Aida Y, Pabst M J
Department of Biochemistry, University of Tennessee, Memphis 38163.
J Immunol. 1991 Feb 15;146(4):1271-6.
We studied neutrophil responses to LPS using three methodologic refinements: Teflon bags or serum-coated glass tubes that did not directly trigger neutrophils, LPS-free cytochrome c to measure O2- release, and heat-inactivated serum to inhibit inactivation of LPS by neutrophils. Neutrophils incubated in uncoated glass or plastic tubes adhered to the glass and released O2-, but were not primed for enhanced release of O2- in response to triggering by FMLP. Triggering by the glass or plastic surface did not occur if the neutrophils were stirred to prevent adherence. Adherence to glass or plastic and O2- release were not affected by a mAb (IB4) directed against the beta-chain of the leukocyte adhesion family of surface glycoproteins (CD11/CD18). Neutrophils incubated in glass or plastic did not show enhanced expression of alkaline phosphatase on their surface. When neutrophils were incubated in serum-coated glass tubes or in Teflon bags, there was no O2- release. However, adherence, expression of alkaline phosphatase, and release of O2- were triggered by adding 1 ng/ml LPS plus 1% serum, but not by either LPS or serum alone. In the presence of LPS and serum, O2- release was much higher when the cells were unstirred (adherent) rather than stirred. However, both unstirred and stirred cells expressed a similar elevated level of alkaline phosphatase. LPS-triggered O2- release and adherence were inhibited by antibody IB4. In contrast, priming by LPS for enhanced FMLP-triggered O2- release was greater in stirred cells than in unstirred cells. The antibody enhanced priming of unstirred neutrophils. These results suggested that uncoated glass or plastic triggered O2- release without involvement of leukocyte adhesion glycoproteins. However, neutrophils incubated with LPS and serum expressed alkaline phosphatase and IB4-inhibitable adherence glycoproteins that allowed neutrophils to interact with serum-coated glass or Teflon to trigger O2- release. Priming by LPS for enhanced response to FMLP was suppressed in adherent neutrophils, and this suppression was partly released by IB4. Thus, triggering and priming were reciprocally regulated by neutrophil glycoproteins interacting with surfaces.
我们采用了三种方法改进来研究中性粒细胞对脂多糖（LPS）的反应：使用不会直接触发中性粒细胞的聚四氟乙烯袋或血清包被的玻璃管；使用无LPS的细胞色素c来测量超氧阴离子（O₂⁻）释放；使用热灭活血清来抑制中性粒细胞对LPS的灭活。在未包被的玻璃管或塑料管中孵育的中性粒细胞会黏附于玻璃并释放O₂⁻，但对甲酰甲硫氨酸-亮氨酸-苯丙氨酸（FMLP）触发的O₂⁻增强释放未产生预刺激。如果搅拌中性粒细胞以防止黏附，则不会发生玻璃或塑料表面的触发。针对表面糖蛋白白细胞黏附家族的β链（CD11/CD18）的单克隆抗体（IB4）不会影响对玻璃或塑料的黏附及O₂⁻释放。在玻璃管或塑料管中孵育的中性粒细胞表面碱性磷酸酶表达未增强。当在血清包被的玻璃管或聚四氟乙烯袋中孵育中性粒细胞时，未出现O₂⁻释放。然而，添加1 ng/ml LPS加1%血清可触发黏附、碱性磷酸酶表达及O₂⁻释放，单独的LPS或血清则不会。在存在LPS和血清的情况下，细胞未搅拌（黏附）时的O₂⁻释放远高于搅拌时。然而，未搅拌和搅拌的细胞碱性磷酸酶表达水平均有类似升高。LPS触发的O₂⁻释放和黏附受到抗体IB4的抑制。相反，LPS对增强FMLP触发的O₂⁻释放的预刺激在搅拌的细胞中比未搅拌的细胞中更强。该抗体增强了未搅拌中性粒细胞的预刺激。这些结果表明，未包被的玻璃或塑料触发O₂⁻释放，且不涉及白细胞黏附糖蛋白。然而，与LPS和血清一起孵育的中性粒细胞表达碱性磷酸酶和IB4可抑制的黏附糖蛋白，这使得中性粒细胞能够与血清包被的玻璃或聚四氟乙烯相互作用以触发O₂⁻释放。LPS对增强FMLP反应的预刺激在黏附的中性粒细胞中受到抑制，且这种抑制部分被IB4解除。因此，触发和预刺激由中性粒细胞糖蛋白与表面的相互作用进行相互调节。
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