Abstract-Although the biophysical fingerprints (ion selectivity, voltage-dependence, kinetics, etc) of Ca 2ϩ -activated Cl Ϫ currents are well established, their molecular identity is still controversial. Several molecular candidates have been suggested; however, none of them has been fully accepted. We have recently characterized a cGMP-dependent Ca 2ϩ -activated Cl Ϫ current with unique characteristics in smooth muscle cells. This novel current has been shown to coexist with a "classic" (cGMP-independent) Ca 2ϩ -activated Cl Ϫ current and to have characteristics distinct from those previously known for Ca 2ϩ -activated Cl Ϫ currents. Here, we suggest that a bestrophin, a product of the Best gene family, is responsible for the cGMP-dependent Ca 2ϩ -activated Cl Ϫ current based on similarities between the membrane currents produced by heterologous expressions of bestrophins and the cGMP-dependent Ca 2ϩ -activated Cl Ϫ current. This is supported by similarities in the distribution pattern of the cGMP-dependent Ca 2ϩ -activated Cl Ϫ current and bestrophin-3 (the product of Best-3 gene) expression in different smooth muscle. Furthermore, downregulation of Best-3 gene expression with small interfering RNA both in cultured cells and in vascular smooth muscle cells in vivo was associated with a significant reduction of the cGMP-dependent Ca 2ϩ -activated Cl Ϫ current, whereas the magnitude of the classic Ca 2ϩ -activated Cl Ϫ current was not affected. Ϫ channel, which results in depolarization in vascular smooth muscle. Furthermore, the current is of similar magnitude as "classic" Ca 2ϩ -activated Cl Ϫ currents in most vascular beds and even larger in some vascular smooth muscles. 3 It is, therefore, highly desirable to know the molecular structure of the channel responsible for this current because it is likely to play an important role in smooth muscle function.Although their biophysical fingerprints (ion selectivity, voltage-dependence, kinetics, etc) are well established, 4 -6 the molecular identity of Ca 2ϩ -sensitive Cl Ϫ channels is still controversial. 7 Recently, the gene responsible for vitelliform macular dystrophy 8 and its homologs that code for bestrophin proteins have been suggested as candidates. 9,10 Four bestrophin family members in the mammalian genome and many homologues in genomes of invertebrates and even prokaryotes have been identified. 11-13 Two different nomenclatures for mammalian bestrophins were previously devel- The majority of suggestions that bestrophins function as Cl Ϫ channels are based on the findings that expression of the gene in different cell types leads to the appearance of a Cl Ϫ conductance 9,10 and that mutations or chemical modifications of the predicted channel pore change this conductance. [15][16][17][18] Although downregulation by small interfering (si)RNA in recent studies demonstrated a direct association between the endogenous Cl Ϫ current in epithelial cell culture and Best-1 expression, 19 -21 the exact role of the bestrophins in native tissues remains questionable...
Best3 is localized intracellularly in cell bodies and primary processes of mouse podocytes and is co-localized with nestin. Two splice variants of Best3 are expressed in glomeruli and in cultured podocytes, and their expression is differentially regulated in ER stress.
Background: Preterm infants exposed to chorioamnionitis and with a fetal inflammatory response are at risk for neonatal morbidity and adverse outcome. Alarmins S100A8, S100A9, and S100A12 are expressed by myeloid cells and have been associated with inflammatory activation and monocyte modulation.Aim: To study S100A alarmin expression in cord blood monocytes from term healthy and preterm infants and relate results to clinical findings, inflammatory biomarkers and alarmin protein levels, as well as pathways identified by differentially regulated monocyte genes.Methods: Cord blood CD14+ monocytes were isolated from healthy term (n = 10) and preterm infants (<30 weeks gestational age, n = 33) by MACS technology. Monocyte RNA was sequenced and gene expression was analyzed by Principal Component Analysis and hierarchical clustering. Pathways were identified by Ingenuity Pathway Analysis. Inflammatory proteins were measured by Multiplex ELISA, and plasma S100A proteins by mass spectrometry. Histological chorioamnionitis (HCA) and fetal inflammatory response syndrome (FIRS) were diagnosed by placenta histological examination.Results: S100A8, S100A9, and S100A12 gene expression was significantly increased and with a wider range in preterm vs. term infants. High S100A8 and S100A9 gene expression (n = 17) within the preterm group was strongly associated with spontaneous onset of delivery, HCA, FIRS and elevated inflammatory proteins in cord blood, while low expression (n = 16) was associated with impaired fetal growth and physician-initiated Golubinskaya et al.Preterm Infants S100A Alarmin Expression delivery. S100A8 and S100A9 protein levels were significantly lower in preterm vs. term infants, but within the preterm group high S100A gene expression, spontaneous onset of labor, HCA and FIRS were associated with elevated protein levels. One thousand nine hundred genes were differentially expressed in preterm infants with high vs. low S100A alarmin expression. Analysis of 124 genes differentially expressed in S100A high as well as FIRS and HCA groups identified 18 common pathways and S100A alarmins represented major hubs in network analyses.
Conclusion:High expression of S100A alarmins in cord blood monocytes identifies a distinct clinical risk group of preterm infants exposed to chorioamnionitis and with a fetal inflammatory response. Gene and pathway analyses suggest that high S100A alarmin expression also affects monocyte function. The connection with monocyte phenotype and inflammation-stimulated S100A expression in other cell types (e.g., neutrophils) warrants further investigation.
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