Oral administration of the KATP channel opener diazoxide ameliorates disease progression in a murine model of multiple sclerosis

Background Multiple Sclerosis (MS) is an acquired inflammatory demyelinating disorder of the central nervous system (CNS) and is the leading cause of nontraumatic disability among young adults. Activated microglial cells are important effectors of demyelination and neurodegeneration, by secreting cytokines and others neurotoxic agents. Previous studies have demonstrated that microglia expresses ATP-sensitive potassium (KATP) channels and its pharmacological activation can provide neuroprotective and anti-inflammatory effects. In this study, we have examined the effect of oral administration of KATP channel opener diazoxide on induced experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. Methods Anti-inflammatory effects of diazoxide were studied on lipopolysaccharide (LPS) and interferon gamma (IFNy)-activated microglial cells. EAE was induced in C57BL/6J mice by immunization with myelin oligodendrocyte glycoprotein peptide (MOG35-55). Mice were orally treated daily with diazoxide or vehicle for 15 days from the day of EAE symptom onset. Treatment starting at the same time as immunization was also assayed. Clinical signs of EAE were monitored and histological studies were performed to analyze tissue damage, demyelination, glial reactivity, axonal loss, neuronal preservation and lymphocyte infiltration. Results Diazoxide inhibited in vitro nitric oxide (NO), tumor necrosis factor alpha (TNF-¿) and interleukin-6 (IL-6) production and inducible nitric oxide synthase (iNOS) expression by activated microglia without affecting cyclooxygenase-2 (COX-2) expression and phagocytosis. Oral treatment of mice with diazoxide ameliorated EAE clinical signs but did not prevent disease. Histological analysis demonstrated that diazoxide elicited a significant reduction in myelin and axonal loss accompanied by a decrease in glial activation and neuronal damage. Diazoxide did not affect the number of infiltrating lymphocytes positive for CD3 and CD20 in the spinal cord. Conclusion Taken together, these results demonstrate novel actions of diazoxide as an anti-inflammatory agent, which might contribute to its beneficial effects on EAE through neuroprotection. Treatment with this widely used and well-tolerated drug may be a useful therapeutic intervention in ameliorating MS disease

AAV-mediated expression of secreted and transmembrane αKlotho isoforms rescues relevant aging hallmarks in senescent SAMP8 mice

Senescence represents a stage in life associated with elevated incidence of morbidity and increased risk of mortality due to the accumulation of molecular alterations and tissue dysfunction, promoting a decrease in the organism's protective systems. Thus, aging presents molecular and biological hallmarks, which include chronic inflammation, epigenetic alterations, neuronal dysfunction, and worsening of physical status. In this context, we explored the AAV9-mediated expression of the two main isoforms of the aging-protective factor Klotho (KL) as a strategy to prevent these general age-related features using the senescence-accelerated mouse prone 8 (SAMP8) model. Both secreted and transmembrane KL isoforms improved cognitive performance, physical state parameters, and different molecular variables associated with aging. Epigenetic landscape was recovered for the analyzed global markers DNA methylation (5-mC), hydroxymethylation (5-hmC), and restoration occurred in the acetylation levels of H3 and H4. Gene expression of pro- and anti-inflammatory mediators in central nervous system such as TNF-α and IL-10, respectively, had improved levels, which were comparable to the senescence-accelerated-mouse resistant 1 (SAMR1) healthy control. Additionally, this improvement in neuroinflammation was supported by changes in the histological markers Iba1, GFAP, and SA β-gal. Furthermore, bone tissue structural variables, especially altered during senescence, recovered in SAMP8 mice to SAMR1 control values after treatment with both KL isoforms. This work presents evidence of the beneficial pleiotropic role of Klotho as an anti-aging therapy as well as new specific functions of the KL isoforms for the epigenetic regulation and aged bone structure alteration in an aging mouse model. Intraventricular administration of AAV vectors expressing secreted and transmembrane Klotho isoforms, rescued accelerated aging phenotype of SAMP8 mice. An improvement in cognitive and physical performance, recovery of epigenetic, inflammatory and senescence markers, as well as structural changes in long bones of these mice was detected

