30 and 300 U/ml), interleukin-1 beta (10 and 100 U/ml) and interferon-gamma (500 U/ml) and PECAM-1 expression was analyzed by a comparative quantitative cell enzyme immunoassay (EIA). The positive cell fraction was treated in the same manner. Both fractions of isolated cells showed strong positivity for cytokeratins 8, 18, 7 and 19, as well as vimentin. CD68, a monocyte marker, was not detected on mesothelial cells. In addition, EIA analysis confirmed the constitutive expression of PECAM-1 obtained from previous studies. This expression on HOMES was not inducible, irrespective of the type and concentration of cytokine studied. These data confirm PECAM-1 expression on mesothelial cells obtained from human omental tissue and suggest a critical role in transmigration of leukocytes during peritoneal inflammation.

Cell Separation↗

Pentamidine congeners. 3. Crystal structure and molecular modeling studies of trans-1,4-bis(4-amidinophenoxy)-2-butene.

X-ray diffraction was used to confirm the geometry of trans-1,4-bis(4-amidinophenoxy)-2-butene dihydrochloride dihydrate (trans-butenamidine). trans-Butenamidine is a semirigid analogue of pentamidine that has demonstrated good anti-Pneumocystis carinii activity in rats. Molecular modeling studies revealed that unlike pentamidine or propamidine, trans-butenamidine does not discriminate between AT and TA sequences in its binding to the minor groove of DNA. Crystal data: [C18H22N4O2(2+)][Cl(-)]2[H2O]2, triclinic space group, P1, a = 9.443(1) A, b = 11.400(1) A, c = 11.919(1) A, alpha = 62.19(1) degree, beta = 81.10(1) degree, gamma = 72.19(1) degree, V = 1080.3(3) A3, Z = 2, R = 0.054 for 1149 observed reflections with I > 3 sigma (1).

Base Sequence↗

Synthesis and antimicrobial activity of 6,7-annulated pyrido[2,3-d]pyrimidines.

Four new 6H-indeno[2',1':5,6]pyrido[2,3-d]pryimidines (10-13) were synthesized via cyclocondensation reactions involving chlorovinyl aldehyde 1 or ketoaldehyde 3 and appropriately substituted 6-aminopyrimidines. The regiochemistry of the compounds was established by 1H NMR and 13C NMR spectral data as well as X-ray crystal data. Compounds 10 and 11 and previously reported homologues 14 and 15 were screened for antimicrobial activity. Moderate antimicrobial activity was observed for some of these compounds. Compound 14 was especially active against Staphylococcus aureus. Crystal data for 13 (C14H7N3Cl2) follows: monoclinic space group, P21/n; Unit cell dimensions, a = 7.284(1) A, b = 12.800(1) A, c = 13.108(1) A, beta = 93.98(1) degree, V = 1219.2(2) A3, Z = 4.

Anti-Bacterial Agents↗

Tissue factor expression during coculture of endothelial cells and monocytes.

The role of monocytes as initiators of coagulation through the expression of tissue factor has been well documented in vitro, and the relationship of monocyte tissue factor to the thrombotic complications of atherosclerosis has been suggested. Tissue factor antigen has been identified in the plasma membranes of monocytes adherent to the vascular endothelium overlying atherosclerotic plaques and the presence of tissue factor in adherent mononuclear cells correlates with the polymerization of fibrin at these same sites. To further understand the relationship of cellular adhesion to tissue factor expression, human monocytes were cocultured for periods ranging from 30 min to 24 hr with endothelial cells isolated from human umbilical veins (HUVEC). Tissue factor antigen, as assayed by both ELISA and immunogold electron microscopy, was minimal on either monocytes or HUVEC maintained in homogeneous cultures or on the cells when cocultured for 1 hr or less. This was true whether the HUVEC were in a native state or if they had been stimulated with interleukin-1 (IL-1 beta) or lipopolysaccharide (LPS) prior to monocyte adhesion. Typically, less than 13% of the cells in short-term cocultures were positively labeled through anti-tissue factor immunogold microscopy. The level of tissue factor, however, was increased 3-fold above baseline when monocytes were cocultured with unstimulated HUVEC for 4 hr, and it was more than double this if the HUVEC had been exposed to IL-1 beta or LPS (7-fold increase). By 24 hr, the expression of tissue factor antigen was nearly 50-fold higher in cocultures involving stimulated HUVEC, and at later times greater than 70% of the cells were labeled with immunogold. Through the use of quantitative immunogold electron microscopy, the increase in tissue factor was most pronounced on monocytes which had three times greater increase in tissue factor than HUVEC in the same cultures. These studies document the stimulation of tissue factor expression by monocytes upon coculture with endothelial cells, and the data document an enhancement of this coculture effect upon HUVEC stimulation with cytokines. These observations have relevance to atherosclerotic disease by suggesting that interaction of monocytes with dysfunctional endothelial cells overlying atherosclerotic plaques would be sufficient to induce tissue factor and by so doing predispose to localized thrombotic events.

