Serum IgG levels and mortality in patients with severe sepsis and septic shock : The SBITS data.

BACKGROUND: The role of intravenous immune globulin (Ig) therapy in patients with severe sepsis and septic shock is discussed controversially. Low initial IgG levels could help to identify those patients who might benefit from an adjunctive Ig treatment. OBJECTIVES: To investigate the effect of initial serum IgG levels on 28-day mortality in patients with severe sepsis and septic shock. MATERIALS AND METHODS: In this retrospective analysis of the SBITS trial data, 543 patients were allocated to four groups (quartiles) depending on their initial serum IgG levels (1: IgG ≤ 6.1 g/l; 2: IgG 6.2-8.4 g/l; 3: IgG 8.5-11.9 g/l; 4: IgG > 11.9 g/l). The third quartile was taken as the reference quartile. For the applied logistic regression model clinically relevant confounders were defined and integrated into further risk-adjusted calculations. RESULTS: Patients with the lowest IgG levels had a mortality rate similar to those patients with initial IgG levels in the second and third quartile, representing the physiological IgG range in healthy people. Surprisingly, patients with the highest IgG levels even showed a significantly higher mortality in a risk-adjusted calculation compared to the reference quartile (OR 1.69, CI 1.01-2.81, p = 0.05). Subgroup analyses revealed that initial IgG levels were of no prognostic value in patients presenting with vasopressor-dependent septic shock on admission as well as in patients with either gram-positive or gram-negative sepsis. CONCLUSIONS: Initially low IgG levels do not discriminate between survival and nonsurvival in patients with severe sepsis and septic shock. Therefore, low IgG cannot help to identify those patients who might benefit from an adjunctive IgG sepsis therapy. Whether a high initial IgG serum level is an independent mortality risk factor needs to be investigated prospectively.

[Pathophysiology of sepsis and possible influence of statins]. / Pathophysiologie der Sepsis und deren mögliche Beeinflussung durch HMG-CoA-Reduktase-Inhibitoren.

Severe sepsis and septic shock are common complications in the intensive care unit and associated with high mortality. Early antimicrobial therapies together with organ-supportive measures are the major therapeutic approaches. However in the last decades immunomodulatory therapies have been investigated due to the notion of a compromised inflammatory response in septic patients. In addition to lowering circulating cholesterol, statins (HMG-CoA-reductase-inhibitors) have also been shown to possess pleiotropic anti-inflammatory potential. Recent studies indicate that these anti-inflammatory effects also modulate acute inflammatory response and therefore may play a protective role in septic patients. In this review, the pathophysiological background and first clinical trials of statins as a new adjuvant therapy in sepsis are summarized.

[Coronary heart disease in childhood in familial hypercholesteremia. Maximum therapy with LDL apheresis]. / Koronare Herzkrankheit im Kindesalter bei familiärer Hypercholesterinämie. Maximaltherapie unter Einsatz der LDL-Apherese.

In children with familial hypercholesterolemia, coronary heart disease requires both medical theraphy and LDL apheresis. We report a case of verified occlusion of the anterior descending branch of the left coronary artery in a 10-year-old patient. The pathological findings revealed by ergometry established the diagnosis. The goal was to achieve the greatest possible reduction of lipid parameters and fibrinogen by lowering plasma viscosity employing LDL apheresis. It is astonishing that this treatment is also well tolerated by children. The basic vascular approaches suffice and shunt operations are not absolutely necessary. The efficacy of this method became vividly apparent by the changes in the skin lesions. Additional angiographic follow-up and further clinical course wil provide information on the usefulness of this treatment strategy with maximum lipid theraphy and the expected improvement in prognosis.

Determination of regional bone blood flow by means of fluorescent microspheres using an automated sample-processing procedure.

The determination of regional blood flow utilizing fluorescent microspheres (FMs) is an established method for numerous organs. Recent progress, in particular the automation of sample processing, has further improved this method. However, the FM method (reference sample technique), which allows repetitive measurement of regional organ blood flow, has so far not been used for the determination of blood flow in bone. The aim of the present study was to establish FM for the quantification of regional bone blood flow (RBBF). Female, anesthetized New Zealand rabbits (n = 6) received left ventricular injections of different amounts of FM at six subsequent time points. In order to examine the precision of RBBF determination, two different FM species were injected simultaneously at the sixth injection. At the end of the experiments the femoral and tibial condyles of each hind limb were removed and the fluorescence intensity in the tissue samples was measured by an automated procedure. In an in vitro study we have shown that acid digestion of the crystalline matrix has no effect on the fluorescence characteristics of FM. The determination of the number of spheres per tissue sample revealed that depending on the tissue sample size up to 3 x 10(6) spheres/injection were necessary to obtain about 400 microspheres in the individual bone samples. RBBF values of the tibial and femoral condyles did not differ at various injection intervals. The tibial blood flow values varied between 6.6 +/- 1.1 and 8.5 +/- 1.4 ml/min/100 g and were significantly higher than those of the femur (4.3 +/- 1.1 to 6.0 +/- 1.8 ml/min/100 g). The bone blood flow values obtained by simultaneous injection of two FM species correlated significantly (r = 0.96, slope = 1.06, intercept = 0.05), the mean difference was 0.39 +/- 1.11 ml/min/100 g. Our data demonstrate that the measurement of RBBF by means of FM allows a valid determination of RBBF.

