Cigarette smoke induces pulmonary arterial dysfunction through an imbalance in the redox status of the soluble guanylyl cyclase.

Chronic obstructive pulmonary disease (COPD), whose main risk factor is cigarette smoking (CS), is one of the most common diseases globally. Some COPD patients also develop pulmonary hypertension (PH), a severe complication that leads to premature death. Evidence suggests reactive oxygen species (ROS) involvement in COPD and PH, especially regarding pulmonary artery smooth muscle cells (PASMC) dysfunction. However, the effects of CS-driven oxidative stress on the pulmonary vasculature are not completely understood. Herein we provide evidence on the effects of CS extract (CSE) exposure on PASMC regarding ROS production, antioxidant response and its consequences on vascular tone dysregulation. Our results indicate that CSE exposure promotes mitochondrial fission, mitochondrial membrane depolarization and increased mitochondrial superoxide levels. However, this superoxide increase did not parallel a counterbalancing antioxidant response in human pulmonary artery (PA) cells. Interestingly, the mitochondrial superoxide scavenger mitoTEMPO reduced mitochondrial fission and membrane potential depolarization caused by CSE. As we have previously shown, CSE reduces PA vasoconstriction and vasodilation. In this respect, mitoTEMPO prevented the impaired nitric oxide-mediated vasodilation, while vasoconstriction remained reduced. Finally, we observed a CSE-driven downregulation of the Cyb5R3 enzyme, which prevents soluble guanylyl cyclase oxidation in PASMC. This might explain the CSE-mediated decrease in PA vasodilation. These results provide evidence that there might be a connection between mitochondrial ROS and altered vasodilation responses in PH secondary to COPD, and strongly support the potential of antioxidant strategies specifically targeting mitochondria as a new therapy for these diseases.

AG490 promotes HIF-1α accumulation by inhibiting its hydroxylation.

AG490 is a tyrphostin originally described as a Janus Activated Kinase (JAK) 2 inhibitor. AG490 also inhibits epidermal growth factor receptor (EGFR) and guanylyl cyclases (GC). More recently, AG490 was associated with oxidative stress protection in experimental acute kidney injury models. We now show that AG490 is also a strong activator of the Hypoxia Inducible Factor (HIF)-1. Under normoxic conditions HIF-1α is degraded through hydroxylation, von Hippel Lindau protein (VHL)-mediated ubiquitin tagging and proteasomal degradation. AG490 increased HIF-1α protein, but not HIF-1α mRNA levels, dose- and time-dependently in cultured endothelial, vascular smooth muscle and kidney proximal tubular epithelial cells. AG490 increased HIF-1α protein half-life, suggesting that HIF-1α protein accumulation resulted from a decreased degradation. In this regard, AG490 prevented HIF-1α hydroxylation and increased HIF-1α protein levels in human renal carcinoma cells expressing VHL, but did not further increase HIF-1α in VHL negative cells. AG490 did not prevent the proteasomal degradation of other proteins. HIF-1α was not upregulated by dominant negative JAK2constructs, tyrphostin AG9, the EGFR inhibitors erbstatin and genistein, the GC inhibitor Ly83583 or cGMP analogues. Finally, AG490 also increased HIF-1α transcriptional activity evidenced by the increased HIF-1α-dependent VEGF expression. In conclusion, AG490 is a novel HIF-1α activator that increases HIF-1α half-life and protein levels through interference with HIF-1α hydroxylation and VHL-mediated degradation. This action may contribute to the cell and tissue protective effects of AG490.

Von Hippel-Lindau syndrome: molecular mechanisms of the disease

Inactivation of the von Hippel-Lindau (VHL) tumour suppressor gene is responsible for the development of renal carcinomas, pheochromocytomas and tumours in other organs. The gene product (pVHL) is a central component in the oxygen-sensing pathway through its role in the regulation of the hypoxia-inducible factor (HIF). Loss of pVHL leads to activation of the HIF pathway in normoxia with the concomitant increase in tumour vascularisation due to the up-regulation of pro-angiogenic genes. However, although the role of pVHL in the regulation of HIF has proved to be important for tumour growth, other pVHL functions independent of HIF have been reported and help to explain why loss of VHL leads to renal cancer. Studies aimed to characterise other molecular pathways that shed light on its physiological roles as a gatekeeper gene in kidney and other organs will be very helpful for the development of novel anticancer therapies (AU)

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Hypoxia-inducible factors and cancer.

