Glucagon-induced changes in fructose 2,6-bisphosphate and 6-phosphofructo-2-kinase in cultured rat foetal hepatocytes

5 pages, 3 figures, 1 table.The sensitivity of 6-phosphofructo-2-kinase to glucagon and cyclic AMP was studied during the perinatal period. In liver homogenates from foetal and neonatal rats, incubation with cyclic AMP produced inactivation of 6-phosphofructo-2-kinase 3 h after birth. The maximal effect was obtained 12 h after birth. In primary cultures of hepatocytes from 22-day-old foetuses, glucogon induced an inhibition of 6-phosphofructo-2-kinase that required 45 min to reach the half-maximal effect. Cycloheximide prevented the glucagon-induced changes in this activity from cultured foetal hepatocytes. These results suggest that the adult form of 6-phosphofructo-2-kinase is rapidly induced after birth, probably by the hormonal changes that occur in this period.This work was supported by grants from Fondo de Investigaciones Sanitarias and Comisión Asesora de Investigación Científica y Técnica.Peer reviewe

Differential substrate specificity could be exploited

Abbreviations: H7 1-(5-isoquinoiine sulphonyl)-2-methylpiperazine dichloride K252b (8R∗; 9s∗; 11∗)-(−)-9-hydroxy-9-methoxycarbonyl-8-methyl-2; 3; 9; 10-tetra-hydro-8; 11-epoxy-1H; 8H; 11H-2; 7b; 11a-triazadibenzo(a; g)cydoocta(c; d; e)trindenl-one-9-carboxylic acid.-- Letter to the editor.Peer Reviewe

Identification of conserved domains in the promoter regions of nitric oxide synthase 2: implications for the species-specific transcription and evolutionary differences

<p>Abstract</p> <p>Background</p> <p>The majority of the genes involved in the inflammatory response are highly conserved in mammals. These genes are not significantly expressed under normal conditions and are mainly regulated at the transcription and prost-transcriptional level. Transcription from the promoters of these genes is very dependent on NF-κB activation, which integrates the response to diverse extracellular stresses. However, in spite of the high conservation of the pattern of promoter regulation in κB-regulated genes, there is inter-species diversity in some genes. One example is nitric oxide synthase 2 (NOS-2), which exhibits a species-specific pattern of expression in response to infection or pro-inflammatory challenge.</p> <p>Results</p> <p>We have conducted a comparative genomic analysis of NOS-2 with different bioinformatic approaches. This analysis shows that in the NOS-2 gene promoter the position and the evolutionary divergence of some conserved regions are different in rodents and non-rodent mammals, and in particular in primates. Two not previously described distal regions in rodents that are similar to the unique upstream region responsible of the NF-κB activation of NOS-2 in humans are fragmented and translocated to different locations in the rodent promoters. The rodent sequences moreover lack the functional κB sites and IFN-γ response sites present in the homologous human, rhesus monkey and chimpanzee regions. The absence of κB binding in these regions was confirmed by electrophoretic mobility shift assays.</p> <p>Conclusion</p> <p>The data presented reveal divergence between rodents and other mammals in the location and functionality of conserved regions of the NOS-2 promoter containing NF-κB and IFN-γ response elements.</p

Phorbol 12,13-dibutyrate and mitogens increase fructose 2,6-bisphosphate in lymphocytes. Comparison of lymphocyte and rat-liver 6-phosphofructo-2-kinase

