Modulatory in vitro effects of interleukin-1 receptor antagonist (IL-1Ra) or antisense oligonucleotide to interleukin-1β converting enzyme (ICE) on acute myeloid leukaemia (AML) cell growth

We investigated the effects of interleukin-1 receptor antagonist (IL-1Ra) on the spontaneous proliferation and AML colony forming unit (CFU-AML) formation of bone marrow and peripheral blood cells in 50 acute myeloid leukaemia (AML) patients. Exposure to IL-1Ra (10 μg/ml) caused either decreased, unaltered or increased AML cell proliferation, as well as of CFU-AML colony formation, depending on the individual patient, but the inhibitory effects were dominant. To evaluate the involvement of IL-1β converting enzyme (ICE) in the autonomous AML cell growth, the effects of an antisense oligonucleotide on ICE were examined in 19 of these patients. In a majority of patients, antisense ICE suppressed both AML cell proliferation and CFU-AML although a stimulatory effect was sometimes evident. The proportion of AML patients with suppression obtained by antisense ICE was higher than with IL-1Ra, suggesting the involvement of additional ICE-dependent cytokine(s) in AML cell growth besides IL-1. The presence of IL-1Ra or antisense ICE also suppressed the endogenous IL-1β production of AML cells, at both the level of pro-IL-1β and mature IL-1β. Although inhibition by IL-1Ra or antisense ICE on growth parameters of AML cells in vitro prevailed, indicating the importance of IL-1 activity in autonomous AML cell growth, stimulatory effects on the cells of some patients suggest that AML is a heterogenous disorder regarding IL-1β regulation.     

Serum concentrations of E-selectin, P-selectin, ICAM-1 and interleukin 6 in acute leukaemia patients with chemotherapy-induced leucopenia and bacterial infections

Summary. Serum concentrations of E-selectin (CD62E), P-selectin (CD62P), ICAM-1 (CD54) and interleukin 6 were investigated in acute leukaemia patients with chemotherapy-induced leucopenia and complicating bacterial infections. Serum concentrations of both E-selectin and P-selectin were decreased in the leucopenic patients without infections when compared with levels before chemotherapy; and serum concentrations of both E-selectin and P-selectin showed a further decrease during complicating bacterial infections. In contrast to the leukaemia patients, previously healthy individuals with meningococcal disease showed markedly elevated serum concentrations of E-selectin and normal levels of P-selectin during infection. Serum concentrations of ICAM-1 and interleukin 6 increased during bacterial infections in the acute leukaemia patients with chemotherapy-induced leucopenia. The alterations in serum concentrations of soluble adhesion molecules and interleukin 6 reversed when clinical signs of bacterial infections resolved during antibiotic therapy. Our results demonstrate that acute leukaemia patients with chemotherapy-induced cytopenia show altered levels of both soluble adhesion molecules and interleukin 6 during complicating bacterial infections.     

Interleukin-1 receptor antagonist is detectable in human carotid artery plaques and is related to triglyceride levels and Chlamydia pneumoniae IgA antibodies

