中性粒细胞在脓毒症引起多器官功能衰竭中的作用

Neutrophils in development of multiple organ failure in sepsis

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多器官功能衰竭（ MOF ）是威胁脓毒症或系统性感染患者生命的主要原因。在英国和美国，其病死率与心肌梗死相当，甚至高于后者。尽管人体免疫系统可以抵抗微生物感染，但在重度脓毒症患者中，受损的免疫系统似乎有助于器官功能障碍的发生。本文作者认为，中性粒细胞的异常活化及其在微脉管系统内的聚集，是引起 MOF 病理表现的原因。作者进一步提出，靶向针对中性粒细胞及其与血管壁相互作用的治疗方案将成为有价值的脓毒症治疗策略。

脓毒症是一种由感染所引发的系统性炎症，严重时会引起器官功能障碍，即组织低灌注与缺氧、乳酸酸中毒、少尿或脑功能改变^[1-2]。即便立即采用抗生素进行治疗、输液支持，并应用相应技术维持器官功能，病死率仍接近 35 %^[3]。多数感染由细菌引起^[4]，并且多数患者出现与心血管系统不稳定及肾功能恶化有关的肺功能障碍^[5]。器官功能衰竭的发生机制与诸多因素有关，如内分泌^[6]与免疫系统^[7]、弥漫性血管内凝血（ DIC ）^[8]、遗传易感性^[9]，及可能发生在线粒体内的能量代谢紊乱^[10]。

脓毒症发生时，通常是细菌成分促发系统性炎症反应，细菌成分主要引起人体免疫系统细胞释放促炎症细胞因子 ― 如白细胞介素（ IL ）-1β、 IL-6 及肿瘤坏死因子（ TNF ）-α 。循环中此类细胞因子的高浓度有时预示着高的死亡风险^[11]，但抑制细胞因子活性的治疗并不能改善患者的存活率^[12]。早期的高细胞因子血症会使循环中抗炎细胞因子（如 IL-10 ）浓度发生代偿性增加，这会导致患者免疫应答能力下降（免疫麻痹）^[13]，并影响预后。这一结论源于研究发现脓毒症患者循环及组织中淋巴细胞功能受损且数目减少^[14]。

中性粒细胞对于防止细菌感染具有至关重要的作用，例如中性粒细胞减少（如化疗后）会使机体对感染及脓毒症的易感性增加。然而，中性粒细胞的大量活化则会引起组织损伤。本文对针对患者的调查研究而非大规模临床干预研究的结果进行了综述，这些结果提示重度脓毒症引起器官功能衰竭时会发生异常的中性粒细胞活化。作者假设器官功能衰竭是由于在感染及炎症环境中，中性粒细胞被过度、异常活化所引发的；并探讨了调节中性粒细胞功能是否对脓毒症患者具有治疗作用的问题。

中性粒细胞与器官功能衰竭

中性粒细胞含有大量蛋白水解酶并可快速生成活性氧，从而降解吞入的病原，因此理论上可以清除致病细菌^[15]。若浸润组织的中性粒细胞向胞外释放细胞溶解因子^[16]或促炎症因子^[17]，将引起局部损伤 ^[18-21]。实际上，尽管这种情况下大范围的组织浸润及器官损伤很罕见；但细菌感染的局部常会发生由中性粒细胞引起的组织损伤，严重时有脓肿形成。相反地，重度脓毒症时，局部感染会引起系统性中性粒细胞活化。

对 MOF 患者尸检标本的检查揭示：中性粒细胞既可聚集在肾血管局部^[22-24]，也可在肺内发生大范围组织浸润^[24]。在急性呼吸窘迫综合征（ARDS ) -可能伴发脓毒症的急性肺损伤的一种更严重的形式 ^[25]，中性粒细胞的浸润程度与肺功能损伤的程度及支气管肺泡灌洗液中中性粒细胞蛋白水解酶浓度的高低有关^[26]。

器官功能衰竭并不常引起大体形态的改变。一项研究表明，细胞死亡与器官功能衰竭无关^[27]。根据此结果，研究人员提出：细胞休眠是预防过度的系统性炎症反应的一种保护机制^[6]。这一假说并未排除微脉管系统发生中性粒细胞介导的组织损伤的可能性。采用直角偏振光谱成像可直接观察血管床，从而监测体内中性粒细胞与内皮细胞间的相互作用。然而，截至目前的多数研究均是基于动物实验，检测技术则因其分辨率不足且仅限于检测舌下及皮肤微脉管系统而无法监测脓毒症时的器官功能衰竭，因而在脓毒症患者的使用受限^[28-29]。

