scholarly journals In Vivo and In Vitro Analysis of Age-Associated Changes and Somatic Cellular Senescence in Renal Epithelial Cells

PLoS ONE ◽  
2014 ◽  
Vol 9 (2) ◽  
pp. e88071 ◽  
Author(s):  
Birgit Berkenkamp ◽  
Nathan Susnik ◽  
Arpita Baisantry ◽  
Inna Kuznetsova ◽  
Christoph Jacobi ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cellular Senescence ◽  
Renal Epithelial Cells ◽  
Vitro Analysis ◽  
In Vitro Analysis

scholarly journals FHIT suppresses inflammatory carcinogenic activity by inducing apoptosis in esophageal epithelial cells

2010 ◽  
Vol 1 (1) ◽  
pp. 7
Author(s):  
Koshi Mimori ◽  
Takehiko Yokobori ◽  
Masaaki Iwatsuki ◽  
Tomoya Sudo ◽  
Fumiaki Tanaka ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cyclooxygenase 2 ◽  
Clinical Samples ◽  
Prostaglandin E ◽  
In Vivo Analysis ◽  
Esophageal Epithelial Cells ◽  
Vitro Analysis ◽  
In Vitro Analysis

We focused on the mechanism by which FHIT suppresses neoplastic transformation in normal but damaged esophageal epithelial cells exposed to inflammatory stimuli in vivo and to chemo-radiotherapy in clinical samples. For in vitro analysis, Adenoviral-FHIT (Ad-FHIT) in TE4 and TE2 were used for microarray analysis. For in vivo analysis, wild-type (WT) FHIT and FHIT-deficient (KO) C57BL/6 mice were exposed to N-nitrosomethylbenzylamine (NMBA) and to a cyclooxygenase-2 inhibitor (COXI). Considering DNA damage on clinical samples, expressions of FHIT, BAX and PCNA were evaluated by comparing between 3 cases of esophageal cancer cases of the chemo-radiotherapy responder and 7 cases of the non-responder. In in vitro analysis, we listed the down-regulated genes in Ad-FHIT that significantly control Lac-Z infected cells, such as prostaglandin E receptor 4, cyclooxygenase-1 and cyclooxygenase-2. In in vivo analysis, FHIT-KO mice were much more susceptible to tumorigenesis than were FHIT-WT mice. A significant difference in PGE2 activation was observed between FHIT-WT mice (5.2 ng/mL) and FHIT-KO mice (28.4 ng/mL) after exposure to NMBA in the absence of COXI as determined by ELISA assay (P less than 0.01). BAX expression was significantly higher in FHIT-WT (1.0±0.43) than in FHIT-KO (0.17±0.17) (P less than 0.05). The IHC score for FHIT and BAX expression was significantly higher in responders than the others (P less than 0.05). FHIT possesses tumor suppressor activity by induction of apoptosis in damaged cells after exposure to inflammatory carcinogens and DNA damaging chemo-radiotherapy.

INTERLEUKIN 4 RECEPTORS ON HUMAN BRONCHIAL EPITHELIAL CELLS. AN IN VIVO AND IN VITRO ANALYSIS OF EXPRESSION AND FUNCTION

Cytokine ◽  
1998 ◽  
Vol 10 (10) ◽  
pp. 803-813 ◽  
Author(s):  
Vincent H.J. van der Velden ◽  
Brigitta A.E. Naber ◽  
Annet F. Wierenga-Wolf ◽  
Reno Debets ◽  
Huub F.J. Savelkoul ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Interleukin 4 ◽  
Human Bronchial Epithelial Cells ◽  
Bronchial Epithelial ◽  
Vitro Analysis ◽  
And Function ◽  
In Vitro Analysis

Three-dimensional Growth of Renal Epithelial Cells in Vitro: A Tool in Toxicity Testing

