Abstract
Diabetes Mellitus is an increasingly prevalent problem worldwide. It is characterised by a dysregulation in the production of insulin or the metabolism of glucose resulting in high plasma glucose levels. Type II diabetes is the more common form making up over 90% of the diabetic population. Type II Diabetes is often the result of the failure of insulin action at the receptor preventing use of glucose as energy. This causes an increase in glucose synthesis to compensate reduction in glucose. The high glucose levels can cause serious complications including retinal damage, cardiovascular dysfunction, and loss of peripheral nerve function. Another common symptom faced by diabetic patients is chronic wounds poor wound healing and amputations. Diabetics are 15% more likely to require amputations than non-diabetic patients due to the reduction in regenerative ability. The mechanism behind the diminished response to wound healing in obese and diabetic patients remains largely unknown. Type II diabetes is commonly thought to be caused by increased visceral fat deposition and obese BMI however there are many genetic components that contribute to the development of type II diabetes. Dysregulation of leptin signalling, and leptin resistance is often seen in combination with insulin resistance. Leptin is a hormone that controls food intake and energy expenditure therefore dysregulation of leptin signalling can cause uncontrolled weight gain. It can also lead to dysregulation of the immune system. Leptin resistance may be a potential mechanism behind the poor regeneration. This thesis examines the effect of leptin on the regeneration of zebrafish caudal fin.
Method: 20-day old zebrafish strains will be used in this experiment. Following anesthetisation with tricaine solution a tail clipping procedure will be performed using a microscope and a sharp scalpel to remove the end portion of the tail. Photos will be taken prior to the tail clipping procedure to establish a baseline, then also immediately following the producer, and day 2, day 4 and day 6. The zebrafish are returned to tank with 300mL of fresh water and dosed with 100nM of leptin, and saline as a control.
Results: Insulin and leucine did not increase the rate of regeneration of wildtype zebrafish. The results show that leptin causes an increase in the regeneration of wild type and Wnt reporter fish zebrafish. This increase in regeneration is not seen in the leptin receptor knockout fish or the leptin receptor crossed with Wnt reporter zebrafish. There is no difference between the rate of regeneration in either the leptin A knockout or the leptin B knockout zebrafish. Leptin has increased the fluorescence in the Wnt reporter zebrafish which is not seen in the leptin receptor crossed with Wnt reporter zebrafish. There was no difference in fluorescence in the leptin A knockout crossed Wnt reporter zebrafish and in the leptin B knockout crossed Wnt reporter zebrafish.
Conclusion: Leucine and insulin have no impact on zebrafish caudal fin regeneration. Leptin increases the rate of regeneration by signalling via through the leptin receptor in zebrafish. Leptin increases regeneration via activation of the Wnt pathway.