在系统性炎症反应过程中，微循环的稳态机制遭到破坏，包括：内皮高反应性、纤维素沉积、血管闭塞以及有时发生的进一步妨碍充分氧合的组织渗出。中性粒细胞通过增强与血管壁的黏附及形成白细胞聚集而参与血液流变学改变^[10^，^30]。表明中性粒细胞活性可能是致病原因的进一步证据源于作者的研究结果，即：采用可清除白细胞的过滤机清除系统性炎症患者血液中的中性粒细胞，可改善肺与肾的功能^[31]。另外，过滤出的中性粒细胞易于与内皮细胞发生黏附^[32]，提示中性粒细胞和血管壁间的相互作用与器官功能衰竭有关。对脓毒症动物模型的研究结果与这些临床观察结果一致。大量中性粒细胞聚集在功能衰竭的器官，一个器官出现功能衰竭将引起其他器官内更大规模的中性粒细胞活化、聚集，最终导致 MOF^[33-34^］。采用试验性干预措施，清除中性粒细胞或抑制其活性，可改善器官功能衰竭^{[ 35-36}^］。在脓毒症时，抑制中性粒细胞功能可以减轻器官功能衰竭程度，但常规建议却认为保持甚至增强中性粒细胞活性对清除初始病原是必须的，这使人们陷人了两难境地。有关脓毒症时中性粒细胞功能状态的报道相互矛盾，有关其在循环中数量的报道也不一致，因此更难以抉择了。

血中性粒细胞数量异常

在骨髓内经历 14d 的发育后，中性粒细胞暂时贮存于储备池，直至释放人血。在血液中，中性粒细胞仅用 12 - 14h 便可由循环池（轴流）进入外周池（与血管壁接触）。在无细菌感染的情况下，中性粒细胞会进入网状内皮器官（如肝脏）^[37]，或返回骨髓^[38]经历凋亡（程序性细胞死亡）^[39]。老化的中性粒细胞皱缩为凋亡小体，最终被周围的巨噬细胞吞噬，由此避免了衰老的中性粒细胞释放溶解酶而导致组织损伤的发生。

脓毒症的诊断标准包括：中性粒细胞计数高、低或包含＞ 10 ％的未成熟细胞（框表）^[1]。血中性粒细胞数量多是因为从骨髓募集的细胞过多和（或）细胞由外周池返回循环池^[40-41]。目前，尚未提出通过使增加的中性粒细胞数量降低来治疗脓毒症的策略，大概是由于人们认为细菌感染时循环中中性粒细胞数量较多总归优于过少。

1. 集落刺激因子

可刺激骨髓释放中性粒细胞的细胞因子包括粒细胞集落刺激因子（ G-CSF ）及粒细胞-巨噬细胞（ GM ）-CSF 。两种细胞因子均可使循环中的中性粒细胞数量增加，促进其成熟与活化^[42]，并延长其寿命^[43]。健康个体血中 G-CSF 的浓度很低，这与感染的急性期不同。在后一种情况下，血中 G-CSF 的浓度增加了数倍并导致中性粒细胞数量增加^[44^］。在脓毒症时，血中 G-CSF 的浓度通常增加，在中性粒细胞数量增多^[45]及呼吸爆发活力增强^[46]之前，便达到峰浓度。

脓毒症预后差与中性粒细胞减少之间的相关性导致了采用 G-CSF 或 GM-CSF 来增加中性粒细胞数量或加速其成熟的治疗方法的出现 ^[47]。采用重组 G-CSF 使中性粒细胞数恢复正常水平，对未成熟中性粒细胞比例低且内源性 G-CSF 浓度低的脓毒症患者尤为有效^[48]。但是，动物模型研究的结果与之相矛盾，影响了 G-CSF 在脓毒症患者治疗中的应用。最初的报道表明， G-CSF 可使中性粒细胞数大大增加^[49]，并可改善患者存活率^［⁵⁰^］。这一研究结果受到了另一些研究的质疑，这些研究发现： G-CSF 的应用会使血中胜粒细胞数减少，加重组织损伤及死亡率^[51]，并会加剧因使中性粒细胞数减少恢复而引发的肺损伤 ^[52]。 2 项多中心试验表明，重组 G-CSF 的应用并不能降低脓毒症患者的死亡率^［^53-54^］。

