Purinergic Signalling (2008) 4:331–338 DOI 10.1007/s11302-008-9108-0 ORIGINAL ARTICLE Purinergic receptors are part of a signalling system for proliferation and differentiation in distinct cell lineages in human anagen hair follicles Aina V. H. Greig & Claire Linge & Geoffrey Burnstock Received: 5 March 2008 / Accepted: 17 April 2008 / Published online: 27 May 2008 # Springer Science + Business Media B.V. 2008 Abstract We investigated the expression of P2X5, P2X7, P2Y1 and P2Y2 receptor subtypes in adult human anagen hair follicles and in relation to markers of proliferation [proliferating cell nuclear antigen (PCNA) and Ki-67], keratinocyte differentiation (involucrin) and apoptosis (anticaspase-3). Using immunohistochemistry, we showed that P2X5, P2Y1 and P2Y2 receptors were expressed in spatially distinct zones of the anagen hair follicle: P2Y1 receptors in the outer root sheath and bulb, P2X5 receptors in the inner and outer root sheaths and medulla and P2Y2 receptors in living cells at the edge of the cortex/medulla. P2X7 receptors were not expressed. Colocalisation experi- ments suggested different functional roles for these recep- tors: P2Y1 receptors were associated with bulb and outer root sheath keratinocyte proliferation, P2X5 receptors were associated with differentiation of cells of the medulla and inner root sheaths and P2Y2 receptors were associated with early differentiated cells in the cortex/medulla that contrib- ute to the formation of the hair shaft. The therapeutic potential of purinergic agonists and antagonists for control- ling hair growth is discussed. A. V. H. Greig : G. Burnstock (*) Autonomic Neuroscience Centre, Royal Free and University College Medical School, Rowland Hill Street, London NW3 2PF, UK e-mail: g.burnstock@ucl.ac.uk C. Linge RAFT Institute of Plastic Surgery, Mount Vernon Hospital, Leopold Muller Building, Northwood, Middlesex HA6 2RN, UK Keywords Receptors . Purinergic . Hair follicle . Proliferation . Differentiation Abbreviations AA amino acids ATP adenosine 5’-triphosphate ADP adenosine 5’-diphosphate DAB diaminobenzidine NGS normal goat serum NHS normal horse serum IRS inner root sheath ORS outer root sheath PBS phosphate buffered saline PCNA proliferating cell nuclear antigen Introduction The cells of the lower portion of the hair follicle bulb are undifferentiated matrix cells. These are rapidly dividing cells that give rise to eight different cell lineages [1]. From within outwards, these include the medulla, cortex and hair cuticle cell lineages that make up the hair shaft; the inner root sheath cuticle, Huxley’s and Henle’s layers that make up the inner root sheath (IRS); and the companion layer and the outer root sheath (ORS) [1]. The hair cycle is divided into periods of hair growth (anagen), which are followed by a regression phase (catagen), when the lower part of the hair follicle undergoes programmed cell death [2], and a resting phase (telogen), before onset of a new growth phase. The growing anagen hair follicle is an attractive system for studying proliferation and differentiation. There is increas- ing evidence that purinergic signalling can have long-term,