Abstract. Gottsäter A, Forsblad J, Mätzsch T, Persson K, Ljungcrantz I, Ohlsson K, Lindgärde F (Malmö University Hospital, University of Lund, Malmö, Sweden). Interleukin‐1 receptor antagonist is detectable in human carotid artery plaques and is related to triglyceride levels and Chlamydia pneumoniae IgA‐antibodies. J Intern Med 2002; 251: 61–68. Objectives. To investigate whether the interleukin‐1 receptor antagonist (Il‐1ra) and interleukin‐1β (Il‐1β) can be detected in human carotid artery tissue, and whether their presence is related to evidence of Chlamydia pneumoniae infection, risk factors for atherosclerosis, and clinical data. Setting. Departments of Vascular Diseases and Surgical Pathophysiology, University Hospital, Malmö, Sweden. Subjects. A total of 66 patients undergoing carotid endarterectomy (median age 74, range 53–89 years, 26 women). Il‐1β and Il‐1ra were studied in carotid artery plaques and in Il‐1ra in serum. Results. Interleukin‐1 receptor antagonist was detected in mononuclear cells in plaques from 37/66 (56%) patients. Patients with Il‐1ra in plaques showed higher [2.04 (1.70–3.14) mmol L^–1 vs. 1.69 (1.09–1.99) mmol L^–1; P < 0.05] serum(s‐)triglyceride(tg) levels, and a higher frequency of IgA seropositivity for C. pneumoniae (76% vs. 52%; P < 0.05) than those without. S‐Il‐1ra levels correlated with s‐tg levels (r=0.38; P=0.047). There were no differences between patients with and without Il‐1ra in plaques concerning s‐Il‐1ra, blood(b‐)haemoglobin or leucocyte count, s‐cholesterol, b‐glucose, blood pressure, IgG seropositivity for C. pneumoniae, prevalence of neurological symptoms preceding operation, smoking, or diabetes mellitus. There were no differences in frequency of Il‐1ra in plaques or in s‐Il‐1ra levels between patients with symptomatic and asymptomatic stenosis, between smokers and nonsmokers, or between diabetic and nondiabetic patients. Il‐1β was not detected in plaques in the current study. Conclusion. Interleukin‐1 receptor antagonist can be detected in human atherosclerotic carotid artery plaques, and is related to s‐triglyceride levels and IgA seropositivity for C. pneumoniae, but not to prevalence of neurological symptoms related to embolization.     

Expression of matrix metalloproteinases (MMP-2 and -9) and tissue inhibitors of metalloproteinases (TIMP-1 and -2) in acute myelogenous leukaemia blasts: comparison with normal bone marrow cells

We compared the expression of matrix metalloproteinases (MMP-2 and MMP-9) and tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) in bone marrow acute myelogenous leukaemia (AML) blasts and leukaemic cell lines (HEL, HL-60, K-562 and KG-1) with their expression in normal bone marrow cells. All AML samples and leukaemic cell lines tested expressed MMP-9 and/or MMP-2 mRNA and, accordingly, these gelatinases were secreted into media. Moreover, TIMP-1 and TIMP-2 mRNA and secreted proteins were demonstrated in all the AML samples. Although all the leukaemic cell lines expressed TIMP-1, the HL-60 cells also expressed TIMP-2. In contrast, normal steady-state bone marrow immature progenitor cells (CD34+ cells) did not express or secrete either MMP-2 or MMP-9, but more mature mononuclear cells from normal bone marrow expressed and secreted MMP-9. Also, normal bone marrow CD34+ cells and mononuclear cells expressed TIMP-1 and TIMP-2 mRNA, but these proteins were not detectable by reverse zymography. Furthermore, whereas bone marrow fibroblasts and endothelial cells secreted only latent MMP-2, the activated form of this enzyme was found in media conditioned by cells obtained from long-term cultures of normal and AML bone marrow adherent layers. Our finding of up-regulated production of gelatinases, TIMP-1 and TIMP-2 by leukaemic cells suggests that these proteins may be implicated in the invasive phenotype of AML.     

All-trans-retinoic acid (ATRA) responsive skin relapses of acute promyelocytic leukaemia followed by ATRA-induced pseudotumour cerebri

A 30-year-old woman with acute promyelocytic leukaemia (APL) went into complete remission following idarubicin and cytarabine chemotherapy; 18 months later she developed repeated skin relapse, with no bone marrow involvement. DNA and RNA analysis of skin lesions revealed the presence of the PML/RARα hybrid gene, which was not detected at the same time in bone marrow. The skin relapses were successfully treated by all-trans-retinoic acid (ATRA) as single agent over 2 years. However, prolonged administration of ATRA caused pseudotumour cerebri, which disappeared upon drug withdrawal. The absence of the hybrid gene in the bone marrow by RT-PCR analysis led to the patient being autografted.     