2. 白细胞减少与器官功能衰竭

中性粒细胞减少的患者会发生由中性粒细胞介导的肺损伤，这表明器官功能障碍可能是由聚集在微脉管系统中的为数不多的中性粒细胞所诱发的^［^55-56^］。相应地，对于重度脓毒症，致病的关键可能并不是血中中性粒细胞的总数，而是表型及活化状态易于诱导组织损伤的一种细胞亚型的存在^[57]。在脓毒症动物模型中，未成熟中性粒细胞最易聚集在肺部微脉管系统，活化后释放防御素（蛋白水解酶）而使组织受损^[58-59]。

因此，虽然是用血中性粒细胞计数异常来诊断系统性炎症反应的发生，但在脓毒症的发病机制中，只是中性粒细胞中的特定亚群（如易与内皮细胞相互作用的亚群）发挥作用。相关理论认为：中性粒细胞在功能上并非同源，而是由具有不同表型与分泌特征的亚群组成的^[60]。

组织渗出

对细菌的清除有赖于血中性粒细胞向感染局部迅速聚集。首先，中性粒细胞必须黏附于血管壁；然后，在化学物质（趋化因子）的介导下主动迁移至周围组织。下面，将讨论在脓毒症时中性粒细胞在各阶段的表现。

1. 黏附于血管内皮

在两种黏附分子家族（选择素与整合素）的相继作用下，中性粒细胞黏附于血管内皮。选择素促使中性粒细胞在血流切应力的作用下延内皮滚动或圈合，同时整合素使其与血管内皮紧密黏附^[61]。整合素有 2 个亚家族（β₁与β ₂ ) ，是由一条α链和一条β链构成的异二聚体，亚家族由β链决定。许多黏附分子间可发生互相转化（表 1 ）。

在中性粒细胞表面， L -选择素（ CD62L ）与内皮细胞表面糖蛋白特异性寡糖发生相互作用；同时，在内皮， E -选择素（ CD62E ）及 P -选择素（ CD62P ）均识别中性粒细胞表面的特定糖基。短暂地滚动可协助中性粒细胞与内皮细胞表面表达的炎症因子（如 IL-8 和血小板活化因子）相接触（图 1 ）。这种相互作用会使中性粒细胞表面β ₂ 整合素（ CD11a 与 CD11b ）的表达迅速上调，并增加其与内皮细胞配体 ― ICAM-l （细胞间黏附分子-1 , CD54 ) ― 的亲和力。细菌感染局部释放的各种因子（如 TNF -α、 IL-1 及脂多糖）也会增加内皮细胞表面 CD54 的表达。脓毒症患者中性粒细胞表面 L -选择素与 CDllb 的分布异常。 L -选择素的表达下调可能是由于循环中炎症介质的刺激或与血管壁的短暂接触引起细胞自溶所致^[62-63]。曾有报道称，脓毒症时中性粒细胞表面CDllb 的表达可以上调^[63-69]、下调^［^{70 -71}^］（提示预后差^[67]或正常（表 2 )^[62,72]。由于活化的中性粒细胞表型为 L -选择素下调及 CDllb 上调，那么脓毒症时，这些细胞可能已经发生了与内皮的紧密黏附（图 1 ）。但是，截至目前，尚无有关报道表明阻断脓毒症患者中性粒细胞表面 L -选择素或β ₂整合素的表达可改变中性粒细胞与内皮细胞间的黏附。本实验室研究发现：抗 CDllb 抗体并不能有效抑制脓毒症患者中性粒细胞与单层内皮的黏附，似乎细胞表面还存在其他起决定性作用的分子，参与介导脓毒症时中性粒细胞与内皮细胞间超常的黏附作用^[32]。

β ₁整合素主要与淋巴细胞及单核细胞有关^[61]。但近期，其中一个成员 VLA-4 ( CD49d ）被证实存在于脓毒症患者近 30 ％的中性粒细胞表面^[73]。 VLA-4 分子可与纤维粘连蛋白及 CD1O6 结合，因此，表达β₁_、β ₂整合素的中性粒细胞可与许多分布于血管内皮的配体发生黏附（图 1 ）。促炎症细胞因子（如 TNF -α及 IL-l ）可诱导内皮细胞表达 VCAM-1 。在脓毒症的急性肺损伤模型中，中性粒细胞的聚集与浸润涉及β₁_、β ₂整合素依赖性途径，对整合素的选择说明其各自血管配体的表达具有时间依赖性，且根据病原菌的刺激有所不同^[88-89]。理论上，脓毒症患者的中性粒细胞通过在其表面表达β₁_、β ₂整合素而适合发生组织渗出；但恰恰相反，事实上并非如此。

