PO.TB04.02 · 肿瘤生物学
新型三重工程化 NCG-hIL6-mTSLP-mFlt3-KO 小鼠模型中人类免疫系统的植入增强
Enhanced human immune system engraftment in a novel triple-engineered NCG-hIL6-mTSLP-mFlt3-KOmouse model
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
稳健的人源化小鼠模型对于评估人类造血干细胞(HSC)生物学、免疫系统发育及治疗疗效至关重要。NCG 品系因其严重免疫缺陷,为人类 HSC 植入提供了卓越的平台。为进一步增强人类免疫系统重建,我们通过以下方式构建了新一代品系 NCG-mTSLP-hIL6-mFlt3 KO:1. 敲入人类 IL6(hIL6)以支持人类髓系细胞的存活与炎症反应;2. 转基因(Tg)表达小鼠 TSLP(mTSLP)以增强人类 T 细胞和 B 细胞的发育及淋巴器官形成;3. 敲除(KO)小鼠 Flt3(mFlt3)以消除对 FLT3 配体(FLT3L)的细胞因子竞争,并促进表达 FLT3 的人类树突状细胞(DC)和自然杀伤(NK)细胞的扩增。将人类 CD34+ HSC 移植入 NCG 和 NCG-mTSLP-hIL6-mFlt3 KO 小鼠。通过流式细胞术对小鼠外周血中人类免疫细胞的重建进行纵向监测(直至移植后 18 周)。通过 ELISA 定量检测人类免疫球蛋白(IgG 和 IgM)水平。选定的 HSC-NCG-mTSLP-hIL6-mFlt3 KO 小鼠队列在移植后 18 周接受重组人类 FLT3L 蛋白的重复腹腔注射。对淋巴组织发育进行解剖学评估。与 HSC-NCG 对照相比,HSC-NCG-mTSLP-hIL6-mFlt3 KO 小鼠表现出更佳的人类免疫细胞植入与重建。流式细胞术显示,在多个时间点外周血中人类 T 细胞、NK 细胞、浆细胞样 DC(pDC)和常规 DC(cDC)的水平显著更高。此外,通过人类 IgG 和 IgM 血清水平升高证实了功能性人类 B 细胞的成熟。解剖学分析显示各类淋巴结(抗原呈递和适应性免疫启动的关键部位)的发育更为完善。外源性人类 FLT3L 的给药进一步放大了人类 NK 细胞、pDC 和 cDC 的重建水平,验证了对人类细胞因子信号的响应性。NCG-mTSLP-hIL6-mFlt3 KO 小鼠模型为人类 HSC 植入及功能性、多谱系人类免疫系统(包括淋巴系[T、B]、髓系[DC 亚群]和 NK 细胞)的发育提供了强大平台。结合增强的淋巴器官发育,该模型有望成为转化免疫学和免疫疗法临床前评估的宝贵工具。
查看英文原文 English abstract
Robust humanized mouse models are critical for evaluating human hematopoietic stem cell (HSC) biology, immune system development, and therapeutic efficacy. The NCG strain provides a superior platform for human HSC engraftment due to severe immunodeficiency. To further enhance human immune system reconstitution, we engineered a next generation strain NCG-mTSLP-hIL6-mFlt3 KO by: 1. knocking in human IL6 (hIL6) to support human myeloid cell survival and inflammation, 2. transgenic (Tg) expression of mouse TSLP (mTSLP) to enhance human T and B cell development and lymphoid organogenesis, and 3. knocked-out (KO) mouse Flt3 (mFlt3) to eliminate cytokine competition for FLT3 ligand (FLT3L) and promote the expansion of FLT3-expressing human dendritic cells (DCs) and natural killer (NK) cells. Human CD34+ HSCs were transplanted into NCG and NCG-mTSLP-hIL6-mFlt3 KO mice. Human immune cell reconstitution was longitudinally monitored in the peripheral blood of mice by flow cytometry (up to 18 weeks post-transplantation). Human immunoglobulin (IgG and IgM) levels were quantified by ELISA. Selected cohorts of HSC-NCG-mTSLP-hIL6-mFlt3 KO mice received repeated intraperitoneal injections of recombinant human FLT3L protein at 18 weeks post-transplantation. Lymphoid tissue development was assessed anatomically. HSC-NCG-mTSLP-hIL6-mFlt3 KO mice exhibited improved engraftment and reconstitution of human immune cells compared to HSC-NCG controls. Flow cytometry revealed significantly higher levels of human T cells, NK cells, plasmacytoid DCs (pDCs), and conventional DCs (cDCs) in peripheral blood at multiple time points. Furthermore, functional human B cell maturation was confirmed by elevated serum levels of human IgG and IgM. Anatomical analysis revealed advanced development of various lymph nodes, which are key sites for antigen presentation and the initiation of adaptive immunity. Administration of exogenous human FLT3L further amplified reconstitution levels of human NK cells, pDCs, and cDCs, validating the responsiveness to human cytokine signaling. The NCG-mTSLP-hIL6-mFlt3 KO mouse model offers a powerful platform for human HSC engraftment and development of a functional, multi-lineage human immune system, including lymphoid (T, B), myeloid (DC subsets), and NK cells. Together with enhanced lymphoid organ development, this model is positioned to be a valuable tool for translational immunology and preclinical evaluation of immunotherapies.
利益披露 Disclosure
H. Sun, None..
J. Xing, None..
S. Guo, None..
Y. Zhang, None..
H. Yang, None..
X. Gao, None.