JEM期刊氯膦酸盐脂质体清除肺泡巨噬细胞专业论文
时间：2024-11-20 阅读：206
中文摘要：
肺泡巨噬细胞 （AM） 是专门的组织驻留巨噬细胞，可在过敏性炎症和哮喘中协调免疫反应。然而，是什么信号指示 AM 与其他免疫细胞进行串扰仍不清楚。在这里，我们报道了自分泌运动因子受体 （AMFR），一种内质网驻留的 E3 泛素连接酶，在哮喘的 AM 中上调，对这种情况至关重要。AMFR 缺乏显著降低过敏诱导的辅助性 T 细胞 2 （Th2） 和嗜酸性粒细胞炎症，AM 中粒细胞-巨噬细胞集落刺激因子 （GM-CSF） 的产生较少。从机制上讲，在胸腺基质淋巴细胞生成素 （TSLP） 刺激后，AMFR 与细胞因子诱导的含 SH2 的蛋白 （CIS） 直接相关，诱导 CIS 的 Lys48 连接的多泛素化的泛素化，从而阻断了 CIS 对信号转导和转录激活因子 5 （STAT5） 磷酸化和 AM 下游途径激活的抑制作用。总之，我们的结果表明，AMFR 通过调节 AM 功能在促进哮喘炎症中起关键作用，并可能成为哮喘治疗的新潜在药物靶点。
英文摘要：
Alveolar macrophages (AMs) are specialized tissue-resident macrophages that orchestrate the immune response in allergic inflammation and asthma. However, what signals direct AMs to cross talk with other immune cells remains unclear. Here, we report that autocrine motility factor receptor (AMFR), an endoplasmic reticulum–resident E3 ubiquitin ligase, is upregulated in AMs of asthma and is critical for this condition. AMFR deficiency significantly decreased allergy-induced T helper 2 (Th2) and eosinophilic inflammation, with less granulocyte-macrophage colony-stimulating factor (GM-CSF) production in AMs. Mechanistically, following thymic stromal lymphopoietin (TSLP) stimulation, AMFR associated directly with cytokine-inducible SH2-containing protein (CIS), induced the ubiquitination of Lys48-linked polyubiquitination of CIS, and consequently blocked the inhibitory effect of CIS on signal transducer and activator of transcription 5 (STAT5) phosphorylation and the downstream pathway activation in AMs. In conclusion, our results demonstrate that AMFR serves a crucial role in promoting inflammation in asthma through regulating AM function, and may emerge as a new potential drug target for asthma therapy.
论文信息：
论文题目： AMFR drives allergic asthma development by promoting alveolar macrophage–derived GM-CSF production
期刊名称：JEM- J Exp Med
时间期卷： (2022) 219 (5): e20211828
在线时间：2022年3月25日
DOI： doi.org/10.1084/jem.20211828
Liposoma巨噬细胞清除剂氯膦酸盐脂质体Clodronate Liposomes见刊于JEM：
Liposoma巨噬细胞清除剂氯膦酸盐脂质体Clodronate Liposomes的材料和方法：
JEM期刊率膦酸盐脂质体清除肺泡巨噬细胞专业论文：肺泡巨噬细胞清除解决方案
Reagents
OVA (A5503), papain (76216), chitin (C9752), collagenase D (11088866001), and DNase I (10104159001) were obtained from Sigma-Aldrich. The Imject Alum adjuvant (77161) and ER-TrackerTM Blue-White DPX (E12353) were purchased from Thermo Fisher Scientific. Clodronate liposomes (CP-005-005) were purchased from Liposoma. The recombinant murine (555-TS) and human (1398-TS) TSLP cytokines were purchased from R&D. Recombinant murine GM-CSF (315-03) and M-CSF (315-02) were from PeproTech. The anti-AMFR (ab76841) antibody was obtained from Abcam. The anti-CIS antibody (sc-166326) was obtained from Santa Cruz Biotechnology. The anti-CD68 antibody (14-0681-80) was purchased from Invitrogen Thermo Fisher Scientific. Antibodies for Myc-Tag (2272S and 2276S), Flag-Tag (14793S), HA-Tag (3724S), β-actin (8457S), Ub (3936S), STAT5 (94205S), phospho-STAT5 (9351L), phospho-JAK1 (74129T), JAK1 (3344T), phospho-JAK2 (8082T), JAK2, (3230T), SOCS1 (3950T), SOCS2 (2779P), SOCS3 (2932P), Alexa Fluor 594 anti-mouse IgG (8890S), and Alexa Fluor 488 anti-rabbit IgG (4412S) were obtained from Cell Signaling Technology. The secondary antibodies peroxidase-conjugated anti-rabbit (111-035-003) and anti-mouse (115-035-003) were purchased from Jackson ImmunoResearch Laboratories. The flow cytometry antibodies, including APC anti-mouse CD11c (117310), FITC anti-mouse Siglec-F (155504), PE anti-mouse Siglec-F (155506), APC anti-mouse/human CD11b (101212), PE/Cyanine7 anti-mouse CD45 (103114), PerCP/Cyanine5.5 anti-mouse CD64 (139307), APC/Fire 750 anti-mouse Ly-6G (127652), Brilliant Violet 650 anti-mouse F4/80 (123149), Brilliant Violet 421 anti-mouse/human CD11b (101235), FITC anti-mouse I-A/I-E (MHC class II; 107605), PE anti-human GM-CSF (502305), and PE/Cyanine7 anti-mouse GM-CSF (505411), were from BioLegend. The BCA protein assay kit (P0012S) and DAPI (C1002) were obtained from Beyotime.
AM adoptive transfer
Adoptive transfer of AMs was performed as previously reported (Miki et al., 2021; Qian et al., 2015). For in vivo deletion of macrophages in lung tissues, mice were sensitized with OVA as described above and treated with 40 μl of clodronate liposome i.t. for two successive days (days 18 and 19). For the AM adoptive transfer study, AMs derived from WT or AMFR knockout mice were then transferred by i.t. injection into the lungs of clodronate liposome-treated and OVA-sensitized WT mice at a density of 5 × 105 cells/mouse (40 μl) on day 20. 24 h after AM delivery, the mice were i.t. challenged with OVA for three days (days 21, 22, and 23). On day 25, the mice were sacrificed to analyze allergic asthmatic inflammation (Fig. 3 A).