2. 趋化作用

在感染灶释放的趋化因子[如补体片段 C5a 、白三烯 B₄ 、血小板活化因子、细菌肽甲酰甲硫氨酰-亮氨酰-苯丙氨酸（ fMLP ）及 IL-8 ］作用下，血液中性粒细胞由趋化因子的低浓度区（如血管壁）向高浓度区（感染灶或炎症区域）迁移，趋化因子由此成为中性粒细胞的强活化因子（图 2 ）^[90]。

在脓毒症患者中，血中性粒细胞向试验诱导的皮肤水疱区的迁移能力缺陷（图 2 ) ^[62^，^80-81]。尽管对中性粒细胞在 fMLP 诱导下的迁移能力受损的报道存在争议，但白三烯 B₄诱导的中性粒细胞的迁移能力的确受到了损害（表 2 ) ^[78-79]。

IL-8 与高亲和力受体 CXCR1 、 CXCR2相结合。CXCR2也是其他一些趋化因子的受体。脓毒症患者中性粒细胞表面CXCR2的表达下调， CXCR1的表达则未下调^［^{69 ,79]}；并且，由 IL-8 介导的趋化反应可能正常^［⁷⁹^］或受阻^［⁶⁹^］。这些分子对于脓毒症的发病机制很关键，脓毒症小鼠模型表明，阻断 CXCR1 及 CXCR2 的信号转导可抑制 MOF 与 DIC 的发生^［⁹¹^］。

在 ARDS 患者中，中性粒细胞大规模迁移至肺部，支气管肺泡灌洗液 IL-8 的浓度与死亡率具有相关性^[92]。在体循环，中性粒细胞通过毛细血管后微静脉进入组织内；但在肺循环，中性粒细胞则通过毛细血管迁移^[93]。肺毛细血管管腔很狭窄，使中性粒细胞与血管壁的接触加强，延长了中性粒细胞穿行时间，并阻止其延内皮细胞表面滚动^[94-95]。因此，肺部中性粒细胞的聚集可能更依赖于整合素，而非选择素，这与脓毒症患者中性粒细胞表面 L -选择素表达较低^[62-63]而 CD11b ^{[ 63-69}^］及 CD49d^{[ 73}^］表达增加的结论-致。并且，是中性粒细胞的一种亚型与趋化因子或引起肺部感染的微生物发生反应，也是因为动物模型发现了肺毛细血管内皮的独特表型，因此与其他器官相比，中性粒细胞更易于进入肺部^[96]。

影响中性粒细胞活化、生存与循环的因素

中性粒细胞以 3 种状态存在：静息（未受刺激）、致敏（在炎症激活因素或能降低中性粒细胞活化所需刺激阈值的微生物产物作用下）、活化（执行特定功能）。中性粒细胞由循环中的静息状态向炎症部位的活化状态的转变是由致敏刺激所诱导产生的一系列信号分子 ― 如 C5a 、脂多糖及细胞因子 ― 相继作用的结果^[97-99]。这种效应对感染区域血管外的中性粒细胞是有利的；但已经和血管内皮相结合的致敏的中性粒细胞，若再次受到刺激，则会引起血管壁损伤 ^[100]。脓毒症患者血液中的中性粒细胞是已被致敏的，其氧化活性增高，细胞内转录基因产物 ― 核因子 KB ( NFKB ) ― 的表达增加^{[74 , 85-87]}。脓毒症患者中性粒细胞 NFKB 活化减少与存活率改善相关^[101]。

中性粒细胞通过经历凋亡来减少数量，这种稳态机制可阻止正常组织受破坏，否则坏死细胞溶解便可导致正常组织损伤。这一过程是预防及消除炎症的中心环节。系统性炎症、系统性感染、重度脓毒症及 MOF 高危患者的中性粒细胞凋亡受抑^{[45 , 74-77]}，多认为这是由于包括脂多糖、脂磷壁酸及促炎症细胞因子^[45^，^102-105]在内的循环因子被活化所致；即便与受促炎症细胞因子活化的内皮黏附会延长中性粒细胞寿命，而与未活化内皮的黏附则会加速其死亡^[106]。浸润组织的中性粒细胞寿命的延长，会增加其通过氧自由基与蛋白水解酶的失控释放而造成胞外损害的可能性。

