Light-Touch-Induced Afterdischarge Firing in the Superficial Spinal Dorsal Horn Neurons in Hairless Mice with Irritant Contact Dermatitis.

The skin is an important barrier that protects against invasion by foreign substances, including irritants and harmful microorganisms, and holds water in the body. Washing the skin with cleansers and shampoos containing anionic surfactants, for example sodium dodecyl sulfate (SDS), is important for maintaining skin homeostasis. However, surfactants can cause dermatitis, cutaneous hypersensitivity (e.g., alloknesis), and pruritus in humans. Our previous studies revealed an alloknesis response in the skin with SDS-induced dermatitis in C57BL/6 mice. In addition, we found that alloknesis responses and afterdischarge responses following stimulation with light touch are related because they are observed contemporaneously. In this study, we used Hos:HR-1 hairless mice to establish a mouse model to evaluate long-term drug application for alloknesis responses. Alloknesis was observed in HR-1 mice with SDS-induced dermatitis. The mean number of c-Fos (a marker of neural activity) immunopositive neurons was increased in the lamina 1-2 (L1-2) spinal dorsal horn, but not in L3-4, of SDS-treated HR-1 mice compared to vehicle-treated mice. We also discovered that afterdischarge responses were observed in neurons in L1-2. There was also a correlation between the intensity of the afterdischarge responses and depth of the recording site. Thus, the following were suggested: 1) neurons that mediate these afterdischarge responses are located on the superficial layer of the spinal cord; 2) afterdischarge responses can be an index of alloknesis responses, and 3) the mouse model of SDS-induced dermatitis is an appropriate alloknesis model.

Pulse Dipolar Electron Paramagnetic Resonance Spectroscopy Reveals Buffer-Modulated Cooperativity of Metal-Templated Protein Dimerization.

Self-assembly of protein monomers directed by metal ion coordination constitutes a promising strategy for designing supramolecular architectures complicated by the noncovalent interaction between monomers. Herein, two pulse dipolar electron paramagnetic resonance spectroscopy (PDS) techniques, pulse electron-electron double resonance and relaxation-induced dipolar modulation enhancement, were simultaneously employed to study the CuII-templated dimerization behavior of a model protein (Streptococcus sp. group G, protein G B1 domain) in both phosphate and Tris-HCl buffers. A cooperative binding model could simultaneously fit all data and demonstrate that the cooperativity of protein dimerization across α-helical double-histidine motifs in the presence of CuII is strongly modulated by the buffer, representing a platform for highly tunable buffer-switchable templated dimerization. Hence, PDS enriches the family of techniques for monitoring binding processes, supporting the development of novel strategies for bioengineering structures and stable architectures assembled by an initial metal-templated dimerization.

Histamine enhances ATP-induced itching and responsiveness to ATP in keratinocytes.

Mechanical stimulation of cultured keratinocytes and a living epidermis increases intracellular calcium ion concentrations ([Ca2+]i) in stimulated cells. This action propagates a Ca2+ wave to neighboring keratinocytes via ATP/P2Y2 receptors. Recent behavioral, pharmacological studies revealed that exogenous ATP induces itching via P2X3 receptors in mice. We previously showed that alloknesis occurs when an external stimulus is applied to the skin with increased epidermal histamine in the absence of spontaneous pruritus. Based on these results, we investigated the effects of histamine at a concentration that does not cause itching on ATP-induced itching. The mean number of scratching events induced by the mixture of ATP and histamine increased by 28% over the sum of that induced by histamine alone or ATP alone. A317491, a P2X3 receptor antagonist, suppressed the mixture-induced scratching more often than the ATP-induced scratching. Next, we examined the ATP-induced [Ca2+]i change before and after histamine stimulation using normal human epidermal keratinocytes. Some cells did not respond to ATP before histamine stimulation but responded to ATP afterward, the phenomenon suppressed by chlorpheniramine maleate. These findings suggest that histamine enhances ATP-induced itching and that a potential mechanism could involve increased responsiveness to ATP in keratinocytes.

Expression of histidine decarboxylase in melanocytes of the human skin.

Histamine-producing cells include storage-type cells (e.g., mast cells and basophils), which store histamine intracellularly, and inducible-type cells (e.g., keratinocytes and macrophages), which induce histidine decarboxylase (HDC, a key enzyme for histamine biosynthesis) activity but do not have a storage pool of histamine. Most of the studies focused on identifying HDC-expressing cells by using cultured cells, and few on investigating the localization of HDC by using skin tissues. Hence, this study conducted immunohistochemical studies using human healthy skin samples. HDC-positive and cytokeratin 14 (a marker of basal keratinocytes)-negative cells were present around the basal layer of the epidermis. These cells did not immunohistochemically react with mast cell tryptase but expressed tyrosinase (a key enzyme for melanin biosynthesis) and microphthalmia-associated transcription factor (MITF, a transcription factor controlling the expression of tyrosinase genes). Melanin granules were clearly observed around HDC-positive and MITF-positive cells. Moreover, HDC mRNA and protein were both detected in cultured normal human epidermal melanocytes. In conclusion, HDC-positive and cytokeratin 14-negative cells around the basal layer of the epidermis are melanocytes.