Serum levels of helper factors (IL-1 alpha, IL-1 beta and IL-6), T-cell products (sCD4 and sCD8), sIL-2R and beta 2-microglobulin in patients with B-CLL and benign B lymphocytosis.

Chronic B-lymphocytic leukemia (B-CLL) cells may be regulated by immune functions. In an attempt to analyze such functions, helper factors (IL-1 alpha, IL-1 beta and IL-6), T-cell products (sCD4 and sCD8) and sIL-2R and beta 2-microglobulin were measured in serum of patients at different stages of the disease. Patients were classified as having monoclonal lymphocytosis of undetermined significance (MLUS), stable or progressive B-CLL respectively. A significant, but modest, increase of IL-1 alpha was found in B-CLL as well as in MLUS patients whereas IL-6 levels were increased in MLUS only. sCD8 levels were increased both in MLUS and B-CLL but augmented sCD4 concentrations were found statistically significant only in progressive B-CLL. beta 2-microglobulin and sIL-2R were related to the extent of the monoclonal B-cell fraction. The data indicate an increased T-suppressor activity in both MLUS and B-CLL patients and a selective increase of helper T-cell activity in progressive B-CLL. A possible immunoregulatory influence of helper T cells on disease progression is discussed.

Increased interleukin-6 levels in cerebrospinal fluid following subarachnoid hemorrhage.

Serum and cerebrospinal fluid (CSF) samples from 12 patients were analyzed for interleukin (IL)-6, soluble IL-2 receptor (IL-2R), and soluble CD8 levels in order to determine the immune activation profile following subarachnoid hemorrhage (SAH). Dramatically increased levels of IL-6 and moderate increases of soluble IL-2R were detected in the CSF in 11 of the 12 patients; slightly elevated levels of soluble CD8 were observed in six patients. The IL-6 levels were higher on Day 6 than on Days 3 and 9. The increases in IL-6, soluble IL-2R, and soluble CD8 levels in the CSF samples were not paralleled by increased values in the serum samples, and thus probably reflected an intrathecal synthesis of the cytokine. Passive transfer of IL-6 across the blood-brain barrier seemed not to occur since the serum and CSF levels of IL-6 showed a negative correlation. The findings suggest a severe inflammatory affection of the central nervous system that could be of importance in understanding the clinical course in patients following SAH.

Postoperative insulin resistance and circulating concentrations of stress hormones and cytokines.

Insulin sensitivity was determined before and after elective surgery in 31 otherwise healthy patients undergoing elective surgery for open cholecystectomy (n = 24) or inguinal hernia repair (n = 7) and compared with concomitant plasma concentrations of stress hormones and cytokines. Insulin sensitivity was determined employing the normoglycaemic, hyperinsulinaemic clamp at a plasma insulin concentration of 380 pmol/I and a blood glucose concentration of 4.5 mmol/I. Five of the patients undergoing cholecystectomy were studied again on days 5, 9 and 20 after surgery. Preoperative insulin sensitivity ranged from 2.2 to 14.3 mg/kg/min. All patients exhibited reduced insulin sensitivity on the first postoperative day and the mean value fell from 4.7 (0.4) to 2.7 (0.5) mg/kg/min. More pronounced reductions were found after cholecystectomy. A significant increase was found in plasma concentrations of interleukin-6 (IL-6) postoperatively as compared to preoperative values. However, no significant changes were seen in the postoperative plasma concentrations of any of the hormones studied in patients undergoing hernia repair, while minor increments were seen in patients undergoing open cholecystectomy. There was a significant (r = 0.50, P = 0.005) linear relationship between the reduction in relative insulin sensitivity and the concomitant plasma levels of IL-6. However, no such relation could be confirmed between the changes in plasma hormone concentrations (neither absolute nor relative changes) and the simultaneous alteration in relative insulin sensitivity. In addition, after including three patients who had undergone ileo-anal pouch construction surgery, the relationship between postoperative insulin sensitivity and IL-6 levels was even stronger (r = 0.62, P = 0.001). These results suggest that the immunomodulating effects of endogenous IL-6 is of importance in the acute response after surgery and are associated with the development of insulin resistance, while simultaneous plasma concentrations of stress hormones seem to be less sensitive markers of the degree of postoperative metabolic disturbance.

Acute exposure to mercury from amalgam: no short-time effect on the peripheral blood lymphocytes in healthy individuals.