Now known as FEBS Journal: Open Access content older than 1 year.The influence of tumour promoters and growth factors on glycolysis and on fructose-2,6-bisphosphate concentration was studied in isolated mouse spleen lymphocytes and in purified B-cells. The intracellular concentration of fructose 2,6-bisphosphate and the rate of lactate release were increased 2–3-fold in spleen lymphocytes exposed to active phorbol esters, mitogenic lectins, interleukin 4 or lipopolysaccharide. The maximal effect was observed after 1 h of exposure. In these cells hexose 6-phosphates increased 2-fold and 6-phosphofructo-2-kinase activity remained unchanged after treatment with phorbol 12,13-dibutyrate or with lectins. Exposure of B-cells to phorbol 12,13-dibutyrate, interleukin 4 or lipopolysaccharide increased the glycolytic flux and the concentration of fructose 2,6-bisphosphate without relation to their mitogenic activity. Lymphocytes and rat liver 6-phosphofructo-2-kinase were partially purified using the same procedure. The lymphocyte enzyme was not inhibited by sn-glycerol 3-phosphate in contrast to the potent inhibition observed in liver. Treatment of both enzymes with the catalytic subunit of the cyclic-AMP-dependent protein kinase failed to inactivate 6-phosphofructo-2-kinase from lymphocytes. These differences suggest that lymphocytes and liver contain different forms of this enzyme.This work was supported by a grant from Consejo Superior de Investigaciones Científicas (603/816).Peer Reviewe

Thromboxane A2-Induced Inhibition of Voltage-Gated K+ Channels and Pulmonary Vasoconstriction

8 pages, 6 figures.Voltage-gated K+ channels (KV) and thromboxane A2 (TXA2) play critical roles in controlling pulmonary arterial tone under physiological and pathological conditions. We hypothesized that TXA2 might inhibit KV channels, thereby establishing a link between these two major pathogenic pathways in pulmonary hypertension. The TXA2 analogue U46619 inhibited IK(V) (Emax=56.1±3.9%, EC50=0.054±0.019 µmol/L) and depolarized pulmonary artery smooth muscle cells via activation of TP receptors. In isolated pulmonary arteries, U46619 simultaneously increased intracellular Ca2+ concentration and contractile force, and these effects were inhibited by nifedipine or KCl (60 mmol/L). U46619-induced contractions were not altered by the inhibitors of tyrosine kinase genistein or Rho kinase Y-27632 but were prevented by the nonselective protein kinase C (PKC) inhibitors staurosporine and calphostin C. Furthermore, these responses were sensitive to Gö-6983 but insensitive to bisindolylmaleimide I and Gö-6976. Based on the specificity of these drugs, we suggested a role for an atypical PKC in U46619-induced effects. Thus, treatment with a PKC{zeta} pseudosubstrate inhibitor markedly prevented the vasoconstriction, the inhibition of IK(V), and the depolarization induced by U46619. Western blots showed a transient translocation of PKC{zeta} from the cytosolic to the particulate fraction on stimulation with U46619. These results indicate that TXA2 inhibits IK(V), leading to depolarization, activation of L-type Ca2+ channels, and vasoconstriction of rat pulmonary arteries. We propose PKC{zeta} as a link between TP receptor activation and KV channel inhibition.This work was supported by grants from the Comisión Interministerial de Ciencia y Tecnología (SAF 2002/02304) and Comunidad Autónoma de Madrid (08.4/0036.2001 and 08.3/0008/2001). A.C. and L.M. are supported by grants from Red Temática de Investigación Cardiovascular and Ministerio de Educación Cultura y Deporte, respectively.Peer reviewe

Epigenetics override pro-inflammatory PTGS transcriptomic signature towards selective hyperactivation of PGE 2 in colorectal cancer