1993 ◽  
Vol 21 (2) ◽  
pp. 191-195 ◽  
Author(s):  
Knut-Jan Andersen ◽  
Erik Ilsø Christensen ◽  
Hogne Vik
Keyword(s):  
Epithelial Cells ◽  
Three Dimensional ◽  
Toxicity Testing ◽  
Renal Tissue ◽  
Epithelial Cell Line ◽  
Multicellular Spheroids ◽  
Renal Epithelial Cell ◽  
Renal Epithelial Cells

The tissue culture of multicellular spheroids from the renal epithelial cell line LLC-PK1 (proximal tubule) is described. This represents a biological system of intermediate complexity between renal tissue in vivo and simple monolayer cultures. The multicellular structures, which show many similarities to kidney tubules in vivo, including a vectorial water transport, should prove useful for studying the potential nephrotoxicity of drugs and chemicals in vitro. In addition, the propagation of renal epithelial cells as multicellular spheroids in serum-free culture may provide information on the release of specific biological parameters, which may be suppressed or masked in serum-supplemented media.

scholarly journals In vitro analysis of elongation and termination by mutant RNA polymerases with altered termination behavior.

1996 ◽  
Vol 16 (11) ◽  
pp. 6468-6476 ◽  
Author(s):  
S A Shaaban ◽  
E V Bobkova ◽  
D M Chudzik ◽  
B D Hall
Keyword(s):  
Rna Polymerase Iii ◽  
Multiple Roles ◽  
Wild Type ◽  
Protein Motifs ◽  
Study Support ◽  
Vitro Analysis ◽  
Pol Iii ◽  
In Vitro Analysis

We have studied the in vitro elongation and termination properties of several yeast RNA polymerase III (pol III) mutant enzymes that have altered in vivo termination behavior (S. A. Shaaban, B. M. Krupp, and B. D. Hall, Mol. Cell. Biol. 15:1467-1478, 1995). The pattern of completed-transcript release was also characterized for three of the mutant enzymes. The mutations studied occupy amino acid regions 300 to 325, 455 to 521, and 1061 to 1082 of the RET1 protein (P. James, S. Whelen, and B. D. Hall, J. Biol. Chem. 266:5616-5624, 1991), the second largest subunit of yeast RNA pol III. In general, mutant enzymes which have increased termination require a longer time to traverse a template gene than does wild-type pol III; the converse holds true for most decreased-termination mutants. One increased-termination mutant (K310T I324K) was faster and two reduced termination mutants (K512N and T455I E478K) were slower than the wild-type enzyme. In most cases, these changes in overall elongation kinetics can be accounted for by a correspondingly longer or shorter dwell time at pause sites within the SUP4 tRNA(Tyr) gene. Of the three mutants analyzed for RNA release, one (T455I) was similar to the wild type while the two others (T455I E478K and E478K) bound the completed SUP4 pre-tRNA more avidly. The results of this study support the view that termination is a multistep pathway in which several different regions of the RET1 protein are actively involved. Region 300 to 325 likely affects a step involved in RNA release, while the Rif homology region, amino acids 455 to 521, interacts with the nascent RNA 3' end. The dual effects of several mutations on both elongation kinetics and RNA release suggest that the protein motifs affected by them have multiple roles in the steps leading to transcription termination.

Interaction of plakophilins with desmoplakin and intermediate filament proteins: an in vitro analysis

2000 ◽  
Vol 113 (13) ◽  
pp. 2471-2483 ◽  
Author(s):  
I. Hofmann ◽  
C. Mertens ◽  
M. Brettel ◽  
V. Nimmrich ◽  
M. Schnolzer ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intermediate Filament ◽  
Solid Phase ◽  
Specific Interaction ◽  
Binding Assays ◽  
Intermediate Filament Proteins ◽  
Amino Terminal ◽  
Vitro Analysis

