Scientists led by a team at the University of Cambridge used human induced pluripotent stem cells (hIPSCs) derived from patients with a genetic mutation that increases the risk of cerebral small vessel disease (SVD) to grow small blood vessel-like models in the lab. The team used these models to show how damage to the scaffolding that supports the vessels can cause them to leak, leading to conditions such as vascular dementia and stroke. The study results identified matrix metalloproteinases (MMP) as a possible drug target to “plug” these leaks and potentially prevent small vessel disease in the brain.
Headed by Alessandra Granata, PhD, at the University of Cambridge Department of Clinical Neurosciences, the team reported on its studies in Stem Cell Reports, in a paper titled “A novel human iPSC model of COL4A1/A2 small vessel disease unveils a key pathogenic role of matrix metalloproteinases.” In their paper the authors noted, “These data provide a basis for targeting MMP as a therapeutic opportunity in SVD.”
Cerebral small vessel disease is a leading cause of age-related cognitive decline and contributes to 45% of dementia cases worldwide, the authors wrote. It is also a common pathology underlying intracerebral haemorrhages (ICH), and is responsible for one in five ischemic strokes, the most common type of stroke, where a blood clot prevents the flow of blood and oxygen to the brain. “SVD refers to the sum of all pathological processes that affect the small vessels of the brain and with an aging population, SVD has major and growing global socio-economic impact,” they continued.
The majority of cases of SVD are associated with conditions such as hypertension and type 2 diabetes and tend to affect people in their middle age. However, there are some rare, inherited forms of the disease that can strike people at a younger age, often in their mid-thirties. Both the inherited and ‘spontaneous’ forms of the disease share similar characteristics. And, they noted, “Emerging evidence supports a role of the extracellular matrix (ECM), at the interface between blood and brain, in the progression of SVD pathology but this remains poorly characterized.”
Despite these clues, the investigators added, “… therapeutic approaches for SVD remain limited due to the lack of mechanistic understanding and relevant models required for target identification and drug discovery.” Granata further pointed out, “Despite the number of people affected worldwide by small vessel disease, we have little in the way of treatments because we don’t fully understand what damages the blood vessels and causes the disease.”
Mutations in collagen type IV, which is a major component of the microvascular ECM, have been implicated as causal in SVD. “COL4A1 and COL4A2 mutations cause highly penetrant multi-system disorders, by disrupting the ECM homeostasis and leading to ICH and porencephaly in human and mouse models,” the investigators noted. For the reported work, scientists at the University of Cambridge Victor Phillip Dahdaleh Heart and Lung Research Institute used cells taken from skin biopsies of two patients with rare forms of SVD caused by mutations in the COL4 gene, one with a mutation in COL4A1, and the other in COL4A2 gene.
By reprogramming the skin cells the scientists were able to create induced pluripotent stem cells, which are cells that have the capacity to develop into almost any type of cell within the body. The team then used these stem cells to brain blood vessels cells, and create a model of the disease that mimics defects seen in patients’ brain vessels. “We used COL4A1/A2 patient derived hiPSC-MC in a co-culture system with brain microvascular endothelial-like cells to mimic the changes seen in patients’ small vessels and to investigate underlying pathological mechanisms,” they explained.
“Most of what we know about the underlying causes tends to come from animal studies, but they are limited in what they can tell us,” Granata noted. “That’s why we turned to stem cells to generate cells of the brain blood vessels and create a disease model ‘in a dish’ that mimics what we see in patients.”
Blood vessels are built around a type of scaffolding, the extracellular matrix, which is a net-like structure that lines and supports the small blood vessels in the brain. The COL4 gene is important for the health of this matrix.
In their disease model, the team found that the extracellular matrix is disrupted, particularly at the tight junctions that “zip” cells together. This disruption leads to the small blood vessels becoming leaky—a key characteristic seen in SVD, where blood leaks out of the vessels and into the brain.
The researchers identified a class of molecules called metalloproteinases that play a key role in this damage. Ordinarily, MMPs are important for maintaining the extracellular matrix, but if too many are produced, they can damage the structure. The COL4A1/A2 models expressed high levels of MMPs.
The team then showed that treating the blood vessels with two MMP inhibitors—the antibiotic doxycycline and the anticancer drug marimastat – reversed the damage and stopped the leakage. Commenting on the study, and noting its limitations, the researchers nevertheless concluded, “These data establish a role for ECM remodelling due to MMPs caused by COL4A1/A2 mutations and provide in vitro evidence that modulating specific MMPs may represent a therapeutic target for SVD.”
Granata acknowledged that the two drugs tested in the study wouldn’t be suitable for clinical use against SVD, but they do demonstrate feasibility of the therapeutic approach. “These particular drugs come with potentially significant side effects so wouldn’t in themselves be viable to treat small vessel disease. But they show that in theory, targeting MMPs could stop the disease. Our model could be scaled up relatively easily to test the viability of future potential drugs.”