The MDM2-p53 pathway is involved in preconditioning-induced neuronal tolerance to ischemia

Article number: 1610 (2018)[EN]Brain preconditioning (PC) refers to a state of transient tolerance against a lethal insult that can be evoked by a prior mild event. It is thought that PC may induce different pathways responsible for neuroprotection, which may involve the attenuation of cell damage pathways, including the apoptotic cell death. In this context, p53 is a stress sensor that accumulates during brain ischemia leading to neuronal death. The murine double minute 2 gene (MDM2), a p53-specific E3 ubiquitin ligase, is the main cellular antagonist of p53, mediating its degradation by the proteasome. Here, we study the role of MDM2-p53 pathway on PC-induced neuroprotection both in cultured neurons (in vitro) and rat brain (in vivo). Our results show that PC increased neuronal MDM2 protein levels, which prevented ischemia-induced p53 stabilization and neuronal death. Indeed, PC attenuated ischemia-induced activation of the p53/PUMA/caspase-3 signaling pathway. Pharmacological inhibition of MDM2-p53 interaction in neurons abrogated PC-induced neuroprotection against ischemia. Finally, the relevance of the MDM2-p53 pathway was confirmed in rat brain using a PC model in vivo. These findings demonstrate the key role of the MDM2-p53 pathway in PC-induced neuroprotection against a subsequent ischemic insult and poses MDM2 as an essential target in ischemic tolerance

Gadd45b Acts as Neuroprotective Effector in Global Ischemia-Induced Neuronal Death

Purpose Transient global ischemia arising in human due to cardiac arrest causes selective, delayed neuronal death in hippocampal CA1 and cognitive impairment. Growth arrest and DNA-damage-inducible protein 45 beta (Gadd45b) is a wellknown molecule in both DNA damage-related pathogenesis and therapies. Emerging evidence suggests that Gadd45b is an anti-apoptotic factor in nonneuronal cells and is an intrinsic neuroprotective molecule in neurons. However, the mechanism of Gadd45b pathway is not fully examined in neurodegeneration associated with global ischemia. Methods Rats were subjected to transient global ischemia by the 4-vessel occlusion or sham operation. The animals were sacrificed at 24 hours, 48 hours, and 7 days after ischemia. The hippocampal CA1 was microdissected and processed to examine mRNA and protein level. To assess neuronal death, tissue sections were cut and processed for Fluoro-Jade and Nissl staining. Results Here we show that ischemic insults increase abundance of Gadd45b and brain-derived neurotrophic factor, a known target of Gadd45 mediated demethylation, in selectively-vulnerable hippocampal CA1 neurons. We further show that knockdown of Gadd45b increases abundance of a pro-apoptotic Bcl-2 family member Bax while decreasing the antiapoptotic protein Bcl-2, which together promote neuronal death. Conclusions These findings document a protective role of Gadd45b against neuronal insults associated with global ischemia and identify Gadd45b as a potential therapeutic target for the amelioration of hippocampal neurodegeneration

Neuroprotective Actions of Estradiol and Novel Estrogen Analogs in Ischemia: Translational Implications

This review highlights our investigations into the neuroprotective efficacy of estradiol and other estrogenic agents in a clinically relevant animal model of transient global ischemia, which causes selective, delayed death of hippocampal CA1 neurons and associated cognitive deficits. We find that estradiol rescues a significant number of CA1 pyramidal neurons that would otherwise die in response to global ischemia, and this is true when hormone is provided as a long-term pretreatment at physiological doses or as an acute treatment at the time of reperfusion. In addition to enhancing neuronal survival, both forms of estradiol treatment induce measurable cognitive benefit in young animals. Moreover, estradiol and estrogen analogs that do not bind classical nuclear estrogen receptors retain their neuroprotective efficacy in middle-aged females deprived of ovarian hormones for a prolonged duration (8 weeks). Thus, non-feminizing estrogens may represent a new therapeutic approach for treating the neuronal damage associated with global ischemia

The MDM2-dependent p53 degradation signaling pathway is involved in neuronal ischemic tolerance

trabajo presentado en el 3rd European Stroke Organisation Conference ​(ESOC 2017), celebrado en Praga (República Checa), del 16 al 18 de mayo de 2017Peer reviewe

Novel pathways involved in the control of survival and death in neuronal ischemic tolerance