Plakophilin 1 and 2 (PKP1, PKP2) are members of the arm-repeat protein family. They are both constitutively expressed in most vertebrate cells, in two splice forms named a and b, and display a remarkable dual location: they occur in the nuclei of cells and, in epithelial cells, at the plasma membrane within the desmosomal plaques. We have shown by solid phase-binding assays that both PKP1a and PKP2a bind to intermediate filament (IF) proteins, in particular to cytokeratins (CKs) from epidermal as well as simple epithelial cells and, to some extent, to vimentin. In line with this we show that recombinant PKP1a binds strongly to IFs assembled in vitro from CKs 8/18, 5/14, vimentin or desmin and integrates them into thick (up to 120 nm in diameter) IF bundles extending for several microm. The basic amino-terminal, non-arm-repeat domain of PKP1a is necessary and sufficient for this specific interaction as shown by blot overlay and centrifugation experiments. In particular, the binding of PKP1a to IF proteins is saturable at an approximately equimolar ratio. In extracts from HaCaT cells, distinct soluble complexes containing PKP1a and desmoplakin I (DPI) have been identified by co-immunoprecipitation and sucrose density fractionation. The significance of these interactions of PKP1a with IF proteins on the one hand and desmoplakin on the other is discussed in relation to the fact that PKP1a is not bound - and does not bind - to extended IFs in vivo. We postulate that (1) effective cellular regulatory mechanisms exist that prevent plakophilins from unscheduled IF-binding, and (2) specific desmoplakin interactions with either PKP1, PKP2 or PKP3, or combinations thereof, are involved in the selective recruitment of plakophilins to the desmosomal plaques.

Multifactorial Regulation of Cardiac Gene Expression: an In Vivo and In Vitro Analysis

1995 ◽  
Vol 752 (1 Cardiac Growt) ◽  
pp. 370-386 ◽  
Author(s):  
J. L. SAMUEL ◽  
I. DUBUS ◽  
F. FARHADIAN ◽  
F. MAROTTE ◽  
P. OLIVIERO ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cardiac Gene Expression ◽  
Cardiac Gene ◽  
Vitro Analysis ◽  
In Vitro Analysis

scholarly journals Tristetraprolin regulates IL-8 mRNA stability in cystic fibrosis lung epithelial cells

2009 ◽  
Vol 296 (6) ◽  
pp. L1012-L1018 ◽  
Author(s):  
Nagaraja Sethuraman Balakathiresan ◽  
Sharmistha Bhattacharyya ◽  
Usha Gutti ◽  
Robert P. Long ◽  
Catherine Jozwik ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Epithelial Cells ◽  
Mrna Stability ◽  
Posttranscriptional Regulation ◽  
Lung Epithelial Cells ◽  
Cellular Phenotype ◽  
Lung Epithelial ◽  
Vitro Analysis ◽  
In Vitro Analysis

Cystic fibrosis (CF) is due to mutations in the CFTR gene and is characterized by hypersecretion of the proinflammatory chemokine IL-8 into the airway lumen. Consequently, this induces the highly inflammatory cellular phenotype typical of CF. Our initial studies revealed that IL-8 mRNA is relatively stable in CF cells compared with those that had been repaired with [WT]CFTR (wild-type CFTR). Relevantly, the 3′-UTR of IL-8 mRNA contains AU-rich sequences (AREs) that have been shown to mediate posttranscriptional regulation of proinflammatory genes upon binding to ARE-binding proteins including Tristetraprolin (TTP). We therefore hypothesized that very low endogenous levels of TTP in CF cells might be responsible for the relative stability of IL-8 mRNA. As predicted, increased expression of TTP in CF cells resulted in reduced stability of IL-8 mRNA. An in vitro analysis of IL-8 mRNA stability in CF cells also revealed a TTP-induced enhancement of deadenylation causing reduction of IL-8 mRNA stability. We conclude that enhanced stability of IL-8 mRNA in TTP-deficient CF lung epithelial cells serve to drive the proinflammatory cellular phenotype in the CF lung.

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