The Laboratory of Molecular Immunology, Department of Microbiology & Immunology, Rega Institute, Katholieke Universiteit Leuven, Leuven, Belgium.
PLoS One. 2011;6(8):e23913. doi: 10.1371/journal.pone.0023913. Epub 2011 Aug 31.
Posttranslational modification of chemokines is one of the mechanisms that regulate leukocyte migration during inflammation. Multiple natural NH(2)-terminally truncated forms of the major human neutrophil attractant interleukin-8 or CXCL8 have been identified. Although differential activity was reported for some CXCL8 forms, no biological data are available for others.
METHODOLOGY/PRINCIPAL FINDINGS: Aminopeptidase-cleaved CXCL8(2-77) and CXCL8(3-77), the product of alternative cleavage of the signal peptide CXCL8(-2-77) and the previously studied forms containing 77 and 72 amino acids, CXCL8(1-77) and CXCL8(6-77), were prepared by solid-phase peptide synthesis, purified and folded into active proteins. No differences in binding and calcium signaling potency were detected between CXCL8(1-77), CXCL8(-2-77), CXCL8(2-77) and CXCL8(3-77) on cells transfected with one of the human CXCL8 receptors, i.e. CXCR1 and CXCR2. However, CXCL8(-2-77) was more potent compared to CXCL8(1-77), CXCL8(2-77) and CXCL8(3-77) in signaling and in vitro chemotaxis of peripheral blood-derived human neutrophils. Moreover, CXCL8(-2-77) was less efficiently processed by plasmin into the more potent CXCL8(6-77). The truncated forms CXCL8(2-77) and CXCL8(3-77) had higher affinity for heparin than CXCL8(1-77), a property important for the presentation of CXCL8 on endothelial layers. Upon intraperitoneal injection in mice, elongated, truncated and intact CXCL8 were equally potent to recruit neutrophils to the peritoneal cavity.
In terms of their ability to induce neutrophil recruitment in vivo, the multiple CXCL8 forms may be divided in three groups. The first group includes CXCL8 proteins consisting of 75 to 79 amino acids, cleaved by aminopeptidases, with intermediate activity on neutrophils. The second group, generated through proteolytic cleavage (e.g. by Ser proteases), contains 69 to 72 amino acid forms which are highly potent neutrophil attractants in vivo. A third category is generated through the modification of the arginine in the NH(2)-terminal region into citrulline by peptidylarginine deiminases and has weak potency to induce neutrophil extravasation.
趋化因子的翻译后修饰是调节炎症期间白细胞迁移的机制之一。已经鉴定出主要人中性粒细胞趋化因子白细胞介素-8 或 CXCL8 的多种天然 NH2-末端截断形式。虽然报道了一些 CXCL8 形式的差异活性，但其他形式没有生物学数据。
方法/主要发现：通过固相肽合成、纯化和折叠成活性蛋白来制备氨基肽酶切割的 CXCL8(2-77)和 CXCL8(3-77)、信号肽 CXCL8(-2-77)的替代切割产物和之前研究的包含 77 和 72 个氨基酸的形式，CXCL8(1-77)和 CXCL8(6-77)。在转染一种人 CXCL8 受体（即 CXCR1 和 CXCR2）的细胞上，CXCL8(1-77)、CXCL8(-2-77)、CXCL8(2-77)和 CXCL8(3-77)之间在结合和钙信号转导效力方面没有差异。然而，与 CXCL8(1-77)、CXCL8(2-77)和 CXCL8(3-77)相比，CXCL8(-2-77)在信号转导和体外趋化性方面对人外周血中性粒细胞更有效。此外，CXCL8(-2-77)被纤溶酶更有效地加工成更有效的 CXCL8(6-77)。截短形式 CXCL8(2-77)和 CXCL8(3-77)比 CXCL8(1-77)对肝素具有更高的亲和力，这一特性对于 CXCL8 在血管内皮层的呈现很重要。在小鼠的腹腔内注射中，延长的、截断的和完整的 CXCL8 同样能够将中性粒细胞募集到腹腔中。
就其在体内诱导中性粒细胞募集的能力而言，多种 CXCL8 形式可分为三组。第一组包括由氨基肽酶切割的 75 到 79 个氨基酸的 CXCL8 蛋白，对中性粒细胞具有中等活性。第二组通过蛋白水解切割（例如通过丝氨酸蛋白酶）产生，包含 69 到 72 个氨基酸的形式，在体内是高度有效的中性粒细胞趋化因子。第三类是通过肽基精氨酸脱亚氨酶将 NH2-末端区域的精氨酸修饰为瓜氨酸产生的，对诱导中性粒细胞渗出的作用较弱。