Trabajo presentado en el XL Congreso de la Sociedad Española de Bioquímica y Biología Molecular. FEBS3+1st Joint Meeting of the French-Portuguese-Spanish Biochemical and Molecular, celebrado en Barcelona (España), del 23 al 26 de octubre de 2017Brain preconditioning (PC) refers to a state of transient tolerance against a lethal insult that can be evoked by a prior mild event. The endogenous molecular mechanisms underlying PC-induced neuroprotection and PC-increased brain ischemic tolerance (IT) are still unknown. However, several studies suggest that PC promotes the attenuation of cell damage pathways, including the apoptotic cell death. In this context, p53 is a stress sensor that accumulates during brain ischemia leading to neuronal death. Our work is focused on the study of the role of p53-pathway on PC-induced IT. For this purpose, we used a validated model of PC in vitro: primary cultured cortical neurons were exposed to low doses of N-methyl-D-aspartate (20 μM NMDA; NMDA-PC) for further 2 h either prior ischemic insult (PC+OGD) or not (oxygen and glucose deprivation; OGD for 90 min). After 4 hours, our results revealed that NMDA-PC promotes cell survival through the control of the p53/PUMA/caspase-3 signaling pathway. Indeed, NMDA-PC increased the murine double minute 2 gene (MDM2), the main cellular antagonist of p53, which prevented ischemia-induced p53 stabilization and apoptotic neuronal death. Pharmacological inhibition of MDM2-p53 interaction in neurons abrogated NMDA-PC-induced neuroprotection against ischemia. Finally, the relevance of the MDM2-p53 pathway was confi rmed in rat brain using an PC model in vivo. The results showed that PC increased MDM2 protein levels, promoted p53 destabilization and attenuated ischemia-induced cerebral infarction. These fi ndings demonstrate that PC promotes cell survival through the control of the p53 signaling pathway against a subsequent ischemic insult and poses MDM2 as an essential target in IT.Funded by grants from the Instituto de Salud Carlos III (Miguel Servet I CP0014/00010; RD12/0014/0007); BIO/SA35/15 JCyL and FEDER (European regional development fund).Peer reviewe

Cerebroprotective Effect of 17β-Estradiol Replacement Therapy in Ovariectomy-Induced Post-Menopausal Rats Subjected to Ischemic Stroke: Role of MAPK/ERK1/2 Pathway and PI3K-Independent Akt Activation

Despite the overwhelming advances in the understanding of the pathogenesis of stroke, a devastating disease affecting millions of people worldwide, currently there are only a limited number of effective treatments available. Preclinical and clinical studies show that stroke is a sexually dimorphic disorder, affecting males and females differently. Strong experimental evidence indicates that estrogen may play a role in this difference and that exogenous 17β-estradiol (E2) is neuroprotective against stroke in both male and female rodents. However, the molecular mechanisms by which E2 intervenes in ischemia-induced cell death, revealing these sex differences, remain unclear. The present study was aimed to determine, in female rats, the molecular mechanisms of two well-known pro-survival signaling pathways, MAPK/ERK1/2 and PI3K/Akt, that mediate E2 neuroprotection in response to acute ischemic stroke. E2 pretreatment reduced brain damage and attenuated apoptotic cell death in ovariectomized female rats after an ischemic insult. Moreover, E2 decreased phosphorylation of ERK1/2 and prevented ischemia/reperfusion-induced dephosphorylation of both Akt and the pro-apoptotic protein, BAD. However, MAPK/ERK1/2 inhibitor PD98059, but not the PI3K inhibitor LY294002, attenuated E2 neuroprotection. Thus, these results suggested that E2 pretreatment in ovariectomized female rats modulates MAPK/ERK1/2 and activates Akt independently of PI3K to promote cerebroprotection in ischemic stroke. A better understanding of the mechanisms and the influence of E2 in the female sex paves the way for the design of future successful hormone replacement therapies

Diabetes impairs the atrial natriuretic peptide relaxant action mediated by potassium channels and prostacyclin in the rabbit renal artery

Diabetes is associated with increased prevalence of hypertension, cardiovascular and renal disease. Atrial natriuretic peptide (ANP) plays an important role in cardiovascular pathophysiology and is claimed to have cardioprotective and renoprotective effect in diabetic patients. The working hypothesis was that alloxan-induced diabetes might modify the vascular effects of ANP in isolated rabbit renal arteries and the mechanisms involved in such actions. Plasma ANP levels were higher in diabetic rabbits than in control rabbits. ANP (10−12–10−7 M) induced a relaxation of precontracted renal arteries, which was lower in diabetic than in control rabbits. In arteries from both groups of animals, endothelium removal decreased the ANP-induced relaxation but inhibition of NO-synthesis did not modify ANP-induced relaxations. In KCl-depolarised arteries, relaxation to ANP was almost abolished both in control and diabetic rabbits. Tetraethylammonium (TEA) partly inhibited the relaxation to ANP in control rabbits but did not modify it in diabetic rabbits. Glibenclamide and 4-aminopyridine inhibited the relaxation to ANP, and these inhibitions were lower in diabetic than in control rabbits. Indomethacin potentiated the relaxation to ANP, more in control than in diabetic rabbits. In the presence of ANP the renal artery released thromboxane A2 and prostacyclin, and the release of prostacyclin resulted decreased in diabetic rabbits. The present results suggest that diabetes produces hyporeactivity of the rabbit renal artery to ANP by mechanisms that at least include the reduced modulation by prostacyclin and a lower participation of ATP-sensitive K+ channel (KATP), voltage-sensitive K+ channels (KV) and TEA-sensitive K+ channels (KCa).Medicin