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Biomedical subjects

G Lonnemann

Publications and source records attributed to G Lonnemann.

At least 73 records · Page 4Linked to original sources
PubMed1988

Measurement of immunoreactive interleukin-1 beta from human mononuclear cells: optimization of recovery, intrasubject consistency, and comparison with interleukin-1 alpha and tumor necrosis factor.

Numerous studies have reported altered levels of in vitro production of the cytokines interleukin-1 (IL-1) and tumor necrosis factor (TNF) from blood leukocytes in various human disease states. Most of these studies have used bioassays which are vulnerable to inhibitors produced by these cells. Furthermore in vitro cytokine production is often assessed on a single occasion. The present study was designed to standardize stimulation conditions for in vitro IL-1 beta production and to employ a competitive radioimmunoassay (RIA) to demonstrate reproducibility and long-term variation of in vitro cytokine production in a cohort of healthy human subjects. We also examined relative amounts of immunoreactive IL-1 beta, IL-1 alpha, and TNF induced by the stimuli endotoxin, phytohemagglutinin, or Staphylococcus epidermidis. We show that the RIA can reliably detect IL-1 beta produced from mononuclear cells in concentrations as low as 115 pg/ml. Lysing cells by repeated freeze-thawing yields maximal recovery of total (i.e., secreted plus cell-associated) immunoreactive IL-1 beta, when compared to extraction with the detergent CHAPS or addition of protease inhibitors. Repeated measurement of in vitro cytokine production on different days within 1 week shows good reproducibility for a given individual and a given stimulus (variation coefficient 20 to 30%). Over a long time period (6 months) in vitro cytokine production is stable in some individuals but changes considerably in others. The soluble stimulus endotoxin induces twofold more IL-1 alpha than IL-1 beta or TNF; in contrast the phagocytic stimulus heat-killed S. epidermidis induces fourfold more IL-1 beta and TNF than IL-1 alpha. This distinct pattern of cytokine response indicates differential stimulation of the mononuclear cells by different stimuli. The results form the basis for studying in vitro cytokine production in different human disease states.

Animals↗
PubMed1988

Detection of endotoxin-like interleukin-1-inducing activity during in vitro dialysis.

In order to study the integrity of dialysis membranes to pyrogens, the dialysate side of a closed loop hemodialysis (HD) circuit was challenged with E. coli microfiltrate containing 500 ng/ml endotoxin. Three solutions, a) tissue culture medium/saline, b) 5% human serum albumin, and c) 10% fresh human plasma, were circulated in the blood loop for five hours. Samples drawn from the blood side were assayed for interleukin-1 (IL-1)-inducing activity on human mononuclear cells (MNC) in vitro. No IL-1-inducing substances were detected when saline or culture medium was circulated in the blood loop. Circulating 5% human serum albumin revealed IL-1-inducing activity in the samples drawn only after five hours of HD. However, the addition of 10% fresh human plasma to the blood side resulted in the appearance of an IL-1-inducing substance(s) after 15 minutes of HD. After 30 minutes, maximum IL-1-inducing activity was observed (control stimulation index, 3.30 +/- 0.67 SEM vs. 7.59 +/- 1.50, P less than 0.02). The IL-1-inducing activity of the samples was completely inhibited by polymyxin B, a cationic antibiotic which blocks the IL-1-inducing activity of endotoxin. Additional experiments demonstrated that in vitro MNC IL-1-production induced by the same E. coli microfiltrate is enhanced in the presence of 10% plasma. These studies demonstrate that: (a) in the presence of plasma, IL-1-inducing factors pass into the blood compartment of a dialysis system challenged with bacterial pyrogen; and (b) MNC production of IL-1 is enhanced in the presence of plasma.(ABSTRACT TRUNCATED AT 250 WORDS)

Endotoxins↗
PubMed1988

Plasma interleukin-1 activity during hemodialysis: the influence of dialysis membranes.

Plasma interleukin-1 (IL-1) activity was measured in 7 stable ESRD patients on regular hemodialysis for no less than 5 months. Predialysis levels were significantly raised compared to 8 normal control subjects. During hemodialysis with four different membranes, plasma IL-1 activity rose with Cuprophan and Hemophan and was unchanged or reduced with Gambrane and Polysulfon. In spite of these differences, body temperature rose in all forms of hemodialysis. Factors responsible for the predialysis elevation included the absence of renal function and/or the repeated stimulus of human blood monocytes by hemodialysis. In view of the uniform increase of body temperature during hemodialysis, the differences in changes of plasma IL-1 activity observed with the various membranes may not be caused by a variable stimulation of monocytes but rather by the presence or absence of the membrane's ability to remove and/or absorb IL-1. Thus, the consequences of monocyte hemodialysis stimulation may be obtained locally, even in the presence of unchanged or reduced plasma IL-1 activity.

