Wohlrab D, Wohlrab J, Markwardt F
Department of Orthopaedics, Martin Luther University, Halle-Wittenberg, Germany.
Exp Dermatol. 2000 Jun;9(3):219-23. doi: 10.1034/j.1600-0625.2000.009003219.x.
The living, not-cornified part of the epidermis consists mainly of keratinocytes. The control mechanisms of proliferation and differentiation are only partly understood. Similarly, the influence of ionic channels of the cell membrane on the proliferation of keratinocytes remains unclear. Preliminary investigations point to a relation between the opening of ionic channels and keratinocyte proliferation. Therefore, voltage clamp experiments were performed to gain further knowledge of the electrophysiological characteristics of human keratinocyte cellular membranes. In-vitro cultured keratinocytes of the cell line HaCaT were characterized by means of the voltage-clamp technique. As measured in the whole-cell configuration, changing the extracellular K+ or Cl--concentration shifted the membrane potential of HaCaT cells. Application of the patch-clamp-technique in the cell-attached and inside-out configuration revealed an ionic channel with a conductance of multiples of 200 pS. The reversal potential of the single channel current was shifted by substituting of intracellular Cl- by aspartate-. Channel openings disappeared after addition of 0.1 mM of the anion channel blocker 4',4' diisothiocyanato-stilbene-2-2' disulfonic acid (DIDS). It is concluded that this channel contributes to the Cl--conductance of the cellular membrane and is a determinant of the membrane potential of human keratinocytes. This channel may represent a target for pharmacological manipulation of the membrane potential and possibly the growth of human keratinocytes in dermatological proliferation disorders.
表皮的活的、未角化部分主要由角质形成细胞组成。增殖和分化的控制机制仅部分为人所知。同样，细胞膜离子通道对角质形成细胞增殖的影响仍不清楚。初步研究表明离子通道的开放与角质形成细胞增殖之间存在关联。因此，进行了电压钳实验以进一步了解人角质形成细胞膜的电生理特性。采用电压钳技术对体外培养的HaCaT细胞系角质形成细胞进行了特性分析。在全细胞模式下测量发现，改变细胞外K+或Cl-浓度会使HaCaT细胞的膜电位发生变化。在细胞贴附式和内面向外式模式下应用膜片钳技术揭示了一种电导为200 pS倍数的离子通道。通过用天冬氨酸替代细胞内Cl-，单通道电流的反转电位发生了偏移。加入0.1 mM阴离子通道阻滞剂4',4'-二异硫氰酸根合芪-2,2'-二磺酸（DIDS）后，通道开放消失。得出的结论是，该通道有助于细胞膜的Cl-电导，并且是人类角质形成细胞膜电位的一个决定因素。该通道可能是在皮肤病增殖性疾病中对膜电位以及可能对角质形成细胞生长进行药理调控的一个靶点。
