Santucci Kimberly L, Baust John M, Snyder Kristi K, Van Buskirk Robert G, Baust John G
Department of Biological Sciences, State University of New York at Binghamton, Binghamton, NY, USA Institute for Biomedical Technology, State University of New York at Binghamton, Binghamton, NY, USA CPSI Biotech, Owego, NY, USA
Institute for Biomedical Technology, State University of New York at Binghamton, Binghamton, NY, USA CPSI Biotech, Owego, NY, USA.
Technol Cancer Res Treat. 2016 Aug;15(4):609-17. doi: 10.1177/1533034616648059. Epub 2016 May 9.
Cryoablation, an effective means of ablating cancer, is often used in conjunction with adjuvants that target cancer cells in a specific cell cycle stage to increase treatment efficacy. The objective of this study was to investigate the impact of cell cycle stage on cancer freeze response as well as investigate the potential cellular kinetic effect of calcitriol, the active metabolic of vitamin D3, when used as a cryosensitizing adjuvant in order to maximize prostate cancer cell death.
Cell cycle distribution of PC-3 cells was analyzed via flow cytometry to compare gap 1, synthesis, and gap 2/mitosis phase subpopulations pre- and postfreeze as well as changes elicited by calcitriol pretreatment. Distinct gap 1, synthesis, and gap 2/mitosis phase populations were obtained through fluorescence-activated cell sorting and synthesis phase thymidine synchronization. Posttreatment viability was assessed using alamarBlue and fluorescence microscopy to assess live, apoptotic, and necrotic subpopulations.
A small but statistically significant increase in synthesis phase and decrease in gap 2/mitosis phase populations was noted at 6 hours postfreeze in asynchronous samples. Synchronization in synthesis phase yielded an increase in cell death when combined with freezing to both -15°C and -20°C. Calcitriol pretreatment increased the gap 1 phase population by 20% and a synergistic decrease in viability following freezing. However, gap 1-sorted populations combined with calcitriol treatment did not exhibit this synergistic effect. Fluorescence microscopy of fluorescence-activated cell sorting-sorted cells revealed necrosis as the predominant form of cell death in all phases, though apoptosis did play a role.
Although initial results suggested a potential sensitivity, PC-3 cells exposed to freezing as sorted populations did not reveal significant differences in cell death. As such, the data from this study suggest that there is no difference in cell cycle stage sensitivity to freezing injury.
冷冻消融是一种有效的癌症消融手段，常与靶向特定细胞周期阶段癌细胞的佐剂联合使用，以提高治疗效果。本研究的目的是调查细胞周期阶段对癌症冷冻反应的影响，并研究维生素D3的活性代谢产物骨化三醇作为冷冻增敏佐剂时的潜在细胞动力学效应，以最大限度地增加前列腺癌细胞死亡。
通过流式细胞术分析PC-3细胞的细胞周期分布，以比较冷冻前后的G1期、S期和G2/M期亚群，以及骨化三醇预处理引起的变化。通过荧光激活细胞分选和S期胸腺嘧啶同步化获得不同的G1期、S期和G2/M期群体。使用alamarBlue和荧光显微镜评估治疗后的活力，以评估活细胞、凋亡细胞和坏死亚群。
在冷冻后6小时，异步样本中S期有小幅但具有统计学意义的增加，G2/M期群体减少。S期同步化与冷冻至-15°C和-20°C相结合时，细胞死亡增加。骨化三醇预处理使G1期群体增加20%，冷冻后活力协同下降。然而，G1期分选群体与骨化三醇治疗相结合并未表现出这种协同效应。荧光激活细胞分选分选细胞的荧光显微镜检查显示，尽管凋亡确实起作用，但坏死是所有阶段细胞死亡的主要形式。
尽管初步结果表明存在潜在敏感性，但作为分选群体暴露于冷冻的PC-3细胞在细胞死亡方面未显示出显著差异。因此，本研究的数据表明，细胞周期阶段对冷冻损伤的敏感性没有差异。
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