Post-transcriptional regulation of inducible nitric oxide in chronic lymphocytic leukemia B cells in pro and antiapoptotic culture conditions

Functional inducible NOS(INOS) may be involved in the prolonged lifespan of chronic lymphocytic leukemia cells (B-CLL), although the exact mechanisms implicated remain elusive as yet. In this work, we have examined INOS expression in normal B lymphocytes and B-CLL in pro and antiapoptotic conditions. Our results demonstrate: 1) The existence of a new splice variant charaterized by a complete deletion of exon 14 (INOS 13-16) which was preferentially detected in normal B lymphocytes and may represent an isoform that could be play a role in the regulation of enzyme activity. 2) The existence of another alternatively spliced INOS mRNA transcript involving a partial deletion of the flavodoxin region (INOS 13-16) was correlated to a decreased B-Cll cell viability. The 9-B-D arabinofuranosyl-2-fluoradenine or fludarabine (F-ara) treatment induced INOS 13-16) and the expression of a 122kDa INOS protein. These results suggest that in B-CLL, a regulation process involving nitric oxide (NO) levels could occur by a post-transcriptional mechanism mediated by soluble factors. Our results also provide an insight into a new complmentary proapoptotic action of F-ara in B-CLL by the induction of particular INOS splice variants, leading to the activation of a caspase-3-dependent apoptotic pathway

Peroxynitrite inhibits T lymphocyte activation and proliferation by promoting impairment of tyrosine phosphorylation and peroxynitrite-driven apoptotic death

Peroxynitrite (ONOO-) is a potent oxidizing and nitrating agent produced by the reaction of nitric oxide with superoxide. It readily nitrates phenolic compounds such as tyrosine residues in proteins, and it has been demonstrated that nitration of tyrosine residues in proteins inhibits their phosphorylation. During immune responses, tyrosine phosphory lation of key substrates by protein tyrosine kinases is the earliest of the intracellular signaling pathways folowing activation through the TCR complex. This work was aimed to evaluate the effects of ONOO- on lymphocyte tyrosine phosphorylation, proliferation, and survival. Additionally, we studied the generation of nitrating species in vivo and in vitro during immune activation. Our results demonstrate that ONOO-, through nitration of tyrosine residues, is able to inhibit activation-induced protein tyrosine phosphorylation in purified lymphocytes and prime them to undergo apoptotic cell death after PHA- or CD3-mediated activation but non upon phorbol ester.mediated stimulation. We also provide evidence indicating that peroxynitrite is produced during in vitro immune activation, mainly by cell of the monocyte/macrophage lineage. Furthermore, immunohistochemical studies demonstrate the in vivo generation of nitrating species in human lymph nodes undergoing mild to strong immune activation. Our results point to a physiological role for ONOO- as a down-modulator of immune responses and also as key mediator in cellular and tissue injury associated with chronic activation of the immune system

Estadificación preoperatoria del cáncer colo-rectal mediante la asociación de cuatro marcadores tumorales

Los autores cuantificaron los niveles séricos de antígeno carcinoembrionario (CEA), fosfohexosa isomerasa (FHI), ácido siálico y proteínas solubles en ácido perclórico (APC) en 32 pacientes con cáncer colo-rectal, a los efectos de determinar la utilidad de la asociación de estos marcadores tumorales para la estadificación preoperatoria. El ácido siálico de la fracción APC fue el más sensible para la enfermedad localizada (62,5 porciento) y para la afectación loco-regional 78,6 porciento), mientras que en la enfermedad metastásica los más sensibles fueron la FHI (100 porciento) y el CEA (90 porciento). Sólo 1 de kis 32 pacientes, portador de un cáncer localizado, presentó los cuatro parámetros en el rango de normalidad. Se observó que existe relación entre el número de marcadores elevados y el estadio de la enfermedad, existiendo patrones que tienen valor predictivo positivo para la estadifiación preoperatoria: a) para la enfermedad localizada, la ausencia de marcadores elevados (100 porciento); b) para la afectación loco-regional, la elevación de tres marcadores (77,8 porciento) y c) para la enfermedad metastásica, el aumento de los cuatro marcadores utilizados (87,5 porciento). Se conbcluye que ante la ausencia de un marcador tumoral ideal para el cáncer colo-rectal, con la determinación simultánea de estos cuatro marcadores bioquímicos, se mejoran notoriamente las posibilidades diagnósticas, por la mayor probabilidad de detección de la población afectada y, fundamentalmente, porqu en muchos casos se puede predecir el estado de la enfermedad