Key points• The mechanisms of anion and fluid transport by airway submucosal glands are not well understood and may differ from those in surface epithelium.• The Calu-3 cell line is often used as a model for submucosal gland serous cells and has cAMP-stimulated fluid secretion; however, it does not actively transport chloride under short-circuit conditions.• In this study we show that fluid secretion requires chloride, bicarbonate and sodium, that chloride is the predominant anion in Calu-3 secretions, and that a large fraction of the basolateral chloride loading during cAMP stimulation occurs by Cl − /HCO 3 − exchange.• The results suggest a novel cellular model for anion and fluid secretion by Calu-3 and submucosal gland acinar cells Abstract Anion and fluid secretion are both defective in cystic fibrosis (CF); however, the transport mechanisms are not well understood. In this study, Cl − and HCO 3 − secretion was measured using genetically matched CF transmembrane conductance regulator (CFTR)-deficient and CFTR-expressing cell lines derived from the human airway epithelial cell line Calu-3. Forskolin stimulated the short-circuit current (I sc ) across voltage-clamped monolayers, and also increased the equivalent short-circuit current (I eq ) calculated under open-circuit conditions. I sc was equivalent to the HCO 3 − net flux measured using the pH-stat technique, whereas I eq was the sum of the Cl − and HCO 3 − net fluxes. I eq and HCO 3 − fluxes were increased by bafilomycin and ZnCl 2 , suggesting that some secreted HCO 3 − is neutralized by parallel electrogenic H + secretion. I eq and fluid secretion were dependent on the presence of both Na + and HCO 3 − . The carbonic anhydrase inhibitor acetazolamide abolished forskolin stimulation of I eq and HCO 3 − secretion, suggesting that HCO 3 − transport under these conditions requires catalysed synthesis of carbonic acid. Cl − was the predominant anion in secretions under all conditions studied and thus drives most of the fluid transport. Nevertheless, 50-70% of Cl − and fluid transport was bumetanide-insensitive, suggesting basolateral Cl − loading by a sodium-potassium-chloride cotransporter 1 (NKCC1)-independent mechanism. Imposing a transepithelial HCO 3 − gradient across basolaterally permeabilized Calu-3 cells sustained a forskolin-stimulated current, which was sensitive to CFTR inhibitors and drastically reduced in CFTR-deficient cells. Net HCO 3 with Cl − , and the resulting HCO 3 − -dependent Cl − transport provides an osmotic driving force for fluid secretion.

Although evidence shows that victims of sudden infant death syndrome (SIDS) suffer repetitive episodes of hypoxemia, only subtle abnormalities have been found in their brains by light microscopy. The aim of the present study was to determine whether apoptosis, a form of cell death that can be triggered by hypoxemia and that leaves no scarring detectable by light microscopy, would be present in hypoxia-sensitive brain regions of SIDS victims. We looked for the presence of apoptosis with an in situ end-labeling method that detects DNA fragmentation. We studied 29 SIDS victims who were age-matched to nine control cases. We found significant neuronal apoptosis in 79% of the SIDS cases: 55% of the cases positive in the hippocampus and 96% positive in the brainstem. Whereas the distribution of apoptosis in the hippocampus was in hypoxia-sensitive subregions, the distribution in the brainstem was mostly in dorsal nuclei, including those involved with sensation in the face and position of the head (nucleus of the spinal trigeminal tract and vestibular nuclei). The control cases showed no significant apoptosis in the hippocampus and a mild degree in the brainstem in three cases. Our results indicate the occurrence of an acute insult at least several hours before death, an insult from which the infants had apparently recuperated. This suggests that SIDS victims suffered repeated apoptosis resulting in significant neuronal damage and, thus, functional loss in key brain regions. The involvement of specific nuclei in the brainstem may be linked to the fact that prone sleeping is a significant risk factor for SIDS. Enhanced neuronal death by apoptosis may thus have major implications for understanding the sequence of events leading to SIDS.

PRMT5 is a type II protein arginine methyltranferase that catalyzes monomethylation and symmetric dimethylation of arginine residues. PRMT5 is functionally involved in a variety of biological processes including embryo development and circadian clock regulation. However, the role of PRMT5 in oligodendrocyte differentiation and central nervous system myelination is unknown. Here we show that PRMT5 expression gradually increases throughout postnatal brain development, coinciding with the period of active myelination. PRMT5 expression was observed in neurons, astrocytes, and oligodendrocytes. siRNA-mediated depletion of PRMT5 in mouse primary oligodendrocyte progenitor cells abrogated oligodendrocyte differentiation. In addition, the PRMT5-depleted oligodendrocyte progenitor and C6 glioma cells expressed high levels of the inhibitors of differentiation/DNA binding, Id2 and Id4, known repressors of glial cell differentiation. We observed that CpG-rich islands within the Id2 and Id4 genes were bound by PRMT5 and were hypomethylated in PRMT5-deficient cells, suggesting that PRMT5 plays a role in gene silencing during glial cell differentiation. Our findings define a role of PRMT5 in glial cell differentiation and link PRMT5 to epigenetic changes during oligodendrocyte differentiation. Protein arginine methyltransferases (PRMTs)3 transfer methyl groups from S-adenosylmethionine to the nitrogen atoms of arginine, a positively charged amino acid that mediates hydrogen bonding and amino aromatic interactions (1). Arginine can be monomethylated or dimethylated (DMA). If two methyl groups are both placed on one of the terminal nitrogen atoms of the guanidino group, the derivative is an asymmetric DMA. Symmetric DMA is where one methyl group is located on each of the two nitrogens. There are currently nine mammalian PRMTs that are grouped as type I and type II enzymes (1, 2). Type I enzymes form monomethylated arginine and asymmetric DMA and include PRMT1, PRMT3, CARM1, PRMT6, and PRMT8. Type II enzymes, including PRMT5, PRMT7, and PRMT9, catalyze the production of monomethylated arginine and symmetric DMA. The enzyme activity of PRMT2 is not yet identified.Symmetrical arginine dimethylation occurs less frequently than asymmetrical dimethylation, and among the type II enzymes, PRMT5 has been the most characterized. It is functionally involved in a broad spectrum of biological processes, but most importantly in early embryo development and later cellular differentiation as well as the circadian clock (3-6). Accumulating evidence reveals that PRMT5 recurrently exerts its biological functions through transcriptional repression of a large group of genes involved in these processes. PRMT5 associates with the cyclin E1 promoter and represses cyclin E1 transcription, leading to inhibition of cell proliferation (7). PRMT5 associates with BRG1-based SWItch/Sucrose NonFermentable (hSWI/SNF) chromatin remodeling complexes and promotes transcriptional repression of the Myc target gene cad (8). BRG1-and hBRM-associated PRMT5 also repress...