Flavonoids have an ability to suppress various ion channels. We determined whether one of flavonoids, cyanidin-3-glucoside, affects adenosine 5'-triphosphate (ATP)-induced calcium signaling using digital imaging methods for intracellular free Ca2+ concentration ([Ca2+]i), reactive oxygen species (ROS) and mitochondrial membrane potential in PC12 cells. Treatment with ATP (100µM) for 90 sec induced [Ca2+]i increases in PC12 cells. Pretreatment with cyanidin-3-glucoside (1µ g/ml to 100µg/ml) for 30 min inhibited the ATP-induced [Ca2+]i increases in a concentration-dependent manner (IC50=15.3µg/ml). Pretreatment with cyanidin-3-glucoside (15µg/ml) for 30 min significantly inhibited the ATP-induced [Ca2+]i responses following removal of extracellular Ca2+ or depletion of intracellular [Ca2+]i stores. Cyanidin-3-glucoside also significantly inhibited the relatively specific P2X2 receptor agonist 2-MeSATP-induced [Ca2+]i responses. Cyanidin-3-glucoside significantly inhibited the thapsigargin or ATP-induced store-operated calcium entry. Cyanidin-3-glucoside significantly inhibited the ATP-induced [Ca2+]i responses in the presence of nimodipine and ω-conotoxin. Cyanidin-3-glucoside also significantly inhibited KCl (50 mM)-induced [Ca2+]i increases. Cyanidin-3-glucoside significantly inhibited ATP-induced mitochondrial depolarization. The intracellular Ca2+ chelator BAPTA-AM or the mitochondrial Ca2+ uniporter inhibitor RU360 blocked the ATP-induced mitochondrial depolarization in the presence of cyanidin-3-glucoside. Cyanidin-3-glucoside blocked ATP-induced formation of ROS. BAPTA-AM further decreased the formation of ROS in the presence of cyanidin-3-glucoside. All these results suggest that cyanidin-3-glucoside inhibits ATP-induced calcium signaling in PC12 cells by inhibiting multiple pathways which are the influx of extracellular Ca2+ through the nimodipine and ω-conotoxin-sensitive and -insensitive pathways and the release of Ca2+ from intracellular stores. In addition, cyanidin-3-glucoside inhibits ATP-induced formation of ROS by inhibiting Ca2+-induced mitochondrial depolarization.
Background: Crouzon syndrome (CS), which results from fibroblast growth factor receptor 2 mutations, is associated with craniosynostosis, exophthalmos, and other symptoms. Herein, we report the genetic abnormalities detected in a Chinese family with autosomal dominant CS, combined with luxation of the eyeball. This luxation was a consequence of the trauma to the shallow orbits. Case presentation: The proband was a 4-year-old boy. He accidentally fell, following which luxation of the bulbus oculi occurred immediately. Computed tomography and magnetic resonance imaging clearly revealed ocular proptosis. Upon physical examination, the proband, his father, and grandfather had ocular proptosis, shallow orbits, and mid-face hypoplasia. However, their hands and feet were clinically normal. Genomic DNA was extracted from the peripheral blood through a polymerase chain reaction performed for the target sequence. Genetic assessments revealed a heterozygous missense mutation (c.1012G > C, p.G338R) in exon 10 of the human FGFR2, cosegregated with the disease phenotype in this family. These findings confirmed the diagnosis of CS. Discussion: CS is usually caused by FGFR2 mutations. While there are a few reports of luxation of the bulbus oculi in Chinese families with CS, the ocular proptosis, shallow orbits, combined with luxation of eyeball after trauma observed in this patient were particularly interesting. Our findings enhance the current knowledge of traumatic luxation concomitant with CS.
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