A new in vivo model for imaging axon dynamics in injured diabetic zebrafish
Diabetes mellitus currently affects over 26 million adults in the United States and 285 million people worldwide. These numbers are expected to nearly double in the next two decades. Neuropathy-mediated wound healing complications are a major problem in diabetes, with diabetic foot ulcerations (DFU) preceding approximately 84% of lower limb amputations. Long-term intensive care and consequences of amputation place immense burdens on individuals and their families as well as healthcare systems. While interdisciplinary clinical groups have made certain advances in DFU prevention and control of the disease, higher success could be achieved through a better understanding of the underlying causes. This lack of knowledge mostly derives from the complex etiology of DFU. Although the use of mammalian model organisms has proven to be highly important for the investigation of diabetes-associated complications, they are subject to technical limitations and high costs. New low-cost model systems are required allowing for in vivo analyses of the complexity of axon-wound interactions in live animals. With the power of in vivo imaging new concepts underlying the etiology of DFU could emerge. Here we propose to develop zebrafish larvae as a new in vivo model for elucidating basic mechanisms of DFU. Zebrafish larvae have several advantages over other model organisms in that they are optically clear and develop rapidly. This makes them suitable for genetic and pharmacological manipulations and for in vivo imaging. In addition, glucose metabolism in larval and adult zebrafish is highly similar to humans. Thus zebrafish larvae are ideal for investigations of diabetic complications. The goal of this proposal is to combine our previously established axon regeneration model in zebrafish larvae with genetic ablation of insulin-secreting pancreatic b-cells to analyze the behavior of acutely injured and neuropathic cutaneous axons under hyperglycemic conditions. This model will allow us to address basic questions that are key to understanding DFU etiology: 1) is there a temporal relationship between b-cell death and onset of neuropathy? 2) Do acutely injured cutaneous axons regenerate under hyperglycemic conditions? And 3) does acute injury initiate or aggravate neuropathy? Understanding these basic mechanisms will be fundamental for investigations on neuropathy-related chronic wound formation. This work has the potential for future pharmacological studies in zebrafish larvae with the goal to identify drugs that suppress the onset of neuropathy and prolong axon health.

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