Papers by Dieter Gruenert
Journal of Biological Chemistry, 2007
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American Journal of Respiratory Cell and Molecular Biology, Mar 1, 2002
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Proceedings of The National Academy of Sciences, 1992
The gene responsible for the lethal disorder cystic fibrosis encodes a 1480-amino acid glycoprote... more The gene responsible for the lethal disorder cystic fibrosis encodes a 1480-amino acid glycoprotein, CFTR. Using polyclonal antibodies directed against separate phosphorylation sites in the pre-nucleotide-binding fold (exon 9) and the R domain (exon 13), we have identified a 165-kDa protein in Xenopus laevis oocytes injected with recombinant CFTR cRNA transcribed from the full-length CFTR plasmid pBQ4.7. A protein of
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Proceedings of the National Academy of Sciences of the United States of America, Jun 15, 1991
Cystic fibrosis (CF) involves a profound reduction of Cl^- permeability in several exocrine tissu... more Cystic fibrosis (CF) involves a profound reduction of Cl^- permeability in several exocrine tissues. A distinctive, outwardly rectifying, depolarization-induced Cl^- channel (ORDIC channel) has been proposed to account for the Cl^- conductance that is defective in CF. The recently identified CF gene is predicted to code for a 1480-amino acid integral membrane protein termed the CF transmembrane conductance regulator (CFTR). The CFTR shares sequence similarity with a superfamily of ATP-binding membrane transport proteins such as P-glycoprotein and STE6, but it also has features consistent with an ion channel function. It has been proposed that the CFTR might be an ORDIC channel. To determine if CFTR and ORDIC channel expression are correlated, we surveyed various cell lines for natural variation in CFTR and ORDIC channel expression. In four human epithelial cell lines (T84, CaCo2, PANC-1, and 9HTEo-/S) that encompass the full observed range of CFTR mRNA levels and ORDIC channel density we found no correlation.
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Molecular therapy. Nucleic acids, 2016
Cystic fibrosis (CF) is a recessive inherited disease associated with multiorgan damage that comp... more Cystic fibrosis (CF) is a recessive inherited disease associated with multiorgan damage that compromises epithelial and inflammatory cell function. Induced pluripotent stem cells (iPSCs) have significantly advanced the potential of developing a personalized cell-based therapy for diseases like CF by generating patient-specific stem cells that can be differentiated into cells that repair tissues damaged by disease pathology. The F508del mutation in airway epithelial cell-derived CF-iPSCs was corrected with small/short DNA fragments (SDFs) and sequence-specific TALENs. An allele-specific PCR, cyclic enrichment strategy gave ~100-fold enrichment of the corrected CF-iPSCs after six enrichment cycles that facilitated isolation of corrected clones. The seamless SDF-based gene modification strategy used to correct the CF-iPSCs resulted in pluripotent cells that, when differentiated into endoderm/airway-like epithelial cells showed wild-type (wt) airway epithelial cell cAMP-dependent Cl ion...
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Biotechniques, 1987
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Cancer Research, Nov 1, 1985
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American Journal of Respiratory Cell and Molecular Biology, Dec 20, 2012
Airway epithelial cells contribute to the inflammatory response of the lung, and their innate imm... more Airway epithelial cells contribute to the inflammatory response of the lung, and their innate immune response is primarily mediated via Toll-like receptor (TLR) signaling. Cystic fibrosis (CF) airways are chronically infected with Pseudomonas aeruginosa, suggesting a modified immune response in CF. We investigated the TLR-4 expression and the inflammatory profile (IL-8 and IL-6 secretion) in CF bronchial epithelial cell line CFBE41o- and its CF transmembrane ion condcutance regulator (CFTR)-corrected counterpart grown under air-liquid interface conditions after stimulation with lipopolysaccharide (LPS) from gram-negative bacteria. In CFTR-corrected cells, IL-8 and IL-6 secretions were constitutively activated but significantly increased after LPS stimulation compared with CFBE41o-. Blocking TLR-4 by a specific antibody significantly inhibited IL-8 secretion only in CFTR-corrected cells. Transfection with specific siRNA directed against TLR-4 mRNA significantly reduced the response to LPS in both cell lines. Fluorescence-activated cell sorter analysis revealed significantly higher levels of TLR-4 surface expression in CFTR-corrected cells. In histologic lung sections of patients with CF, the TLR-4 expression in the bronchial epithelium was significantly reduced compared with healthy control subjects. In CF the loss of CFTR function appears to decrease innate immune responses, possibly by altering the expression of TLR-4 on airway epithelial cells. This may contribute to chronic bacterial infection of CF airways.
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BioTechniques
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ABSTRACT
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The American journal of physiology
Inherited phenotypic changes in cultured cells, as observed during differentiation and transforma... more Inherited phenotypic changes in cultured cells, as observed during differentiation and transformation, reflect alterations in gene expression and have both a genetic and epigenetic basis. The causes of specific changes are often difficult to define especially when observing phenomenological end points. Although such observations are an important step in defining the phenotypic changes that endure for multiple generations, it is necessary to analyze cells at the molecular level to characterize the pathways leading to changes in phenotype. Gene expression can be regulated at multiple levels, i.e., DNA structure, gene transcription, and/or posttranscriptional modifications. Four genetic mechanisms (DNA point mutations, deletion, rearrangement, and amplification) and two epigenetic mechanisms (DNA methylation and the preservation of DNA-protein complexes) can account for the majority of enduring changes observed in cultured cells. Genetic alterations in DNA sequence appear to be largely responsible for altered growth regulation associated with transformation, but there is also evidence to suggest that epigenetic mechanisms play a role in transformation. Differentiation of cultured cells is often associated with lack of growth, and has been ascribed in part to epigenetic mechanisms. However, differentiation and transformation are not mutually exclusive but may be regulated by parallel multistep pathways. Analysis of the causes underlying transformation has been complicated by the use of aneuploid cells, but it is clear that the tools for overcoming the ambiguities associated with phenomenological analysis are available.
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Advances in Experimental Medicine and Biology
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The American journal of physiology
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The American journal of physiology
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The American journal of physiology
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The American journal of physiology
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Papers by Dieter Gruenert