Cell Adhesion↗

Comparative studies on vascular endothelium in vitro. 2. Hypoxia: its influences on endothelial cell proliferation and expression of cell adhesion molecules.

Recent studies have presented evidence that the processes of hypoxaemia and reperfusion are involved in several pathogenetic mechanisms of atherosclerotic lesions. The ability of hypoxaemia to activate circulating white blood cells (WBCs) and enhance WBC-endothelial cell (EC) interactions is suspected to be a major factor in deleterious processes in the blood vessel wall. Various groups have suggested that cell adhesion molecules (CAMs), such as ICAM-1, VCAM-1 and E-selectin and their leukocyte ligands are involved in intercellular activities of the relevant cell types. We studied the effects of different oxygen tensions, simulating normoxic conditions, hypoxia and hyperoxia in vitro with the help of an umbilical vein EC model in order to determine the effects of oxygenation on CAM presentation on vascular ECs with and without further cytokine and endotoxin (lipopolysaccharides; LPS) stimulation. Semiquantitative analysis of ICAM-1, E-selectin and VCAM-1 was performed using cell enzyme immunoassay techniques. The presentation of ICAM-1, E-selectin and VCAM-1 remained on the whole unaffected by both hypoxia and hyperoxic conditioning after both 7 and 24 h. Stimulation of ICAM-1 by cytokines and LPS was only marginally influenced by the oxygen tension. Cytokine induction of E-selectin was not affected after 7 h and was even reduced under hypoxia, compared to the control culture after 24 h, while stimulation was increased by hyperoxia. VCAM-1 was reduced in both the hypoxic and hyperoxic culture, while being maximally stimulated by cytokines and LPS after 7 h. In general, an effect of hypoxia was not found without any further stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Adhesion Molecules↗

Comparative studies on vascular endothelium in vitro. 3. Effects of cytokines on the expression of E-selectin, ICAM-1 and VCAM-1 by cultured human endothelial cells obtained from different passages.

Endothelial cells (EC) are very responsive to proinflammatory cytokines, e.g. interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF alpha), as well as to bacterial lipopolysaccharide. EC are stimulated by these substances to secrete chemotactic factors and to increase expression of cell adhesion molecules (CAM), leading to dramatically altered interactions with leukocytes, e.g. granulocytes and monocytes. In these interactions E-selectin, ICAM-1 and VCAM-1 are known to play an important role, as they are presented by the EC and interact with corresponding ligands on the white blood cell membranes. These adhesion molecules have been studied worldwide in a variety of in vitro experiments using cultured EC. Different passages and mixtures of passages have been used in these experiments, often without any regard to the comparability of the results. In this study the expression of E-selectin, ICAM-1 and VCAM-1 on cultured human umbilical vein EC (HUVEC) obtained from different passages (passages 1-6) was studied after 4, 8 and 24 h of exposure to IL-1 beta and TNF alpha. In previous studies, we have shown that IL-1 beta and TNF alpha increase the expression of E-selectin and ICAM-1 on the cytoplasmatic membranes of HUVEC and human adult EC from the saphenous vein and femoral artery in a similar fashion. Using a comparative quantitative cell enzyme immunoassay, we found that the expression of the adhesion molecules was significantly reduced with increasing passages. There was also a decreased persistence of CAM comparing different periods of stimulation between 6 and 24 h in the different passages. These data indicate that the number of passages plays an important role in the expression of adhesion molecules on EC. The results are relevant for the meaningful planning of comparative in vitro studies on EC presentation of CAM.

Cells, Cultured↗

Effects of cytokines on the expression of cell adhesion molecules by cultured human omental mesothelial cells.