Adult↗
PubMed1988

Interleukin-1 beta in human plasma: optimization of blood collection, plasma extraction, and radioimmunoassay methods.

Current immunoassays for interleukin-1 beta (IL-1 beta) are effective for analyzing fluids derived from cultured cells. However, IL-1 beta determinations in human plasma or serum samples are technically complicated by higher protein and lipid concentrations, physicochemical differences which exist between samples from healthy subjects and those experiencing acute phase responses, and by the fact that IL-1 beta can be produced and degraded in the blood collection tube after the sample is drawn. A simple chloroform extraction process has been developed which eliminates several of the interfering factors from plasma samples and increases the amount of IL-1 beta detected by radioimmunoassay and lymphocyte activation assay. In the radioimmunoassay, rabbit sera was found to influence the accuracy and variability of plasma measurements. Improvements in radioimmunoassay reagents and methods are reported which reduce this influence. Finally, different concentrations of IL-1 beta were measured depending on whether serum or plasma was tested. We propose that plasma samples collected with EDTA and aprotinin provide a better determination of free circulating IL-1 beta in vivo than serum samples, which may contain IL-1 beta secreted from blood leukocytes during the clotting process.

Acute-Phase Reaction↗
PubMed1988

Studies on the ability of hemodialysis membranes to induce, bind, and clear human interleukin-1.

Interleukin-1 (IL-1) is a polypeptide cytokine predominantly produced by monocytes in response to injury or infection. Effects of IL-1 such as fever, acute phase protein synthesis, hypotension, and loss of body mass are complications of hemodialysis. Endotoxin-contaminated dialysate fluid, sodium acetate as a dialysate buffer, and activated complement can induce IL-1 during hemodialysis. In the present study, we investigated the intrinsic property of hemodialysis membranes to stimulate human blood mononuclear cells (MNC) to produce IL-1. Incubation of MNC on sheets of two commonly used hemodialysis membranes, regenerated cellulose (RC) and polyacrylonitrile (PAN) resulted in significant induction of IL-1 in the absence of endotoxin or complement. Production of intracellular and extracellular IL-1 was greater in MNC exposed to PAN compared with RC (p less than 0.01). Similar results were obtained when MNC were exposed to strips of hemodialysis membranes in rotating tubes. However, compared with RC, PAN binds significant amounts of human IL-1. In a model of in vitro hemodialysis, radiolabeled recombinant human IL-1 was added to the blood compartment of a PAN and a RC dialyzer. Forty percent of radiolabeled IL-1 bound to the PAN dialyzer membrane compared with 10% to the membrane of a RC dialyzer. In addition, 22% of radiolabeled IL-1 was found in the dialysate compartment of a PAN dialyzer after 1 hour whereas 1% was found in the dialysate side of a RC dialyzer; these results were confirmed by measuring bioactivity of IL-1. These studies demonstrate the intrinsic property of hemodialysis membranes to stimulate human IL-1 production; in addition they establish that dialysis membranes differ in their ability to bind and clear IL-1.

Acrylic Resins↗
PubMed1988

A radioimmunoassay for human interleukin-1 alpha: measurement of IL-1 alpha produced in vitro by human blood mononuclear cells stimulated with endotoxin.

A specific radioimmunoassay (RIA) for human interleukin-1 alpha (IL-1 alpha) which detects less than 25-50 pg/ml IL-1 alpha is described. Although human IL-1 alpha shares structural homology, receptors and multiple biological properties with IL-1 beta, this RIA does not detect human IL-1 beta or other human cytokines. We recovered nearly 100% of IL-1 alpha added to fresh human heparinized blood or freshly voided urine; in contrast, using a specific RIA for IL-1 beta, recovery of IL-1 beta added to fresh blood is approximately 50% reduced by nonspecific factors. In the present study, we employed this RIA to measure the amount of total (extracellular and cell-associated) immunoreactive IL-1 alpha produced by human blood mononuclear cells stimulated in vitro by different concentrations of endotoxin. Using ultrafiltered culture medium to reduce endotoxin content, there was no detectable (less than 50 pg/ml) IL-1 alpha produced after 24 hours. Endotoxin (0.5 ng/ml) induced a mean concentration of 900 pg/ml (range 180-1660 pg/ml). At higher concentrations of endotoxin (500 ng/ml), a mean of 6,990 pg/ml (range 415-11,900 pg/ml) was produced. These levels were comparable to the amount of IL-1 beta produced under similar culture conditions. The results indicate that IL-1 alpha can be measured independently of IL-1 beta in human body fluids and from human mononuclear cells.

