促血小板生成素模拟物刺激骨髓血管和基质龛以减轻急性放射综合征。

BACKGROUND: Acute radiation syndrome (ARS) manifests after exposure to high doses of radiation in the instances of radiologic accidents or incidents. Facilitating regeneration of the bone marrow (BM), namely the hematopoietic stem and progenitor cells (HSPCs), is key in mitigating ARS and multi-organ failure. JNJ-26366821, a PEGylated thrombopoietin mimetic (TPOm) peptide, has been shown as an effective medical countermeasure (MCM) to treat hematopoietic-ARS (H-ARS) in mice. However, the activity of TPOm on regulating BM vascular and stromal niches to support HSPC regeneration has yet to be elucidated. METHODS: C57BL/6J mice (9-14 weeks old) received sublethal or lethal total body irradiation (TBI), a model for H-ARS, by Cs or X-rays. At 24 h post-irradiation, mice were subcutaneously injected with a single dose of TPOm (0.3 mg/kg or 1.0 mg/kg) or PBS (vehicle). At homeostasis and on days 4, 7, 10, 14, 18, and 21 post-TBI with and without TPOm treatment, BM was harvested for histology, BM flow cytometry of HSPCs, endothelial (EC) and mesenchymal stromal cells (MSC), and whole-mount confocal microscopy. For survival, irradiated mice were monitored and weighed for 30 days. Lastly, BM triple negative cells (TNC; CD45, TER-119, CD31) were sorted for single-cell RNA-sequencing to examine transcriptomics after TBI with or without TPOm treatment. RESULTS: At homeostasis, TPOm expanded the number of circulating platelets and HSPCs, ECs, and MSCs in the BM. Following sublethal TBI, TPOm improved BM architecture and promoted recovery of HSPCs, ECs, and MSCs. Furthermore, TPOm elevated VEGF-C levels in normal and irradiated mice. Following lethal irradiation, mice improved body weight recovery and 30-day survival when treated with TPOm after Cs and X-ray exposure. Additionally, TPOm reduced vascular dilation and permeability. Finally, single-cell RNA-seq analysis indicated that TPOm increased the expression of collagens in MSCs to enhance their interaction with other progenitors in BM and upregulated the regeneration pathway in MSCs. CONCLUSIONS: TPOm interacts with BM vascular and stromal niches to locally support hematopoietic reconstitution and systemically improve survival in mice after TBI. Therefore, this work warrants the development of TPOm as a potent radiation MCM for the treatment of ARS.

背景：急性辐射综合征（ARS）是在放射事故或放射事件中暴露于高剂量辐射后出现的。促进骨髓（BM）的再生，即造血干细胞和祖细胞（HSPCs），是减轻 ARS 和多器官衰竭的关键。JNJ-26366821 是一种聚乙二醇化血小板生成素模拟物（TPOm）肽，已被证明是治疗小鼠造血性 ARS（H-ARS）的有效医疗对策（MCM）。然而，TPOm 调节 BM 血管和基质龛以支持 HSPC 再生的活性尚未阐明。

方法：C57BL/6J 小鼠（9-14 周龄）接受 Cs 或 X 射线亚致死或致死全身照射（TBI），建立 H-ARS 模型。照射后 24 小时，小鼠皮下注射单次剂量的 TPOm（0.3mg/kg 或 1.0mg/kg）或 PBS（载体）。在 TBI 后和没有 TPOm 治疗的情况下，在第 4、7、10、14、18 和 21 天的平衡期和平衡期，采集 BM 进行组织学、BM 造血干细胞、内皮（EC）和间充质基质细胞（MSC）的流式细胞术以及全层共聚焦显微镜检查。为了生存，用辐射监测和称重小鼠 30 天。最后，对 BM 三阴性细胞（TNC；CD45、TER-119、CD31）进行单细胞 RNA 测序，以检查 TBI 后有无 TPOm 治疗的转录组学。

结果：在平衡期，TPOm 增加了循环血小板和 HSPCs、ECs 和 MSC 的数量。亚致死性 TBI 后，TPOm 改善了 BM 结构，促进了 HSPCs、ECs 和 MSC 的恢复。此外，TPOm 提高了正常和照射小鼠的 VEGF-C 水平。在致死性照射后，用 TPOm 治疗 Cs 和 X 射线照射后的小鼠，体重恢复和 30 天存活率提高。此外，TPOm 减少了血管扩张和通透性。最后，单细胞 RNA-seq 分析表明，TPOm 增加了 MSC 中胶原蛋白的表达，以增强其与 BM 中其他祖细胞的相互作用，并上调了 MSC 中的再生途径。

结论：TPOm 与 BM 血管和基质龛相互作用，局部支持造血重建，系统改善 TBI 后小鼠的生存。因此，这项工作证明了 TPOm 作为一种有效的辐射 MCM，可用于治疗 ARS。