We previously introduced the concept that intrahepatic bile duct epithelial cells, or cholangiocytes, are functionally heterogeneous. This concept is based on the observation that secretin receptor (SR) gene expression and secretin-induced cAMP synthesis are present in cholangiocytes derived from large (Ͼ15 µm in diameter) but not small (Ͻ15 µm in diameter) bile ducts. In work reported here, we tested the hypothesis that cholangiocytes are heterogeneous with regard to proliferative capacity. We assessed cholangiocyte proliferation in vivo by measurement of [ 3 H]thymidine incorporation and in vitro by both [ 3 H]thymidine incorporation and H 3 histone gene expression in small (fraction 1) and large (fraction 2) cholangiocytes isolated from rats after bile duct ligation (BDL). In the two cholangiocyte subpopulations, we also studied basal somatostatin receptor (SSTR 2) gene expression as well as the effects of somatostatin on 1) SR gene expression and secretin-induced cAMP synthesis and 2) [ 3 H]thymidine incorporation and H 3 histone gene expression. In normal rat liver, cholangiocytes, unlike hepatocytes, were mitotically dormant; after BDL, incorporation of [ 3 H]thymidine markedly increased in cholangiocytes but not hepatocytes. When subpopulations of cholangiocytes were isolated after BDL, DNA synthesis assessed by both techniques was limited to large cholangiocytes, as was SSTR 2 steady-state gene expression. In vitro, somatostatin inhibited SR gene expression and secretin-induced cAMP synthesis only in large cholangiocytes. Moreover, compared with no hormone, somatostatin inhibited DNA synthesis solely in large cholangiocytes. These results support the concept of the heterogeneity of cholangiocytes along the biliary tree, extend this concept to cholangiocyte proliferative activity, and imply that the proliferative compartment of cholangiocytes after BDL is located principally in the cholangiocytes lining large (Ͼ15 µm) bile ducts.

Secretion stimulates ductular bile secretion by binding to receptors on intrahepatic bile duct epithelial cells (i.e., cholangiocytes). In the rat, this choleretic effect increases after bile duct ligation (BDL). Although cholangiocyte proliferation induced by BDL contributes to secretin-induced hypercholeresis, the mechanisms modulating these alterations in secretin-induced ductular bile secretion are obscure. Thus we studied the expression of secretin receptor mRNA (SR-mRNA) in purified liver cells from normal and BDL rats. Northern blot analysis and RNase protection assays with mRNA from purified liver cells demonstrated SR-mRNA only in cholangiocytes; moreover, SR gene expression showed a seven- to ninefold increase in individual cholangiocytes from BDL rats compared with controls. This increase in SR-mRNA expression was related to a similar increase in the rate of transcription of SR-mRNA in cholangiocytes from BDL rats. Thus our studies indicate that 1) SR-mRNA is detected in liver only in cholangiocytes; 2) BDL causes an increase in SR-mRNA in individual cholangiocytes; and 3) the increase in SR-mRNA after BDL is partly related to an increase in the rate of transcription of SR-mRNA by cholangiocytes after BDL. Our data suggest that upregulation of the SR gene may contribute to secretin-induced hypercholeresis.

BAF (Brg/Brm-associated factors) complexes play important roles in development and are linked to chromatin plasticity at selected genomic loci. Nevertheless, a full understanding of their role in development and chromatin remodeling has been hindered by the absence of mutants completely lacking BAF complexes. Here, we report that the loss of BAF155/BAF170 in double-conditional knockout (dcKO) mice eliminates all known BAF subunits, resulting in an overall reduction in active chromatin marks (H3K9Ac), a global increase in repressive marks (H3K27me2/3), and downregulation of gene expression. We demonstrate that BAF complexes interact with H3K27 demethylases (JMJD3 and UTX) and potentiate their activity. Importantly, BAF complexes are indispensable for forebrain development, including proliferation, differentiation, and cell survival of neural progenitor cells. Our findings reveal a molecular mechanism mediated by BAF complexes that controls the global transcriptional program and chromatin state in development.