Endotoxins↗
PubMed1987

Interleukin 1 induces interleukin 1. I. Induction of circulating interleukin 1 in rabbits in vivo and in human mononuclear cells in vitro.

Interleukin 1 (IL-1) plays an important role in host defense mechanisms by increasing body temperature, inducing the synthesis of a variety of lymphokines and hepatic acute phase proteins and acting as a chemoattractant for lymphocytes. However, in some microenvironments such as injured tissue or joint spaces, elevated IL-1 levels may contribute to pathologic processes, for example, proliferation and fibrosis of tissue involved in pannus formation as well as degradation of matrix and abnormal tissue architecture. To investigate potential mechanisms that may lead to excessive production of IL-1, we have examined the ability of IL-1 to participate in an amplification event by inducing its own gene expression leading to synthesis of biologically active IL-1. When injected into rabbits, recombinant human IL-1-alpha induced biphasic fevers, and during the second temperature elevation 3 hr later, a circulating pyrogenic material was detected by passive transfer of plasma to other rabbits. Induction of the biphasic fever was not caused by endotoxin contamination of the recombinant IL-1. The 3-hr circulating pyrogen was heat-labile and was not residual injected IL-1-alpha. Chromatographic separation of this plasma and biologic assay suggested that it was new IL-1 of rabbit origin. We next incubated human blood mononuclear cells with recombinant IL-1-alpha and measured the intracellular and extracellular levels of IL-1 by bioassay using the D10.G4.1 murine T cell line. In order to control for the carryover of recombinant IL-1-alpha used to stimulate the mononuclear cells (MNC), we used neutralizing antibodies that were specific for IL-1-alpha or IL-1-beta. The results of these neutralizations showed that recombinant human IL-1-alpha induces the synthesis of IL-1-beta in human MNC in vitro. These results were verified with a radioimmunoassay specific for IL-1-beta. At concentrations of 100 ng/ml, IL-1-alpha induced prostaglandin E2 production in the MNC culture, and this was associated with decreased production of immunoreactive IL-1-beta. Adding indomethacin to the cultures prevented the decreased production of IL-1-beta induced by high concentrations of IL-1-alpha. Using nonadherent MNC, we observed an increase in IL-1-beta as well as IL-1-alpha mRNA after 4 hr of exposure to recombinant IL-1-alpha. These results demonstrate that IL-1-alpha induces biologically active and immunoreactive IL-1-beta from MNC in vitro and that the same concentrations of IL-1-alpha induce gene expression for both forms of IL-1.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Enhancement of in-vitro human interleukin-1 production by sodium acetate.

Human blood monocytes were incubated in vitro in the presence of various concentrations of sodium acetate or sodium chloride or with medium alone. Intracellular and extracellular levels of interleukin-1 (IL-1) were measured. The production of intracellular IL-1 and the release of extracellular IL-1 were higher in the presence of acetate than in the presence of chloride or in medium alone. The concentrations of acetate used were comparable to those encountered by blood monocytes on the surface of haemodialysis membranes. Since complications of peritoneal dialysis, such as loss of ultrafiltration and progressive fibrosis of the peritoneum, have been associated with the use of sodium acetate as the exchange-fluid buffer, these results suggest that the widespread use of sodium acetate as a buffer during haemodialysis may be contraindicated.

Acetates↗
PubMed1987

Ultrafiltration to reject human interleukin-1-inducing substances derived from bacterial cultures.

Interleukin-1 (IL-1), a polypeptide cytokine, is an important mediator of host responses to infection and injury. Picogram per milliliter concentrations of bacterial products (endo- or exotoxins) stimulate human monocytes to produce IL-1 in vitro. The design of this study was based on the clinical model of bacterial contamination of fluid intended to be directly injected into humans. Physiologic saline contaminated with bacterial toxins was passed through a hollow fiber ultrafilter, and the ultrafiltrates were tested for their ability to induce human IL-1 production. The ultrafiltrates were added directly to freshly obtained human blood mononuclear cells, and after 24 h of incubation the supernatant media were assayed for the presence of IL-1. The results indicate that the IL-1-inducing material(s) present in bacterial cultures of gram-negative organisms is rejected by a factor of 100 to 100,000 by molecular size exclusion and by absorption; rejection is sustained for at least 32 liters of fluid; the rejection of Limulus-reactive material by the ultrafilter is greater for purified endotoxin than for native endotoxins derived from live bacterial cultures; and nonendotoxin IL-1-inducing toxins (molecular weight, 24,000) from Staphylococcus aureus are not rejected or absorbed. These results demonstrate that there is a considerable margin of safety with the ultrafiltration method and that it can be applied to clinical situations.

