DNA methylation in ciliates: implications in differentiation processes

Much experimental evidence on the role of DNA methylation in gene expression has been reported. Here we review reports on DNA methylation in ciliated protozoa, emphasizing its implications in cell differentiation processes. Both types of methylated bases (adenine and cytosine) can be found in macronuclear DNA. The division cycle and conjugation have been studied with regard to adenine methylation, and several different functions have been assigned to the methylation changes detected in these processes. Cytosine methylation changes were analyzed during stomatogenesis of Paramecium and encystment of Colpoda inflata. A comparative analysis with other similar microbial eukaryotic differentiation processes is carried out

Mitochondrial dysfunction in human pathologies

The integrity of mitochondrial function is fundamental to cell life. The cell demands for mitochondria and their complex integration into cell biology, extends far beyond the provision of ATP. It follows that disturbances of mitochondrial function lead to disruption of cell function, expressed as disease or even death. Mitochondria are major producers of free radical species and also possibly of nitric oxide, and are, at the same time, major targets for oxidative damage. In this review we consider recent developments in our knowledge of how the mitochondrial production of reactive oxygen species (ROS) plays a critical role in several major human pathologies. We will also consider recent advances in our understanding of the molecular mechanisms involved in mitochondrial ROS detoxification.This work was supported by an institutional grant from the CNIC, by Plan Nacional de I+D+I grants SAF2003-01039 and SAF2003-04901 and grant-in-aid from the Spanish Society of Nephrology to Dr. Santiago Lamas. Sara Borniquel is holder of grant SAF2003-04901 predoctoral fellowship. Inmaculada Valle is holder of a CNIC-Bancaja predoctoral fellowship. Maria Monsalve is holder of a Ramon y Cajal contract from the Ministerio de Educación y Ciencia.Peer Reviewe

Preventive and therapeutic effects of nitrite supplementation in experimental inflammatory bowel disease

BACKGROUND: Inorganic nitrate and nitrite have emerged as alternative substrates for nitric oxide (NO) generation in the gastrointestinal tract, and have shown to be protective against drug-induced gastric injury. The aim of this study was to investigate the preventive and therapeutic effects of nitrate and nitrite in a model of experimental colitis. METHODS: Colitis was induced in mice by administrating dextran sulfate sodium (DSS) with concurrent administration of nitrite (1 mM) or nitrate (10 mM) in the drinking water for 7 days. A therapeutic approach was also investigated by initiating nitrite treatment 3 days after DSS-induced colitis. Clinical and inflammatory markers were assessed and the colonic mucus thickness was measured in vivo. The effect of nitrite on wound healing was evaluated using colon epithelial cells. RESULTS: Concurrent administration of DSS and nitrite (1 mM) alleviated inflammation as determined by reduced disease activity index score (DAI) and increased colon length, while nitrate (10 mM) only reduced the DAI-score. Nitrite also displayed therapeutic effects by ameliorating established colonic inflammation with reduced colonic expression of iNOS and improving histopathology. DSS-induced decrease in colonic mucus thickness was completely prevented by nitrite administration. In addition, goblet cell abundance was lower by DSS treatment, but was increased by addition of nitrite. Further studies using colon epithelial cells revealed an NO-dependent improvement in wound healing with nitrite administration. CONCLUSION: Nitrite exerts both preventive and therapeutic effects in colonic inflammation. The protective effects involve preservation of an intact adherent mucus layer and regulation of epithelial cell restitution

Nitric oxide regulates mitochondrial oxidative stress protection via the transcriptional coactivator PGC-1α

Nitric oxide (NO) has both prooxidant and antioxidant activities in the endothelium; however, the molecular mechanisms involved are still a matter of controversy. PGC-1α [peroxisome proliferators-activated receptor (PPAR) γ coactivator 1-α] induces the expression of several members of the mitochondrial reactive oxygen species (ROS) detoxification system. Here, we show that NO regulates this system through the modulation of PGC-1α expression. Short-term (24 h) treatment up-regulates it. Treatment with the NOS inhibitor L-NAME has the opposite effect. Downregulation of PGC-1α by NO is mediated by protein kinase G (PKG). It is blocked by the soluble guanylate cyclase (sGC) inhibitor ODQ and the PKG inhibitor KT5823, and mimicked by the cGMP analog 8-Br-cGMP. Changes in PGC-1α expression are in all cases paralleled by corresponding variations in the mitochondrial ROS detoxification system. Cells that transiently overexpress PGC-1α from the cytomeglovirus (CMV) promoter respond poorly to NO donors. Analysis of tissues from eNOS-/- mice showed reduced levels of PGC-1α and the mitochondrial ROS detoxification system. These data suggest that NO can regulate the mitochondrial ROS detoxification system both positively and negatively through PGC-1α.This work was supported by an institutional grant from the CNIC, Plan Nacional de I+D+I grants SAF2003–01039, SAF2003–04901, BFI2003–03493, and grant-in-aid from the Spanish Society of Nephrology to S.L. S.B. is a holder of a predoctoral fellowship under grant SAF2003–04901. Inmaculada Valle is a holder of a CNIC-Bancaja predoctoral fellowship. M.M. and S.C. are holders of a Ramon&Cajal contract from the Ministerio de Educación y Ciencia.Peer Reviewe