脓毒症时，局部诱导产生的抗凋亡因子有利于组织内中性粒细胞的存活。例如， ARDS 时，肺泡内中性粒细胞的低水平凋亡与支气管肺泡灌洗液中 IL-2 的浓度相关^[107]。凋亡抑制与复杂的细胞内信号转导网络及细胞器功能失调有关^[108]，包括酪氨酸磷酸化增强及持续的线粒体跨膜电位^[109-110]。脓毒症患者中性粒细胞寿命延长，而淋巴组织内淋巴细胞凋亡增加，进而引起免疫麻痹^[14]。

循环因素可能也参与脓毒症时血中性粒细胞表型及功能状态的改变。用 IL-8 （脓毒症时其血浓度常增加^[111]）对中性粒细胞进行预处理，可抑制其在内皮单细胞层的迁移^[112]，而给兔静脉注射 IL-8 ，可阻止中性粒细胞从肠系膜小静脉移出^[113]。类似地，脓毒症患者血 C5a 浓度升高，使体外试验的中性粒细胞对趋化因子失去反应性^[97]；脓毒症患者血中高浓度的 TNFa 也可阻止中性粒细胞迁移^[114]；并且将正常中性粒细胞与 TNF-a 共孵育可抑制细胞凋亡^[115]、增加活性氧的生成， ^[116]并抑制 CXCR2表达 ^[117]。这些发现均表明，重度脓毒症时中性粒细胞功能障碍并非是一种原发效应，而是一种系统性活化引起的继发效应。通过延长中性粒细胞的寿命、妨碍其在脉管系统内的迁移能力，循环因子可增强中性粒细胞与内皮细胞间的相互作用，并加重血管损伤。

对细菌的识别与吞噬

尽管对于动物感染局部的中性粒细胞的吞噬作用到底是增强还是减弱尚无定论^[118-119]，但脓毒症患者中性粒细胞对微生物的内吞及破坏作用确是增强的（表 2 )^{[74 ,82-83]}。截至目前，大多数关于脓毒症患者中性粒细胞吞噬作用的研究主要集中于参与细菌识别与吞噬的中性粒细胞受体的分布与表达，而不是细胞的功能活性方面。

当病原被 IgG 包被，中性粒细胞与病原的结合能力大大增强。 CD64 是 IgG 的高亲和力受体，在静息态中性粒细胞表面不表达，因此可作为一种中性粒细胞活化标记。干扰素（ IFN ）一y及 GM-CSF 可诱导 CD64 的表达^[120-121]。脓毒症患者的多数中性粒细胞表面表达 CD64^[84]，新生中性粒细胞表面 CD64 表达的上调可提示脓毒症的存在^[122]。 CD64 表达上调与呼吸爆发活动增加有关^[46]，这种分子存在于与培养的内皮黏附的大多数中性粒细胞表面，内皮与中性粒细胞间的相互作用可被抗 CD64 抗体抑制^[123]。中性粒细胞与细菌的黏附也可由 CD14 介导， CD14 是脂多糖受体，存在于所有单核细胞表面，但在中性粒细胞表现表达较弱^[124]，细菌感染时表达上调^[125]。其他可加强吞噬作用及细菌识别的受体包括 C3b 受体（与补体肽 C3b 结合）、 CD16 及 CD32 （类似于 CD64 ，与 IgG 的 Fc段相结合），这些受体均在脓毒症患者的中性粒细胞表面充分表达。

Toll 样受体（ TLRs ）因与果蝇 Toll 蛋白同源而得名，是模式识别受体，介导对各种微生物配体的天然免疫应答。目前已发现了 11 种： TLR4 与 CD14 密切相关，是脂多糖的信号转导受体； TLR2 专门识别革兰染色阳性细菌^[126]。健康个体未受刺激的中性粒细胞表面几乎不表达这 2 种受体，不过在 G-CSF 与 GM-CSF 刺激后， TLR2表达上调^[127]。这2 种受体中的任何一种受到激活剂刺激后，会诱导细胞发生呼吸爆发、释放 IL-8 、分泌 L 一选择素，并上调 CD11b 的表达^[128-129]。 TLR2^[130]与 TLR4 的激活剂均可直接延缓中性粒细胞的凋亡，但单核-巨噬细胞介导的间接作用对中性粒细胞寿命的延长可能更为重要^[129]。尽管 TLR2 与 TLR4 的活化会使中性粒细胞趋化因子受体表达下调，特别是 CXCR2^[128^，^130]，但细胞仍然保持一定的 IL-8 刺激下的迁移能力^[131]。 TLRs 与趋化因子的相互作用很复杂，小鼠实验发现 TLR4 的活化会通过下调 G 蛋白受体激酶（参与 CXCR 脱敏）而加强中性粒细胞对趋化因子的反应性^[132]。对 TLR2 或 TLR4 反应性的不同可能由其他表面受体介导，如通过 CD18 对革兰阴性细菌发生应答，对革兰阳性细菌则通过不依赖 CD18 的途径发生应答^[133]。对 TLR_S 进行药理学修饰已被提出可在将来作为脓毒症休克患者的治疗策略^[134]。

