Cholangiocyte anion exchange and biliary bicarbonate excretion

Primary canalicular bile undergoes a process of fluidization and alkalinization along the biliary tract that is influenced by several factors including hormones, innervation/neuropeptides, and biliary constituents. The excretion of bicarbonate at both the canaliculi and the bile ducts is an important contributor to the generation of the so-called bile-salt independent flow. Bicarbonate is secreted from hepatocytes and cholangiocytes through parallel mechanisms which involve chloride efflux through activation of Cl- channels, and further bicarbonate secretion via AE2/SLC4A2-mediated Cl-/HCO3- exchange. Glucagon and secretin are two relevant hormones which seem to act very similarly in their target cells (hepatocytes for the former and cholangiocytes for the latter). These hormones interact with their specific G protein-coupled receptors, causing increases in intracellular levels of cAMP and activation of cAMP-dependent Cl- and HCO3- secretory mechanisms. Both hepatocytes and cholangiocytes appear to have cAMP-responsive intracellular vesicles in which AE2/SLC4A2 colocalizes with cell specific Cl- channels (CFTR in cholangiocytes and not yet determined in hepatocytes) and aquaporins (AQP8 in hepatocytes and AQP1 in cholangiocytes). cAMP-induced coordinated trafficking of these vesicles to either canalicular or cholangiocyte lumenal membranes and further exocytosis results in increased osmotic forces and passive movement of water with net bicarbonate-rich hydrocholeresis

In situ detection of AE2 anion-exchanger mRNA in the human liver

Na+-independent anion exchangers, a family of membrane proteins that mediate electroneutral exchanges of chloride and bicarbonate ions across the cell membrane, are considered to be involved in intracellular pH regulation as well as in transepithelial acid/base transport. Previous immunohistochemical data have shown that anion-exchanger-2 (AE2) protein is expressed in the liver parenchyma, localizing at both the canaliculi and the luminal surfaces of intrahepatic bile ducts, where it may have a role in the biliary secretion of bicarbonate. In the present study, we have carried out in situ hybridization experiments on biopsies of human liver using three overlapping antisense anion-exchanger-2 riboprobes. Anion-exchanger-2 mRNA signals were localized mainly in the cytoplasm of terminal and interlobular bile-duct cells, whereas weaker signals were observed in bile-duct cells of larger intrahepatic ducts. Furthermore, some hepatocytes, mostly periportal, contained detectable anion-exchanger-2 mRNA signals in their cytoplasm. No hybridization signals were observed in controls with sense riboprobes, with omission of the antisense probe, or with treatment of the sections with RNase before hybridizations. Finally, intense anion-exchanger-2 hybridization signals were observed in lymphomononuclear cells in sinusoids and in portal infiltrates. Immunocytochemical data from reverse-phase sections suggest that these cells correspond to some of the CD45R+ (UCHL1+) T lymphocytes resident in the liver

A deep neural network to assess spontaneous pain from mouse facial expressions

Grimace scales quantify characteristic facial expressions associated with spontaneous pain in rodents and other mammals. However, these scales have not been widely adopted largely because of the time and effort required for highly trained humans to manually score the images. Convoluted neural networks were recently developed that distinguish individual humans and objects in images. Here, we trained one of these networks, the InceptionV3 convolutional neural net, with a large set of human-scored mouse images. Output consists of a binary pain/no-pain assessment and a confidence score. Our automated Mouse Grimace Scale integrates these two outputs and is highly accurate (94%) at assessing the presence of pain in mice across different experimental assays. In addition, we used a novel set of “pain” and “no pain” images to show that automated Mouse Grimace Scale scores are highly correlated with human scores (Pearson’s r = 0.75). Moreover, the automated Mouse Grimace Scale classified a greater proportion of images as “pain” following laparotomy surgery when compared to animals receiving a sham surgery or a post-surgical analgesic. Together, these findings suggest that the automated Mouse Grimace Scale can eliminate the need for tedious human scoring of images and provide an objective and rapid way to quantify spontaneous pain and pain relief in mice

