Auteur Topic: RNA sequencing used to discover novel genes and pathways in celiac disease  (gelezen 386 keer)

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Volledig artikel in PlosOne - zie onder bericht MedicalExpress
(correspondentie:
Alessio Fasano, MD, director of the Center for Celiac Research and Treatment at MGHfC)




Bericht MedicalExpress
Citaat
RNA sequencing used to discover novel genes and pathways in celiac disease
by Massachusetts General Hospital

Researchers at the Mucosal Immunology and Biology Research Center at MassGeneral Hospital for Children (MGHfC) have discovered novel genes and pathways related to early stages in the development of celiac disease and the ongoing inflammation and comorbidities associated with the condition.
The findings, published in PLOS One, include analyses of RNA sequences in duodenal biopsies from individuals with and without celiac disease and are consistent with many previously described pathways in the development of celiac disease.


In collaboration with Regeneron Pharmaceuticals, Inc. - a biopharmaceutical company based in Tarrytown, New York—researchers performed whole-transcriptome shotgun sequencing of 12 patients with active celiac disease, 15 celiac patients in remission with no intestinal damage, and 15 individuals without celiac disease.
By analyzing participants' transcriptome—the total sum of transcribed RNA equences—researchers discovered which genes were expressed and which genes were not expressed to determine genetic signatures linked to celiac disease.


"We know that celiac disease is a multifactorial disease with about 57 genes associated with this autoimmune condition. By performing RNA sequencing, we have uncovered additional genetic 'signatures' and moved closer to identifying targets for future therapeutic agents—in celiac disease and possibly other autoimmune conditions," says Maureen Leonard, MD, clinical director of the Center for Celiac Research and Treatment at MGHfC, an instructor in Pediatrics at Harvard Medical School and first author of the study.


The results showed clear differences in gene expressions among the three groups.
Researchers found significant differences in the expression of 945 genes between people with active celiac disease and non-celiac controls;
290 genes between people with celiac disease in remission and the non-celiac group; and 538 genes between the active celiac group and the celiac disease in remission group.


"The identified genes activated three major pathways: innate immunity, gut permeability and differentiation in cell maturation," says Alessio Fasano, MD, director of the Center for Celiac Research and Treatment at MGHfC, a professor of Pediatrics at Harvard Medical School and senior author of the PLOS One report.
"We can confirm that these functions are instrumental when you develop celiac disease."


Expression of some of the genes returned to normal when patients were placed on a gluten-free diet, he notes, and some did not.
Fasano notes that this finding could provide some insight into why some people have persistent intestinal damage even after following a strict gluten-free diet.
From the enormous array of data generated, researchers chose to focus on functional pathways that were significantly different between the active celiac group and those in remission.


Two of the three top perturbed pathways in the active celiac group involved signaling by cytokines and chemokines, which are known as the immune system's "first responders" and markers of inflammation in innate immunity in the early stages of disease development.
The researchers also found evidence to suggest that the risk of co-morbid autoimmune disorder may be high in active celiac disease, as pathways for type 1 diabetes, lupus and autoimmune thyroid disease also were upregulated.


Identifying potential targets for therapeutic intervention in celiac disease and other autoimmune conditions is a long-term goal of the research group.
"This study is only the beginning. Our findings provide the framework for future validation studies to investigate the early steps in celiac disease pathogenesis and to examine the remission state," says Leonard.
Research is underway at other centers for the use of the transcriptome in targeted therapy in Inflammatory Bowel Disease, and Leonard and Fasano hope to see this replicated for celiac disease.



Volledig artikel in PlosOne
Published: April 18, 2019
Citaat
RNA sequencing of intestinal mucosa reveals novel pathways functionally linked to celiac disease pathogenesis

Maureen M. Leonard1,2,3, Yu Bai4, Gloria Serena2,3, Kourtney P. Nickerson2, Stephanie Camhi2, Craig Sturgeon5, Shu Yan2, Maria R. Fiorentino2,
Aubrey Katz1, Barbara Nath6, James Richter6, Matthew Sleeman4, Cagan Gurer4, Alessio FasanoID1,2,3*
1 Mass General Hospital for Children and Division of Pediatric Gastroenterology and Nutrition, Harvard Medical School, Boston, Massachusetts,
   United States of America,
2 Center for Celiac Research and Treatment, Mucosal Immunology and Biology Research Center, Boston, Massachusetts, United States of America,
3 Celiac Research Program, Harvard Medical School, Boston, Massachusetts, United States of America,
4 Regeneron Pharmaceuticals, Tarrytown, New York, United States of America,
5 Graduate Program in Life Sciences, University of Maryland, Baltimore, Maryland, United States of America,
6 Department of Gastroenterology, Massachusetts General Hospital, Boston, Massachusetts, United States of America
* afasano@mgh.harvard.edu