Mercury, released from dental amalgam, has been considered to adversely affect the human immune system. This study has been performed in order to evaluate if an acute low-dose mercury exposure, achieved by total amalgam removal in 10 healthy individuals, would affect the immunocompetent cells in human blood when the mercury level in blood and plasma was increasing. Induction of lymphocyte proliferation, measured as spontaneous de novo DNA synthesis, and total T cells, CD4+ T cells, CD8+ T cells, and B cells, was studied prior to and 7, 31, and 48 h after amalgam removal. In addition, the levels of interleukin-6 (IL-6) and C-reactive protein (CRP) in serum/plasma were measured. Despite a significant increase of the plasma mercury levels within 24 h after intervention, no significant influence on the peripheral blood lymphocytes could be detected during the first 48 h. The serum IL-6 levels increased significantly within 48 h after intervention, but were still low and within normal range. No influence on the CRP levels up to 7 d after amalgam removal was detected.

Differential regulation of in vitro cytokine production by human blood cells in response to methylmethacrylate.

The effect of methylmethacrylate (MMA) on human whole blood cultures (WBC) obtained from healthy donors was investigated. Lymphocyte transformation and cytokine production, that is, interleukin 6 (IL-6), interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha), were used to evaluate the immunological activities of MMA. Primary cytotoxicity testing of MMA in Jurkat cells showed that this compound decreased the cell proliferation to 50% at a concentration of >60 mmol/L. Similarly, MMA significantly decreased lymphocyte transformation in either phytohemagglutinin (PHA) or Staphylococcus aureus protein A (SpA) activated WBC at 100 mmol/L. In contrast to activated WBC, MMA had no observed effect on resting blood cells. Cytokine expression in WBC seemed differentially modulated by MMA. There was a tendency for IL-6 production in both resting and PHA-stimulated WBC to be upregulated, while IL-6 induced in SpA stimulated cultures was downregulated. TNF-alpha was slightly increased by MMA in resting WBC at early incubation periods, and it was slightly downregulated in response to PHA or SpA activation. Suppression of IFN-gamma secretion was observed in WBC with or without PHA or SpA stimulation. The overall results demonstrated that MMA at physiological levels could influence the cytokine production in normal human blood cells in vitro. Alterations of cytokine production patterns by MMA indicate that this compound has multiple regulatory effects on immune reactions in normal human blood.

In vitro effects of mercuric chloride (HgCl2) on human mononuclear cells.

Due to the release of the toxic compounds of mercury from amalgam fillings, dental amalgam has been questioned as an adequate restoration material for tooth fillings. HgCl2 has been found to be mitogenic for human blood lymphocytes in vitro. However, activation required much higher concentrations than are ever found in vivo. This study has been initiated to evaluate further the influence of HgCl2 on human immunocompetent cells in vitro. It is found that HgCl2 in a narrow concentration range has the ability to preferentially stimulate the CD4+ T cell subset to blast transformation and DNA synthesis. The reaction, when monitored during days 2-6, is maximal at day 6, and most blasts express the IL-2 receptor (IL-2R), indicating in vitro activation. The CD8+ T cell subset is not affected to the same extent. In addition, HgCl2-induced lymphocyte reactivity is dependent on accessory cells, i.e. CD14+ cells.

HgCl(2)-induced human lymphocyte activation in vitro: a superantigenic mechanism?

Mercuric chloride (HgCl(2)) has been proposed to be a mitogen for human blood lymphocytes in vitro. In our previous study, we demonstrated that HgCl(2) preferentially stimulates the CD4+ T cell subset to blast transformation and DNA synthesis and that the reaction is dependent on CD14+ accessory cells. In order to characterise the responding cells further and to elucidate the mechanism of T cell activation, the T cell receptor (TCR) Vbeta chain expression of the blast-transformed cells was analysed by monoclonal antibodies and flow cytometry. The 22 TCR-Vbeta-specific antibodies used were found to react with 55-80% of the naive CD4+ and CD8+ blood T cells from the different donors. Six to 18% of the lymphocytes, mainly CD4+ T cells, were blast transformed after addition of HgCl(2). The distribution of the lymphoblasts carrying certain TCR Vbeta chains were skewed, and 15-40% expressed the TCR Vbeta2 chain. Furthermore, if cells were pretreated for 5 days with HgCl(2), whereafter recombinant interleukin-2 in fresh medium was added, the TCR Vbeta7+ T cell subset was also stimulated to blast transformation. The superantigen staphyloccal enterotoxin B, as a control, induced blast transformation in 10-26% of the lymphocytes, mainly CD4+ T cells, which were, as expected, positive for Vbeta3, Vbeta12, Vbeta14 or Vbeta17. We conclude that HgCl(2) has characteristics of a superantigen, activating the human lymphocytes in a Vbeta-chain-selective manner in vitro.