Preventive and therapeutic effects of nitrite supplementation in experimental inflammatory bowel disease

BACKGROUND: Inorganic nitrate and nitrite have emerged as alternative substrates for nitric oxide (NO) generation in the gastrointestinal tract, and have shown to be protective against drug-induced gastric injury. The aim of this study was to investigate the preventive and therapeutic effects of nitrate and nitrite in a model of experimental colitis. METHODS: Colitis was induced in mice by administrating dextran sulfate sodium (DSS) with concurrent administration of nitrite (1 mM) or nitrate (10 mM) in the drinking water for 7 days. A therapeutic approach was also investigated by initiating nitrite treatment 3 days after DSS-induced colitis. Clinical and inflammatory markers were assessed and the colonic mucus thickness was measured in vivo. The effect of nitrite on wound healing was evaluated using colon epithelial cells. RESULTS: Concurrent administration of DSS and nitrite (1 mM) alleviated inflammation as determined by reduced disease activity index score (DAI) and increased colon length, while nitrate (10 mM) only reduced the DAI-score. Nitrite also displayed therapeutic effects by ameliorating established colonic inflammation with reduced colonic expression of iNOS and improving histopathology. DSS-induced decrease in colonic mucus thickness was completely prevented by nitrite administration. In addition, goblet cell abundance was lower by DSS treatment, but was increased by addition of nitrite. Further studies using colon epithelial cells revealed an NO-dependent improvement in wound healing with nitrite administration. CONCLUSION: Nitrite exerts both preventive and therapeutic effects in colonic inflammation. The protective effects involve preservation of an intact adherent mucus layer and regulation of epithelial cell restitution

Ciliate cryptobiosis: a microbial strategy against environmental starvation

This review outlines the main features of ciliate resting-cyst formation or encystment. It represents a strategy against several environmental stresses such as starvation), which involves a highly gene-regulated cell di€erentiation process and originates a more resistant, differentiated form or resting cyst. This process is mainly characterized by drastic cytoplasmic dehydration that induces a general metabolic rate decrease, intense autophagic activity, the formation of a permeable cyst wall protecting the cell against the adverse environmental conditions, and a gene-silencing mechanism after opening the specific encystment genes

Inactivation of Foxo3a and Subsequent Downregulation of PGC-1α Mediate Nitric Oxide-Induced Endothelial Cell Migration▿

In damaged or proliferating endothelium, production of nitric oxide (NO) from endothelial nitric oxide synthase (eNOS) is associated with elevated levels of reactive oxygen species (ROS), which are necessary for endothelial migration. We aimed to elucidate the mechanism that mediates NO induction of endothelial migration. NO downregulates expression of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), which positively modulates several genes involved in ROS detoxification. We tested whether NO-induced cell migration requires PGC-1α downregulation and investigated the regulatory pathway involved. PGC-1α negatively regulated NO-dependent endothelial cell migration in vitro, and inactivation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway, which is activated by NO, reduced NO-mediated downregulation of PGC-1α. Expression of constitutively active Foxo3a, a target for Akt-mediated inactivation, reduced NO-dependent PGC-1α downregulation. Foxo3a is also a direct transcriptional regulator of PGC-1α, and we found that a functional FoxO binding site in the PGC-1α promoter is also a NO response element. These results show that NO-mediated downregulation of PGC-1α is necessary for NO-induced endothelial migration and that NO/protein kinase G (PKG)-dependent downregulation of PGC-1α and the ROS detoxification system in endothelial cells are mediated by the PI3K/Akt signaling pathway and subsequent inactivation of the FoxO transcription factor Foxo3a