Abstract
Background & aims
The early steps in the pathophysiology of celiac disease (CD) leading to loss of tolerance to gluten are poorly described. Our aim was to use RNA sequencing of duodenal biopsies in patients with active CD, CD in remission, and non-CD controls to gain insight into CD pathophysiology, identify additional genetic signatures linked to CD, and possibly uncover targets for future therapeutic agents.

 
Methods
We performed whole transcriptome shotgun sequencing of intestinal biopsies in subjects with active and remission CD and non-CD controls. We also performed functional pathway analysis of differentially expressed genes to identify statistically significant pathways that are up or down regulated in subjects with active CD compared to remission CD.

 
Results
We identified the upregulation of novel genes including IL12R, ITGAM and IGSF4 involved in the immune response machinery and cell adhesion process in the mucosa of subjects with active CD compared to those in remission. We identified a unique signature of genes, related to innate immunity, perturbed exclusively in CD irrespective of disease status. Finally, we highlight novel pathways of interest that may contribute to the early steps of CD pathogenesis and its comorbidities such as the spliceosome, pathways related to the innate immune response, and pathways related to autoimmunity.

 
Conclusions
Our study confirmed previous findings based on GWAS and immunological studies pertinent to CD pathogenesis and describes novel genes and pathways that with further validation may be found to contribute to the early steps in the pathogenesis of CD, ongoing inflammation, and comorbidities associated with CD.


S1 Table. 
Pathways unique to patients with active CD.
https://doi.org/10.1371/journal.pone.0215132.s001
(DOCX)

S1 Fig. 
Gene Expression Signatures Unique to Active CD.
Heatmap shows non-CD control subjects (blue), active CD subjects (green) and remission CD subjects (pink) with each subject’s data represented vertically. The color scale show gene expression with dark blue indicating downregulation, light blue indicating decreased expression, light red indicating increased expression, and red indicating upregulation.
https://doi.org/10.1371/journal.pone.0215132.s002
(TIF)

S2 Fig.
Gene Expression Common to Active and Remission CD.
Heatmap shows non-CD control subjects (blue), active CD subjects (green) and remission CD subjects (pink) with each subject’s data represented vertically. The color scale show gene expression with dark blue indicating downregulation, light blue indicating decreased expression, light red indicating increased expression, and red indicating upregulation.
https://doi.org/10.1371/journal.pone.0215132.s003
(TIF)

S3 Fig.
RTPCR Results in a subset of genes in healthy control patients, patients with active CD, and patients with CD in remission.
https://doi.org/10.1371/journal.pone.0215132.s004
(TIF)

S4 Fig.
Immunohistochemistry results in a subset of healthy control patients, patients with active CD, and patients with CD in remission.
https://doi.org/10.1371/journal.pone.0215132.s005
(TIF)

S5 Fig.
KEGG pathway cytokine-cytokine receptor interaction. We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission.
Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s006
(TIF)

S6 Fig.
KEGG pathway chemokine signaling pathway.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s007
(TIF)

S7 Fig.
KEGG pathway cell cycle.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s008
(TIF)

S8 Fig.
KEGG pathway cell adhesion molecules.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s009
(TIF)

S9 Fig.
KEGG pathway spliceosome.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s010
(TIF)

S10 Fig.
KEGG pathway retinol metabolism.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s011
(TIF)

S11 Fig.
KEGG pathway systemic lupus erythematosus.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s012
(TIF)

S12 Fig.
KEGG pathway type 1 diabetes mellitus.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s013
(TIF)

S13 Fig.
KEGG pathway autoimmune thyroid disease.
We performed a subtraction analysis using GSEA for patients with active CD compared to those with CD in remission. Genes identified as significant are highlighted in red against a background of other known genes in the pathway.
https://doi.org/10.1371/journal.pone.0215132.s014
(TIF)


Volledig:
https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0215132&type=printable