The University of New South Wales, Sydney, Australia.
Biomacromolecules. 2010 Feb 8;11(2):412-20. doi: 10.1021/bm901129x.
The RAFT technique has been increasingly used to generate polymers for potential biological applications. However, to-date, the toxicity of the RAFT-polymers has received limited attention. In this study, the in vitro cytotoxicity of three different, RAFT-synthesized, water-soluble polymers was investigated using three different adherent cell lines via CellTiter-Blue cell viability and the cytosolic enzyme lactate dehydrogenase (LDH) cytotoxicity assays. In brief, P(OEG-A) and P(OEG-MA) samples bearing omega-dithiobenzoate or omega-trithiocarbonate end groups and varying P(HPMA) samples bearing omega-dithiobenzoate, omega-trithiocarbonate, or non-RAFT end groups, were investigated using Chinese hamster ovary cells (CHO-K1), mouse macrophage cells (Raw264.7), and mouse fibroblast cells (NIH3T3). Any changes in the morphology of the cells after treatment with polymers were monitored via microscopy. The cytotoxicity of the polymers after treatment with metabolic liver enzymes was also evaluated. The average viability of CHO-K1 and NIH3T3 cells treated with dithiobenzoate- and trithiocarbonate-ended OEG-based polymers (1000 microM) for 24 h was close to 100%. The RAW264.7 cells were slightly more sensitive when incubated with dithiobenzoate-ended polymers (cell viability above 73%) for 24 h. The viability of the cells after 3 days of incubation with the polymers either slightly decreased or showed no change with respect to the viabilities obtained after 1 day of incubation. Analyses of cell morphology and cell membrane integrity via microscopy and a LDH assay confirmed the cell viability results obtained via CellTiter-Blue Assay. Unexpectedly, dithiobenzoate-ended P(HPMA) (at 1000 microM) exhibited high cytotoxicity after 24 h with all three cells lines. Further investigation of various P(HPMA) samples revealed that trithiocarbonate-ended and HPMA-capped P(HPMA)s at the same concentration were nontoxic over the same period of time. Also, dithiobenzoate-ended P(HPMA) at low concentrations (< or = 200 microM) can be tolerated by the cells tested.
RAFT 技术已被越来越多地用于生成潜在生物应用的聚合物。然而，迄今为止，RAFT 聚合物的毒性受到了有限的关注。在这项研究中，通过 CellTiter-Blue 细胞活力测定和细胞质酶乳酸脱氢酶 (LDH) 细胞毒性测定，使用三种不同的贴壁细胞系研究了三种不同的 RAFT 合成的水溶性聚合物的体外细胞毒性。简而言之，研究了具有 ω-二硫代苯甲酸酯或 ω-三硫代碳酸酯端基的 P(OEG-A)和 P(OEG-MA) 样品以及具有 ω-二硫代苯甲酸酯、ω-三硫代碳酸酯或非 RAFT 端基的不同 P(HPMA) 样品，使用中国仓鼠卵巢细胞 (CHO-K1)、小鼠巨噬细胞 (Raw264.7) 和小鼠成纤维细胞 (NIH3T3)。通过显微镜监测用聚合物处理后细胞形态的任何变化。还评估了用代谢性肝酶处理后聚合物的细胞毒性。用二硫代苯甲酸酯和三硫代碳酸酯端基的 OEG 基聚合物（1000 μM）处理 24 小时后，CHO-K1 和 NIH3T3 细胞的平均活力接近 100%。用二硫代苯甲酸酯端基聚合物孵育 24 小时时，RAW264.7 细胞的敏感性略高（细胞活力高于 73%）。用聚合物孵育 3 天后，细胞活力与孵育 1 天后的活力相比略有下降或没有变化。通过显微镜和 LDH 测定分析细胞形态和细胞膜完整性，证实了通过 CellTiter-Blue 测定获得的细胞活力结果。出乎意料的是，用二硫代苯甲酸酯端基的 P(HPMA)（在 1000 μM 时）在三种细胞系中均在 24 小时后表现出高细胞毒性。对各种 P(HPMA)样品的进一步研究表明，在同一时间段内，三硫代碳酸酯端基和 HPMA 封端的 P(HPMA)在相同浓度下无毒性。此外，低浓度（≤200 μM）的二硫代苯甲酸酯端基的 P(HPMA)可以被测试的细胞耐受。
