Dose-dependent effects of Allopurinol on human foreskin fibroblast cell and human umbilical vein endothelial cell under hypoxia

Allopurinol, an inhibitor of xanthine oxidase, has been used in clinical trials of patients with cardiovascular and chronic kidney disease. These are two pathologies with extensive links to hypoxia and activation of the transcription factor hypoxia inducible factor (HIF) family. Here we analysed the effects of allopurinol treatment in two different cellular models, and their response to hypoxia. We explored the dose-dependent effect of allopurinol on Human Foreskin Fibroblasts (HFF) and Human Umbilical Vein Endothelial Cells (HUVEC) under hypoxia and normoxia. Under normoxia and hypoxia, high dose allopurinol reduced the accumulation of HIF-1α protein in HFF and HUVEC cells. Allopurinol had only marginal effects on HIF-1α mRNA level in both cellular systems. Interestingly, allopurinol effects over the HIF system were independent of prolyl-hydroxylase activity. Finally, allopurinol treatment reduced angiogenesis traits in HUVEC cells in an in vitro model. Taken together these results indicate that high doses of allopurinol inhibits the HIF system and pro-angiogenic traits in cells

Biochemical mechanisms implemented by human acute myeloid leukemia cells to suppress host immune surveillance

Acute myeloid leukaemia (AML) is a blood/bone marrow cancer originating from myeloid cell precusors capable of self-renewing. AML cells implement biochemical mechanisms which allow them not only to survive, but also to successfully escape immune surveillance. ln this work, we discuss crucial molecular mechanisms used by human AML cells in order to evade immune attack

Characteristics and treatment regimens across ERS SHARP severe asthma registries

Little is known about the characteristics and treatments of patients with severe asthma across Europe but both are likely to vary. This is the first study in the ERS Severe Heterogeneous Asthma Research collaboration, Patient-centred (SHARP) and it is designed to explore these variations. Therefore, we aimed to compare characteristics of patients in European severe asthma registries and treatments before starting biologicals. This was a cross-sectional retrospective analysis of aggregated data from 11 national severe asthma registries that joined SHARP with established patient databases. Analysis of data from 3233 patients showed many differences in characteristics and life style factors. Current smokers ranged from 0% (Poland, PL, Sweden, SE) to 9.5% (Belgium, BE), mean BMI ranged from 26.2 (Italy) to 30.6 kg·m-2 (UK) and the largest difference in mean pre-bronchodilator FEV1% pred. was 20.9% (Netherlands versus Hungary). Before starting biologicals patients were treated differently between countries: mean ICS dose ranged from 700 to 1335 µg·day-1 between those from Slovenia (SL) versus PL when starting anti-IL-5 antibody and from 772 to 1344 µg·day-1 in those starting anti-IgE (SL versus Spain). Maintenance OCS use ranged from 21.0% (BE) to 63.0% (SE) and from 9.1% (Denmark) to 56.1% (UK) in patients starting anti-IL-5 and anti-IgE, respectively. The severe asthmatic population in Europe is heterogeneous and differs in both clinical characteristics and treatment, often appearing not to comply with the current ERS/ATS guidelines definition of severe asthma. Treatment regimens before starting biologicals were different from inclusion criteria in clinical trials and varied between countries

Apoptosis signal-regulating kinase 1 (ASK1) and HIF-1 alpha protein are essential factors for nitric oxide-dependent accumulation of p53 in THP-1 human myeloid macrophages