Cultured mesothelial cells (HOMES) are very responsive to the proinflammatory cytokines, interleukin (IL)-1 and tumor necrosis factor-alpha (TNF-alpha). E-selectin, ICAM-1 and VCAM-1 are known to play an important role, because they are presented by diverse cell types, for example endothelial cells (ECs), and interact with co-responding ligands on white blood cell membranes. In this study, the expression of ICAM-1, VCAM-1, E-selectin as well as PECAM-1 on cultured HOMES was studied over 5, 24, 48 and 72 h exposure to IL-1 beta, interferon-gamma and TNF-alpha. In previous studies we have shown that IL-1 beta and TNF-alpha increase the expression of ICAM-1, E-selectin and VCAM-1 on the cytoplasmatic membranes of HUVECs, HSVECs and HAFECs (ECs from human umbilical vein, saphenous vein and femoral artery, respectively). Using a comparative quantitative cell enzyme immunoassay, we found that expression of the adhesion molecules ICAM-1 and VCAM-1 was significantly increased on HOMES in a dose- and time-dependent manner, compared to nonstimulated cells. Thus, ICAM-1 increased dramatically after 5 h incubation with TNF-alpha. Values of about 450% of the control level were measured. VCAM-1 was similarly stimulated after 24 h incubation with the same cytokine, although its level of expression was significantly lower than that of ICAM-1. In contrast to findings in the literature, VCAM-1 was not found to be expressed constitutively. E-selectin was neither constitutively expressed nor markedly inducible on HOMES. Only weak expression was found after 24 h incubation with high-dose IL-1 beta. PECAM-1 was expressed constitutively, as became evident in antibody dilution studies. These data indicate that HOMES respond to inflammatory stimuli, in some ways in a similar fashion to vascular endothelial cells, but also show a specific pattern of antigen presentation. The results are important for a better understanding of inflammatory processes in serous cavities. The data are also relevant for the improvement of antithrombogenous surfaces of the lumina of vascular prostheses by cell seeding.

Cell Adhesion Molecules↗

(-)-Norcocaine.

The title compound, [2R,3S-(2 beta,3 beta)]-methyl 3-(benzoyloxy)-8-azabicyclo[3.2.1]octane-2-carboxylate, C16H19NO4, is a metabolite of the tropane alkaloid cocaine. The molecule crystallized as the free base with the piperidine ring in a chair conformation. The tropane ring system and its methoxycarbonyl and benzoyl groups are rigid; only rotational flexibility is allowed in the conformation of the substituents.

Cocaine↗

2'-hydroxy-5,9-dimethyl-2-(3-methyl-2-butenyl)-6,7-benzomorphan (pentazocine) hydrochloride hydrate.

The title compound [1,2,3,4,5,6-hexahydro-6,11-dimethyl-3-(3-methyl-2-butenyl)-2,6-methano- 3- benzazocin-8-ol hydrochloride hydrate, C19H28NO+.-Cl-.1/8H2O, (I)] crystallizes with two molecules per asymmetric unit. A comparison of the bond lengths and torsion angles shows a side-chain orientation similar to that of cyclazocine but unlike that of naloxone. The two molecules are linked through a hydrogen-bond network to the interlayer Cl ions, forming a linear chain extending along the ac diagonal of the unit cell.

Crystallography, X-Ray↗

Crystal and molecular structures of tricyclic neuroleptics.

The X-ray crystal structures of three tricyclic neuroleptics have been completed and are reported herein. These include amitriptyline hydrochloride (I), chlorprothixene hydrochloride (II), and ethopropazine hydrochloride (III). The structures were compared with the structures of similar molecules with similar pharmacological activity. It was found that there is a great deal of flexibility in the structures of these molecules, which makes it difficult to determine the pharmacologically active conformations. Crystallographic data: (I) C20H24NCl, monoclinic space group P2(1)/a, a = 13.812(4) A, b = 9.140(2) A, c = 14.345(4) A, beta = 96.82(2) degrees, Z = 4, final R = 0.059 for 1630 observed (l > 3 sigma(l)) reflections. (II) C18H20NSCl2.CHCl3, monoclinic space group C2/c, a = 29.416(4) A, b = 6.986(1) A, c = 23.374(3) A, beta = 109.78(1) degrees, Z = 8, final R = 0.046 for 1436 observed reflections. (III) C19H25N2SCl, monoclinic space group P2(1)/c, a = 8.849(1) A, b = 14.475(2) A, c = 14.832(1) A, beta = 98.72(1) degrees, Z = 4, final R = 0.039 for 2583 observed reflections.

