Kidneys perform the vital role of filtering waste products from the blood. Yet the complete catalogue of constituents that comprise these filters is not known. In new work, analyses of extracellular proteins present in specialised filtration units called glomeruli reveal a composition far more complex than previously appreciated.

Extracellular proteins of the glomerulus // Image by Adam Byron

Extracellular proteins of the glomerulus

Kidneys have the unenviable, but life-sustaining, task of filtering about 100 ml of blood per minute: that’s around 150 litres of fluid every day. This filtering gets rid of waste products of the body’s metabolism. (Kidneys also have other important jobs, such as keeping the body chemically balanced and supplied with hormones.)

Glomeruli, the kidneys’ specialised sieving structures, ensure that unwanted substances are removed from the bloodstream while necessary ones are retained. This selectivity is controlled by a combination of cellular and extracellular components of glomeruli, which act together to maintain the integrity of the filtration barrier.

When the integrity of this barrier fails, proteins, which are normally retained in the blood, are excreted into the urine. It was Richard Bright, Bristol born and a University of Edinburgh alumnus, who first described in 1827 the presence of serum albumin (a blood plasma protein) in the urine of patients with oedema. Bright linked this symptom to structural changes in the patients’ kidneys (and went on to earn the affectionate title of the Father of Nephrology). Indeed, the appearance of large amounts of protein in the urine, known as proteinuria, is strongly associated with the progression of kidney disease.

There are many causes of chronic kidney disease, including inherited conditions, such as polycystic kidney disease, diseases such as hypertension (high blood pressure) and diabetes, infections and long-term drug use. In common to all these potential causes is damage to the kidneys’ filtration apparatus.

The main components of the glomerular filtration barrier are glomerular endothelial cells, which line the blood vessels, podocytes, cells that wrap around the outside of blood vessels and control the rate of filtration, and the glomerular basement membrane, a layer of extracellular matrix that separates the blood from the filtrate that will become urine.

The complexity of components is clearly complicated.

A need for a better understanding of how the filtration barriers work in health and disease motivated a research team led by Rachel Lennon (University of Manchester) to examine the extracellular components of glomeruli. In collaboration with researchers from the University of Manchester (including me, in my former role), University of Bristol and Vanderbilt University Medical Center, the team set out to analyse the set of extracellular proteins that make up glomeruli – the glomerular extracellular matrix proteome.

The extracellular matrix is a relatively understudied collection of molecules in general, especially using approaches that enable most or all proteins to be studied together – termed proteomics. Yet extracellular proteins are essential for building and maintaining the structure of tissues and organs and for interacting with and signalling to cells to control their behaviour. For the glomerular filtration barrier in particular, of course, the extracellular basement membrane, separating blood from urine, is critical for its function.

So the team isolated extracellular matrix from glomeruli as well as glomerular endothelial cells and podocytes, and this was analysed using mass spectrometry, a powerful technique that can identify and quantify proteins in samples such as this.

The experiments generated a lot of data on the composition of glomerular extracellular matrix, which are reported in two articles, published online (and open access) in the Journal of the American Society of Nephrology today.

The first paper reports the previously unappreciated complexity of the extracellular matrix in glomerular tissue. The findings suggest that many more extracellular proteins may contribute to the functioning of the glomerular filtration barrier, which has implications for our understanding of kidney function in health and disease. To work out the relative importance of these extracellular components, their precise roles in glomeruli must now be the subjects of future studies.

The second paper shows that there is a complex interplay between the two main cell types of the glomerular filtration barrier. Glomerular endothelial cells and podocytes contributed distinct subsets as well as a shared subset of proteins to the assembly of glomerular extracellular matrix. Interestingly, when the cells were grown together (co-cultured) in the lab, the composition and organisation of the extracellular matrix changed to resemble the glomerular basement membrane better, suggesting an important role for crosstalk between the two cell types in the kidney.

Glomerular cell co-culture could not fully recapitulate the composition of glomerular matrix seen in tissue, however, so there was clearly something missing from the experimental system. This was not surprising, given the complex environment in which glomeruli reside in the body and the abundance of stimuli that they encounter. It will be very interesting to see if improved cell culture systems that mimic more accurately kidney physiology can add insights to the co-culture model of glomerular extracellular matrix.

Taking the data from both papers together, it also appears that a core network of structural extracellular proteins are central to glomerular extracellular matrix assembly. It would be interesting to know whether these proteins have distinct jobs in the matrix or whether the network properties generated by many multimolecular interactions dominate. Insights such as this may enable mathematical modelling of the assembly and integrity of the filtration barrier, which could pave the way to predicting new therapeutic approaches to tackle barrier failure and kidney disease.

As a coda, the complexity of components is clearly complicated here. But biology is complicated, life is complicated. Only by considering all of the components of a system together can we expect to understand how biology works, how life works. Systems biology and systems medicine, as fields that attempt to address this challenge in a quantitative way – informed by approaches such as proteomics – offer hope to understand fundamentally the complexity of life, in health and in disease. But more on that another time…

Funding
This work was supported by the Wellcome Trust.

Citations
Lennon et al. Global analysis reveals the complexity of the human glomerular extracellular matrix. J. Am. Soc. Nephrol. doi:10.1681/ASN.2013030233 (2014). HTML | PDF

Byron et al. Glomerular cell crosstalk influences composition and assembly of extracellular matrix. J. Am. Soc. Nephrol. doi:10.1681/ASN.2013070795 (2014). HTML | PDF

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