Centre for Human Development, Stem Cells and Regenerative Medicine, Bone and Joint Research group, University of Southampton, United Kingdom; Bioengineering Sciences Group, Institute for Life Sciences, University of Southampton, United Kingdom.
Bioengineering Sciences Group, Institute for Life Sciences, University of Southampton, United Kingdom.
Biochim Biophys Acta Gen Subj. 2023 Dec;1867(12):130481. doi: 10.1016/j.bbagen.2023.130481. Epub 2023 Oct 5.
Ultrasound-responsive microbubbles offer a means of achieving minimally invasive, localised drug delivery in applications including regenerative medicine. To facilitate their use, however, it is important to determine any cytotoxic effects they or their constituents may have. The aim of this study was to test the hypothesis that phospholipid-shelled microbubbles are non-toxic to human bone-derived cells at biologically-relevant concentrations.
Microbubbles were fabricated using combinations of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dibehenoyl-sn-glycero-3-phosphocholine (DBPC), polyoxyethylene(40) stearate (PEG40S) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene-glycol)-2000] (DSPE-PEG). Microbubble size and concentration were measured as a function of time and temperature by optical microscopy. Effects on MG63 osteosarcoma and human bone marrow stromal cells (BMSCs) were measured for up to 72 h by assay for viability, metabolic activity and proliferation.
DBPC:DSPE-PEG microbubbles were significantly more stable than DSPC:PEG40S microbubbles under all conditions tested. Serum-containing medium had no detrimental effect on microbubble stability, but storage at 37 °C compared to at 4 °C reduced stability for both preparations, with almost complete dissolution of microbubbles at times ≥24 h. DSPC:PEG40S microbubbles had greater inhibitory effects on cell metabolism and growth than DBPC:DSPE-PEG microbubbles, with PEG40S found to be the principle inhibitory component. These effects were only evident at high microbubble concentrations (≥20% (v/v)) or with prolonged culture (≥24 h). Increasing cell-microbubble contact by inversion culture in a custom-built device had no inhibitory effect on metabolism.
These data indicate that, over a broad range of concentrations and incubation times, DBPC:DSPE-PEG and DSPC:PEG40S microbubbles have little effect on osteoblastic cell viability and growth, and that PEG40S is the principle inhibitory component in the formulations investigated.
超声响应微泡为实现再生医学等应用中的微创、局部药物输送提供了一种手段。然而，为了方便使用，确定它们或其成分可能具有的任何细胞毒性作用非常重要。本研究旨在验证以下假设：在生物相关浓度下，磷脂壳微泡对人源性骨细胞无毒。
使用 1,2-二硬脂酰-sn-甘油-3-磷酸胆碱（DSPC）、1,2-二油酰基-sn-甘油-3-磷酸胆碱（DBPC）、聚氧乙烯（40）硬脂酸酯（PEG40S）和 1,2-二硬脂酰-sn-甘油-3-磷酸乙醇胺-N-[甲氧基（聚乙二醇）-2000]（DSPE-PEG）组合来制备微泡。通过光学显微镜测量微泡的大小和浓度随时间和温度的变化。通过活力、代谢活性和增殖测定，在长达 72 小时内，检测 MG63 骨肉瘤细胞和人骨髓基质细胞（BMSCs）的变化。
在所有测试条件下，DBPC:DSPE-PEG 微泡均明显比 DSPC:PEG40S 微泡稳定。含血清的培养基对微泡稳定性没有不利影响，但与 4°C 相比，37°C 下储存会降低两种制剂的稳定性，微泡在≥24 小时时几乎完全溶解。DSPC:PEG40S 微泡对细胞代谢和生长的抑制作用大于 DBPC:DSPE-PEG 微泡，发现 PEG40S 是主要的抑制成分。这些作用仅在高微泡浓度（≥20%（v/v））或长时间培养（≥24 小时）时才明显。通过定制设备中的倒置培养增加细胞-微泡接触，对代谢没有抑制作用。
这些数据表明，在广泛的浓度和孵育时间范围内，DBPC:DSPE-PEG 和 DSPC:PEG40S 微泡对成骨细胞活力和生长几乎没有影响，PEG40S 是所研究制剂中的主要抑制成分。
