Yvette Mándi

Diagnostic relevance of procalcitonin, IL-6, and sICAM-1 in the prediction of infected necrosis in acute pancreatitis

SummaryBackground. Infected pancreatic necrosis (IPN) is an absolute indication for surgical intervention, therefore an early and accurate laboratory diagnosis is necessary to confirm the infection. The aim of the study was to analyze the clinical value of procalcitonin (PCT) for the prediction of infected necrosis, in comparison with interleukin-6 (IL-6) and sICAM-1.Patients and Methods. A total of 30 patients were investigated; 10 patients with sterile pancreatic necrosis (SPN), 10 with IPN, and 10 with sepsis of different origin. The concentrations of PCT in the patients’ sera were measured by immunoluminometric assay (BRAHMS Diagnostica, Berlin, Germany, PCTLumitest), the IL-6 concentrations by bioassay, applying the B-9 cell line, and the sICAM-1 levels by enzyme-linked immunosorbent assay (ELISA) (R&D). PCT was determined in cell lysates by ECL Western blot.Results. PCT was found in relatively high concentrations (8.5±4.8 ng/mL) only in patients with infected pancreatic necrosis, and in patients with sepsis of different origin (15±5.4 ng/mL). Positive values (>1 ng/mL) preceded positive bacterial results from either blood or surgical samples. None of the serum samples of patients with SPN exhibited PCT concentrations higher than 1.2 ng/mL. In contrast, IL-6 and sICAM-1 were overproduced in both types (infected and sterile) of pancreatic necrosis, and their levels remained elevated for several days even after surgical elimination of the infected focus (widespread necrosectomy and continuous lavage). Sensitivity, specificity, and positive predictive values for discriminating IPN from SPN was 90, 100, and 100% for PCT (p<0.0001); 100, 20, and 55% for IL-6 (p 0.474 n.s.) and 90, 10, and 50% for sICAM-1 (p 1.000 n.s.). Immunoblotting revealed no PCT in patients’ leukocytes, or in human endothelial cell lines. Conclusion. Elevated serum IL-6 and sICAM-1 levels are characteristic in systemic inflammatory response syndrome (SIRS) of either infectious or noninfectious origin. In contrast, the PCT level is an accurate, readily available parameter that allows the discrimination of IPN, and is a helpful marker facilitating a decision concerning surgical intervention.

Histamine and histamine-receptor antagonists modify gene expression and biosynthesis of interferon gamma in peripheral human blood mononuclear cells and in CD19-depleted cell subsets

The effect of histamine and histamine antagonists was examined on gene expression and biosynthesis of bacterial endotoxin (LPS) induced interferon gamma (IFNgamma) both in human peripheral mononuclear cells (PMBC) and in T-cell enriched fractions. We found, that histamine inhibited the LPS induced transcription of IFNgamma gene and biosynthesis of IFNgamma protein in PMBC and also in CD19-depleted cell populations. The inhibitory effect of histamine could be reversed by the H2 histamine receptor (HR2) antagonists cimetidine and ranitidine both in PMBC and in CD19-depleted cells, but not with triprolidine, an H1 receptor antagonist, suggesting that the inhibition of IFNgamma production is mediated through H2 receptors in these cell populations. In contrast to the inhibitory effect of histamine, cimetidine alone (in the absence of exogenous histamine) strongly stimulated both the IFNgamma mRNA and protein production, whereas this effect was hardly seen by and other H2 receptor blocker, ranitidine. This superinduction of IFNgamma gene by cimetidine disappeared if the CD19+ cells are removed from PMBC. These results suggest, that inhibition of IFNgamma gene expression by histamine is a direct effect of histamine on H2 receptor of T lymphocytes; however, the superinduction of IFNgamma by cimetidine requires the presence of other (probably primarily B) cell subsets.

The pathogenesis of L-arginine-induced acute necrotizing pancreatitis: Inflammatory mediators and endogenous cholecystokinin