Validity of fluorescent microspheres method for bone blood flow measurement during intentional arterial hypotension.

In this study, we compared bone blood flow values obtained by simultaneously injected fluorescent (FM) and radiolabeled microspheres (RM) at stepwise reduced arterial blood pressure. Ten anesthetized female New Zealand White rabbits received simultaneous left ventricular injections of FM and RM at 90, 70, and 50 mmHg mean arterial blood pressure (MAP). After the experiments, both kidneys and long bones of all four limbs were removed and dissected in a standardized manner. Radioactivity (corrected for decay, background, and spillover) and fluorescence were determined, and blood flow values were calculated. Relative blood flow values estimated for each bone sample by RM and FM were significantly correlated (r = 0.98, slope = 0.99, and intercept = 0.04 for 90 mmHg; r = 0.98, slope = 0.94, and intercept = 0.09 for 70 mmHg; r = 0.98, slope = 0.96, and intercept = 0.07 for 50 mmHg). Blood flow values (ml x min-1 x 100 g-1) of right and left bone samples determined at the different arterial blood pressures were identical. During moderate hypotension (70 mmHg MAP), blood flow in all bone samples remained unchanged compared with 90 mmHg MAP, whereas a significant decrease of bone blood flow was observed at severe hypotension (50 mmHg MAP). Our results demonstrate that the FM technique is valid for measuring bone blood flow. Differences in bone blood flow during altered hemodynamic conditions can be detected reliably. In addition, changes in bone blood flow during hypotension indicate that vasomotor control mechanisms, as well as cardiac output, play a role in setting bone blood flow.

Binding kinetics of triclosan (Irgasan) to alloplastic vascular grafts: an in vitro study.

The aim of this study was to investigate the binding kinetics of triclosan (Irgasan) to alloplastic vascular grafts and to examine its antimicrobial activity against various microbial pathogens in vitro. Vascular grafts made by Intergard (Intervascular), Fluoropassiv (Vascutek), and Gore-tex (Gore) were examined. Grafts were incubated in 10 g/L triclosan (Irgasan), dried, sterilized, and incubated in RPMI medium. One-centimeter segments of the grafts were resected under sterile conditions at intervals of minutes, then hours, followed by days and up to 4 weeks. Samples were stored frozen at -20 degrees C for the measurement of triclosan bound to the vascular graft by high-performance liquid chromatography (HPLC). The binding kinetics under perfusion conditions were determined for Intergard grafts, which were perfused with 50 mL of nutrient medium for 24 hr. Samples were taken at various time intervals for the measurement of triclosan. The antimicrobial activity of triclosan against Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans as well as Enterococcus faecium was determined. Triclosan effectively binds to vascular graft without the use of intermediate binding substances. It stayed on the graft for the duration of 4 weeks. Under both static and perfusion conditions, the binding kinetics are similar. Triclosan binds most effectively to Intergard grafts, less so to Fluoropassiv grafts, and not at all to Gore-tex material. Antimicrobial activity of triclosan is very effective against S. aureus and E. faecium but not against P. aeruginosa.

Oxidant stress in hyperlipidemia-induced renal damage.

