TY  - JOUR
AU  - Vukmirović, Milica
AU  - Herazo-Maya, Jose D.
AU  - Blackmon, John
AU  - Skodrić-Trifunović, Vesna
AU  - Jovanović, Dragana
AU  - Pavlović, Sonja
AU  - Stojsić, Jelena
AU  - Zeljković, Vesna
AU  - Yan, Xiting
AU  - Homer, Robert
AU  - Stefanović, Branko
AU  - Kaminski, Naftali
PY  - 2017
UR  - https://imagine.imgge.bg.ac.rs/handle/123456789/1065
AB  - Background: Idiopathic Pulmonary Fibrosis (IPF) is a lethal lung disease of unknown etiology. A major limitation in transcriptomic profiling of lung tissue in IPF has been a dependence on snap-frozen fresh tissues (FF). In this project we sought to determine whether genome scale transcript profiling using RNA Sequencing (RNA-Seq) could be applied to archived Formalin-Fixed Paraffin-Embedded (FFPE) IPF tissues. Results: We isolated total RNA from 7 IPF and 5 control FFPE lung tissues and performed 50 base pair paired-end sequencing on Illumina 2000 HiSeq. TopHat2 was used to map sequencing reads to the human genome. On average similar to 62 million reads (53.4% of similar to 116 million reads) were mapped per sample. 4,131 genes were differentially expressed between IPF and controls (1,920 increased and 2,211 decreased (FDR  lt  0.05). We compared our results to differentially expressed genes calculated from a previously published dataset generated from FF tissues analyzed on Agilent microarrays (GSE47460). The overlap of differentially expressed genes was very high (760 increased and 1,413 decreased, FDR  lt  0.05). Only 92 differentially expressed genes changed in opposite directions. Pathway enrichment analysis performed using MetaCore confirmed numerous IPF relevant genes and pathways including extracellular remodeling, TGF-beta, and WNT. Gene network analysis of MMP7, a highly differentially expressed gene in both datasets, revealed the same canonical pathways and gene network candidates in RNA-Seq and microarray data. For validation by NanoString nCounter (R) we selected 35 genes that had a fold change of 2 in at least one dataset (10 discordant, 10 significantly differentially expressed in one dataset only and 15 concordant genes). High concordance of fold change and FDR was observed for each type of the samples (FF vs FFPE) with both microarrays (r = 0.92) and RNA-Seq (r = 0.90) and the number of discordant genes was reduced to four. Conclusions: Our results demonstrate that RNA sequencing of RNA obtained from archived FFPE lung tissues is feasible. The results obtained from FFPE tissue are highly comparable to FF tissues. The ability to perform RNA-Seq on archived FFPE IPF tissues should greatly enhance the availability of tissue biopsies for research in IPF.
PB  - Biomed Central Ltd, London
T2  - BMC Pulmonary Medicine
T1  - Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis
VL  - 17
DO  - 10.1186/s12890-016-0356-4
ER  - 

@article{
author = "Vukmirović, Milica and Herazo-Maya, Jose D. and Blackmon, John and Skodrić-Trifunović, Vesna and Jovanović, Dragana and Pavlović, Sonja and Stojsić, Jelena and Zeljković, Vesna and Yan, Xiting and Homer, Robert and Stefanović, Branko and Kaminski, Naftali",
year = "2017",
abstract = "Background: Idiopathic Pulmonary Fibrosis (IPF) is a lethal lung disease of unknown etiology. A major limitation in transcriptomic profiling of lung tissue in IPF has been a dependence on snap-frozen fresh tissues (FF). In this project we sought to determine whether genome scale transcript profiling using RNA Sequencing (RNA-Seq) could be applied to archived Formalin-Fixed Paraffin-Embedded (FFPE) IPF tissues. Results: We isolated total RNA from 7 IPF and 5 control FFPE lung tissues and performed 50 base pair paired-end sequencing on Illumina 2000 HiSeq. TopHat2 was used to map sequencing reads to the human genome. On average similar to 62 million reads (53.4% of similar to 116 million reads) were mapped per sample. 4,131 genes were differentially expressed between IPF and controls (1,920 increased and 2,211 decreased (FDR  lt  0.05). We compared our results to differentially expressed genes calculated from a previously published dataset generated from FF tissues analyzed on Agilent microarrays (GSE47460). The overlap of differentially expressed genes was very high (760 increased and 1,413 decreased, FDR  lt  0.05). Only 92 differentially expressed genes changed in opposite directions. Pathway enrichment analysis performed using MetaCore confirmed numerous IPF relevant genes and pathways including extracellular remodeling, TGF-beta, and WNT. Gene network analysis of MMP7, a highly differentially expressed gene in both datasets, revealed the same canonical pathways and gene network candidates in RNA-Seq and microarray data. For validation by NanoString nCounter (R) we selected 35 genes that had a fold change of 2 in at least one dataset (10 discordant, 10 significantly differentially expressed in one dataset only and 15 concordant genes). High concordance of fold change and FDR was observed for each type of the samples (FF vs FFPE) with both microarrays (r = 0.92) and RNA-Seq (r = 0.90) and the number of discordant genes was reduced to four. Conclusions: Our results demonstrate that RNA sequencing of RNA obtained from archived FFPE lung tissues is feasible. The results obtained from FFPE tissue are highly comparable to FF tissues. The ability to perform RNA-Seq on archived FFPE IPF tissues should greatly enhance the availability of tissue biopsies for research in IPF.",
publisher = "Biomed Central Ltd, London",
journal = "BMC Pulmonary Medicine",
title = "Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis",
volume = "17",
doi = "10.1186/s12890-016-0356-4"
}

Vukmirović, M., Herazo-Maya, J. D., Blackmon, J., Skodrić-Trifunović, V., Jovanović, D., Pavlović, S., Stojsić, J., Zeljković, V., Yan, X., Homer, R., Stefanović, B.,& Kaminski, N.. (2017). Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis. in BMC Pulmonary Medicine
Biomed Central Ltd, London., 17.
https://doi.org/10.1186/s12890-016-0356-4

Vukmirović M, Herazo-Maya JD, Blackmon J, Skodrić-Trifunović V, Jovanović D, Pavlović S, Stojsić J, Zeljković V, Yan X, Homer R, Stefanović B, Kaminski N. Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis. in BMC Pulmonary Medicine. 2017;17.
doi:10.1186/s12890-016-0356-4 .

Vukmirović, Milica, Herazo-Maya, Jose D., Blackmon, John, Skodrić-Trifunović, Vesna, Jovanović, Dragana, Pavlović, Sonja, Stojsić, Jelena, Zeljković, Vesna, Yan, Xiting, Homer, Robert, Stefanović, Branko, Kaminski, Naftali, "Identification and validation of differentially expressed transcripts by RNA-sequencing of formalin-fixed, paraffin-embedded (FFPE) lung tissue from patients with Idiopathic Pulmonary Fibrosis" in BMC Pulmonary Medicine, 17 (2017),
https://doi.org/10.1186/s12890-016-0356-4 . .