Grabham Peter, Sharma Preety, Bigelow Alan, Geard Charles
Center for Radiological Research, Columbia University, VC 11-205A/243, 630 West 168th street, New York, NY 10032, USA.
Vasc Cell. 2013 Sep 17;5(1):16. doi: 10.1186/2045-824X-5-16.
Charged particle radiation is known to be more biologically effective than photon radiation. One example of this is the inhibition of the formation of human blood vessels. This effect is an important factor influencing human health and is relevant to space travel as well as to cancer radiotherapy. We have previously shown that ion particles with a high energy deposition, or linear energy transfer (LET) are more than four times more effective at disrupting mature vessel tissue models than particles with a lower LET. For vasculogenesis however, the relative biological effectiveness between particles is the same. This unexpected result prompted us to investigate whether the inhibition of vasculogenesis was occurring by distinct mechanisms.
Using 3-Dimensional human vessel models, we developed assays that determine at what stage angiogenesis is inhibited. Vessel morphology, the presence of motile tip structures, and changes in the matrix architecture were assessed. To confirm that the mechanisms are distinct, stimulation of Protein Kinase C (PKC) with phorbol ester (PMA) was employed to selectively restore vessel formation in cultures where early motile tip activity was inhibited.
Endothelial cells in 3-D culture exposed to low LET protons failed to make connections with other cells but eventually developed a central lumen. Conversely, cells exposed to high LET Fe charged particles extended cellular processes and made connections to other cells but did not develop a central lumen. The microtubule and actin cytoskeletons indicated that motility at the extending tips of endothelial cells is inhibited by low LET but not high LET particles. Actin-rich protrusive structures that contain bundled microtubules showed a 65% decrease when exposed to low LET particles but not high LET particles, with commensurate changes in the matrix architecture. Stimulation of PKC with PMA restored tip motility and capillary formation in low but not high LET particle treated cultures.
Low LET charged particles inhibit the early stages of vasculogenesis when tip cells have motile protrusive structures and are creating pioneer guidance tunnels through the matrix. High LET charged particles do not affect the early stages of vasculogenesis but they do affect the later stages when the endothelial cells migrate to form tubes.
已知带电粒子辐射比光子辐射具有更高的生物学效应。其中一个例子是对人类血管形成的抑制作用。这种效应是影响人类健康的一个重要因素，与太空旅行以及癌症放射治疗都相关。我们之前已经表明，具有高能量沉积或线性能量转移（LET）的离子粒子在破坏成熟血管组织模型方面比具有较低LET的粒子有效四倍以上。然而，对于血管生成，不同粒子之间的相对生物学效应是相同的。这一意外结果促使我们研究血管生成的抑制是否通过不同机制发生。
使用三维人类血管模型，我们开发了测定血管生成在哪个阶段受到抑制的试验。评估了血管形态、活动的尖端结构的存在以及基质结构的变化。为了确认机制不同，使用佛波酯（PMA）刺激蛋白激酶C（PKC），以选择性地恢复早期活动尖端活性受到抑制的培养物中的血管形成。
暴露于低LET质子的三维培养中的内皮细胞未能与其他细胞建立连接，但最终形成了中央管腔。相反，暴露于高LET铁带电粒子的细胞伸出细胞突起并与其他细胞建立连接，但没有形成中央管腔。微管和肌动蛋白细胞骨架表明，低LET而非高LET粒子抑制了内皮细胞延伸尖端的运动性。当暴露于低LET粒子而非高LET粒子时，含有成束微管的富含肌动蛋白的突出结构减少了65%，同时基质结构也有相应变化。用PMA刺激PKC可恢复低LET而非高LET粒子处理的培养物中的尖端运动性和毛细血管形成。
低LET带电粒子在尖端细胞具有活动的突出结构并在基质中创建先驱引导通道时抑制血管生成的早期阶段。高LET带电粒子不影响血管生成的早期阶段，但它们确实影响内皮细胞迁移形成管的后期阶段。
