Impaired NAD+ metabolism as a driver of liver fibrosis
Morten Dall has high hopes for the possible findings of his postdoc project. The overall aim of the project is to shed light on the link between NAD+ metabolism and liver fibrosis (scar tissue in the liver that can cause liver failure), and the hope is that this will make it possible to develop a treatment capable of curbing the development of liver fibrosis, as he writes in his funding application to the Danish Diabetes Academy.
The academy’s international judging panel believes in his ideas and has provided DKK 1.2 million of support – and he will get started next year.
Non-alcoholic steatohepatitis (NASH) is a serious condition that develops from fatty liver. Apart from lifestyle changes, there is currently no effective cure for NASH.
‘We therefore need to understand why the diseases develops from fatty liver, so that we can develop new methods of treatment’, he says.
One piece of information Morten Dall has to go on is that he and the researchers around him have discovered that mice with impaired NAD+ production in the liver develop fibrosis. ‘We have experimentally knocked out the gene that codes for the protein NAMPT, which converts the B3 vitamin nicotinamide to NAD+ in the liver. We have also shown that, if we give the mice a different B3 vitamin - nicotinamide riboside (NR), which is not dependent on NAMPT – in their drinking water, this prevents liver fibrosis. We are currently looking at whether NR can also be used to reverse liver fibrosis. The overall aim of the project is to identify the mechanisms and cell types that are activated during the development of fibrosis’, he explains.
Component parts of the project:
- In the project, the researchers will investigate whether there are differences in the liver’s vitamin B3 metabolism depending on where in the liver’s microstructures the cells are located, and whether this can explain the pattern of liver damage observed in the knock-out mice.
- They will give NR to liver-specific NAMPT knock-out mice over a period from 0 to 6 weeks and monitor whether the liver regenerates when the mice receive NR in their drinking water. This will be evaluated using histological staining of the liver fibrosis.
- They will take blood samples from the mice and test whether their plasma proteins are normalized by the NR treatment. This is because they know from preliminary experiments that the plasma protein profile of NAMPT knock-out mice strongly resembles that of mice and humans suffering from NASH.
- They will use mass spectrometry to measure the protein composition in the liver, providing a general picture of how the liver is affected by NR. These results will go towards understanding the mechanisms initiated in the liver after NR treatment.
- They will analyse tissue sections from the mice using single-molecule fluorescence in situ hybridization (smFISH), a technique in which individual RNA molecules are stained, and which shows where in the tissue a particular gene is expressed. This technique will be set up in collaboration with Professor Shalev Itzkovitz of the Weizmann Institute in Rehovot, Israel, as part of a research visit. Data suggests that NAD+ use in the liver is not homogeneous but varies according to where in the liver’s microstructures the liver cells are located. smFISH will enable precise quantification of this variation, something entirely new in NAD+ research. The same technique can also be used to study where in the liver’s microstructures the effect of NR occurs. This may affect whether NR is usable as a treatment. The researchers will supplement this microscopy data with single-nucleus RNA sequencing, a technique available at the University of Copenhagen’s Center for Basic Metabolic Research.
- By integrating data from these two methods, they will be able to pinpoint where in the liver NR takes effect, and identify the mechanisms activated in individual cells after NR treatment.
- To identify the liver cell populations causing fibrosis and inflammation, they will inactivate specific cell populations by means of glucocorticoids administered with specific antibodies and see what this does to fibrosis development in their mouse model. This is a technique established at the University of Southern Denmark (SDU), and they have therefore made an arrangement with Associate Professor Jonas Graversen of SDU to spend time at his laboratory learning to set up the technique in their own lab.
- They will inactivate different fibrosis-causing cell populations to learn which cells are driving the damage in the mouse model.
NASH is characterized by inflammation, cell death and fibrosis in the liver. It is prevalent in obese people and especially in diabetes patients.
Morten Dall MSc, PhD
Novo Nordisk Foundation Center for Basic Metabolic Research
+45 35 33 70 88