Overexpression of brain-derived neurotrophic factor increases Merkel cell number in murine skin.

Merkel cells (MC) are specialized epithelial cells, having sensory and neuroendocrine functions, and present within basal layers of epidermis (disseminated as well as clustered in Pinkus’s hair disc), and outer root sheath of hair follicles of men (Moll et al, 1986, 1993, 1995; Halata, 1993). In mice, numerous MC are present in the foot pad epidermis, whereas in the epidermis of back skin MC are exclusively arranged in small clusters within so-called Haarscheiben (hairdisc, tactile disc), which are associated with tylotrich hair follicles in animals (Moll et al, 1996a). MC within tactile discs form intimate contacts with sensory nerve terminals and function as slowly adapting type I mechanoreceptors (cf. Lewin and Barde, 1996). Increasing evidence suggests that neurotrophins control the development, differentiation, and also postnatal physiology of MC. Neurotrophin-3 regulates MC number and their innervation during postnatal development (Airaksinen et al, 1996; Albers et al, 1996), whereas brain-derived neurotrophic factor (BDNF) stimulates the mechanotransducing properties of slowly adapting type I mechanoreceptors (Caroll et al, 1998). Recently we have shown that BDNF overexpression in murine skin is associated with sympathetic hyperinnervation of the arrector pili muscle, and with alterations of hair follicle cycling, i.e., premature catagen induction (Botchkarev et al, 1998a, 1999); however, it is as yet unknown whether or not BDNF influences the MC number. Therefore, we have studied the distribution and number of MC in the foot pad epidermis of transgenic mice overexpressing BDNF (Botchkarev et al, 1998a) by immunocytochemistry, using the expression of cytokeratin (CK) 18 and 20 as MC markers (Moll et al, 1984, 1995, 1996a). BDNF overexpression was directed by the alpha-myosin heavy chain promoter, which leads to the ectopic expression of BDNF mRNA in the subcutaneous panniculus carnosus muscle and three-fold elevated steady-state levels of BDNF protein in murine back skin (Botchkarev et al, 1998a). Foot pad skin of adolescent BDNF transgenic and corresponding age-matched wild-type mice (n54) was harvested and processed for the generation of 8 μm, acetone-fixed cryosections, which were then incubated with primary antibodies against CK18 and CK20 (Progen Biotechniks, Heidelberg, Germany) and secondary peroxidase-conjugated antibody for indirect immunoperoxidase microscopy (Dako) as described before (Moll et al, 1996a). The number of CK18or CK20-positive MC per mm of epidermis length was calculated in 100–120 microscopic fields (3200) derived from four BDNF transgenic mice, and was compared with that of four age-matched wild-type controls, and means and SEM were calculated from pooled data. Differences were judged as significant if p,0.05, as determined by independent Student’s t-test for unpaired samples. Photo-documentation was performed with the