Abstract 1504: Increased Expression of Stem Cell Factor and its Receptor Following LVAD: A Potential Novel Target for Therapeutic Interventions In Heart Failure

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Jama Jahanyar ◽  
Keith A Youker ◽  
George P Noon ◽  
Guillermo Torre-Amione ◽  
Michael M Koerner ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heart Failure ◽  
Stem Cell ◽  
In Situ Hybridization ◽  
Mast Cells ◽  
Cardiac Remodeling ◽  
Stem Cell Factor ◽  
Therapeutic Interventions ◽  
Circulatory Support

Introduction: Previous studies have demonstrated the involvement of mast cells (MCs) in cardiac remodeling during heart failure. LVADs cause an influx of MCs into the failing heart, but the underlying mechanism is unknown. We hypothesize that stem cell factor (SCF) induces migration of MCs to the heart. This study investigates the potential role of SCF and its receptor (C-Kit) in promoting the recruitment of stem cell derived MCs during heart failure and following LVAD support. Methods: Myocardial samples were collected from 10 patients undergoing LVAD implantation (Pre-LVAD) paired with samples taken at the time of heart transplantation (Post-LVAD). Biopsies of 4 normal hearts served as controls. We assessed gene expression of SCF and C-Kit. Additionally, we stained for SCF, C-Kit and tryptase protein and utilized In-situ hybridization to determine the origin of SCF. Results: Real-time PCR: SCF mRNA is significantly increased (P<0.01) following mechanical circulatory support as compared to paired heart failure tissue. C-Kit mRNA was significantly increased post-LVAD, as compared to normal tissues (p<0.05). Immunohistochemistry: The C-Kit protein was only expressed on cardiac mast cells. In-Situ hybridization: SCF mRNA was found in endothelial cells, myocytes and interstitial cells. This was confirmed by antibody staining for the SCF protein. Conclusions: LVADs cause an increase of SCF and C-Kit gene expression during unloading. SCF appears to be an important mechanism for the recruitment and maturation of MCs involved in cardiac remodeling, and we suggest that pharmacologic or biologic modification of SCF may provide a new therapeutic path for heart failure treatment.

scholarly journals RNA In Situ Hybridization for Detecting Gene Expression Patterns in the Abdomens and Wings of Drosophila Species

2021 ◽  
Vol 4 (1) ◽  
pp. 20
Author(s):  
Mujeeb Shittu ◽  
Tessa Steenwinkel ◽  
William Dion ◽  
Nathan Ostlund ◽  
Komal Raja ◽  
...  
Keyword(s):  
Gene Expression ◽  
In Situ Hybridization ◽  
Expression Patterns ◽  
Drosophila Species ◽  
Gene Expression Patterns ◽  
Probe Design ◽  
Tissue Preparation ◽  
Design And Synthesis ◽  
Rna In Situ Hybridization

RNA in situ hybridization (ISH) is used to visualize spatio-temporal gene expression patterns with broad applications in biology and biomedicine. Here we provide a protocol for mRNA ISH in developing pupal wings and abdomens for model and non-model Drosophila species. We describe best practices in pupal staging, tissue preparation, probe design and synthesis, imaging of gene expression patterns, and image-editing techniques. This protocol has been successfully used to investigate the roles of genes underlying the evolution of novel color patterns in non-model Drosophila species.

scholarly journals Role of the lncRNA BACE1-AS in shared disease mechanisms between heart failure and Alzheimer's disease

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
S Greco ◽  
A Made' ◽  
A.S Tascini ◽  
J Garcia Manteiga ◽  
S Castelvecchio ◽  
...  
Keyword(s):  
Heart Failure ◽  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
In Situ Hybridization ◽  
Cell Apoptosis ◽  
Common Disease ◽  
Funding Source ◽  
Mrna Levels ◽  
Rna Seq