Decreased oxygen availability is a common feature during embryonic development as well of malignant tumours. Hypoxia regulates many transcription factors, and one of the most studied is the hypoxia-inducible factor (HIF). As a consequence of HIF stabilisation, the cell constitutively upregulates the hypoxic programme resulting in the expression of genes responsible for global changes in cell proliferation, angiogenesis, metastasis, invasion, de-differentiation and energy metabolism. Of the three known alpha subunits of HIF transcription factors, HIF-1alpha and HIF-2alpha have been the most studied. Their differential expression and function have been widely discussed, however no clear picture has been drawn on how these two transcription factors differently regulate common and unique target genes. Their role as oncogenes has also been suggested in several studies. In this review we provide an overview of the current knowledge on some of the most important aspects of HIFalpha regulation, its role in tumour angiogenesis and energetic metabolism. We also give an overview of how the modulation of HIF regulating pathways is a potential therapeutic target that may have benefits in the treatment of cancer.

Functional characterization of GPIIb- and GPIIIa-specific monoclonal antibodies. Further evidence for the existence of agonist-specific activated states of the platelet fibrinogen receptor.

In this work human platelet aggregation induced in vitro by ADP, collagen, arachidonic acid and U-46619 (a thromboxane A(2) analogue) was used as a functional test to characterize 19 anti-GPIIb (M series) and anti2 GPIIIa (P series) monoclonal antibodies whose epitope location is known for most of them. Additionally, flow cytofluorimetry was applied to study the epitope expression of these antibodies in resting, EDTA-treated and SFLLRN peptide (thrombin receptor agonist)-activated platelets. Antibodies M6 (epitope located at GPIIbH 657-665), P23-7 (GPIIIa 114-122) and P40 (GPIIIa 262-303) bind weakly to only 43%, 70% and 66%, respectively, of the resting platelet population. This binding was enhanced in EDTA-treated and in activated platelets. Platelet activation enhances the apparent binding of most of the other antibodies. Further evidence on the existence of agonist-specific activated states of GPIIb/IIIa was provided by the agonist-dependent immunochemical inhibition in vitro of platelet aggregation by some of the anti-subunit antibodies studied here. The most notable cases are those of P40 and M6, which at 140 nM inhibit most, the platelet aggregation induced by arachidonic acid and U-46619. On the other hand, three of the most strong and agonist-independent inhibitors, P37 (GPIIIa 101-109), P97 and P95-2 (GPIIIa N-terminal half) bind to resting platelets with high affinity (5-8 nM), compete with each other for binding to GPIIb-IIIa and their epitopes are located at the N-terminal domain of GPIIIa, where the receptor ligand binding site(s) have been found. Given that the formation of activated GPIIb-IIIa (GPIIb-IIIa*) is the first step at which the anti-subunit antibodies can intervene as inhibitors and that agonist-specific inhibitors should block only agonist-specific steps, while nonspecific inhibitors should block steps common to all the agonists, then our present work support the hypothesis that there are different agonist-specific GPIIb-IIIa*s or, alternatively, different receptor environments, that can be specifically blocked by some of the antibodies. These results add to earlier evidence on agonist-dependent ligand specificity and activated states found for this and other integrins. Finally, the correlation between the in vitro inhibition of platelet aggregation and the antithrombotic activity in vivo is discussed for these antibodies.

[Hepatitis B virus vaccination in the province of Saragossa]. / Vacunación de hepatitis B en la provincia de Zaragoza.

We have studied 3001 hepatitis B vaccination protocols, during the period 1982-89, in three hospitals of Zaragoza. 87.24% of the vaccinations have been conducted on health staff, meanwhile other epidemiological important groups (drug users) don't have been attended. The seroprevalence of HBsAg was 0.96% at the prevaccination study. The cleaning staff and the hemodialyzed patients had the greatest ones (5.69% and 3.33%). The administration of the vaccine failed due to rejection in 25.50% and to withdrawal in 8.09%. Only 8.65% don't have developed antibodies after vaccination.

[Epidemiology of brucellosis. Retrospective study of 246 hospital cases]. / Epidemiología de la brucelosis. Estudio retrospectivo de 246 casos hospitalarios.

246 cases of brucellosis registered by the Medicine Preventive Service of Miguel Servet Hospital for the period 1981-87 are retrospectively studied. It emphasizes the consumption of non controlled dairy products as the most frequent (54.5%) risk factor. A greater prevalence of professional and contact with cattle risk factors are observed in the male patients coming from rural areas.