Antidepressive Agents, Tricyclic↗

Application of X-ray microanalysis to study of the expression of endothelial adhesion molecules on human umbilical vein endothelial cells in vitro.

A semi-quantitative procedure is described, which allows the evaluation of expression levels of endothelial adhesion molecules on cultured human umbilical vein endothelial cells (HUVEC) using energy dispersive X-ray microanalysis (EDX). As a model two adhesion molecules, E-selection (CD62E; ELAM-1/endothelial leukocyte adhesion molecule-1) and ICAM-1 (intercellular adhesion molecule-1; CD54), were localized by the use of the silver-enhancement colloidal gold method after stimulation of HUVEC with endotoxin lipopolysaccharide (LPS), tumour necrosis factor (TNF) or a phorbol ester (PMA). The analysis was performed in a scanning electron microscope (SEM) at an accelerating voltage of 15 kV with scanned areas of 200 x 400 microns. The semi-quantitative data indicated that in LPS-treated groups both adhesion molecules were expressed at a significantly higher level than in all other groups (P < 0.01). In addition, after a 4 h treatment the expression levels of E-selectin in all groups were higher compared to ICAM-1. The experimental data from X-ray microanalysis were compared with data obtained from an enzyme-linked immunosorbent assay (ELISA) and similar values were found for both types of preparation. This microanalytical method is relatively simple and seems to be suitable for immunogold labelling studies on different types of endothelial cells in vitro.

Cell Adhesion Molecules↗

Application of immunogold labelling for light and electron microscopic localization of endothelial leukocyte adhesion molecule 1 (ELAM-1) on cultured human endothelial cells.

This study describes the expression characteristics of E-selectin molecules using immunogold histochemical techniques on cultured human umbilical vein endothelial cells (HUVEC). The expression of E-selectin was induced by tumour necrosis factor-alpha (TNF-alpha, 300 U/ml), phorbol ester (PMA, 10 ng/ml) and bacterial lipopolysaccharide (LPS, 4 micrograms/ml). No expression was demonstrated on control cells. Using the silver-enhanced colloidal gold-labelling technique, at the light microscopical level, HUVEC could be distinctively subdivided into three staining types. The cell labelling index, expressed as the number of 'positively' stained cells as a proportion of all viewed cells was the highest in the LPS group. For transmission electron microscopy (TEM) the preembedding immunocytochemical staining method and embedding in epoxy resin (Agar 100) according to standard procedures was used. In TEM gold particles were localized in close association with the apical plasma membrane, as well as on the surface of microvillus-like projections (the latter by TNF-alpha group). For high resolution scanning electron microscopy (HR-SEM) the secondary (SEI) and the backscattered electron imaging (BEI) modes were used. Gold particles were randomly distributed over the whole cell surface, although they appeared to be denser in the perinuclear zone. The quantitative evaluation on SE and BE viewing (the number of gold particles per cell area in microns 2) demonstrated the highest density of labelling in the LPS-treated group, but there was only a significant difference between LPS and TNF-alpha groups (P < 0.01, t-test). Furthermore, the ultrastructural studies indicated that treatment with substances which up-regulate E-selectin expression was not related to toxic cell damage or significant alterations of cellular ultrastructure.

Cell Adhesion Molecules↗

Increased adhesion and activation of polymorphonuclear neutrophil granulocytes to endothelial cells under heavy metal exposure in vitro.

Heavy metals have been implicated in the mechanisms of endothelial damage. Influences of heavy metal ions on diverse cell types have been studied using a variety of in vitro and in vivo methods. Polymorphonuclear neutrophil granulocytes (PMNs) have physiological and pathological functions, including the modulation of adhesion to and destruction of endothelial cells (ECs). PMNs were studied during interaction with human umbilical vein ECs under exposure to zinc, nickel and cobalt using an in vitro model. We studied adhesion processes with the help of a computer-controlled image-analyzing system and examined the activation of PMNs by quantification of leukotriene B4 (LTB4) release. The biphasic effects of the evaluated heavy metals on PMN-EC adhesion, with stimulation at very high and very low molar concentrations, were observed. The release of LTB4 by PMNs increased during exposure to very low metal concentrations. The initiation of these important pathogenetic mechanisms of inflammation at very low metal ion concentrations, which give no morphological changes, must be regarded as potentially significant with respect to the toxic effects of heavy metals.

Cell Adhesion↗