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microRNA and diabetes

miRCURY LNA™ microRNA Inhibitors
 
Dr. Jan-Wilhelm Kornfeld
Dr. Jan-Wilhelm Kornfeld

Introduction

Dr. Jan-Wilhelm Kornfeld works in the lab of Prof. Jens C. Brüning in the Department of Mouse Genetics and Metabolism at the University of Cologne and the Max-Planck-Institute for Neurological Research (MPI-nF). They have recently published an article in Nature detailing the role of miR-802 in glucose metabolism and insulin resistance in Type2 Diabetes (Kornfeld et al., Nature 2013). Their work using in vivo LNA™ microRNA inhibitors towards miR-802 in mice demonstrates the great future potential for oligonucleotide-based therapeutics for this complex disease.


What is the main focus of the research conducted in your lab?

Our lab aims to define the biology of insulin action and resistance in the central nervous system, especially in the setting of obesity-associated insulin resistance. We try to decipher which neuronal subpopulations translate reduced sensitivity towards insulin actions into the regulation of peripheral glucose homeostasis. In addition we are interested in the cell-autonomous regulation of metabolism via non-coding RNAs such as microRNAs.


How did your research lead to the study of microRNAs?

Dysregulated protein expression and posttranslational modifications cannot fully explain the clinically observed association between obesity and insulin resistance. Genome-wide association studies revealed that the majority of disease-associated variants lie outside of protein-coding genes. We thus reasoned that non-coding RNAs such as microRNAs could be causally involved in the development of obesity-associated insulin resistance.


What was the aim of the project?

We conducted whole-miRNome profiling to identify novel microRNAs possibly involved in the regulation of liver glucose metabolism. Transgenic overexpression of a newly defined candidate microRNA, miR-802, revealed the impairment of hepatic glucose homeostasis under conditions of miR-802 excess. To complement our findings we aimed at specifically depleting the levels of hepatic miR-802 using Locked Nucleic Acids (LNA™) containing inhibitors and hypothesized that this might ameliorate hepatic insulin action.


How did you perform the experiments and analyze the results?

We designed an experimental setup in which we treated obese mice twice on two consecutive days using anti-miR-802 LNA™ inhibitors at a dose previously published. The animals tolerated the LNA™ oligos well and we observed a profound repression of 80-90% of hepatic miR-802 levels. miR-802 was also significantly reduced in kidneys but no other metabolically relevant tissues. Following this, we applied glucose-tolerance tests as well euglycemic-hyperinsulinemic clamp analyses and could demonstrate that reducing miR-802 levels in vivo indeed improved glucose homeostasis in obese mice and led to an amelioration of insulin responsiveness.


How do you feel about your results?

We are optimistic that by showing the efficacy and specificity of anti-microRNA treatment using LNA™ oligonucleotides in obese mice we are one step further towards designing sequence-specific anti-RNA treatment of complex disease such as obesity-associated type 2 diabetes mellitus.


Why did you choose to use the LNA™ enhanced inhibitors from Exiqon?

We chose to use LNA™ inhibitors from the beginning on because of the extensive body of literature showing the successful use of LNA™ microRNA inhibitors for in vitro and in vivo depletion of microRNAs.


What do you find to be the main advantage of the LNA™ microRNA inhibitors from Exiqon?

The rapid generation and delivery of LNA™ microRNA inhibitors allowed for the quick execution of in vivo experiments and offered an attractive alternative to the generation of conventional microRNA knockout models.The addition, the LNA™ inhibitors are highly effective. We routinely apply LNA™ oligonucleotide-mediated depletion of non-coding RNAs in vitro and in vivo and usually end up with a suppression of >60% of cellular RNA levels, irrespective of the initial RNA expression levels.


What would be your advice to colleagues about getting started with in vivo microRNA inhibition?

Using in vitro experiments, the best concentration as well as the specificity and efficacy of the LNA™ microRNA inhibitors should be determined. Also proof-of-principle approaches for determining the best pharmacological dose for microRNA silencing in vivo should be assessed for each inhibitor.


What are the future perspectives for this research?

In the future we would like to address the role of other non-coding RNAs in the development of obesity-associated insulin resistance. Here RNA-Sequencing efforts will hopefully identity novel (long) non-coding RNAs regulating glucose metabolism, the molecular mode-of-action of which will be elucidated using RNA pull-down experiments as well as LNA ™ oligonucleotide -mediated depletion approaches.



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