Abstract Background BACE1 encodes for β-secretase, the key enzyme involved in β-amyloid (βA) generation, a peptide well known for its involvement in Alzheimer's disease (AD). Of note, heart failure (HF) and AD share several risk factors and effectors. We recently showed that, in the heart of ischemic HF patients, the levels of both BACE1, its antisense RNA BACE1-AS and βA are all increased. BACE1-AS positively regulates the expression of BACE1, triggering βA intracellular accumulation, and its overexpression or βA administration induce cardiovascular-cell apoptosis. Aim To characterize the transcripts of the BACE1 locus and to investigate the molecular mechanisms underpinning BACE1-AS regulation of cell vitality. Methods By PCR and sequencing, we studied in the heart the expression of a variety of antisense BACE1 transcripts predicted by FANTOM CAT Epigenome. We studied BACE1 RNA stability by BrdU pulse chase experiments (BRIC assay). The cellular localization of BACE1-AS RNA was investigated by in situ hybridization assay. BACE1-AS binding RNAs were evaluated by BACE1-AS-MS2-Tag pull-down in AC16 cardiomyocytes followed by RNA-seq. Enriched RNAs were validated by qPCR and analysed by bioinformatics comparison with publicly available gene expression datasets of AD brains. Results We readily detected several antisense BACE1 transcripts expressed in AC16 cardiomyocytes; however, only BACE1-AS RNAs overlapping exon 6 of BACE1 positively regulated BACE1 mRNA levels, acting by increasing its stability. BACE1 silencing reverted cell apoptosis induced by BACE1-AS expression, indicating that BACE1 is a functional target of BACE1-AS. However, in situ hybridization experiments indicated a mainly nuclear localization for BACE1-AS, which displayed a punctuated distribution, compatible with chromatin association and indicative of potential additional targets. To identify other BACE1-AS binding RNAs, a BACE1-AS-MS2-tag pull-down was performed and RNA-seq of the enriched RNAs identified 698 BACE1-AS interacting RNAs in cardiomyocytes. Gene ontology of the BACE1-AS binding RNAs identified categories of relevance for cardiovascular or neurological diseases, such as dopaminergic synapse, glutamatergic synapse, calcium signalling pathway and voltage-gated channel activity. In spite of the differences between brain and heart transcriptomes, BACE1-AS-interacting RNAs identified in cardiomyocytes were significantly enriched in transcripts differentially expressed in AD brains as well as in RNAs expressed by enhancer genomic regions that are significantly hypomethylated in AD brains. Conclusions These data shed a new light on the complexity of BACE1-AS locus and on the existence of RNAs interacting with BACE1-AS with a potential as enhancer-RNAs. Moreover, the dysregulation of the BACE1-AS/BACE1/βA pathway may be a common disease mechanism shared by cardiovascular and neurological degenerative diseases. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Italian Health Ministery_Ricerca Corrente 2020

scholarly journals FISHing for chick genes: Triple-label whole-mount fluorescence in situ hybridization detects simultaneous and overlapping gene expression in avian embryos

2004 ◽  
Vol 229 (3) ◽  
pp. 651-657 ◽  
Author(s):  
Nathaniel Denkers ◽  
Pilar García-Villalba ◽  
Christopher K. Rodesch ◽  
Kandice R. Nielson ◽  
Teri Jo Mauch
Keyword(s):  
Gene Expression ◽  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Avian Embryos ◽  
Overlapping Gene ◽  
Whole Mount

scholarly journals Localization of stromelysin gene expression in atherosclerotic plaques by in situ hybridization.

1991 ◽  
Vol 88 (18) ◽  
pp. 8154-8158 ◽  
Author(s):  
A. M. Henney ◽  
P. R. Wakeley ◽  
M. J. Davies ◽  
K. Foster ◽  
R. Hembry ◽  
...  
Keyword(s):  
Gene Expression ◽  
In Situ Hybridization ◽  
Atherosclerotic Plaques
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