Hyperlipoproteinemia can aggravate glomerulosclerosis and chronic tubulointerstitial (ti) damage in kidneys without primary immunologic disease. We evaluated whether the effect of hyperlipidemia on progression of renal damage differed between kidneys without preexisting glomerular disease and kidneys with mesangioproliferative glomerulonephritis and whether the renal actions of hyperlipidemia were dependent on oxidant-antioxidant balance. Hyperlipidemia was induced by high-fat and high-cholesterol diet in uninephrectomized rats. In rats without glomerulonephritis, hyperlipidemia led to a rise in glomerular and ti generation of reactive oxygen species (ROS). Oxygen radicals were mainly generated by enhanced xanthine oxidoreductase (XO), which rose with protein concentration and activity during hyperlipidemia; concurrently, glomerulosclerosis and chronic ti injury were noticed during hyperlipidemia [ti damage (% of total tubulointerstitium (TI) after 150 days): normolipidemia 0.1 +/- 0% vs. hyperlipidemia 3.4 +/- 0. 9%; P < 0.05]. In mesangioproliferative Thy-1 nephritis, ti injury was significantly accelerated by hyperlipidemia (ti damage after 150 days: normolipidemic Thy-1 nephritis 2.5 +/- 0.6% vs. hyperlipidemic Thy-1 nephritis 12.5 +/- 3.1%; P < 0.05). Antioxidant enzyme activities decreased and XO activity rose markedly in the TI (XO activity in TI after 150 days: normolipidemic Thy-1 nephritis 2.2 +/- 0.5 vs. hyperlipidemic Thy-1 nephritis 4.5 +/- 0.7 cpm/microg protein; P < 0.05). In hyperlipidemic Thy-1 nephritis rats, which had a higher urinary protein excretion than normolipidemic rats, hypochlorite-modified proteins, an indirect measure for enhanced myeloperoxidase activity, were detected in renal tissue and in urine, respectively. During hyperlipidemia, chronic damage increased in renal TI. Enhanced generation of ROS, rise in oxidant enzyme activity, and generation of hypochlorite-modified proteins in renal tissue and urine were noticed. These data suggest that oxidant stress contributed to the deleterious effects of hyperlipidemia on the renal TI.

Reduction of circulating cholesterol and apolipoprotein levels during sepsis.

Sepsis with multiple organ failure is frequently associated with a substantial decrease of cholesterol levels. This decrease of cholesterol is strongly associated with mortality suggesting a direct relation between inflammatory conditions and altered cholesterol homeostasis. The host response during sepsis is mediated by cytokines and growth factors, which are capable of influencing lipid metabolism. Conversely lipoproteins are also capable of modulating cytokine production during the inflammatory response. Therefore the decrease in circulating cholesterol levels seems to play a crucial role in the pathophysiology of sepsis. In this review the interaction between cytokines and lipid metabolism and its clinical consequences will be discussed.

Association of serum tumor necrosis factor levels with decrease of cholesterol during septic shock.

Recent studies suggest that release of cytokines during inflammatory states such as septic shock leads to hypocholesterolemia. To examine whether tumor necrosis factor alpha (TNF), which is the major cytokine in inflammatory disease, causes hypocholesterolemia, we measured serum levels of total (bioactive and receptor-bound) TNF, cholesterol, Apo B, and Apo A1 in seven patients with septic shock over a period of 8 days. Since elevated serum TNF levels are accompanied by the release of soluble TNF receptors, levels of TNF receptors p55 and p75 were also measured. Patients with septic shock had significantly higher serum TNF and TNF receptor levels compared with healthy controls. Increased cytokine levels were accompanied by a significant decline in total serum cholesterol apolipoprotein A1 and B. In vitro studies with cultured human skin fibroblasts, human umbilical vein endothelial cells, and HepG2 hepatoma cells showed that TNF increased the degradation of 125I-labeled low-density lipoprotein in all the cell lines tested. Recombinant soluble TNF receptors inhibited the TNF-induced stimulation of low-density lipoprotein receptor in a concentration-dependent manner. However, the calculated ratio of TNF receptors to total TNF measured in serum of these patients was not able to counteract the stimulatory effect of TNF, possibly due to the higher molar excess of TNF receptors required to achieve this effect in vitro. Our data strengthen the hypothesis that serum values of total TNF determine the extent of hypocholesterolemia during sepsis and septic shock despite the presence of a high concentration of TNF receptors. Studies with recombinant TNF also confirm the role of TNF in hypocholesterolemia in inflammation.

Validation of an automated enzyme immunoassay for Interleukin-6 for routine clinical use.

Serum levels of Interleukin-6 (IL-6), a proinflammatory cytokine, are increased in early stages of inflammatory diseases such as infection and sepsis. Assay systems which permit its measurement within a few hours and as a single measurement have not been reported so far. We therefore evaluated a now commercially available automated method for IL-6 measurement on the Cobas Core immunological analyzer (Roche Diagnostic Systems) which enables single IL-6 measurement within about 1 hour. The automated assay correlates well with an established, manual microtiter plate assay (Biosource GmbH) which uses the same antibodies and reagents (r=0.98). Accuracy of the automated method was established by adding known amounts of IL-6 international reference preparation. Recovery of the international standard was in the range of 92104%. The automated assay had a precision of singletons below 6% and was linear up to 2800 pg/ml. This automated assay provides a suitable, convenient and time saving method for measurement of IL-6 serum levels in the routine clinical laboratory.

