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

• Feline diabetes mellitus shares many similarities with human type 2 diabetes mellitus (T2DM), including clinical, physiological and pathological features of the disease. Domestic cats spontaneously develop diabetes associated with insulin resistance in their middle age or later, with residual but declining insulin secretion. Humans and cats share 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 diabetic cat pancreas, as in humans. Subsequently, studying the molecular mechanisms in the failing beta cells may contribute to a better understanding of the pathophysiology of T2DM in both cats and humans. The aim of the present study was to develop a method to study mRNA expression of islet-specific genes in healthy and diabetic cats. 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. By using the laser microdissection technique, we were able to sample islets that were not contaminated with exocrine tissue, from both healthy and diabetic cats. High RNA quality was confirmed with gel electrophoresis. By quantitative real-time PCR (qRT-PCR), mRNA levels of the islet-specific genes insulin, PDX-1, IAPP, CHGA and IA-2 were detected in both healthy and diabetic cats. We used actin b, GAPDH and RPS7 as internal reference genes for normalizations of our qRT-PCR data. The laser microdissection technique allows studies of islets without contamination of acinar cells, as shown in this study, and is of great advantage since it is difficult to get pure feline islets from collagenase-based isolation. Differences in gene expression in healthy and diabetic cats may reveal underlying mechanisms for beta cell dysfunction and decreased beta cell mass in human and feline type 2 diabetes.

Ämnesord

LANTBRUKSVETENSKAPER  -- Veterinärmedicin -- Medicinsk biovetenskap (hsv//swe)

AGRICULTURAL SCIENCES  -- Veterinary Science -- Medical Bioscience (hsv//eng)

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