Effects of soluble interleukin-1 receptor and tumor-necrosis factor receptor,respectively, on the IL-1- and TNF-α-induced DNA synthesis of acute myeloblastic leukemia blasts in vitro

Abstract: This study demonstrates that soluble interleukin-1 receptor and tumor necrosis factor receptor modulate their corresponding cytokine-induced DNA synthesis of acute myeloblastic leukemia (AML) blasts in a dose-dependent, bimodal fashion; at lower concentrations they enhanced, while at high concentrations they inhibited, the cytokine-mediated effects. Furthermore, the concentrations of endogenously produced IL-1β and TNF-a were found to be significantly (p<0.01) higher in supernatants of AML cells cultured in the presence of corresponding soluble receptors compared to their levels in supernatants of cells growing in the absence of these molecules. Our data might suggest that the attenuation of the spontaneous decay of IL-1β as well as TNF-a activities by soluble receptors may account for their ability to augment some of their effects.     

Protective effect of a human C5a receptor antagonist against hepatic ischaemia-reperfusion injury in rats

BACKGROUND/AIMS: Complement activation is induced by ischaemia-reperfusion (I/R) and the complement factor C5a plays an important role in organ specific I/R injuries. This study investigated the efficacy of a small molecule C5a receptor (C5aR) antagonist against hepatic I/R injury. METHODS: Total hepatic ischaemia or partial hepatic ischaemia were induced in rats, followed by a period of reperfusion. The C5aR antagonist, AcF-[OPdChaWR], was administered at 1 mg/kg i.v. or 10 mg/kg p.o. or s.c. before induction of ischaemia. Total hepatic I/R-induced mortality was measured and partial hepatic ischaemia injury was assessed by measuring the serum levels of liver enzymes, tissue or serum TNFalpha, liver and lung myeloperoxidase activity, the number of infiltrating neutrophils, neutrophilia and liver histopathology. RESULTS: C5aR antagonist treatment reduced total hepatic I/R-induced mortality. In partial hepatic I/R rats, treatment with the C5aR antagonist significantly attenuated the increases in liver enzymes, serum and tissue TNFalpha, myeloperoxidase activity, infiltrating neutrophils, neutrophilia, and also reduced liver histopathology. CONCLUSIONS: This study shows that an orally active, small molecule C5aR antagonist is effective in reducing the markers of tissue damage caused by I/R in the rat, suggesting an important role for C5a in I/R injuries in the liver.     

RUNX3 promoter hypermethylation is frequent in leukaemia cell lines and associated with acute myeloid leukaemia inv(16) subtype

Correlative and functional studies support the involvement of the RUNX gene family in haematological malignancies. To elucidate the role of epigenetics in RUNX inactivation, we evaluated promoter DNA methylation of RUNX1, 2, and 3 in 23 leukaemia cell lines and samples from acute myeloid leukaemia (AML), acute lymphocytic leukaemia (ALL) and myelodysplatic syndromes (MDS) patients. RUNX1 and RUNX2 gene promoters were mostly unmethylated in cell lines and clinical samples. Hypermethylation of RUNX3 was frequent among cell lines (74%) and highly variable among patient samples, with clear association to cytogenetic status. High frequency of RUNX3 hypermethylation (85% of the 20 studied cases) was found in AML patients with inv(16)(p13.1q22) compared to other AML subtypes (31% of the other 49 cases). RUNX3 hypermethylation was also frequent in ALL (100% of the six cases) but low in MDS (21%). In support of a functional role, hypermethylation of RUNX3 was correlated with low levels of protein, and treatment of cell lines with the DNA demethylating agent, decitabine, resulted in mRNA re‐expression. Furthermore, relapse‐free survival of non‐inv(16)(p13.1q22) AML patients without RUNX3 methylation was significantly better (P = 0·016) than that of methylated cases. These results suggest that RUNX3 silencing is an important event in inv(16)(p13.1q22) leukaemias.     

