Stem cell therapies are a promising treatment for many patients suffering from diseases with poor prognosis. However, clinical translation is inhibited by a lack of in vivo monitoring techniques to track stem cells throughout the course of treatment. Ultrasound-guided photoacoustic (PA) imaging of nanoparticle-labeled stem cells may be a solution. To allow PA tracking, stem cells must be labeled with an optically absorbing contrast agent. Gold nanoparticles are one option due to their cytocompatibility and strong optical absorption in the near-infrared region. However, stem cell labeling can require up to 24-h incubation with nanoparticles in culture before use. Although stem cell monitoring is critically needed, the additional preparation time may not be feasible-it is cost prohibitive and stem cell treatments should be readily available in emergency situations as well as scheduled procedures. To remedy this, stem cells can be labeled before freezing and long-term storage. While it is well known that stem cells retain their cellular function after freezing, storage, and thawing, the impact of gold nanoparticles on this process has yet to be investigated. Therefore, we assessed the viability, multipotency, and PA activity of gold nanosphere-labeled mesenchymal stem cells (MSCs) after freezing, storing, and thawing for 1 week, 1 month, or 2 months and compared to unlabeled, naive MSCs which were frozen, stored, and thawed at the same time points. Results indicated no substantial change in viability as assessed by the MTT assay. Differentiation, observed through adipogenesis and osteogenesis, was also comparable to controls. Finally, strong PA signals and similar PA spectral signatures remained. Further studies involving more diverse stem cell types and nanoparticles are required, but our data suggest that function and imaging properties of nanoparticle-labeled stem cells are maintained after freezing and storage, which improve translation of stem cell monitoring techniques by simplifying integration with clinical protocols. Impact statement Although stem cell tracking techniques are critically needed, stem cells must be labeled with contrast agents in advance of procedures, which is not clinically feasible due to increased procedure time. As a solution, a stock of labeled stem cells could be frozen and stored, ready for immediate use. Results showed that gold nanosphere-labeled stem cells can be frozen and stored long-term without impacting cellular function or photoacoustic imaging contrast, supporting further investigation of other contrast agents and cell types. Creating a bank of nanoparticle-labeled stem cells advances translation and scalability of stem cell tracking methods by improving integration with clinical protocols.

中文翻译：

---

冷冻对金纳米颗粒标记的间充质干细胞的影响。

对于许多患有预后差的疾病的患者，干细胞疗法是一种有前途的治疗方法。然而，由于缺乏在整个治疗过程中追踪干细胞的体内监测技术，临床翻译受到了抑制。纳米标记的干细胞的超声引导光声（PA）成像可能是一种解决方案。为了进行PA追踪，必须用光吸收性造影剂标记干细胞。金纳米颗粒是一种选择，因为它们具有细胞相容性，并且在近红外区域具有很强的光吸收能力。但是，干细胞标记在使用前可能需要与培养物中的纳米颗粒进行长达24小时的孵育。尽管迫切需要进行干细胞监测，额外的准备时间可能不可行-成本高昂，并且在紧急情况下以及计划的程序中应易于获得干细胞治疗。为了解决这个问题，可以在冷冻和长期保存之前对干细胞进行标记。众所周知，干细胞在冷冻，储存和解冻后仍能保持其细胞功能，但金纳米颗粒对该过程的影响尚待研究。因此，我们评估了冷冻，储存和解冻1周，1个月或2个月后金纳米球标记的间充质干细胞（MSC）的生存力，多能性和PA活性，并与冷冻的未标记，未加工的MSC进行了比较。 ，存储并在同一时间解冻。结果表明，通过MTT测定评估，生存力没有实质性变化。差异化 通过脂肪形成和成骨作用观察到的结果也与对照相当。最终，保留了强大的PA信号和相似的PA频谱特征。需要对涉及更多种干细胞类型和纳米颗粒的进一步研究，但我们的数据表明，纳米颗粒标记的干细胞在冷冻和保存后仍能保持其功能和成像特性，从而通过简化与临床方案的整合来改善干细胞监测技术的翻译。影响陈述尽管迫切需要干细胞跟踪技术，但干细胞必须在手术前用造影剂标记，由于增加了手术时间，因此在临床上不可行。作为一种解决方案，可以将已标记的干细胞储备液冷冻保存，以备立即使用。结果表明，金纳米球标记的干细胞可以冷冻和长期保存，而不会影响细胞功能或光声成像对比度，从而支持对其他造影剂和细胞类型的进一步研究。通过改善与临床方案的整合，创建纳米级标记的干细胞库可促进干细胞追踪方法的翻译和可扩展性。