Deletion of protein tyrosine phosphatase 1B in vascular fibroblasts promotes proliferation and pathological vascular remodeling by enhancing PDGFRB signaling

Background and Aim: Growth factor signaling via the reversible phosphorylation of receptors or signaling intermediates is catalyzed by protein tyrosine kinases, whose activities are controlled by dephosphorylation. Protein Tyrosine Phosphatase-1B (PTP1B) and Src Homology Region 2-Containing Protein Tyrosine Phosphatase (SHP2) negatively regulate the activity of receptor tyrosine kinases, including key growth factor receptors like Platelet Derived Growth Factor Receptor (PDGFR), Whether PTP1B and SHP2 have specific roles in vascular remodeling in response to injury remains unclear. We have previously shown that lack of PTP1B in vascular mesenchymal progenitor cells promotes adventitial fibrosis. This study explores the role of PTP1B and SHP2 in vascular fibroblasts during pathological vascular remodeling.

Methods: To generate fibroblast PTP1B and SHP2 knockout mice (PTP1B-KO and SHP2-KO), PDGFRB.Cre-ER^T2 transgenic mice were crossed with either PTP1B flox/flox or SHP2 flox/flox mice. Mice expressing membrane-tethered green fluorescent protein (mEGFP) under control of PDGFRB.Cre-ER^T2 were utilized for mesenchymal lineage tracing. Vascular injury was induced in 12-week-old male mice by applying 10% ferric chloride to the common carotid artery. To investigate the regulatory role of PTP1B and SHP2 on specific cellular functions, primary mural cells were isolated from the aorta and carotid arteries of uninjured mice and mice 3 weeks after vascular injury, cultivated in DMEM and analyzed between passage 0 and 1.

Results: Primary murine aortic fibroblasts with genetic PTP1B deficiency exhibited higher PDGFRB mRNA and protein expression and increased proliferation, as shown by significantly elevated cyclin D1 mRNA levels, PCNA protein expression, and numbers of Ki-67-immunopositive cells. Moreover, PTP1B-deficient mural cells exhibited faster wound closure rates in response to a scratch injury, which was significantly reduced in the presence of the PDGFRB inhibitor imatinib and the DNA synthesis inhibitor mitomycin C, suggesting a predominant role of increased proliferation and not migration in this observation. PTP1B inhibitors confirmed these findings in human aortic fibroblasts. On the other hand, transmigration experiments using conditioned medium from primary fibroblast showed that altered paracrine activities did not play a significant role in these results. Immunoprecipation followed by immunoblot uncovered that PTP1B directly interacts with PDGFRB. Morphometric analysis of serial carotid artery cross-sections three weeks after FeCl₃ vascular injury revealed that absence of PTP1B in fibroblasts resulted in significantly larger neointima area, intima-to-media ratio, and vascular lumen stenosis. Lineage tracing experiments using PDGFRB reporter mice showed that the majority of cells within the neointima were derived from PDGFRB-expressing cells. Pathological vascular remodeling, increased proliferation and PDGFRB overexpression was also observed in mice and cells lacking the non-receptor protein tyrosine phosphatase SHP2 (SH2 domain-containing protein tyrosine phosphatase-2) in fibroblasts suggesting common activities and substrates in response to vascular injury regulating remodeling.

Conclusion: Our findings suggest that absence or inactivation of PTP1B in vascular fibroblasts promotes pathological vascular remodeling by enhancing PDGRRB signaling and proliferation.