IntroductionAttenuated viruses with tumor specificity have attracted considerable interest as novel anticancer agents, and clinical testing of several such agents is under way. 1 Tumor selectivity of these viruses has been attributed to various intracellular restrictions to their life cycles that are strongly inhibitory to virus propagation in nontransformed cells but that are overridden by cellular factors present in neoplastic cells. [2][3][4][5][6] Measles is an acute viral disease caused by a negative-strand RNA virus of the family Paramyxoviridae, which remains responsible for approximately 1 million deaths each year. 7 In the early stages of measles virus (MV) infection, the lymphoid organs and tissues are predominant sites for viral replication, leading to the formation of giant reticuloendothelial (Warthin-Finkeldey) cells in the tissues. 7 Measles is prevented by the use of a live attenuated virus vaccine now routinely administered during childhood. The Edmonston-B vaccine strain of measles virus (MV-Edm) was attenuated by serial tissue culture passage of a clinical isolate. 7 Despite its profound attenuation as a human pathogen, MV-Edm replicates more efficiently than nonattenuated measles virus in many primate cell lines, inducing cell cell fusion and the formation of characteristic multinucleated syncytia. 8 Here, we compared the ability of MV-Edm to replicate in neoplastic myeloma cells and normal cells, and we investigated its potential as an antitumor agent in human myeloma xenografts in vivo. We report that the virus replicated selectively in a panel of 6 myeloma cell lines and in CD138-sorted myeloma cells from 6 patients and that it caused potent cytopathic effects. When administered intratumorally or intravenously into mice bearing established myeloma xenografts, MV-Edm caused growth inhibition or total regression of 2 different myeloma xenograft models. Materials and methods Cell cultureThe multiple myeloma ARH-77 cell line (ATCC CRL-1621) was obtained from American Type Culture Collection (Rockville, MD). The rest of the multiple myeloma cell lines were kind gifts of Dr John Lust (RPMI 8226), Dr Diane Jelinek (KAS-6/1), and Dr Rafael Fonseca (KMS-11, MM1, JJN-3) from the Mayo Clinic (Rochester, MN). All the human myeloma cell lines were maintained in RPMI-1640 (Gibco BRL, Rockville, MD) supplemented with 10% fetal bovine serum (FBS) except for KAS-6/1, which was grown in media supplemented with 1 ng/mL interleukin (IL)-6 (Sigma, St Louis, MO). Bone marrow aspirates were obtained after institutional review board approval and informed patient consent. Bone marrow samples were drawn into a tube containing heparin and centrifuged on a Ficoll-Hypaque gradient (Amersham Pharmacia, Piscataway, NJ) to enrich for mononuclear cells. Mononuclear cells were then incubated with MACS CD138 microbeads (Miltenyi Biotech, Auburn, CA) for 15 minutes in a 8°C water bath. CD138 ϩ primary myeloma cells were then isolated, washed, and maintained in 10% FBS-RPMI supplemented with 1 ng/mL IL-6. Peripheral blood lym...

Previous work from our laboratory has implicated hormone-induced plasma membrane movement (i.e., endo- and exocytosis) in water and electrolyte transport by the epithelial cells that line the ducts in the liver (i.e., cholangiocytes). To further explore the cellular mechanisms regulating ductal bile secretion, we infused somatostatin and/or secretin intravenously into rats 2 wk after either bile duct ligation (BDL), a procedure that induces selective proliferation of cholangiocytes, or sham surgery and measured bile flow and biliary constituents. We also determined the effect of somatostatin on basal and secretin-induced exocytosis by purified cholangiocytes isolated from rat liver after BDL. Finally, we studied the expression of the somatostatin receptor gene by both ribonuclease (RNase) protection and nuclear run-on assays using cDNA encoding for two subtypes of the somatostatin receptor gene (i.e., SSTR1 and SSTR2). In vivo, somatostatin infusion caused a dose-dependent bicarbonate-poor decrease (57% maximal decrease below baseline; P < 0.05) in bile flow in BDL but not in sham-operated rats; in contrast, secretin caused a dose-dependent bicarbonate-rich choleresis (228% maximal increase above baseline; P < 0.05) in BDL but not in sham-operated rats. Simultaneous or prior infusion of somatostatin inhibited the secretin-induced hypercholeresis in BDL rats. In vitro, somatostatin had no effect on basal exocytosis by cholangiocytes isolated from BDL rats; however, somatostatin inhitibed (88% maximal inhibition; P < 0.05) secretin-induced exocytosis by cholangiocytes in a dose-dependent fashion. In addition, somatostatin inhibited secretin-induced increases in levels of adenosine 3',5'-cyclic monophosphate (cAMP) in cholangiocytes isolated from BDL rats.(ABSTRACT TRUNCATED AT 250 WORDS)