Attenuation of neutrophil adhesion and formation of neutrophil extracellular traps by pooled human immune globulins.

IF 4.4 2区 医学 Q1 PHARMACOLOGY & PHARMACY
Frontiers in Pharmacology Pub Date : 2024-12-12 eCollection Date: 2024-01-01 DOI:10.3389/fphar.2024.1465776
Vidhya R Rao, Sana Iqbal, Bradford A Young, Christine Mun, Sandeep Jain, Simon Kaja
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引用次数: 0

Abstract

Introduction: This study investigated the efficacy of pooled human immune globulins (Flebogamma® DIF) to combat the formation of neutrophil extracellular traps (NETs) and NETosis, along with neutrophil adhesion to corneal epithelial cells in response to dry eye disease relevant stimuli.

Methods: Human neutrophils were isolated by bead-based immunomagnetic depletion of non-target cells from human whole blood. NETosis was induced using phorbol 12-myristate 13-acetate (PMA) or anti-citrullinated histone 4 R3 antibody (H4R3 ACPA). Extracellular DNA was used as a surrogate biomarker of NETosis, and it was quantified using a 96-well, plate reader-based fluorescent assay and by confocal microscopy in 8-well chambers using the DNA dye, SYTOXTM Green. Neutrophils were labeled with calcein-AM and adhesion to human corneal epithelial cells was measured. The efficacy of a dose-range of pooled human immune globulin (Flebogamma® DIF, 0.01%-5%) was tested in all assays.

Results: Pooled human immune globulins (Flebogamma® DIF) dose-dependently inhibited both PMA and H4R3 ACPA induced NETosis, with concentrations ≥2.5% fully preventing release of extracellular DNA over a 2-16 h time period. Similarly, Flebogamma® 5% DIF prevented NETosis against PMA (20 nM) and a dose range (0.1-10 μg/mL) of H4R3 ACPA. Both PMA and H4R3 ACPA increased adhesion of neutrophils to corneal epithelial cells by 20% and 5%, respectively. Flebogamma® DIF treatment resulted in a dose-dependent reduction of neutrophil adhesion, with Flebogamma® 5% DIF reducing adhesion to baseline levels.

Discussion: These findings show the dose-dependent efficacy of pooled human immune globulins, specifically Flebogamma® DIF against experimentally and pathologically induced NETosis and neutrophil adhesion to corneal epithelial cells, in vitro. The results from this study support the continued clinical development of Flebogamma® 5% DIF as a novel and efficacious treatment for the signs and symptoms of dry eye disease.

人类免疫球蛋白对中性粒细胞粘附的衰减和中性粒细胞胞外陷阱的形成。
本研究探讨了混合人免疫球蛋白(Flebogamma®DIF)对抗中性粒细胞细胞外陷阱(NETs)和NETosis的形成,以及干眼病相关刺激下中性粒细胞粘附角膜上皮细胞的功效。方法:采用磁珠法从人全血中分离非靶细胞,分离中性粒细胞。采用12-肉豆蔻酸酯(PMA)或抗瓜氨酸化组蛋白4r3抗体(H4R3 ACPA)诱导NETosis。细胞外DNA被用作NETosis的替代生物标志物,使用96孔、基于平板读码器的荧光分析和8孔共聚焦显微镜,使用DNA染料SYTOXTM Green进行定量。用钙黄蛋白am标记中性粒细胞,测定其与人角膜上皮细胞的粘附性。所有试验均检测了混合人免疫球蛋白(Flebogamma®DIF, 0.01%-5%)剂量范围的疗效。结果:混合人免疫球蛋白(Flebogamma®DIF)剂量依赖性地抑制PMA和H4R3 ACPA诱导的NETosis,浓度≥2.5%在2-16小时内完全阻止细胞外DNA的释放。同样,Flebogamma®5% DIF可阻止NETosis抗PMA (20 nM)和剂量范围(0.1-10 μg/mL)的H4R3 ACPA。PMA和H4R3 ACPA分别增加了中性粒细胞对角膜上皮细胞的粘附20%和5%。Flebogamma®DIF治疗导致中性粒细胞粘附的剂量依赖性降低,Flebogamma®5% DIF可将粘附降低至基线水平。讨论:这些发现表明,在体外,混合人免疫球蛋白,特别是Flebogamma®DIF,对实验和病理诱导的NETosis和中性粒细胞粘附角膜上皮细胞具有剂量依赖性的功效。这项研究的结果支持Flebogamma®5% DIF作为干眼病体征和症状的新型有效治疗方法的持续临床开发。
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来源期刊
Frontiers in Pharmacology
Frontiers in Pharmacology PHARMACOLOGY & PHARMACY-
CiteScore
7.80
自引率
8.90%
发文量
5163
审稿时长
14 weeks
期刊介绍: Frontiers in Pharmacology is a leading journal in its field, publishing rigorously peer-reviewed research across disciplines, including basic and clinical pharmacology, medicinal chemistry, pharmacy and toxicology. Field Chief Editor Heike Wulff at UC Davis is supported by an outstanding Editorial Board of international researchers. This multidisciplinary open-access journal is at the forefront of disseminating and communicating scientific knowledge and impactful discoveries to researchers, academics, clinicians and the public worldwide.
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