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Abstract：
BACKGROUND: Hematoma leads to progressive neurological deficits and poor outcomes after intracerebral hemorrhage (ICH). Early clearance of hematoma is widely recognized as an essential treatment to limit the damage and improve the clinical prognosis. CD163, alias hemoglobin (Hb) scavenger receptor on microglia, plays a pivotal role in hematoma absorption, but CD163 on neurons permits Hb uptake and results in neurotoxicity. In this study, we focus on how to specially promote microglial but not neuronal CD163 mediated-Hb uptake and hematoma absorption.
METHODS: RNA sequencing was used to explore the potential molecules involved in ICH progression, and hematoma was detected by magnetic resonance imaging (MRI). Western blot and immunofluorescence were used to evaluate the expression and location of fractalkine (FKN) after ICH. Erythrophagocytosis assay was performed to study the specific mechanism of action of FKN in hematoma clearance. Small interfering RNA (siRNA) transfection was used to explore the effect of peroxisome proliferator-activated receptor-γ (PPAR-γ) on hematoma absorption. Enzyme-linked immunosorbent assay (ELISA) was used to determine the serum FKN concentration in ICH patients.
RESULTS: FKN was found to be significantly increased around the hematoma in a mouse model after ICH. With its unique receptor CX3CR1 in microglia, FKN significantly decreased the hematoma size and Hb content, and improved neurological deficits in vivo. Further, FKN could enhance erythrophagocytosis of microglia in vitro via the CD163/ hemeoxygenase-1 (HO-1) axis, while AZD8797 (a specific CX3CR1 inhibitor) reversed this effect. Moreover, PPAR-γ was found to mediate the increase in the CD163/HO-1 axis expression and erythrophagocytosis induced by FKN in microglia. Of note, a higher serum FKN level was found to be associated with better hematoma resolution in ICH patients.
CONCLUSIONS: We systematically identified that FKN may be a potential therapeutic target to improve hematoma absorption and we shed light on ICH treatment.
METHODS: RNA sequencing was used to explore the potential molecules involved in ICH progression, and hematoma was detected by magnetic resonance imaging (MRI). Western blot and immunofluorescence were used to evaluate the expression and location of fractalkine (FKN) after ICH. Erythrophagocytosis assay was performed to study the specific mechanism of action of FKN in hematoma clearance. Small interfering RNA (siRNA) transfection was used to explore the effect of peroxisome proliferator-activated receptor-γ (PPAR-γ) on hematoma absorption. Enzyme-linked immunosorbent assay (ELISA) was used to determine the serum FKN concentration in ICH patients.
RESULTS: FKN was found to be significantly increased around the hematoma in a mouse model after ICH. With its unique receptor CX3CR1 in microglia, FKN significantly decreased the hematoma size and Hb content, and improved neurological deficits in vivo. Further, FKN could enhance erythrophagocytosis of microglia in vitro via the CD163/ hemeoxygenase-1 (HO-1) axis, while AZD8797 (a specific CX3CR1 inhibitor) reversed this effect. Moreover, PPAR-γ was found to mediate the increase in the CD163/HO-1 axis expression and erythrophagocytosis induced by FKN in microglia. Of note, a higher serum FKN level was found to be associated with better hematoma resolution in ICH patients.
CONCLUSIONS: We systematically identified that FKN may be a potential therapeutic target to improve hematoma absorption and we shed light on ICH treatment.
摘要：
背景：脑出血（ICH）后血肿导致进行性神经功能缺损和不良预后。早期清除血肿被广泛认为是限制损伤和改善临床预后的必要治疗方法。小胶质细胞上的CD163，别名血红蛋白（Hb）清道夫受体，在血肿吸收中起着关键作用，但是神经元上的CD163允许Hb摄取并导致神经毒性。在这项研究中,我们关注如何特别促进小胶质细胞而不是神经元CD163介导的Hb摄取和血肿吸收。
方法：RNA测序用于探索参与ICH进展的潜在分子，并通过磁共振成像（MRI）检测到血肿。应用Westernblot和免疫荧光技术评价脑出血后fractalkine(FKN)的表达和定位。通过红细胞吞噬试验研究FKN在血肿清除中的具体作用机制。利用小干扰RNA(siRNA)转染,探讨过氧化物酶体增殖物激活受体-γ(PPAR-γ)对血肿吸收的影响。采用酶联免疫吸附试验(ELISA)测定ICH患者血清FKN浓度。
结果：在ICH后的小鼠模型中，发现血肿周围的FKN明显增加。以其在小胶质细胞中的独特受体CX3CR1，FKN显著降低血肿大小和Hb含量,和改善体内神经功能缺损。Further,FKN可以通过CD163/血红素加氧酶-1（HO-1）轴增强体外小胶质细胞的红细胞吞噬作用，而AZD8797（一种特定的CX3CR1抑制剂）逆转了这种作用。此外，发现PPAR-γ介导小胶质细胞中FKN诱导的CD163/HO-1轴表达和红细胞吞噬作用的增加。值得注意的是，在ICH患者中,较高的血清FKN水平与更好的血肿消退相关.
结论：我们系统确定FKN可能是改善血肿吸收的潜在治疗靶点，并阐明ICH治疗。
方法：RNA测序用于探索参与ICH进展的潜在分子，并通过磁共振成像（MRI）检测到血肿。应用Westernblot和免疫荧光技术评价脑出血后fractalkine(FKN)的表达和定位。通过红细胞吞噬试验研究FKN在血肿清除中的具体作用机制。利用小干扰RNA(siRNA)转染,探讨过氧化物酶体增殖物激活受体-γ(PPAR-γ)对血肿吸收的影响。采用酶联免疫吸附试验(ELISA)测定ICH患者血清FKN浓度。
结果：在ICH后的小鼠模型中，发现血肿周围的FKN明显增加。以其在小胶质细胞中的独特受体CX3CR1，FKN显著降低血肿大小和Hb含量,和改善体内神经功能缺损。Further,FKN可以通过CD163/血红素加氧酶-1（HO-1）轴增强体外小胶质细胞的红细胞吞噬作用，而AZD8797（一种特定的CX3CR1抑制剂）逆转了这种作用。此外，发现PPAR-γ介导小胶质细胞中FKN诱导的CD163/HO-1轴表达和红细胞吞噬作用的增加。值得注意的是，在ICH患者中,较高的血清FKN水平与更好的血肿消退相关.
结论：我们系统确定FKN可能是改善血肿吸收的潜在治疗靶点，并阐明ICH治疗。