Dysregulation of monocytic nuclear factor-kappa B by oxidized low-density lipoprotein.

Nuclear factor-kappa B (NF-kappa B)/Rel transcription factors may be involved in atherosclerosis, as is suggested by the presence of activated NF-kappa B in human atherosclerotic lesions. The aim of the present study was to investigate the effects of oxidized LDL (oxLDL) on the NF-kappa B system in human THP-1 monocytic cells as well as adherent monocytes. Our results demonstrate that short-term incubation of these cells with oxLDL activated p50/p65 containing NF-kappa B dimers and induced the expression of the target gene IL-8. This activation of NF-kappa B was inhibited by the antioxidant and H2O2 scavenger pyrrolidine dithiocarbamate and the proteasome inhibitor PSI. The oxLDL-induced NF-kappa B activation was accompanied by an initial depletion of I kappa B-alpha followed by a slight transient increase in the level of this inhibitor protein. In contrast, long-term treatment with oxLDL prevented the lipopolysaccharide-induced depletion of I kappa B-alpha, accompanied by an inhibition of both NF-kappa B activation and the expression of tumor necrosis factor-alpha and interleukin-1 beta genes. These observations provide additional evidence that oxLDL is a potent modulator of gene expression and suggest that (dys)regulation of NF-kappa B/Rel is likely to play an important role in atherogenesis.

Role of nuclear factor-kappa B in atherogenesis.

Transcription factors of the nuclear factor-kappa B (NF-kappa B)/Rel family have an important function in the regulation of a variety of genes involved in the inflammatory and proliferative responses of cells. Recent studies strongly indicate that the inducible transcription factor NF-kappa B is involved in the pathogenesis of atherosclerosis. Activated NF-kappa B is present in the fibrotic thickened intima-media and atheromatous areas of the atherosclerotic lesion, within smooth muscle cells, macrophages and endothelial cells, whereas little or no activated NF-kappa B can be detected in vessels lacking atherosclerosis. A variety of molecules have been identified in the atherosclerotic environment that are able to activate NF-kappa B in vitro. Furthermore, an increased expression of numerous genes known to be regulated by NF-kappa B has been found in the atherosclerotic lesion. Possible functional implications for activated NF-kappa B in atherogenesis are discussed here. The activation and role of NF-kappa B in atherosclerosis may provide a model for the involvement of the transcription factor in human chronic inflammatory disease.

Determination of LDL- and scavenger-receptor activity in adherent and non-adherent cultured cells with a new single-step fluorometric assay.

Lipoproteins labeled with 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) are widely used to visualize LDL-and scavenger-receptor activity in cultured cells. The purpose of this study was to evaluate a new single-step fluorometric assay with high sensitivity for the quantitative determination of the LDL- or scavenger-receptor activity in adherent and non-adherent cells. We used an aqueous solution of 1 g/l SDS dissolved in 0.1 M NaOH to lyse the cells after incubation with DiI-LDL or DiI-acetylated LDL. This allows for the first time the determination of fluorescence intensity and cell protein in the same sample without prior lipid extraction of the fluorochrome. Fluorescence of the cell lysates was determined in microtiter plates with excitation-emission set at 520 and 580 nm, respectively. This rapid method demonstrates high specificity for determining the LDL- and scavenger-receptor activity in cultured cells (e.g., human skin fibroblasts from patients with and without familial hypercholesterolemia; human U-937 monocyte and murine P388 D1 macrophage cell lines). The validity of our fluorescence assay is demonstrated by comparison of cellular uptake and metabolism of lipoproteins labeled with both, DiI and 125iodine. The rapidity and accuracy of this assay allows its routine application for studying receptor-mediated lipoprotein uptake in various cell types.

Hyperlipoprotein(a)aemia in nephrotic syndrome.