Fluoride augments the mitogenic and antigenic response of human blood lymphocytes in vitro.

It has been shown that fluoride, the agent responsible for reduction of dental caries worldwide and a recognized proliferative agent, is an adjuvant when given intragastrically to rats. Furthermore, plasma fluoride levels increase in humans after various fluoride treatments. The studies presented here show that fluoride also has the ability to affect the cells of the human immune system. This was tested by measuring the effect of sodium fluoride (NaF) on cytokine production by human whole blood cells stimulated in vitro. These studies revealed that NaF augments the human lymphocyte response from human blood to a mitogen (phytohemagglutinin, PHA) or a specific antigen (morbilli antigen from infected cells, MorbAg). The cytokine interferon-gamma (IFN-gamma), released from activated T and/or NK cells, was significantly (p<0.01) increased when whole blood cells were simultaneously incubated with 0.62 mmol/l NaF and PHA compared to PHA alone. This tendency was also true for NaF and MorbAg. The lymphocyte activation marker interleukin-2 receptor (measured in soluble form) increased after simultaneous stimulation of the cells with PHA and 0.62 mmol/l NaF compared to stimulation with PHA only. However, 0.62 mmol/l NaF did not enhance interleukin-6 release, in blood mainly produced by monocytes. The ability to influence the IFN-gamma release during an immune response could be one of the primary means by which the fluoride ion influences the immune system.

Antibody responses in schistosomiasis haematobium in Somalia. Relation to age and infection intensity.

Antibody responses in schistosomiasis haematobium were studied in relation to age and infection intensity in Somalia. The area is highly endemic for Schistosoma haematobium but free of S. mansoni. Antibodies of the IgG class against particulate antigens of S. mansoni adult worms were investigated by immunofluorescence (gut and somatic associated antigens) and against soluble egg and adult worm antigens by ELISA. Total IgE levels were examined by Pharmacia IgE RIA, and specific IgE against soluble adult worm antigen by enzyme immunoassay. The IgG antibody response showed a characteristic pattern with highest reactivity against both gut associated and soluble egg antigens in the age group 10-14 years, when both prevalence and intensity of the infection were highest. Reactivity against somatic associated antigen was also high in this age group, but it increased slightly and remained at high level in the older ages. It is thought that such antigen is exposed mainly after the death of the parasite and that the antigenic stimulation may remain throughout most of the life of infected individuals. On the other hand, the IgG antibody reactivity against soluble adult worm antigen was low during childhood, but it increased significantly with age. It is suggested that repeated booster effects are needed for more potent response against these antigenic components. The finding of high levels of total IgE already in the youngest age groups, together with low specific IgE response, indicates that mainly other antigens are involved in the IgE production. The specific IgE response against soluble adult worm antigen was low but increased significantly with age.

The metabolic response to cholecystectomy: insulin resistance after open compared with laparoscopic operation.

OBJECTIVE: To study the changes in insulin sensitivity and plasma concentrations of interleukin-6 (IL-6) after open compared with laparoscopic cholecystectomy. DESIGN: Prospective open study. SETTING: University hospital, Sweden. SUBJECTS: 12 otherwise healthy patients undergoing either open (n = 6) or laparoscopic (n = 6) cholecystectomy. MAIN OUTCOME MEASURE: Relative insulin sensitivity (compared with preoperative) on the day after operation. Changes in IL-6 concentrations postoperatively. RESULTS: The mean (SEM) relative reduction in insulin sensitivity was significantly smaller after laparoscopic (18 (5)%) compared with the open operation, (58 (4)%) (p < 0.01). There was a significant increase in plasma concentrations of IL-6 postoperatively, but there was no difference between the groups. CONCLUSION: Insulin sensitivity is less affected 24 hours after laparoscopic than after open cholecystectomy, which in this study was not accompanied by a simultaneous difference in the IL-6 response. The small postoperative reduction of insulin sensitivity may be a contributing factor to the clinical benefit of improved wellbeing observed after laparoscopic surgery.

No evidence for specific in vitro lymphocyte reactivity to HgCl2 in patients with dental amalgam-related contact lesions.