Bacterial Toxins↗
PubMed1987

Plasma interleukin-1 activity in humans undergoing hemodialysis with regenerated cellulosic membranes.

Because of the similarity between the acute phase response and the post-hemodialysis syndrome, it has been proposed that human blood monocytes adhering to the hemodialysis membrane are exposed to several inducers of interleukin-1 (IL-1). These include endotoxin from the dialysate fluid and C5a activated on the surface of regenerated cellulosic hemodialysis membranes. In order to test this hypothesis, we measured IL-1 plasma activity by employing gel filtration to remove IL-1 inhibitors and subsequent augmentation of lectin-induced murine thymocyte proliferation in each fraction. To evaluate the acute response of end stage renal disease patients during a single hemodialysis with a regenerated cellulosic membrane, plasmas were tested before and after hemodialysis. Control plasmas were obtained from healthy individuals. Plasma IL-1 activity eluted at a molecular weight of predominantly 15 kD. This material was neutralized by anti-human-IL-1 which does not recognize IL-2. IL-1 activity was detected in the pre-dialysis plasma of 6 out of 7 patients and rose in 6 patients following a 5 hour hemodialysis. This increase was associated with a significant rise in body temperature. This study supports the hypothesis that hemodialysis with regenerated cellulose might augment IL-1 production in endstage renal disease patients maintained on longterm hemodialysis. It demonstrates for the first time that dialysis patients have detectable plasma levels of IL-1 compared to normal individuals.

Adult↗
PubMed1987

Development and use of a radioimmunoassay for human interleukin-1 beta.

Interleukin-1 (IL-1) mediates immunological, physiological, and metabolic changes associated with inflammation and host defense systems. Measurement of IL-1 activity can vary with the bioassay employed and substances have been described in several bioassay systems that either inhibit or mimic IL-1 activity. We now report a sensitive, rapid (24 hour), and specific radioimmunoassay for human IL-1 beta. Using 125I-labeled IL-1 beta and polyclonal rabbit antisera raised against recombinant human IL-1 beta, a competitive inhibition assay is described which detects 250 pg/ml of recombinant human IL-1 beta and 500 pg/ml of pI 7 human monocyte IL-1. The assay does not detect human IL-1 alpha, human interleukin-2, human tumor necrosis factor-alpha or human interferon-gamma. Nearly 100% of IL-1 beta added to human serum or urine can be quantitatively recovered. Substances such as fetal calf serum, phytohemagglutinin, opsonized Staphylococcus albus or E. coli endotoxin do not affect the assay. Using this assay, IL-1 beta was measured in both the intracellular and extracellular compartments of stimulated human blood mononuclear cells. These results were unaffected by the presence of substances that interfere with bioassays for IL-1 such as indomethacin or BW 755C, a lipoxygenase inhibitor. These studies establish the usefulness of quantitating immunoreactive human IL-1 beta produced by human blood cells and that which may be present in human body fluids in the presence of other cytokines.

Animals↗
PubMedDate not supplied

Plasma C-reactive protein is linked to backfiltration associated interleukin-6 production.

Bacterial contamination of dialysate may enhance cytokine production in hemodialysis. The authors tested the hypothesis that C-reactive protein and interleukin-6 (IL-6) may be linked in a large group of patients exposed to backfiltration of dialysate over a long period of observation. Plasmas stored in a recently published multicenter study were reevaluated. Plasma C-reactive protein and IL-6 concentrations in patients with chronic uremia undergoing hemodiafiltration, which is known to be associated with backfiltration (Group II, 12 patients), were compared with those found in patients treated with a modified hemodiafiltration modality without backfiltration (Group I, 16 patients), and in patients shifted from one modality to the other (Group III, 27 patients), and in 10 patients on hemodialysis (Group IV) in a 1 year multicenter study. Plasma C-reactive and IL-6 both increased significantly (p < 0.002), but slowly (after 8 months) in Group II compared with I, and during the 4 month period in hemodiafiltration with backfiltration in Group III. Backfiltration of dialysate with a moderate to low degree of contamination may enhance synthesis of cytokine and C-reactive protein in the long term. Thus, the relevance for dialytic strategies aiming at improving dialysate quality or at reducing backfiltration is highlighted.

C-Reactive Protein↗