School of Biomedical Sciences, Faculty of Health, Queensland University of Technology (QUT), Brisbane, Australia.
Australian Prostate Cancer Research Centre - Queensland (APCCRC-Q) and Queensland Bladder Cancer Initiative (QBCI), Brisbane, Queensland, Australia.
Stem Cell Res Ther. 2021 Apr 12;12(1):231. doi: 10.1186/s13287-021-02297-7.
Direct bone marrow injection of cells into murine marrow cavities is used in a range of cell characterization assays and to develop disease models. While human bone marrow-derived stromal cells (hBMSC, also known as mesenchymal stem cells (MSC)) are frequently described in therapeutic applications, or disease modeling, their behavior following direct injection into murine bone marrow is poorly characterized. Herein, we characterized hBMSC engraftment and persistence within the bone marrow of NOD-scid interleukin (IL)-2γ (NSG) mice with or without prior 2 Gy total-body γ-irradiation of recipient mice.
One day after conditioning NSG mice with sublethal irradiation, 5 × 10 luciferase (Luc) and green fluorescent protein (GFP)-expressing hBMSC (hBMSC-Luc/GFP) were injected into the right femurs of animals. hBMSC-Luc/GFP were tracked in live animals using IVIS imaging, and histology was used to further characterize hBMSC location and behavior in tissues.
hBMSC-Luc/GFP number within injected marrow cavities declined rapidly over 4 weeks, but prior irradiation of animals delayed this decline. At 4 weeks, hBMSC-Luc/GFP colonized injected marrow cavities and distal marrow cavities at rates of 2.5 ± 2.2% and 1.7 ± 1.9% of total marrow nucleated cells, respectively in both irradiated and non-irradiated mice. In distal marrow cavities, hBMSC were not uniformly distributed and appeared to be co-localized in clusters, with the majority found in the endosteal region.
While significant numbers of hBMSC-Luc/GFP could be deposited into the mouse bone marrow via direct bone marrow injection, IVIS imaging indicated that the number of hBMSC-Luc/GFP in that bone marrow cavity declined with time. Irradiation of mice prior to transplant only delayed the rate of hBMSC-Luc/GFP population decline in injected femurs. Clusters of hBMSC-Luc/GFP were observed in the histology of distal marrow cavities, suggesting that some transplanted cells actively homed to distal marrow cavities. Individual cell clusters may have arisen from discrete clones that homed to the marrow, and then underwent modest proliferation. The transient high-density population of hBMSC within the injected femur, or the longer-term low-density population of hBMSC in distal marrow cavities, offers useful models for studying disease or regenerative processes. Experimental designs should consider how relative hBMSC distribution and local hBMSC densities evolve over time.
直接将细胞注入骨髓腔是用于一系列细胞特征分析和疾病模型建立的方法。虽然人骨髓基质细胞（hBMSC，也称为间充质干细胞（MSC））在治疗应用或疾病建模中经常被描述，但它们在直接注入鼠骨髓后的行为特征还没有得到很好的描述。在此，我们描述了 hBMSC 在 NOD-scid 白细胞介素（IL）-2γ（NSG）小鼠骨髓中的植入和持续存在情况，这些小鼠在接受小鼠之前接受了亚致死剂量的全身γ照射或不接受照射。
在使用亚致死剂量照射使 NSG 小鼠接受预处理的一天后，将 5×10 个荧光素酶（Luc）和绿色荧光蛋白（GFP）表达的 hBMSC（hBMSC-Luc/GFP）注入动物的右侧股骨。使用 IVIS 成像在活体动物中追踪 hBMSC-Luc/GFP，并通过组织学进一步描述 hBMSC 在组织中的位置和行为。
在 4 周内，注入骨髓腔中的 hBMSC-Luc/GFP 数量迅速下降，但动物的预先照射延迟了这种下降。在 4 周时，hBMSC-Luc/GFP 定植于注入的骨髓腔和远端骨髓腔的比率分别为照射和未照射小鼠的总骨髓有核细胞的 2.5±2.2%和 1.7±1.9%。在远端骨髓腔中，hBMSC 没有均匀分布，似乎聚集在一起，大多数位于骨内膜区域。
虽然可以通过直接骨髓注射将大量 hBMSC-Luc/GFP 注入到小鼠骨髓中，但 IVIS 成像表明，骨髓腔中的 hBMSC-Luc/GFP 数量随时间而减少。在移植前对小鼠进行照射仅延迟了注入股骨中 hBMSC-Luc/GFP 群体减少的速度。在组织学上，在远端骨髓腔中观察到 hBMSC-Luc/GFP 簇，这表明一些移植细胞主动归巢到远端骨髓腔。单个细胞簇可能来自归巢到骨髓的离散克隆，然后经历适度增殖。在注入股骨中的 hBMSC 的短暂高密度群体，或在远端骨髓腔中的长期低密度群体，为研究疾病或再生过程提供了有用的模型。实验设计应考虑相对 hBMSC 分布和局部 hBMSC 密度如何随时间演变。