Mowat-Wilson syndrome

Mowat-Wilson syndrome (MWS) is a multiple congenital anomaly syndrome characterized by a distinct facial phenotype (high forehead, frontal bossing, large eyebrows, medially flaring and sparse in the middle part, hypertelorism, deep set but large eyes, large and uplifted ear lobes, with a central depression, saddle nose with prominent rounded nasal tip, prominent columella, open mouth, with M-shaped upper lip, frequent smiling, and a prominent but narrow and triangular pointed chin), moderate-to-severe intellectual deficiency, epilepsy and variable congenital malformations including Hirschsprung disease (HSCR), genitourinary anomalies (in particular hypospadias in males), congenital heart defects, agenesis of the corpus callosum and eye anomalies. The prevalence of MWS is currently unknown, but 171 patients have been reported so far. It seems probable that MWS is under-diagnosed, particularly in patients without HSCR. MWS is caused by heterozygous mutations or deletions in the Zinc finger E-box-binding homeobox 2 gene, ZEB2, previously called ZFHX1B (SIP1). To date, over 100 deletions/mutations have been reported in patients with a typical phenotype; they are frequently whole gene deletions or truncating mutations, suggesting that haploinsufficiency is the main pathological mechanism. Studies of genotype-phenotype analysis show that facial gestalt and delayed psychomotor development are constant clinical features, while the frequent and severe congenital malformations are variable. In a small number of patients, unusual mutations can lead to an atypical phenotype. The facial phenotype is particularly important for the initial clinical diagnosis and provides the hallmark warranting ZEB2 mutational analysis, even in the absence of HSCR. The majority of MWS cases reported so far were sporadic, therefore the recurrence risk is low. Nevertheless, rare cases of sibling recurrence have been observed. Congenital malformations and seizures require precocious clinical investigation with intervention of several specialists (including neonatologists and pediatricians). Psychomotor development is delayed in all patients, therefore rehabilitation (physical therapy, psychomotor and speech therapy) should be started as soon as possible

Diazoxide attenuates autoimmune encephalomyelitis and modulates lymphocyte proliferation and dendritic cell functionality

Activation of mitochondrial ATP-sensitive potassium (KATP) channels is postulated as an effective mechanism to confer cardio and neuroprotection, especially in situations associated to oxidative stress. Pharmacological activation of these channels inhibits glia-mediated neuroinflammation. In this way, diazoxide, an old-known mitochondrial KATP channel opener, has been proposed as an effective and safe treatment for different neurodegenerative diseases, demonstrating efficacy in different animal models, including the experimental autoimmune encephalomyelitis (EAE), an animal model for Multiple Sclerosis. Although neuroprotection and modulation of glial reactivity could alone explain the positive effects of diazoxide administration in EAE mice, little is known of its effects on the immune system and the autoimmune reaction that triggers the EAE pathology. The aim of the present work was to study the effects of diazoxide in autoimmune key processes related with EAE, such as antigen presentation and lymphocyte activation and proliferation. Results show that, although diazoxide treatment inhibited in vitro and ex-vivo lymphocyte proliferation from whole splenocytes it had no effect in isolated CD4(+) T cells. In any case, treatment had no impact in lymphocyte activation. Diazoxide can also slightly decrease CD83, CD80, CD86 and major histocompatibility complex class II expression in cultured dendritic cells, demonstrating a possible role in modulating antigen presentation. Taken together, our results indicate that diazoxide treatment attenuates autoimmune encephalomyelitis pathology without immunosuppressive effect

The L-type voltage-gated calcium channel modulates microglial pro-inflammatory activity

Under pathological conditions, microglia, the resident CNS immune cells, become reactive and release pro-inflammatory cytokines and neurotoxic factors. We investigated whether this phenotypic switch includes changes in the expression of the L-type voltage-gated calcium channel (VGCC) in a rat model of N-methyl-d-aspartate-induced hippocampal neurodegeneration. Double immunohistochemistry and confocal microscopy evidenced that activated microglia express the L-type VGCC. We then analyzed whether BV2 microglia express functional L-type VGCC, and investigated the latter's role in microglial cytokine release and phagocytic capacity. Activated BV2 microglia express the CaV1.2 and CaV1.3 subunits of the L-type VGCC determined by reverse transcription-polymerase chain reaction, Western blot and immunocytochemistry. Depolarization with KCl induced a Ca2+ entry facilitated by Bay k8644 and partially blocked with nifedipine, which also reduced TNF-α and NO release by 40%. However, no nifedipine effect on BV2 microglia viability or phagocytic capacity was observed. Our results suggest that in CNS inflammatory processes, the L-type VGCC plays a specific role in the control of microglial secretory activity

Carotid revascularisation using angioplasty and stent in 134 consecutive cases in a reference hospital: A risky technique?

