Amigorena S, Choquet D, Teillaud J L, Korn H, Fridman W H
Laboratoire d'Immunologie Cellulaire et Clinique, INSERM U255, Institut Curie, Paris, France.
J Immunol. 1990 Mar 15;144(6):2038-45.
Lymphocytes express voltage-activated K+ channels in their membrane. Combining the patch-clamp techniques of recording with immunological methods, we have analyzed the expression and the involvement of these channels during defined steps of LPS-induced B cell activation. We show that the number of K+ channels increased strongly when B cells entered in the G1 phase of the cell cycle. The involvement of ion channels in B cell proliferation was assessed using channel blockers that inhibit the K+ current. It was first found that TEA, but not TMA, quinine and verapamil totally suppressed both K+ current and DNA synthesis by stimulated lymphocytes as measured by [3H]TdR uptake or propiedium iodide staining. The drugs affected neither the induction by LPS of activation markers such as Ag of the murine class II MHC and type II receptor for the Fc region of IgG nor the initial cell enlargement that occur early during activation. These data indicate that functional K+ channels are not essential for the transition from the G0 to the G1 phases. In contrast, the same channel antagonists blocked the induction of transferrin receptor expression, characteristic of the final stages of G1. These drugs acted on cells already in G1, because their addition 30 h after LPS still suppressed DNA synthesis, and because they inhibited the proliferation of purified B cell blasts. The effect of tetraethylammonium was reversible, a lag period of 12 h occurring before the cells start DNA synthesis after drug removal. Taken together, these data demonstrate that the proliferation of LPS-stimulated B cells requires functional ion channels at a critical period in the G1 phase, taking place before transferrin receptor expression and the entry into the S phase. The involvement of voltage dependent K+ channels at this particular point is suggested by the parallel effects of the drugs used on K+ currents and DNA synthesis.
淋巴细胞在其细胞膜上表达电压激活的钾离子通道。结合膜片钳记录技术与免疫学方法，我们分析了这些通道在脂多糖（LPS）诱导的B细胞活化特定阶段的表达及作用。我们发现，当B细胞进入细胞周期的G1期时，钾离子通道数量显著增加。使用抑制钾离子电流的通道阻滞剂评估离子通道在B细胞增殖中的作用。首先发现，四乙铵（TEA）能完全抑制刺激淋巴细胞的钾离子电流和DNA合成，而四甲铵（TMA）、奎宁和维拉帕米则无此作用，DNA合成通过[³H]胸腺嘧啶核苷（TdR）摄取或碘化丙啶染色来测定。这些药物既不影响LPS诱导的活化标志物表达，如小鼠II类主要组织相容性复合体（MHC）抗原和IgG Fc区II型受体，也不影响活化早期出现的初始细胞增大。这些数据表明，功能性钾离子通道对于从G0期向G1期的转变并非必需。相反，相同的通道拮抗剂可阻断转铁蛋白受体表达的诱导，而转铁蛋白受体表达是G1期末期的特征。这些药物作用于已处于G1期的细胞，因为在LPS刺激30小时后添加它们仍能抑制DNA合成，且它们能抑制纯化的B细胞母细胞的增殖。四乙铵的作用是可逆的，在去除药物后细胞开始DNA合成前会有12小时的延迟期。综上所述，这些数据表明，LPS刺激的B细胞增殖在G1期的关键时期需要功能性离子通道，该时期发生在转铁蛋白受体表达和进入S期之前。所用药物对钾离子电流和DNA合成的平行作用表明电压依赖性钾离子通道在这一特定点发挥作用。