This is an Open Access article distributed under the terms of the Creative Commons Attribution License.-- et al.[Background]: Misregulation of the PTGS (prostaglandin endoperoxide synthase, also known as cyclooxygenase or COX) pathway may lead to the accumulation of pro-inflammatory signals, which constitutes a hallmark of cancer. To get insight into the role of this signaling pathway in colorectal cancer (CRC), we have characterized the transcriptional and epigenetic landscapes of the PTGS pathway genes in normal and cancer cells. [Results]: Data from four independent series of CRC patients (502 tumors including adenomas and carcinomas and 222 adjacent normal tissues) and two series of colon mucosae from 69 healthy donors have been included in the study. Gene expression was analyzed by real-time PCR and Affymetrix U219 arrays. DNA methylation was analyzed by bisulfite sequencing, dissociation curves, and HumanMethylation450K arrays. Most CRC patients show selective transcriptional deregulation of the enzymes involved in the synthesis of prostanoids and their receptors in both tumor and its adjacent mucosa. DNA methylation alterations exclusively affect the tumor tissue (both adenomas and carcinomas), redirecting the transcriptional deregulation to activation of prostaglandin E 2 (PGE 2 ) function and blockade of other biologically active prostaglandins. In particular, PTGIS, PTGER3, PTGFR, and AKR1B1 were hypermethylated in more than 40 % of all analyzed tumors. [Conclusions]: The transcriptional and epigenetic profiling of the PTGS pathway provides important clues on the biology of the tumor and its microenvironment. This analysis renders candidate markers with potential clinical applicability in risk assessment and early diagnosis and for the design of new therapeutic strategies.IC was funded by Fundação para a Ciência e a Tecnologia (SFRH/BD/28464/2006); JC was funded by a FPI fellowship. ADV was supported in part by a contract from the Ministerio de Economía y Competitividad (MINECO) (PTC2011-1091). This work was supported by the MINECO(SAF2011/23638, SAF2014/52492), the Catalan Institute of Oncology and the Instituto de Salud Carlos III (grant PI11-01439, RD12/0042/0019 and CIBERESP CB06/02/2005), the Generalitat de Catalunya (grant 2014SGR647), and the Asociación Española Contra el Cáncer (AECC).Peer Reviewe

Inflammation in Parkinson's disease: Mechanisms and therapeutic implications

Parkinson's disease (PD) is a common neurodegenerative disorder primarily characterized by the death of dopaminergic neurons that project from the substantia nigra pars compacta. Although the molecular bases for PD development are still little defined, extensive evidence from human samples and animal models support the involvement of inflammation in onset or progression. However, the exact trigger for this response remains unclear. Here, we provide a systematic review of the cellular mediators, i.e., microglia, astroglia and endothelial cells. We also discuss the genetic and transcriptional control of inflammation in PD and the immunomodulatory role of dopamine and reactive oxygen species. Finally, we summarize the preclinical and clinical approaches targeting neuroinflammation in PD.This work was supported by PID2019-110061RB-I00 and SAF2017-82436R of the Spanish Ministry of Economy and Competiveness; P-024-FTPGB 2018 from the Spanish “Tatiana de Guzman el Bueno Foundation” and by the P_37_732/2016 grant (REDBRAIN) financed by the European Regional Development Fund, Competitiveness Operational Program 2014–2020. Comunidad Autónoma de Madrid (grants B2017/BMD-3827, S2017-BMD-3686). MP is recipient of a contract Juan de la Cierva (MICINN)

Role of caveolin 1 on liver regeneration after partial hepatectomy. Mechanisms of signalling and effect on the regulation of hepatocyte proliferation

[ES]: Las caveolas participan en múltiples procesos celulares tales como el transporte vesicular, homeostasis del colesterol, regulación de la señalización intracelular, por integrinas y proliferación celular. Sin embargo, su función en el hígado no está bien establecida. La expresión de caveolina 1 (Cav), la proteína más abundante en las caveolas, está bien descrita en el hígado y en varias líneas de hepatocitos y en hígado cirrótico humano y en carcinoma hepatocelular. Sin embargo, el papel de Cav-1 en la fisiopatología hepática es controvertido, ya que se ha propuesto un papel crítico en el proceso de regeneración tras hepatectomía parcial (HP). Contrariamente a esta observación, nuestros datos sugieren que Cav-1 aumenta en el hígado regenerante, con una re-distribución de la proteína desde las caveolas hacia dominios no caveolares. Además, la Cav-1 localizada en estas fracciones está fosforilada en la tirosina 14. A pesar de ello, el gen de la Cav-1 es dispensable para la regeneración hepática tras HP, tal como se deduce de animales que carecen de este gen. En conjunto, estos datos muestran un papel dinámico de la Cav-1 en la proliferación hepática tras HP y en líneas hepáticas en cultivo, pero con mínimas implicaciones en el proceso regenerativo.[EN]: Although caveolae participate in many cellular processes such as vesicular transport, cholesterol homeostasis, regulation of signal transduction, integrin signaling and cell growth, their role in liver remains elusive. Expression of caveolin 1 (Cav), the most abundant protein of caveolae, has been reported in liver and in different hepatocyte cell lines, in human cirrhotic liver and in hepatocellular carcinomas. However, the role of Cav-1 in liver pathophysiology remains controversial and a critical role in regeneration after partial hepatectomy (PH) has been reported. Opposite to this observation, our data support the view that Cav-1 increases in liver after PH with a redistribution of the protein from the caveolae enriched domain to the noncaveolar fraction. Moreover, the Cav-1 located in the noncaveolar fraction is phosphorylated in tyrosine 14 (Tyr14). Even though, the Cav-1 gene is dispensable for liver regeneration after PH as deduced from data obtained with commercially available animals lacking this gene. Taken together these results support a dynamic role for Cav-1 in liver proliferation both in vivo after PH, and in vitro in cultured hepatic cell lines, but with minimal implications in the liver regeneration process.Peer Reviewe