Introduction: Carotid revascularisation (CR) using angioplasty and stent (ASC) is an effective procedure in the prevention of ischaemic stroke, but with a controversial morbidity and mortality in the different studies conducted in this field. Methods: The results of the ASCs performed in the Virgen de la Arrixaca University Hospital (Murcia) between January 2006 and April 2009 were analysed (epidemiology, indication, grade of residual stenosis and procedure complications). All patients subjected to ASC were pre-selected and followed up by neurologists, and they followed a strict medical protocol for performing the procedure. All ASCs were performed by a team consisting of two surgeons, an anaesthetist and a nurse. Results: A total of 134 ASC were performed. The mean age of our patients was 72.7 years, with the large majority (75%) being male. The most prevalent diseases were, high blood pressure (81%), smoking (66.4%), and diabetes (38.1%). The most common indications for CR were symptomatic carotid stenosis with a level of stenosis of 75-99%, either in the left (33.6%) or right (32.1%), followed by asymptomatic stenosis combined with risk factors (11.2% in the left side and 10.4% in the right side). A level of stenosis less than 30% was achieved in 132 of the 134 ASC (98.5%). performed. Five patients (3.7%) had complications associated with the procedure, of which four were different clinical presentations of a re-perfusion syndrome and one an asymptomatic thrombosis of the stent. Conclusions: ASC is a complex technique that must be performed by appropriately trained specialists. The performing a minimum number of procedures per year and an admission protocol controlled by Neurology are essential conditions for a low rate of complications. Under these conditions, the morbidity and mortality of the technique is no higher than that of endarterectomy. Resumen: Introducción: La revascularización carotídea mediante angioplastia y stent (ASC) es un procedimiento eficaz en la prevención del ictus isquémico, pero con una morbimortalidad periprocedimiento muy discutida en los diferentes estudios realizados al respecto. Métodos: Se analizan los resultados (epidemiología, tipo de indicación, grado de estenosis residual y complicaciones periprocedimiento) de las ASC realizadas en el Hospital Universitario Virgen de la Arrixaca entre enero de 2006 y abril de 2009. Todos los pacientes sometidos a ASC fueron preseleccionados y seguidos por neurólogos, y siguieron un protocolo médico estricto para la realización del procedimiento. Todas las ASC fueron llevadas a cabo por un equipo formado por dos intervencionistas, un anestesista y un enfermero. Resultados: Se realizaron 134 ASC. La edad media de nuestros pacientes fue de 72,7 años, con predominio del sexo masculino (75%). Las patologías más prevalentes fueron la hipertensión arterial (81%), el tabaquismo (66,4%), y la diabetes (38,1%). La indicación de RC más frecuente fue la estenosis carotídea sintomática con grado de estenosis 75-99%, tanto izquierda (33,6%) como derecha (32,1%), seguida de las estenosis asintomáticas asociadas a factores de riesgo (11,2% en el lado izquierdo y 10,4% en el lado derecho). En 132 de las 134 ASC (98,5%) se consiguió un grado de estenosis residual menor al 30%. Cinco pacientes (3,7%) presentaron complicaciones relacionadas con el procedimiento, de las cuales cuatro correspondieron a diferentes presentaciones clínicas del síndrome de reperfusión y una, a una trombosis asintomática del stent. Conclusiones: La ASC es una técnica compleja que debe ser llevada a cabo por especialistas con una capacitación adecuada. La realización de un número mínimo de procedimientos al año, y la existencia de un protocolo de ingreso controlado por Neurología son condiciones imprescindibles para una tasa de complicaciones. En estas circunstancias, la morbimortalidad de la técnica no es superior a la de la endarterectomía. Keywords: Carotid revascularisation, Angioplasty, Stent, Palabras clave: Revascularización carotídea, Angioplastia, Sten