Tratamiento quirúrgico con placa bloqueada en fracturas de fémur distal tipo C en pacientes mayores de 60 años

Objetivo. Determinar el resultado clínico, radiológico y complicaciones asociadas del tratamiento con placa bloqueada en fracturas distales de fémur tipo C en pacientes mayores de 60 años. Material y métodos. Estudio descriptivo retrospectivo de 16 pacientes. Se registraron variables epidemiológicas, comorbilidades, complicaciones y resultados clínicos y radiológicos. Resultados. Edad media de 75,3 años. El seguimiento medio fue de 13,5 meses. La flexión media postoperatoria de la rodilla fue de 61,6º. Sólo un 12,5% de los pacientes no referían gonalgia ni precisaban ayudas para la deambulación al final del seguimiento. Radiológicamente, en un 68,7% la translación femoral era mayor de 5 mm, y en un 31,3% la reducción en el plano lateral no era correcta. Se consiguió la consolidación de la fractura en 13 pacientes (86,6%), con un tiempo medio de 10,2 semanas. Como complicaciones hubo una pseudoartrosis, una consolidación viciosa y una infección. Discusión. El tratamiento de la fractura distal de fémur con placa bloqueada viene avalado en la literatura como un método fiable. Sin embargo, en nuestra serie, hemos tenido un alto porcentaje de malos resultados clínicos con presencia de dolor en la rodilla y necesidad de ayudas para la deambulación.Objective. To evaluate functional and radiographic results, and complications, after surgical treatment with locked plate in fractures of the distal femur in patients 60 years old or older. Material and methods. We reviewed 16 patients. We recorded epidemiological variables, comorbidities, functional and radiographic results. Results. Mean aged of 75,3 years. Mean follow-up of 13,5 months. Mean knee flexion of 61,6º. Only 12,5% of the patients did not refer knee pain and walked unaided. Radiologically, in 68,7% of the patients femoral translation was greater than 5 mm, and in 31,3% lateral reduction was not correct. Fracture healed in 13 patients (86,6%), in a mean time of 10,2 weeks. There were 1 nonunion, 1 malunion, and 1 infection. Discussion. Locked plating is a realiable treatment for distal femoral fractures. But, in our elderly serie, we experienced bad clinical results with high percentage of knee pain and walking aid

Taurocholate-stimulated leukotriene C4 biosynthesis and leukotriene C4-stimulated choleresis in isolated rat liver

BACKGROUND/AIMS: Cysteinyl-containing leukotrienes seem to exert a cholestatic effect. However, leukotriene inhibitors were found to reduce bile salt efflux in isolated rat hepatocytes, suggesting a role for leukotrienes in bile flow formation. METHODS: In the isolated rat liver, the effects of two different concentrations of leukotriene C4 on bile flow and bile salt excretion are analyzed, as well as the possible effect of taurocholate on the hepatic production of cysteinyl-containing leukotrienes. RESULTS: Leukotriene C4 (0.25 fmol) increased bile salt excretion (+22.2%; P < 0.05), whereas a much higher dose (0.25 x 10(6) fmol) showed the known cholestatic effect, reducing bile salt excretion (-25.9%; P < 0.01). These dose-dependent biphasic effects were specific because they could be prevented by the simultaneous administration of cysteinyl-containing leukotriene antagonists. On the other hand, taurocholate administration induced a dose-dependent increase in biliary excretion of cysteinyl-containing leukotrienes. Furthermore, taurocholate increased messenger RNA levels of 5-lipoxygenase, a key enzyme in leukotriene biosynthesis. Taurocholate increase of hepatocyte intracellular calcium was not significant, suggesting that taurocholate effects are not mediated by stimulation of calcium metabolism. CONCLUSIONS: These results constitute evidence for the existence of a positive feedback mechanism by which bile salts stimulate the synthesis of leukotrienes that, in turn, stimulate bile salt excretion

