UNIVERSITY OF OSLO. Cytokine expression profiling of the myocardium reveals a role for CX3CL1 (fractalkine) in heart failure.
Cytokine expression profiling of the myocardium reveals a role for CX3CL1 (fractalkine) in heart failure
Cathrine Husberg (1,8), Ståle Nygård (1,2,8), Alexandra Vanessa Finsen (1,8), Jan Kristian Damås (4), Arnoldo Frigessi(3), Erik Øye(6,7,8), Lars Gullestad(4,8), Pål Aukrust(4,5), Arne Yndestad(4,8), Geir Christensen(1,8)
1.Institute for Experimental Medical Research, Ullevål University Hospital
2.Department of Mathematics, University of Oslo
3.Department of Biostatistics, University of Oslo
4. Research Institute for Internal medicine, Rikshospitalet-Radiumhospitalet Medical Center
5. Section of Clinical Immunology and Infectious Diseases, Rikshospitalet-Radiumhospitalet Medical Center
6. Department of Cardiology, Rikshospitalet-Radiumhospitalet Medical Center
7. Institute for Surgical Research, Rikshospitalet-Radiumhospitalet Medical Center
8. Center for Heart Failure Research, University of Oslo
Identify cytokines imortant for the development of heart failure (HF)
- and that are not previously associated with HF
Gene modified mice
At each time point tissues from 5 mice with myocardial infarction (MI) and 5 SHAM operated mice were used for cDNA microarray screening.
Microarray preprosessing by MAANOVA
Following the MicroArray ANOVA (MAANOVA) model of Kerr et al (2000), log-transformed intensity for a gene g on array i with dye j (Cy5 or Cy3) and treatment k (MI or SHAM) was modelled by
We are mainly interested in the quantity VG1g-VG2g , as it represents the gene-specific effect of MI.
Adding the other (nuisance) parameters results in a model based normalisation of the expression measurements.
R package: maanova
> h<-read.madata("highint.txt", header=FALSE, designfile="des.txt”......
> r<-s.c(l,h) #correct for saturated spots according to Lyng et al (2004)
> n<-transform.madata(d,method="rlowess",draw="off") #”pre-normalisation” using lowess
> m<-makeModel(n,formula=~Dye+Type+Array) #Type=MI or SHAM
4. Read result file using Excel.
HF C HF C HF C HF C HF C HF C HF
Verification by qRT-PCR and in humans.
Exploring fractalkine’s molecular function
Stimulate cells with fractalkine, and screen for differentially expressed genes.
Use CXCR knock-out mice, and screen for differentially expressed genes
Identify signalling pathways significantly affected by fractalkine stimulation/ knock-out using the software Ingenuity Pathway Analysis (enrichment analysis).
Ingenuity Pathway Analysis
Commercial software for understanding complex biological systems.
Uses a knowledge base containing (millions of) biological and chemical relationships (manually) extracted from the scientific literature.
* Signaling and Metabolic Pathways Analysis
* Cellular and Disease Process Analysis
* Molecular Network Analysis
Signaling pathways activated by fractalkine
Signalling pathways in adult cardiomyocytes stimulated by fractalkine (identified by Ingenuity Pathways Analysis software)