另一类模式识别受体是 TREM 家族（骨髓细胞表达的激发受体）。高表达 TREM-1 的中性粒细胞浸润被细菌感染的人体组织，阻断实验诱导的脓毒症模型中 TREM-1 的表达可改善存活情况^[135]。 TREM-1 信号转导可引起 IL-8 释放，并使细胞表面黏附分子表达上调^[136]。有研究发现：仅在急性炎症时，中性粒细胞 TREM-1 表达上调，细菌感染可加速其表达；并且，脓毒症患者血浆可溶性 TREM-1 浓度下降，对预后有利^[137]，这些结果支持这样一个观点，即在脓毒症时， TREM-1 可能具有很重要的致病作用。

对脓毒症时中性粒细胞功能理解的局限性

对脓毒症患者中性粒细胞许多方面（如吞噬作用）的研究尚不充分，甚至有的研究结果相互矛盾（如黏附分子的表达），因此妨碍了对其功能状态的阐明。研究结果间显著存在的不一致可能是由对患者分层的不够细致、干预药物（如类固醇激素）的多样化及研究设计的不同所致。由于循环池中性粒细胞寿命短，而且对持续数天乃至数周的病变而言，在某一时间点的评估只能提供很少的信息，因此，要获得对脓毒症患者中性粒细胞状态的有价值评估，就需要对个体患者进行纵向研究。但对典型的中性粒细胞生命周期的各个方面，仍知之甚少。例如，对 G-CSF 与 GM-CSF 的产生、中性粒细胞由骨髓进入外周的机制，以及是否中性粒细胞进入器官并凋亡是通过与促进中性粒细胞进入感染和炎症部位相同的表面黏附分子及趋化因子受体所介导，仍不明确。

动物模型可帮助识别那些参与实验诱导脓毒症的特定介质与黏附分子，但截至目前，研究结论还没有临床意义。抗 TNF-α、抗 IFN -γ抗体可保护狒狒^[138]及小鼠^[139]免受细菌感染，但抗 TNF- α 的治疗对脓毒症患者却无效^[140-141]，甚至有害 ^[142]。同样地，应用 IFN -γ虽可增强中性粒细胞功能^[143]，却不能改善患者预后^[144]。拮抗 IL-1 及血小板活化因子的活性也无任何临床疗效（表 3 ) ^[145-146]。有一个事实常被忽略，即中性粒细胞是人体血液中白细胞的一个最主要类型；但在常被用作脓毒症实验动物的小鼠体内，中性粒细胞却只占少数。 β ₁整合素，即 CD49d ，仅存在于脓毒症患者的一小部分细胞表面^［^{73 ]}，健康个体中性粒细胞表面不表达，而在小鼠中性粒细胞则呈组成性表达^[153]。对同系杂交动物的药物干预常采用预防性治疗，但对脓毒症患者却不合适。采用基因敲除小鼠模型，可对参与脓毒症病理机制的单个分子的作用进行评价，但研究结果的总体重要性有时却因代偿机制的存在而显得很混乱，令人困惑。例如，在细菌感染的刺激下，浸润 CD62E / CD62P 选择素基因双敲除小鼠肺部的中性粒细胞出乎意料地远多于浸润野生型小鼠肺部的中性粒细胞^[154]。

