Laser microdissection of pancreatic islets allows for quantitative real-time pcr detection of islet-specific gene expression in healthy and diabetic cats

摘要

Background: Feline diabetes mellitus shares many similarities with human type 2 diabetes mellitus, including clinical, physiological and pathological features of the disease. The domestic cat spontaneously develops diabetes associated with insulin resistance in their middle age or later, with residual but declining insulin secretion. Humans and cats share largely the same environment and risk factors for diabetes, such as obesity and physical inactivity. Moreover, amyloid formation and loss of beta cells are found in the islets of the diabetic cat, as in humans. Altogether, the diabetic cat is a good model for type 2 diabetes in humans. The aims of the present study were to isolate feline islets using laser microdissection and to develop a quantitative method for detection of mRNA levels in islets of healthy and diabetic cats.udResults: By using the laser microdissection technique, we were able to meticulously sample islets from both healthy and diabetic cats. Insulin staining of separate sections showed many beta cells in islets from healthy cats, whereas few insulin positive cells were found in islets from diabetic cats. By quantitative real-time PCR, mRNA levels of the islet-specific genes INS, PDX1, IAPP, CHGA and IA-2 could be detected in both healthy and diabetic cats.udConclusions: Laser microdissection allows distinct studies of islets without contamination of acinar cells. Previous attempts in isolating feline islets with different collagenase-based protocols have led to damaged islets or islets coated with exocrine acinar cells, which either way compromise the results obtained from gene expression studies. The use of the laser microdissection technique eliminates these problems as shown in this study. Differences in gene expression between healthy and diabetic cats can reveal underlying mechanisms for beta cell dysfunction and decreased beta cell mass in human type 2 diabetes.udKeywords: Beta cell; Type 2 Diabetes mellitus; Felis catus; Gene expression; Islet isolation; Laser microdissection; Pancreatic islets