This study was aimed at an assessment of the role of oxygen-derived free radicals, cytokines and endogenous cholecystokinin (CCK) in the pathogenesis of L-arginine (Arg)-induced acute pancreatitis in rat. We measured the levels of malonyl dialdehyde (MDA), glutathione peroxidase (GPx), catalase and superoxide dismutase (Mn- and Cu, Zn-SOD) in pancreatic tissue, the serum levels of tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and CCK, and evaluated the protective effect of the xanthine oxidase inhibitor allopurinol and a novel CCK receptor antagonist KSG-504. Acute pancreatitis was induced in male Wistar rats by injecting 2x 250 mg/100 g body weight of Arg intraperitoneally in an 1-h interval, as a 20% solution in 0.15 M NaCl. Control rats received the same quantity of glycine. 200 mg x kg(-1) allopurinol 30 min before the first Arg treatment or 50 mg x kg(-1) KSG-504 30 min before and 6, 18 and 36 h after the first Arg injection was administered subcutaneously. Rats were killed at 6, 12, 24 and 48 h following Arg administration, and acute pancreatitis was confirmed by a serum amylase level elevation and typical inflammatory features observed microscopically. The serum level of amylase reached the peak level at 24 h after the Arg injection (30,800 +/- 3,813 versus 6,382 +/- 184 U x L(-1) in the control) and normalized at 48 h. The tissue concentration of MDA was significantly elevated at 24 h, and reached the peak value at 48 h (5.00 +/- 1.75 versus 0.28 +/- 0.05 nM x mg(-1) protein in the control). The catalase and Mn-SOD activities were significantly decreased throughout the study, while the GPx activity was significantly reduced at 6 and 12 h, and the Cu, Zn-SOD activity was significantly lower at 12 h after the Arg injection as compared with the controls. Both the TNF-alpha and the IL-6 levels were already elevated significantly at 12 h and peak at 24 h versus the controls (19.1 +/- 7.9 U x mL(-1) and 57.6 +/- 11.2 pg x mL(-1) versus 3.1 +/- 0.8 U x mL(-1) and 15.2 +/- 3.1 pg x mL(-1), respectively). No significant changes in plasma CCK levels were observed. Allopurinol treatment markedly reduced the serum amylase elevation (12.631 +/- 2.257 U x L(-1) at 24 h), prevented the increase in tissue MDA concentration (0.55 +/- 0.09 nM x mg(-1) protein at 48 h) and significantly ameliorated the pancreatic edema, necrosis and inflammation at 48 h after Arg administration. KSG-504 administration did not exert any beneficial effect on the development of histopathological changes neither modified the serum amylase or cytokine levels. Oxygen-derived free radicals and cytokines are involved, while endogenous CCK does not seem to play a role in the pathogenesis of Arg-induced acute pancreatitis.

GROWTH HORMONE RECEPTOR GENE EXPRESSION ON HUMAN LYMPHOCYTIC AND MONOCYTIC CELL LINES

The potential effect of growth hormone (GH) in tumorigenesis, particularly in acute leukemia is controversial. Human growth hormone has the ability to influence certain immune functions; the majority of immune cells express growth hormone receptor (GHR) on plasma membranes. We determined GHR gene expression on different human lymphocyte (JURKAT, CESS) and monocyte (U937, THP1) cell lines by reverse transcriptase polymerase chain reaction analysis of GHR mRNA after stimulating the cells with phytohaemagglutinin or phorbolester, human growth hormone and with a combination of these. The receptor gene expression showed differences; in the U937 and CESS cell lines only the stimulants were able to induce GHR mRNA expression; in the case of JURKAT cells even the hormone alone had the ability to express its own receptor gene. Both the increased TNF‐α production of U937 (but not that of THP1 cells), and the decreased proliferation of JURKAT cells in response to GH stimuli also prove the presence of biologically active GHR on the cell surface. Our data suggest asymmetric interaction between GH or phorbolester‐induced signal pathways in U937 cells sharply depending on the temporal sequence of treatments. THP1 monocytes showed no gene expression in response to any of the stimulants. The phenomenon that certain human lymphoid and monocytoid cell lines at different levels of cell differentiation are able to express the GH receptor gene could have importance in the rhGH therapy.

Natural killer cell mediated cytotoxicity against VERO target cells; the suppressive effect of pentoxifylline

The K-562 cell line is widely known and used as a NK cell target. In this study we report that VERO (African green monkey kidney epithelial cell line) is an excellent target of the human NK cell cytotoxicity. Considerable cytotoxicity was observed in a 4 h 51Cr release assay with nonadherent and immunomagnetically separated CD56+ NK cells from PBMC. On the contrary, adding K-562 cells as cold target to the assay the cytotoxicity significantly decreased. Using a standard chromium-release assay the NK cell activity (NKCA) against VERO cells was investigated in a population of healthy volunteers (mean value of cytotoxicity was 26.6%) and compared with the values of cytotoxicity against K-562 target cells (32.6%). The difference was not significant (P > 0.05). The suppressive effect of PTX on in vitro NK cell activity was observed at concentration of 100 microg/ml using VERO target cells as well as K-562 cells. Our studies provide the first evidence that the NK cell activity is suppressed in vitro by PTX using VERO cells as NK target cells.

Inhibition of cytotoxicity of chicken granulocytes by serotonin and ketanserin

Serotonin (5-HT) has been observed to impair the cytotoxicity of human natural killer (NK) cells. A study has now been made of the effect of 5-HT on the cytotoxic activity of chicken granulocytes. 5-HT at concentrations of 10(-3) to 10(-6) M inhibited the cytotoxicity of chicken granulocytes when added at the onset of the short-term cytotoxicity assay. Ketanserin, a 5-HT2 receptor antagonist, did not reverse the inhibitory effect of 5-HT on chicken granulocyte cytotoxicity. Moreover, ketanserin at concentrations of 5 x 10(-4) to 5 x 10(-7) M inhibited the cytotoxicity mediated by chicken granulocytes. Pretreatment of the effector cells for 2 h with chick fibroblast interferon reduced the inhibition of chicken NK cytotoxicity induced by 5-HT and by ketanserin. These data indicate that, in birds, the neurotransmitter 5-HT serves as a link between the central nervous system and the immune system, and that interferon can modulate the inhibitory effect of 5-HT on the function of cytotoxic granulocytes.