Bridging the gap to translation by understanding and preventing diabetic vascular complications using human organoids

Diabetes is a major public health issue with current global prevalence estimated at 8.8% and rising due to the aging population and increasing obesity. The treatment of diabetes is putting a high strain on healthcare resources. 10% of the UK NHS budget is spent on treating diabetes, whilst in Canada the cost of new cases of diabetes diagnosed between 2012-22 is estimated at $15.4 billion. Many of these healthcare costs are due to treating complications associated with diabetes such as kidney disease, blindness, heart attacks, stroke and amputation of lower limbs. These are often caused by changes in the vasculature, therefore, strategies which protect or repair blood vessels have the potential to be new therapies for diabetic patients. Studies using murine models and cultured cells have identified several candidate molecules which protect the diabetic vasculature. However, the challenge is to translate these findings into the clinical setting. For this, we need appropriate experimental models to bridge the gap between rodents and clinical trials.

The research team on this proposal has made a key breakthrough in this area by developing vascular human organoids to model diabetes. Published in Nature (2019), these organoids are derived from human stem cells which are then exposed to a milieu of growth factors to assemble into capillary networks. Exposure of the organoid to high glucose or their transplantation into mice which are subsequently made diabetic leads to structural changes which mimic the changes to blood vessels seen in diabetic patients. Gene expression profiling has revealed molecular pathways altered in the blood vessels when the vascular organoid is exposed to hyperglycaemia. One of these is angiopoietin-2, which modulates blood vessel growth and inflammation and has been shown to drive vascular dysfunction in a murine model of diabetic kidney disease. The second is apelin, a peptide whose blockade has been shown to reduce tumor growth.

This proposal will build on these findings by using the human vascular organoid to examine if blockade of angiopoietin-2 or apelin can prevent the hyperglycaemia induced changes in the human vessel organoid system (Aim 1). Secondly, we will use sophisticated animal models to work out how both angiopoietin-2 (Aim 2) and apelin (Aim 3) in blood vessels precisely effects diabetic vascular complications. Finally, we use the human vessel organoid system to understand why some patients are protected from diabetic vascular complications (Aim 4). To do this, we will use serum samples from type 1 diabetic patients with/without history of albuminuria (protected or susceptible towards the development of vascular disease in their kidneys), a general marker for blood vessel damage. We will expose the human vessel organoids to serum from these two groups of patients and examine changes in organoid structure, cellular composition and gene profile. Finally, we will use non-biased proteomics to assess the composition of the serum to identify protective factors which prevent blood vessel damage in diabetes.

Collectively, our proposal will bridge the gap between rodent studies and clinical trials and test the potential of manipulating angiopoietins and apelin as a therapy for diabetic vascular complications. We predict that using samples from diabetic patients with/without albuminuria will facilitate the discovery of new therapeutic targets to treat diabetic complications in the future.

Aims and Objectives: To (i) determine if modulation of apelin-APLNR or angiopoietin-2 signalling affects hyperglycaemia-induced damage to human vascular organoids; (ii) establish the specific role of endothelial apelin-APLNR and angiopoietin-2 signalling on the diabetic vasculature and (iii) test how human vascular organoids respond to being exposed to serum from diabetic patients with/without vascular complications.

Methods: Apelin-APLNR and angiopoietin-2 signalling will be modulated using recombinant proteins, lentiviral transfections and chemical inhibitors in human vascular organoids cultured in a diabetic medium. Structural and transcriptome analysis will be performed. Novel transgenic mice to modulate endothelial apelin-APLNR and angiopoietin-2 signalling in a spatial- and temporal- specific manner will be used and the effect on the kidney diabetic vasculature explored. Finally, human vascular organoids will be exposed to serum from diabetic patients with/without vascular complications. We will then use proteomics, lipidomics and metabolomics to investigate if there are any protective factors in the serum of patients without vascular complications.

Outcomes: Our proposal will bridge the gap between rodent studies and clinical trials and test the potential of manipulating angiopoietins and apelin as a therapy for diabetic vascular complications. We predict that using samples from diabetic patients with/without albuminuria will facilitate the discovery of new therapeutic targets to treat diabetic complications in the future.

This research has the potential to discover new treatments for diabetic vascular complications. This would have a considerable impact on society by improving the care and quality of life of diabetic patients. Recent figures from the World Health Organisation estimate the global prevalence of diabetes to be 8.8% affecting 422 million individuals and increasing due to the aging population and increasing obesity rates. In the UK 3.8 million individuals are affected by diabetes, whilst in Canada 2.3 million people were diagnosed with diabetes in 2017.

Finding new treatments for diabetic vascular complications will also have a considerable impact on healthcare resources. 10% of the UK NHS budget is spent on treating diabetes, whilst in Canada the cost of new cases of diabetes diagnosed between 2012-22 is estimated at $15.4 billion. Around 80% of this money is spent on complications associated with diabetes such as kidney disease, blindness, heart attacks, stroke and amputation of lower limbs. Our work has the potential to reduce the number of people with diabetic complications and save money on healthcare worldwide.

Our project will also have a significant academic impact. We will provide information on the role of two important molecular pathways that play a critical role in blood vessels in health and disease, apelin-APLNR signalling and angiopoietin-2. We will establish if human vascular organoids can be used as an experimental model to identify underlying molecular mechanisms that cause disease and test new treatments for vascular complications. By using the kidney glomerulus as an experimental model to understand the effect of modulating apelin-APLNR signalling and angiopoietin-2 in the endothelium we will also provide important information as to how these molecules contribute to kidney health.

Our results will also have impact in the pharmaceutical industry who will be interested in generating drugs to target apelin-APLNR signalling and angiopoietin-2. There are several trials on-going examining the effect of apelin on heart failure and pulmonary hypertension. There are also many drugs available to modulate Angpt2 which are being tested in the cancer field which may be helpful for diabetic vascular complications. Using non-biased proteomics, lipidomics, and metabolomics we are also aiming to identify factors that protect patients form diabetic vascular complications. This information is likely to be very informative to the pharmaceutical industry and guide new therapeutic strategies to prevent diabetic vascular complications.

This project will also generate experimental tools which will be of benefit to researchers from a broad range of disciplines including vascular, renal and stem cell biologists. These include the generation of novel mouse models to modulate apelin-APLNR signalling and angiopoietin-2 in the endothelium. These tools will be made publicly available to the research community.

We will establish strong collaborative links between a group of investigators in the UK and Canada with expertise in renal cell biology, vascular biology, developmental biology, stem cells and organoid culture providing a stimulating environment for the research. This is likely to generate new future research directions.