Adenosine triphosphate stimulates phosphoinositide metabolism, mobilizes intracellular calcium, and inhibits terminal differentiation of human epidermal keratinocytes.

Abstract

During wound healing, release of ATP from platelets potentially exposes the epidermis to concentrations of ATP known to alter cellular functions mediated via changes in inositol trisphosphate (IP3) and intracellular calcium (Cai) levels. Therefore, we determined whether keratinocytes respond to ATP with a rise in Cai and IP3 and whether such increases are accompanied by a change in their proliferation and differentiation. Changes in Cai were measured in Indo-l-loaded neonatal human foreskin keratinocytes after stimulation with extracellular ATP. Extracellular ATP evoked a transient and acute increase in Cai of keratinocytes both in the presence and in the absence of extracellular calcium. ATP also induced the phosphoinositide turnover of keratinocytes, consistent with its effect in releasing calcium from intracellular sources. ATP did not permeabilize keratinocytes, nor did it promote Ca influx into the cells. The half-maximal effect of ATP was at 10 MM, and saturation was observed at 30-100 ,M. UTPP, ITP, and ATP-yS were as effective as ATP in releasing Cai from intracellular stores and competed with ATP for their response, whereas AMP and adenosine were ineffective, suggesting the specificity of P2 purinergic receptors in mediating the ATP response in keratinocytes. Single cell measurements revealed heterogeneity in the calcium response to ATP. This heterogeneity did not appear to be due to differences in the initial Cai response but to subsequent removal of increased Cai by these cells. ATP inhibited terminal differentiation of keratinocytes as measured by I35Slmethionine incorporation into cornified envelopes and modestly stimulated incorporation of r3Hlthymidine into DNA.Chelation of Cai by bis-(o-aminophenoxy)-NNN',N'-tetraacetic acid reduced basal Cai, blocked the Cai response to ATP, inhibited the basal rate of DNA synthesis, and blocked the ATP-induced increase in DNA synthesis. We conclude that extracellular ATP may be an important physiological regulator of epidermal growth and differentiation acting via IP3 and Cai. (J. Clin. Invest. 1992. 90:42-51.)

Chemosensory Information Processing between Keratinocytes and Trigeminal Neurons

“…P2 receptors comprise two subgroups, the ionotropic P2X receptors and the metabotropic P2Y receptors (56). Subtypes from both groups show tissue-dependent expression (57)(58)(59)(60). A previous investigation in our laboratory investigated ATP-mediated currents in trigeminal neurons (13).…”

Connexin 26 regulates epidermal barrier and wound remodeling and promotes psoriasiform response

“…In keratinocytes, the increased release of ATP from CX26 hemichannels initially results in a rise of intracellular calcium. However, continual stimulation of keratinocytes by ATP depletes the intracellular calcium stores and desensitizes the ATP receptors from responding to the proper differentiation cues (43). Thus, while elevation of intracellular calcium levels normally promotes the differentiation process, excessive ATP stimulation inhibits terminal differentiation and barrier recovery in the epidermis (43,44).…”

“…However, continual stimulation of keratinocytes by ATP depletes the intracellular calcium stores and desensitizes the ATP receptors from responding to the proper differentiation cues (43). Thus, while elevation of intracellular calcium levels normally promotes the differentiation process, excessive ATP stimulation inhibits terminal differentiation and barrier recovery in the epidermis (43,44). Furthermore, keratinocyte release of ATP exacerbates inflammation by activating CD39 + Langerhans cells, as observed in response to irritant chemicals (45,46).…”

Changes in calcium responsiveness and handling during keratinocyte differentiation. Potential role of the calcium receptor.

“…Cai measurement in suspended keratinocytes was carried out as described previously (7,21,23). Briefly, trypsinized keratinocytes were loaded with 2 M Indo-1 AM in buffer A (20 mM Hepes buffer, pH 7.4, containing 120 mM sodium chloride, 5 mM potassium chloride, 1 mM magnesium chloride, 1 mg/ml pyruvate, 1 mg/ ml glucose, and 0.03 mM calcium chloride).…”

“…This appears to occur by calcium activation of both chloride channels, leading to hyperpolarization of the membrane (19) and voltageindependent nonspecific cation channels permeable to calcium (20). The net result is that Cao leads to an acute but transient rise in Cai followed by a sustained increase in Cai, which appears to be necessary for the differentiation response (6)(7)(8)(9)(10)(11)(12)(21)(22)(23).…”