An important breach exists in our understanding of how the negative regulation of antimicrobial peptides (AMPs) leads to the clinical symptoms associated with skin disease. The persistent lack of knowledge in this area of research signifies an important challenge to develop improved therapies for cutaneous infection and disease. The long-term goal is to identify how acetylcholine nicotinic receptor (nAChR) activation suppresses AMP expression and activity, and relate these changes in AMP regulation to the causal treatment of human skin diseases associated with AMP dysregulation. The objective of the proposed research is to identify the nAChR subtype(s) involved in AMP suppression in keratinocytes, evaluate the AMP profile altered by nAChR activation, and link this mechanism to the pathogenesis of Atopic Dermatitis (AD). Our hypothesis is that excess nAChR activation in keratinocytes impairs the normal processing and function of cutaneous AMPs, which contributes to the pathogenesis of AD. Our hypothesis was formulated based on preliminary data establishing that nAChR activation significantly reduced AMP activity and the resistance to cutaneous infection. Patients with AD develop a compromised skin barrier, which includes decreased AMP expression thought to precipitate inflammation and infection. Stress likely exacerbates the clinical manifestations of AD by increasing epidermal ACh and, consequently, nAChR activation to compromise normal AMP regulation and microbial susceptibility. Our rationale for these studies is that most research has focused on the positive regulation of AMPs to increase their expression and activity, yet identifying those molecules that inhibit or diminish AMP activity is critical to develop better clinical remedies to ameliorate the symptoms associated with AD and other inflammatory skin diseases. Driven by compelling preliminary data, our hypothesis will be evaluated by addressing three Specific Aims: 1) Identify which nAChR subtypes are involved in AMP suppression and identify the AMP profile in primary normal human epidermal keratinocytes (NHEKs) in vitro;2) Determine the role of nAChR activation in the AMP response to infection in vivo;3) Link mechanisms of nAChR signaling to the pathogenesis of AD.
For Aim 1, NHEKs will be stimulated with nAChR agonists and antagonists to assess known AMPs and identify new antimicrobial molecules using established biochemical techniques.
Aim 2 will use mice with altered states of nAChR activation to analyze the AMP response to infection using established proteomic approaches.
Aim 3 will demonstrate that nAChR activation participates in the suppression of the AMP response to infection in skin from AD patients using molecular approaches. Our approach is innovative because it utilizes novel proteomic and molecular techniques to further define how nAChR activation influences the epidermal AMP response to infection. The proposed studies are significant because they are anticipated to identify the major nAChR signaling mechanism that negatively regulates AMP activity in skin, and identify alternative pathways for disease progression.
The proposed studies are directly relevant to improving overall human health given that the identification of alternative pathways for disease progression will ultimately impart new targets for preventative and therapeutic interventions for AD and other dermatoses associated with AMP dysregulation. Understanding how the neural system regulates local cutaneous immune responses will disseminate this knowledge to other areas of inflammatory disease research to improve treatment modalities and preventative measures. Ultimately, the proposed research is relevant to the part of the NIH's mission that intends to reduce both the economical and social burdens, and promote the development of more effective treatments to treat cutaneous infection and inflammatory diseases, ultimately enhancing the quality of global public health.