The nephrotic syndrome is frequently associated with hyperlipidaemia and hyperfibrinogenaemia, leading to an increased coronary and thrombotic risk, which may be enhanced by high lipoprotein (a) [Lp(a)] concentrations. We followed the quantitative and qualitative pattern of plasma lipoproteins over 18 months in a patient with nephrotic syndrome suffering from premature coronary artery disease and with elevated level of Lp(a) (470 mg dL-1). Analysis of kinetic parameters after heparin-induced extracorporeal plasma apheresis revealed a reduced fractional catabolic rate for both low-density lipoprotein (LDL) and Lp(a). After improvement of the nephrotic syndrome, Lp(a) decreased to 169 mg dL-1 and LDL concentrations were normalized. The decrease of Lp(a) was associated with an increase in plasma albumin concentrations. Analysis of apo(a) isoforms in the patient showed the presence of isoform S2 (alleles 10 and 19). Consequently, the authors' present strategy is to normalize the elevated Lp(a) and fibrinogen levels. For this purpose heparin-mediated extracorporeal LDL precipitation (HELP) apheresis is a promising regimen, helping to reduce the thrombotic risk and prevent coronary and graft atherosclerosis as well as the progression of glomerulosclerosis in our patient.

Study of causes underlying the low atherosclerotic response to dietary hypercholesterolemia in a selected strain of rabbits.

We have recently characterized a strain of rabbits that shows a low atherosclerotic response (LAR) to dietary hypercholesterolemia in contrast to the usual high atherosclerotic response (HAR) of rabbits [1]. Presently, we have focused on three well established and important stages of atherogenesis, i.e., monocyte adhesion to endothelium, cell mediated peroxidative modification of lipoproteins and induction of a receptor that recognizes modified low density lipoprotein (LDL). The results obtained show that (1) beta-very low density lipoprotein (beta-VLDL) from LAR and HAR rabbits enhanced monocyte adhesion to endothelial cells to the same extent; (2) Cell mediated peroxidation of LDL and beta-VLDL, tested by loss of alpha-tocopherol and formation of thiobarbituric acid reacting substances (TBARS), was compared using macrophages, fibroblasts and smooth muscle cells (SMC) of LAR and HAR rabbits and no significant differences were found; (3) Induction of scavenger receptor by phorbol ester (phorbol 12-myristate 13-acetate (PMA)) and platelet-derived growth factor-BB (PDGF-BB) was determined in SMC or fibroblasts from LAR and HAR rabbits using 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate-acetylated LDL (DiL-acLDL). We found a significantly higher uptake of DiI-acLDL in SMC and fibroblasts derived from HAR rabbits as compared with cells from LAR rabbits. Similar results were also obtained with [125I]-acLDL in fibroblasts from LAR and HAR rabbits with respect to cellular lipoprotein degradation after PMA pretreatment. Even though the attenuated atherosclerotic response to hypercholesterolemia of LAR rabbits may have multiple underlying causes, the most prominent so far is an apparent difference in inducibility of scavenger receptor in SMC and fibroblasts.

The role of lipids in nephrosclerosis and glomerulosclerosis.

Hyperlipidemia and lipoprotein abnormalities are often encountered in patients with nephrotic syndrome or chronic renal disease and also in those undergoing haemodialysis and with renal transplant. Even though the significance of lipid deposition in renal tissue and the role of lipoproteins in the pathogenesis of renal disease in man is unclear, experimental and clinical data indicate a possible damaging effect of a disturbed lipid metabolism on the kidney. In humans, glomerular lipid deposition is observed in genetic diseases such as Fabry's disease, lecithin:cholesterol acyltransferase activity (LCAT) deficiency and arteriohepatic dysplasia, and in diseases with acquired disturbance of lipid metabolism such as nephrotic syndrome and cholestatic liver disease. Studies on animals with lupus nephritis, aminonucleoside nephrosis, reduced renal mass, diabetes mellitus or systemic hypertension have shown that cholesterol can increase the incidence of glomerulosclerosis. As most of these studies have been performed in the rat, which has a different lipoprotein profile to that of man, these results should be carefully interpreted with regard to their relevance for humans. In vitro cell culture studies on human glomerular cells have given some preliminary insights into the cellular mechanisms of lipid induced glomerular damage. Apo E-containing lipoproteins, which are pathologically elevated in many renal diseases, are avidly taken up by human mesangial cells. These cells seem to play a central role in the initiation of glomerulosclerosis by inducing proliferation and production of excess extracellular matrix. Lipoproteins are able to stimulate DNA synthesis in these cells, and increase the synthesis of mitogens and extracellular matrix protein. The pathogenic role of oxidized lipoproteins has not yet been defined. Human mesangial cells do not seem to take up these modified lipoproteins. However, macrophages infiltrate glomeruli and may constitute the stimulus for the generation of minimally modified lipoproteins and their cellular uptake. The data from animal experiments suggest that treatment that corrects hyperlipidemia may have an ameliorative effect on renal function. Thus, there are strong indications that lipoproteins may play a critical role in mediating the development of glomerulosclerosis.