Unraveling the interplay between iron homeostasis, ferroptosis and extramedullary hematopoiesis

Iron participates in myriad processes necessary to sustain life. During the past decades, great efforts have been made to understand iron regulation and function in health and disease. Indeed, iron is associated with both physiological (e.g., immune cell biology and function and hematopoiesis) and pathological (e.g., inflammatory and infectious diseases, ferroptosis and ferritinophagy) processes, yet few studies have addressed the potential functional link between iron, the aforementioned processes and extramedullary hematopoiesis, despite the obvious benefits that this could bring to clinical practice. Further investigation in this direction will shape the future development of individualized treatments for iron-linked diseases and chronic inflammatory disorders, including extramedullary hematopoiesis, metabolic syndrome, cardiovascular diseases and cancer.Work in the authors laboratory is supported by: Ministerio de Economía, Industria y Competitividad/Agencia Estatal de Investigacion ´ 10.13039/501100011033 (PID2019-108977RB-I00 and PID2020- 113238RB-I00), Centro de Investigacion Biomédica en Red en Enfermedades Cardiovasculares (CB16/11/00222), Consorcio de Investigacion en Red de la Comunidad de Madrid, S2017/BMD-3686 and Fondo Europeo de Desarrollo Regiona

Lipoxin A4 impairment of apoptotic signaling in macrophages: Implication of the PI3K/Akt and the ERK/Nrf-2 defense pathways

El pdf del artículo es la versión pre-print.Lipoxin A4 (LXA4) is an endogenous lipid mediator that requires transcellular metabolic traffic for its synthesis. The targets of LXA4 on neutrophils are well described, contributing to attenuation of inflammation. However, effects of lipoxins on macrophage are less known, particularly the action of LXA4 on the regulation of apoptosis of these cells. Our data show that pretreatment of human or murine macrophages with LXA4 at the concentrations prevailing in the course of resolution of inflammation (nanomolar range) significantly inhibits the apoptosis induced by staurosporine, etoposide and S-nitrosoglutathione or by more pathophysiological stimuli, such as LPS/IFNγ challenge. The release of mitochondrial mediators of apoptosis and the activation of caspases was abrogated in the presence of LXA4. In addition to this, the synthesis of reactive oxygen species induced by staurosporine was attenuated and antiapoptotic proteins of the Bcl-2 family accumulated in the presence of lipoxin. Analysis of the targets of LXA4 identified an early activation of the PI3K/Akt and ERK/Nrf-2 pathways, which was required for the observation of the antiapoptotic effects of LXA4. These data suggest that the LXA 4, released after the recruitment of neutrophils to sites of inflammation, exerts a protective effect on macrophage viability that might contribute to a better resolution of inflammation. © 2010 Macmillan Publishers Limited All rights reserved.PP was supported by a fellowship from Ministerio de Ciencia e Innovación (MICINN; Spain). This work was supported by grant BFU2008-02161 from MICINN, S-BIO-0283/2006 from Comunidad de Madrid and FIS-RECAVA RD06/0014/0025. RECAVA and Ciberehd are funded by the Instituto de Salud Carlos III.Peer Reviewe