中性粒细胞与内皮细胞间的相互作用及脓毒症相关性器官功能衰竭

重度脓毒症时，中性粒细胞对细菌感染的应答功能似乎具有两面性。在肺组织以外，中性粒细胞向感染区域的渗入可能会因为与内皮细胞黏附过于紧密及趋化反应减弱而受阻；而在感染的肺组织，中性粒细胞却大量浸润（图 3 ）。中性粒细胞的“被隔离”是触发 MOF 的关键环节^[155^］。中性粒细胞与血管壁的黏附可能是由与促进中性粒细胞与肺内皮黏附所不同的黏附分子异常表达或与相应的内皮配体具有高亲和力的分子所介导的。黏附作用在趋化因子受体下调的影响下得以加强，可能引起微血管阻塞，继发组织低灌注与缺氧^[10^，^30]。相关发现还有，脓毒症患者反应性充血障碍与组织损伤中脓毒症休克缺陷均与组织灌注障碍有关，也与发病率和病死率的增加有关^[156-157]。如果中性粒细胞与内皮的黏附发生在微脉管部位表明存在纤维素沉积，那么，将有可能发生进一步的伴白细胞聚集的黏附。若由循环因子致敏的中性粒细胞遇到血管壁上其他的致敏物质，中性粒细胞将会释放细胞溶解因子，后者将损伤内皮细胞并增加血管通透性（图 3 ）。

换言之，器官功能衰竭可继发于中性粒细胞对内皮细胞功能的轻微影响，这也可能与脓毒症的其他病理机制有关。中性粒细胞是促炎症细胞因子的重要来源^[10], 促炎症细胞因子的表达由 NFKB 调节，而后者在脓毒症患者中高度表达^[87^，^101]。局限于血管壁的由中性粒细胞分泌的细胞因子会使内皮细胞由非促血栓形成状态转变为促凝状态，从而触发 DIC^[8]，也会导致内皮与平滑肌细胞产生 NO （一氧化氮）^[4]。除引起脓毒症休克时的低血压外^[158]， NO 的释放还会通过抑制线粒体酶的活性^[159]而阻碍组织代谢，中性粒细胞自身所产生的 NO 也会加强这一效应^[160]。肺功能衰竭始自肺泡剥蚀、基底膜破坏^[4]及肺泡液清除机制障碍^[10]。进入肺泡组织与间隙的大量致敏中性粒细胞可能会在局部炎症因子与细菌产物的不良刺激下，释放蛋白水解酶与氧自由基（图 3 ）。

治疗策略

1. 中性粒细胞黏附分子

若中性粒细胞与内皮的紧密黏附是脓毒症器官功能障碍的诱因，那么抑制两者的相互作用是否能获得临床收益呢？在实验诱导的脓毒症模型中， CD18 拮抗剂可逆转中性粒细胞诱导的肺血管损伤^[161]。类似的方法尚未在临床展开应用，但对创伤性休克^[162]或心肌梗死患者^[163]应用抗 CD18 抗体的效果均不令人满意，这可能是由于以 CD18 为目标并不合适，中性粒细胞很大程度上依赖于除 CD18 以外的黏附分子（如β ₁整合素）。脓毒症时，除一些迄今尚未被发现的参与黏附过程的分子或仍待被证实的分子（如 CD64 ）外， β ₁整合素家族的其他成员可能也参与了中性粒细胞与内皮的黏附^[32]。

作者对白细胞所表达的表皮生长因子样 7 个跨膜区段家族进行了研究，其分子结构提示其在黏附与信号传递中发挥一定的作用^[164]。作者注意到此家族的一个成员（ EMR2 ) ― 表达于一些正常中性粒细胞表面 ― 在脓毒症时表达于约 50 ％的中性粒细胞表面（资料未公布）。今后的研究若着眼于脓毒症患者的中性粒细胞，便可能发现：其他黏附分子样分子表达的增多是感染与炎症状态的显著特征。倘若这类分子有望成为新的候选治疗靶向，那么应在疾病的哪一阶段进行干预呢？

鉴于多数出现感染体征和症状的患者住人重症监护病房时已出现某一器官功能障碍，因此，在发生器官功能衰竭前给药并不可行。根据抗整合素抗体可阻断中性粒细胞与内皮的黏附^[165]，各种治疗策略可能均需以阻断聚集在微脉管内的中性粒细胞的黏附或预防可使器官功能障碍加剧的进一步黏附为目标。就如“拯救脓毒症患者行动”最近所强调的那样，如果这种策略可行，则应尽早应用。“拯救脓毒症患者行动”描述了一系列护理措施，包括应在急诊看护病房开始及患者病情尚未恶化到需要重症监护之前便进行抗生素及靶向治疗^[166]。

2. 基因与细胞信号转导

个体对系统性感染及炎症的易感性可能与中性粒细胞黏附分子的多态性有关。据报道， TNF -α^［⁹^］与 IL-1 家族成员^[167]等位基因的多态性均可增加脓毒症患者的死亡风险。鉴于 IgG 的 Fc 受体多态性与脑膜炎球菌感染患者预后有关^[168], CD64 与脓毒症预后间也可能存在类似的相关性。 CD64 是高亲和力 IgG 受体，在脓毒症患著中性粒细胞表面的表达上调^[84]，并与内皮黏附有关^[123]。