Course of infection by Babesia sp. BQ1 (Lintan) and B. divergens in sheep depends on the production of IFNγ and IL10

Ovine babesiosis is an important disease in China and responsible for economic losses. Several Babesia strains are involved, but Babesia sp. BQ1 (Lintan) and Babesia sp. BQ1 (Ningxian) are particularly prevalent in the Guansu region. Babesia divergens, in contrast, can experimentally infect spleen‐intact sheep, but does not induce clinical signs. The immune response of spleen‐intact sheep to Babesia sp. BQ1 (Lintan) and to B. divergens was therefore compared to identify the immune mechanisms involved in pathogenicity. The greater pathogenicity of Babesia sp. BQ1 (Lintan) than that of B. divergens was confirmed: sheep infected with Babesia sp. BQ1 (Lintan), but not with B. divergens, developed hyperthermia and showed patent parasitaemia in Giemsa‐stained blood smears from the ear vein. Furthermore, more parasites were also detected in the blood from the jugular vein of Babesia sp. BQ1 (Lintan)‐infected sheep. Pathogenicity of Babesia spp. involved cellular responses, but not humoral responses. Interferon‐gamma was produced only by specifically activated PBMC from B. divergens‐infected sheep and interleukin‐10 only by specifically activated PBMC from Babesia sp. BQ1 (Lintan)‐infected sheep. The role of these cytokines in the course of infection by Babesia spp. is discussed.     

Effects of 1‐O‐hexyl‐2,3,5‐trimethylhydroquinone on carbon tetrachloride‐induced hepatic cirrhosis in rats

Aim: The present study was undertaken to evaluate the effects of 1‐O‐hexyl‐2,3,5‐trimethylhydroquinone (HTHQ), a synthesized vitamin E derivative, on carbon tetrachloride (CCl₄)‐induced cirrhosis. Methods: Rats were treated with hypodermic injections of CCl₄ twice a week to induce the hepatic cirrhosis, and given drinking water containing HTHQ or solvent. Primary cultures of rat hepatocytes were performed to evaluate the effects of HTHQ on the expression of inducible nitric oxide synthase (iNOS). Results: Masson's staining of rat livers showed fibrosis around pseudo‐lobules in the CCl₄ group, the lesions being reduced in the CCl₄ HTHQ group. Increases in liver tissue hydroxyproline and α₁(I) collagen, α‐smooth muscle actin and iNOS induced by CCl₄, were also markedly diminished by HTHQ. Furthermore, both HTHQ and vitamin E attenuated interleukin‐1β‐induced iNOS protein expression in cultured hepatocytes, the potency of HTHQ being 10‐times higher than that of vitamin E. Conclusion: HTHQ may inhibit development of hepatic cirrhosis in rats, more potently than vitamin E, by inhibiting the iNOS expression in hepatocytes. Because vitamin E has a radical scavenging action, roles of NO and peroxynitrite will be discussed in the effects of HTHQ on the fibrosis.     

The primary mitogen (TCPOBOP)‐induced hepatocyte proliferation is resistant to transforming growth factor‐ β‐1 inhibition

Background: Transforming growth factor (TGF)‐β‐1 is a very efficient inhibitor of hepatocyte proliferation in various in vivo and in vitro experimental systems. However, there are no data on whether it can influence the mitogenic response induced by primary hepatocyte mitogens. Aims: In this study, we compared the proliferative response in the liver between wild‐type and transgenic mice, overexpressing active TGF‐β‐1 in their liver following the treatment by a primary hepatocyte mitogen TCPOBOP (1,4‐bis[2‐(3,5‐dichloropyridyloxy)]benzene). Methods: The proliferative response was characterized by the immunohistochemical examination of pulse and cumulative bromodeoxyuridine labelling and by quantitative real‐time polymerase chain reaction analysis of cell cycle‐related genes. Results: Neither of the applied techniques revealed significant differences between the two groups of mice; furthermore, we observed the upregulation of TGF‐β‐1 expression following the mitogenic treatment. Conclusions: TGF‐β‐1 does not inhibit the primary mitogen‐induced proliferative response of the hepatocytes. This observation may provide an explanation for the divergent consequences of hepatic proliferations induced by partial hepatectomy or primary mitogenic treatment.