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60th Annual Meeting of the German Society of Neurosurgery (DGNC)
Joint Meeting with the Benelux countries and Bulgaria

German Society of Neurosurgery (DGNC)

24 - 27 May 2009, Münster

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CCM3 gene silencing causes apoptosis-resistance and increases proliferation in the endothelium of cerebral cavernous malformations (CCMs)

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  • Y. Zhu - Klinik für Neurochirurgie, Universitätsklinikum Essen der Universität Duisburg-Essen
  • J.F. Xu - Klinik für Neurochirurgie, Universitätsklinikum Essen der Universität Duisburg-Essen
  • D. Miller - Klinik für Neurochirurgie, Universitätsklinikum Essen der Universität Duisburg-Essen
  • L. Benes - Klinik für Neurochirurgie, Universitätsklinikum Essen der Universität Duisburg-Essen
  • U. Sure - Klinik für Neurochirurgie, Universitätsklinikum Essen der Universität Duisburg-Essen

Deutsche Gesellschaft für Neurochirurgie. 60. Jahrestagung der Deutschen Gesellschaft für Neurochirurgie (DGNC), Joint Meeting mit den Benelux-Ländern und Bulgarien. Münster, 24.-27.05.2009. Düsseldorf: German Medical Science GMS Publishing House; 2009. DocP16-09

doi: 10.3205/09dgnc430, urn:nbn:de:0183-09dgnc4301

Published: May 20, 2009

© 2009 Zhu et al.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc-nd/3.0/deed.en). You are free: to Share – to copy, distribute and transmit the work, provided the original author and source are credited.

Outline

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Objective: CCM is a dynamically developing disease involving aberrant angiogenesis. Identification of frequent mutations of CCM1, CCM2 and CCM3 genes in familial CCMs has advanced our understanding of the genetic mechanism of this disease. However, the functions of these genes remain largely unknown. We have recently demonstrated a crucial role of CCM1 in regulating angiogenesis. The CCM3 locus, on the other hand, has been identified as the PDCD10 gene which codes for a protein linked to apoptosis. The present work continues to study the functions of the CCM3 gene in angiogenesis as well as in regulating endothelial apoptosis.

Methods: To carry out the gene functional study, CCM3 was silenced by siRNA in endothelium isolated, purified and cultured from CCMs (CCM-EC), and in two other types of commercially available endothelial cells (HUVEC and HBMEC). Silencing efficiency was confirmed by real-time PCR. After CCM3 silencing, cells were either exposed to apoptotic stimuli (staurosporin, 100nM, 4h) followed by nuclear staining, or underwent a proliferation study using the WST-1 assay, or a trans-well migration assay.

Results: Real-time PCR revealed a 3- to 5-fold reduction of CCM3 gene levels 72h after siRNA transfection in all three types of endothelial cells. Under such a condition of silencing, apoptotic damage significantly decreased by 23% in CCM-EC (p < 0.05). Similar results were obtained from HUVEC and HBMEC. The WST-1 assay indicated a significant increase of cell proliferation in CCM-EC as well as in HUVEC and HBMEC after CCM3 silencing. Furthermore, CCM3 silencing resulted in a 50% decrease of the numbers of migrated CCM-EC (p < 0.001). However, no significant change in migration pattern was shown in HUVEC and HBMEC after siRNA transfection.

Conclusions: For the first time, the present study provides evidence that CCM3 silencing induces resistance against apoptosis and increases proliferation in the endothelium of CCMs, which may potentially contribute to the aberrant angiogenesis in CCMs. The distinct migratory inhibition in CCM-EC after CCM3 silencing suggests an abnormal angiogenetic status of the endothelium derived from CCMs.