另一种调节脓毒症患者中性粒细胞功能的方法是区别维持稳态的信号转导通路与诱发组织损伤的信号转导通路^[169]。在脓毒症实验模型中，中性粒细胞引起的器官损伤有赖于信号转导子与转录激动子（ STAT ) 4 或 6 的活化^[170]；而 IL-10 发挥抗炎活性需要 STAT3^[171]，抗凋亡及趋化过程则需要磷酸肌醇3 激酶^[172]。细胞内的信号转导通路分布广泛且相互作用复杂，但在设计特异性抑制中性粒细胞促炎活性药物时，可望不受上述因素的制约。

3. 当前方法

对脓毒症时中性粒细胞作用的另一种看法可能源自对有效治疗药物作用模式的研究（表 3 ）。例如：脓毒症休克患者应用氟康唑后存活率得以改善，这得益于氟康唑对中性粒细胞功能的调节^[147]；而小剂量氢化可的松的疗效^[148]可能与其对中性粒细胞活性的抑制有关^[149]。采用重组活化蛋白 C 可改善重度脓毒症患者的存活率^[173]，疗效归因于对凝血系统与炎症反应的调节。活化蛋白 C 的潜在抗炎作用包括妨碍中性粒细胞一内皮细胞相互作用 ^[150]及下调血管黏附分子^[151]在内毒素诱导的人肺炎模型中，活化蛋白 C 可在肺局部发挥抗凝作用^[174]，并抑制中性粒细胞向肺组织迁移^[152]。这些结果均说明，中性粒细胞对于凝血和纤溶系统的作用远较以往人们认为的大。中性粒细胞直接参与 DIC 的病理生理过程^[175]，蛋白 C 通路可阻碍中性粒细胞与内皮的相互作用^[176]，而可溶性内皮蛋白 C 受体可与活化的中性粒细胞结合^[177]。

结论

在脓毒症时，淋巴细胞的反应性低，加之因凋亡增加而引起的数量减少，机体免疫应答势必受到抑制。然而，循环血中性粒细胞却常常增多、寿命延长、应答能力（除趋化作用外）增强（表 2 ）。

脓毒症时，中性粒细胞发挥抵挡病原入侵的作用，但同时也可导致器官功能受损。目前，尚无法将中性粒细胞的这两方面作用分隔开来，因此，为保护器官功能而抑制中性粒细胞功能的作法将阻碍机体对病原的清除。可解决这一两难处境的核心问题是：哪一方对患者存活的威胁更大，是感染本身还是免疫反应引起的器官损害？值得注意的是，在非感染状态下（如创伤、胰腺炎及心肺转流手术），系统性炎症反应所引起器官功能衰竭的免疫病理特征与脓毒症类似。另外，在重症监护病房中，所有重度脓毒症患者均能马上通过抗生素治疗来控制细菌感染，这弥补了中性粒细胞作用的不足^［³^］。因此在这一前提下，作者建议采用抑制免疫系统的药物防止器官功能衰竭，这种策略可能会使患者受益。尽管之前所采用的免疫抑制治疗（如 TNF-α 活性拮抗剂，大剂量皮质类固醇激素）并未改善脓毒症患者的预后，作者仍相信选择性抑制中性粒细胞亚群与功能易受损器官内血管壁间的相互作用，可作为今后治疗策略的选择。

Lancet 2006 ; 368 : 157-69

（金洋译李晶校）

Acknowledgments:

Our work is supported by The Trustees of the Henry Smith Charity and The British Heart Foundation. These funding sources had no role in the writing of the Review or in the decision to submit the paper for publication. Division of Medical Education (K A Brown FRCPath) and Cardiovascular Division (Prof S D Brain PhD, Prof J D Pearson FMedSci, S M Lewis BSc), King’s College School of Medicine, London, UK; Department of Infection, Guy’s and St Thomas` Hospital, London, UK (J D Edgeworth li/IFICPath); and Intensive Care Unit, Guy’s and St Thomas’ NHS Foundation Trust (D F Treacher FRCP)

Correspondence to: Dr K Alun Brown, The Rayne Institute, St Thomas' Hospital, London SE1 7EH, UK (e-mail: alun.brown@kcl.ac.uk)

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