Nitric oxide (NO) is a reactive secondary mediator, which has been found to participate in cell cycle regulation and apoptosis in myeloid macrophages, the key effectors of inflammatory and innate immune responses. However, the molecular mechanisms of nitric oxide-induced death of myeloid macrophages are not well understood. In this study we have found that NO derived from S-nitrosoglutathione (GSNO) activates ASK1 in THP-1 human myeloid macrophages in a concentration and time-dependent manner. It also induces accumulation of HIF-1 alpha protein in a concentration-dependent manner, which peaks at 4 h of exposure to 1 mM GSNO. GSNO does not affect the level of HIF-1 alpha mRNA as detected by the RT-PCR. In addition, GSNO was found to induce accumulation of p53 in normal but not HIF-1 alpha knockdown THP-1 cells, where expression of this protein was silenced by specific siRNA. It has also been found that GSNO-mediated accumulation of p53 depends on activation of ASK1 since no GSNO-induced p53 stabilisation was observed in THP-1 cells transfected with dominant-negative form of this kinase. However, in both HIF-1 alpha knockdown THP-1 cells and those transfected with the dominant-negative form of ASK1, GSNO was able to induce cell death as detected by the MTS cell viability assay leading to an increase in release of LDH

Interleukin-1 beta induces the expression and production of stem cell factor by epithelial cells: crucial involvement of the PI-3K/mTOR pathway and HIF-1 transcription complex

Potential crosslinks between inflammation and leukaemia have been discussed for some time, but experimental evidence to support this dogma is scarce. In particular, it is important to understand the mechanisms responsible for potential upregulation of proto-oncogenic growth factor expressions by inflammatory mediators. Here, we investigated the ability of the highly inflammatory cytokine interleukin-1 beta (IL-1?) to induce the production of stem cell factor (SCF), which is a major hematopoietic growth factor that controls the progression of acute myeloid leukaemia upon malignant transformation of haematopoietic myeloid cells. We found that human IL-1? induced the expression/secretion of SCF in MCF-7 human epithelial breast cancer cells and that this process depended on the hypoxia-inducible factor 1 (HIF-1) transcription complex. We also demonstrated a crucial role of the phosphatidylinositol-3 kinase (PI-3K)/mammalian target of rapamycin (mTOR) pathway in IL-1?-induced HIF-1? accumulation in MCF-7 cells. Importantly, mTOR was also found to play a role in IL-1?-induced SCF production. Furthermore, a tendency for a positive correlation of IL-1? and SCF levels in the plasma of healthy human donors was observed. Altogether, our results demonstrate that IL-1?, which normally bridges innate and adaptive immunity, induces the production of the major haematopoietic/proleukaemic growth factor SCF through the PI-3K/mTOR pathway and the HIF-1 transcription complex. These findings strongly support a cross-talk between inflammation and acute myeloid leukaemia

Low levels of endogenous anabolic androgenic steroids in females with severe asthma taking corticosteroids

Rationale Patients with severe asthma are dependent upon treatment with high doses of inhaled corticosteroids (ICS) and often also oral corticosteroids (OCS). The extent of endogenous androgenic anabolic steroid (EAAS) suppression in asthma has not previously been described in detail. The objective of the present study was to measure urinary concentrations of EAAS in relation to exogenous corticosteroid exposure. Methods Urine collected at baseline in the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease outcomes) study of severe adult asthmatics (SA, n=408) was analysed by quantitative mass spectrometry. Data were compared to that of mild-to-moderate asthmatics (MMA, n=70) and healthy subjects (HC, n=98) from the same study. Measurements and main results The concentrations of urinary endogenous steroid metabolites were substantially lower in SA than in MMA or HC. These differences were more pronounced in SA patients with detectable urinary OCS metabolites. Their dehydroepiandrosterone sulfate (DHEA-S) concentrations were <5% of those in HC, and cortisol concentrations were below the detection limit in 75% of females and 82% of males. The concentrations of EAAS in OCS-positive patients, as well as patients on high-dose ICS only, were more suppressed in females than males (p<0.05). Low levels of DHEA were associated with features of more severe disease and were more prevalent in females (p<0.05). The association between low EAAS and corticosteroid treatment was replicated in 289 of the SA patients at follow-up after 12–18 months. Conclusion The pronounced suppression of endogenous anabolic androgens in females might contribute to sex differences regarding the prevalence of severe asthma