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microRNAs and diabetes

miRCURY LNA™ microRNA Arrays

Himawan Harryanto
Himawan Harryanto at the University of Adelaide, Australia, studies the effect of fetal growth restriction on the development of diabetes in adult life. He has used Exiqon's microRNA microarray system to profile microRNAs in the pancreas. 

1. What is the main focus of the research conducted in your lab and how did it lead to the study of microRNAs?

We are studying the mechanistic links between fetal growth restriction and development of diabetes in adult life. Identifying influential links will allow us to design and test for interventions. We have been using a sheep animal model to study this human condition.

One important characteristic of fetal growth restricted offspring is that they do not receive adequate essential nutrients across the placenta from the mother; the reduced and altered abundance of oxygen, glucose and other nutrients, together with abnormal hormone levels in the intrauterine environment could alter gene expression epigenetically and long term in offspring. Epigenetics are the study of heritable changes that occur due to mechanisms other than changes in DNA sequence. Recent studies have shown that microRNA gene expression can be epigenetically regulated and is therefore a strong candidate molecular pathway for early life programming of diabetes.

2. Tell us about how you became interested in the role of microRNA in early life programming of diabetes?

We have previously shown that there were two outcomes that caused the development of diabetes in sheep that were subjected to fetal growth restriction: reduced insulin production from the pancreas in older sheep offspring (Gatford et al. 2008) and lack of insulin sensitivity in the target tissues: skeletal muscle (Jaquet et al. 2001), liver (Thorn et al. 2009) and adipose tissue (Ozanne et al. 2000). The cause of these reductions in the two arms of insulin action is still unknown. In our current study, we are testing whether altered expression and action of microRNAs following fetal growth restriction could play a major role in altering insulin sensitivity of skeletal muscle, liver and adipose, through targeting of the insulin signaling pathway or pancreatic insulin production, by affecting its molecular determinant.

3. What is the aim and future plans for your current study?

The main aim of this study was to identify whether fetal growth restriction would alter pancreatic expression of microRNAs known or predicted to target key molecules involved in insulin production and release, including those directly shown to be essential for this. We then intend to determine if epigenetic changes following fetal growth restriction occur and could be responsible for any altered pancreatic microRNA expression. The long term goal is then determine if normalizing pancreatic microRNA expression or abundance following fetal growth restriction improves the production of insulin and prevents onset of diabetes

4. How far are you in the study process and what are the next steps after expression analysis of your samples?

The results from the miRCURY LNA™ microarrays gave us an important insight into pancreatic microRNA expression in adult offspring, following fetal growth restriction. They showed that microRNA expression in the pancreas of the sheep is similar to that in other species so far studied with pancreatic and islet specific microRNAs present. In addition, candidate microRNAs whose pancreatic expression may be altered by fetal growth restriction long term have been identified. Currently, we are using qPCR to validate the microarray data. Using predicted target database algorithms (miRecords, Xiao et al. 2009), we are identifying the predicted targets of any differentially expressed microRNAs.

This list of targets will be analyzed further with Ingenuity Pathway Analysis (IPA) to find any pathways and networks that may be affected by differentially expressed microRNAs. Especially for those known to be essential to beta cell or islet function in terms of insulin secretion.

5. What do you find to be the main benefits of Exiqon's LNA™ based MicroRNA microarrays?

We found that Exiqon's LNA™ microRNA microarrays were versatile and easy to use. On top of that the user manual provided from Exiqon was detailed and complex but yet easy to understand, even for the first time user. Moreover, the technical support we received from Exiqon was very helpful.

6. How did you run the microarray experiment and how did you analyze the data from the microRNA microarrays?

As suggested by the user manual, we ran the hybridization using NimbleGen/MAUI® 4-Bay hybridizations stations. This station allowed us to run 4 arrays at the same time and able to maintain constant temperature to run this array.

Adelaide Microarray Centre helped us in analyzing the data. Microarray data was analyzed using R software with LIMMA plug in (WEHI, Australia) with the global loess normalization routine. We received a lot of information from Exiqon technical support to analyze this data.

7. What significance will the microarray grant from Exiqon have on your research?

The grant is helping us to set up a very important foundation for our future research in this area. The outcomes of this study also add further evidence that some microRNA genes are heavily conserved, at least, across mammalian species.

8. What are the next steps in your microRNA research, when you complete this specific study?

After we identify which target(s) is (are) affected by differentially expressed microRNAs we will priorities those related to insulin secretion and directly test their interactions using reporter assays in relevant cell lines. This will be followed by manipulating the expression of particular microRNAs in vitro in these cell lines to show and further investigate how these microRNAs may affect insulin production.

9. When and where will we hear more about your studies?

We hope we are able to publish this study in the not to distance future.



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