title: Rap1-mediated nuclear factor-kappaB (NF-κB) activity regulates the paracrine capacity of mesenchymal stem cells in heart repair following infarction.
abstract: Paracrine effect is the major mechanism that underlies mesenchymal stem cells (MSC)-based therapy. This study aimed to examine how Rap1, telomeric repeat-binding factor 2-interacting protein 1 (Terf2IP), which is a novel modulator involved in the nuclear factor-kappaB (NF-κB) pathway, regulates the paracrine effects of MSC-mediated heart repair following infarction. NF-κB activity of stromal cells was increased by Rap1 as measured by pNF-κB-luciferase reporter activity, and this was abolished by IkB-dominant-negative protein. Knockdown of Rap1 with shRap1 resulted in diminished translocation of p65-NF-κB from the cytoplasm to nuclei in response to tumor necrosis factor-α (TNF-α) stimulation. Compared with BM-MSCs, Rap1(-/-)-BM-MSCs displayed a significantly reduced ratio of phosphorylated NF-κB to NF-κB-p65 and of Bax to Bcl-2, and increased resistance to hypoxia-induced apoptosis by the terminal deoxynucleotidal transferase-mediated dUTP nick end labeling (TUNEL) assay. In contrast, re-expression of Rap1 in Rap1(-/-)-BM-MSCs resulted in loss of resistance to apoptosis in the presence of hypoxia. Moreover, absence of Rap1 in BM-MSCs led to downregulation of NF-κB activity accompanied by reduced pro-inflammatory paracrine cytokines TNF-α, IL (interleukin)-6 and monocyte chemotactic protein-1 in Rap1(-/-)-BM-MSCs compared with BM-MSCs. The apoptosis of neonatal cardiomyocytes (NCMCs) induced by hypoxia was significantly reduced when cocultured with Rap1(-/-)-BM-MSC hypoxic-conditioned medium (CdM). The increased cardioprotective effects of Rap1(-/-)-BM-MSCs were reduced when Rap1(-/-)-BM-MSCs were reconstituted with Rap1 re-expression. Furthermore, in vivo study showed that transplantation of Rap1(-/-)-BM-MSCs significantly improved heart function, decreased infarct size, prevented cardiomyocyte apoptosis and inhibited inflammation compared with controls and BM-MSCs (P<0.01). This study reveals that Rap1 has a critical role in the regulation of MSC paracrine actions. Compared with BM-MSCs, Rap1(-/-)-BM-MSCs decreased NF-κB sensitivity to stress-induced pro-inflammatory cytokine production and reduced apoptosis. Selective inhibition of Rap1 in BM-MSCs may be a novel strategy to enhance MSC-based therapeutic efficacy in myocardial infarction.
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: S
authors.initials: S
authors.lastname: Chiu
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: X
authors.initials: X
authors.lastname: Liang
authors.affiliation: Department of Ophthalmology, Li Ka Shing Faculty of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: F
authors.initials: F
authors.lastname: Gao
authors.affiliation: Department of Ophthalmology, Li Ka Shing Faculty of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: Z
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: S
authors.initials: S
authors.lastname: Liao
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Liang
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong , Hong Kong SAR, China.
authors.forename: Y-H
authors.initials: YH
authors.lastname: Chai
authors.affiliation: Institute of Molecular and Cellular Biology , Biopolis, Singapore.
authors.forename: D J H
authors.initials: DJ
authors.lastname: Low
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong, Hong Kong SAR, China; Research Centre of Heart, Brain, Hormone, and Healthy Aging, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China.
authors.forename: H-F
authors.initials: HF
authors.lastname: Tse
authors.affiliation: Institute of Molecular and Cellular Biology , Biopolis, Singapore.
authors.forename: V
authors.initials: V
authors.lastname: Tergaonkar
authors.affiliation: Division of Cardiology, Department of Medicine, The University of Hong Kong, Hong Kong SAR, China; Research Centre of Heart, Brain, Hormone, and Healthy Aging, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China; Department of Ophthalmology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong SAR, China; Shenzhen University Health Science Center, Shenzhen, China.
authors.forename: Q
authors.initials: Q
authors.lastname: Lian