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Transcriptional regulation of Pim-1 kinase in vascular smooth muscle cells and its role for proliferation

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Abstract

The Ser/Thr-protein kinase Pim-1 has been discovered as a novel transducer of survival- and cell cycle promoting signals in the hematopoietic cell system. Although its significance for proliferation of vascular smooth muscle cells (VSMC) in vitro and neointima formation in vivo has been suggested recently, the mechanism has barely been characterized. This study aimed to foster the understanding of Pim-1 expression and regulation in murine VSMC in response to factors typically present within the atherosclerotic plaque. While oxidative stress, VEGF-A165 and angiotensin II did not have any effect on Pim-1 expression, VSMC strongly increased (3-fold) Pim-1 mRNA upon stimulation with PDGFbb, followed by its protein upregulation. Half life of Pim-1 RNA and protein were determined to be 25 min and 6 h, respectively. PDGFbb induced a strong, 10-fold increase in BrdU-uptake, a marker of proliferation. This was effectively blocked by either Pim-1-specific inhibitor quercetagetin or adenovirally introduced Pim-1 shRNA. We further identified the signaling pathways linking PDGFbb to Pim-1 in VSMC: as expected, we determined transcriptional stimulation of Pim-1 via Janus-activated kinase (Jak), but also an additional pathway involving protein kinase C (PKC) and the mitogen-activated protein kinase Mek1/2. Blockade of Akt signaling did, however, not interfere with Pim-1 upregulation, suggesting an independence of either survival system. PDGFbb-induced proliferation of VSMC is partly attributed to transcriptionally upregulated Pim-1 and was assigned to distinct cell signaling. Our findings help to understand the fundamental processes of vasculoproliferative diseases thus opening avenues for its prevention and treatment.

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Acknowledgments

This work was supported by grants from the Deutsche Forschungsgemeinschaft (DFG BR 1603/4, SFB655/A11, and DFG Research Center for Regenerative Therapies, CRTD).

Conflict of interest statement

None declared.

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Authors and Affiliations

  1. Internal Medicine, Department of Cardiology and Intensive Care, University of Technology Dresden, Dresden, Germany

    Manuela Willert, Antje Augstein, David M. Poitz & Ruth H. Strasser

  2. Internal Medicine, Department of Cardiology, Angiology, and Pneumology, Magdeburg University, Leipziger Strasse 44, 39120, Magdeburg, Germany

    Alexander Schmeisser & Ruediger Christian Braun-Dullaeus

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  1. Manuela Willert
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  2. Antje Augstein
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  6. Ruediger Christian Braun-Dullaeus
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Correspondence to Ruediger Christian Braun-Dullaeus.

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395_2009_55_MOESM1_ESM.doc

Supplemental material: Figure 1. Influence of quercetagetin on VSMC cell viability. VSMC were serum starved for 24 h prior to 24 h of stimulation with PDGFbb [10ng/ml] while being exposed to the chemical Pim-1-inhibitor quercetagetin. In contrast to VSMC treated with staurosporin, the cells incubated with quercetagetin did not show increased levels of DNA-fragmentation as demonstrated by Cell Death Detection ELISA. n=3; significance vs. untreated ctl (DOC 123 kb)

395_2009_55_MOESM2_ESM.doc

Supplemental material: Figure 2. Induction of Pim-1 transcription by PDGFbb- stimulation of quiescent VSMC derived from human aorta. Exposure of serum- starved VSMC to PDGFbb [10 ng/ml] transiently upregulates Pim-1 mRNA production by trend after one hour of stimulation as measured in Real-Time RT-PCR. n=4 (DOC 93 kb)

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Willert, M., Augstein, A., Poitz, D.M. et al. Transcriptional regulation of Pim-1 kinase in vascular smooth muscle cells and its role for proliferation. Basic Res Cardiol 105, 267–277 (2010). https://doi.org/10.1007/s00395-009-0055-x

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