Discovery and Observations of ASASSN-13db, an EX Lupi-Type Accretion Event on a Low-Mass T Tauri Star

We discuss ASASSN-13db, an EX Lupi-type ("EXor") accretion event on the young stellar object (YSO) SDSS J051011.01$-$032826.2 (hereafter SDSSJ0510) discovered by the All-Sky Automated Survey for SuperNovae (ASAS-SN). Using archival photometric data of SDSSJ0510 we construct a pre-outburst spectral energy distribution (SED) and find that it is consistent with a low-mass class II YSO near the Orion star forming region ($d \sim 420$ pc). We present follow-up photometric and spectroscopic observations of the source after the $\Delta V \sim-$5.4 magnitude outburst that began in September 2013 and ended in early 2014. These data indicate an increase in temperature and luminosity consistent with an accretion rate of $\sim10^{-7}$ $\rm{M}_\odot$ yr$^{-1}$, three or more orders of magnitude greater than in quiescence. Spectroscopic observations show a forest of narrow emission lines dominated by neutral metallic lines from Fe I and some low-ionization lines. The properties of ASASSN-13db are similar to those of the EXor prototype EX Lupi during its strongest observed outburst in late 2008.Comment: 14 pages, 4 figures, 1 table. Updated May 2014 to reflect changes in the final version published in ApJL. Photometric data presented in this submission are included as ancillary files. For a brief video explaining this paper, see http://youtu.be/yRCCrNJnvt

Promoter hypermethylation of the AE2/SLC4A2 gene in PBC

Primary biliary cholangitis (PBC) is a chronic immuneassociated cholestatic liver disease with unclear complex/multifactorial etiopathogenesis affecting mostly middle-aged women. Patients with PBC exhibit reduced expression of the AE2/SLC4A2 gene. Herein, we found that AE2 promoter regions are hypermethylated in the liver and peripheral blood mononuclear cells of patients with PBC. This increased methylation is associated with downregulated AE2-gene expression, which might contribute to the pathogenesis of PBC. Therefore, novel epigenetic targets may improve treatment in patients with PBC who respond poorly to current pharmacological therapies

Shared apical sorting of anion exchanger isoforms AE2a, AE2b1, and AE2b2 in primary hepatocytes

AE2 (SLC4A2) is the member of the Na(+)-independent anion exchanger (AE) family putatively involved in the secretion of bicarbonate to bile. In humans, three variants of AE2 mRNA have been described: the full-length transcript AE2a (expressed from the upstream promoter in most tissues), and alternative transcripts AE2b(1) and AE2b(2) (driven from alternate promoter sequences in a tissue-restricted manner, mainly in liver and kidney). These transcripts would result in AE protein isoforms with short N-terminal differences. To ascertain their translation, functionality, and membrane sorting, we constructed expression vectors encoding each AE2 isoform fused to GFP at the C-terminus. Transfected HEK293 cells showed expression of functional GFP-tagged AE2 proteins, all three isoforms displaying comparable AE activities. Primary rat hepatocytes transfected with expression vectors and repolarized in a collagen-sandwich configuration showed a microtubule-dependent apical sorting of each AE2 isoform. This shared apical sorting is liver-cell specific, as sorting of AE2 isoforms was basolateral in control experiments on polarized kidney MDCK cells. Hepatocytic apical targeting of AE2 isoforms suggests that they all may participate in the canalicular secretion of bicarbonate to bile

Self-organized single crystal mixed magnetite/cobalt ferrite films grown by infrared pulsed-laser deposition

We have grown mixed magnetite/cobalt ferrite epitaxial films on SrTiO3 by infrared pulsed-laser deposition. Diffraction experiments indicate epitaxial growth with a relaxed lattice spacing. The films are flat with two distinct island types: nanometric rectangular mounds in two perpendicular orientations, and larger square islands, attributed to the two main components of the film as determined by Mössbauer spectroscopy. The origin of the segregation is suggested to be the oxygen-deficiency during growth.Peer Reviewe