Blood lymphocytes from 20 patients with oral contact lesions to dental amalgam and 10 healthy individuals were analyzed for HgCl2-induced proliferation in vitro, using both a modified assay and a conventional assay. The release of interferon-gamma (IFN-gamma) was measured in cell supernatants. Six patients displayed positive reactions in patch tests to mercuric compounds. No significant differences were recorded in HgCl2-induced proliferation in cells from patients and controls, since only few in the whole material responded to submitogenic concentrations. IFN-gamma was detectable in cell supernatants from some patients but also from controls and is not predictive of mercury allergy. Neither the phenotypes of peripheral lymphocyte subsets, the frequency of circulating cells expressing the interleukin-2 (IL-2) receptor, spontaneous lymphocyte proliferation nor concentrations of serum interleukin-6 differed between patient and control samples. In contrast to what has been claimed before, we did not find any evidence for specific in vitro lymphocyte reactivity in patients with oral contact lesions.

Expression of interleukin-1 alpha, interleukin-1 beta and interleukin-6 in chronic B lymphocytic leukaemia (B-CLL) cells from patients at different stages of disease progression.

We have previously found that isolated B-CLL cells from progressive disease produce less interleukin-1 beta (IL-1 beta) as compared with cells from patients with indolent disease. Here we extend that finding to include measurements of IL-1 beta mRNA and secretion of IL-1 alpha and interleukin-6 (IL-6). As before, a lower production of IL-1 beta was found in cells from progressive disease. IL-6 was produced by cells from patients at all stages, with a tendency to follow the IL-1 beta production. Low secretion of IL-1 alpha was noted. When viable cells were permeabilized and analysed at the single cell level with monoclonal antibodies, most B-CLL cells were found to contain IL-1 alpha. A minor fraction of non-permeabilized cells expressed IL-1 alpha at the cell membrane. However, only small fractions of cells were positive for intracellular IL-1 beta (less than 1%) and almost no IL-6-positive cells were found. We conclude that either IL-1 beta and IL-6 are produced by a minor population of undefined cells, or that a more sensitive in situ method is needed to detect production of these cytokines in B-CLL cells. The possible biological significance of secreted, and membrane-expressed helper factors in B-CLL is discussed.

Visceral leishmaniasis in Somalia. Circulating antibodies as measured by DAT, immunofluorescence and ELISA.

Sera from patients with visceral leishmaniasis (VL) (n = 26), healthy residents of Mogadishu (n = 157), inhabitants of a village in an endemic area (n = 276) and healthy Swedes (n = 60) were examined using the direct agglutination test (DAT), immunofluorescence (IF) and ELISA for antibodies against Leishmania donovani. The study was carried out in order to provide baseline data for antibody responses in visceral leishmaniasis as existing in Somalia and to explore which one of these methods would be most suitable for diagnosis of clinical cases as well as for epidemiological population studies in Somalia. All patients had high levels of circulating antibodies, however, lower values were recorded in the early stages of the disease. High reactivity in ELISA was seen first after one year. All three tests distinguished well between sera from VL patients and healthy controls. Approximately 10% of the sera from villagers were reactive above the cut-off levels in the three tests. DAT is the simplest to perform and does not require much equipment. ELISA can be made simple and economic if performed in one serum dilution and read visually. IF requires more expensive and specialized equipment and is not suitable for large scale examination of sera. A complete evaluation of the three tests should also include the analysis of sera from various stages and manifestations of the disease.

Antibodies against Entamoeba histolytica antigens in sera from individuals with amoebiasis of different localization.

The over-all contents and relative component composition of Entamoeba histolytica antigens in abscess fluids and in extracts of cultured amoebae, strain NIH 200, were studied by antigen-catching EIA, counterimmunoelectrophoresis (CIE) and immunoblotting techniques. The antigen contents of liver abscess fluid were determined semiquantitatively by the antigen-catching EIA in four cases. In CIE against a standard "diagnostic" extract of cultured amoebae, sera from cases of acute amoebic liver abscess gave 4-5 precipitation lines while sera from cases of intestinal amoebiasis gave at most 3 lines. In immunoblotting tests with the same antigen, intestinal cases gave blotting bands in the intermediate molecular weight range (25-99 kD) while acute abscess cases, in addition, gave bands in the high (100-175 kD) and low (= less than 25 kD) molecular weight range. These serological differences between clinical forms of amoebiasis were more definite when using amoeba abscess fluid as antigen. Amoeba antigens in high concentrations could be demonstrated in amoeba abscess fluids with all methods employed. In immunoblotting experiments abscess fluids generally gave stronger and more numerous bands with anti-amoeba antibody-containing sera than did the standard "diagnostic" antigen from cultured amoebae. Especially the abscess fluids gave with sera from acute abscess cases a number of prominent bands in the low molecular weight range (less than 25 kD). The experiments in this study were performed with crude amoebic extracts, which contained a multitudes of antigenic components and a still greater diversity of antigenically inert proteins.(ABSTRACT TRUNCATED AT 250 WORDS)