An intravital model to study mechanisms of leukocyte recruitment in experimental crescentic glomerulonephritis (CrGN)

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Glomerulonephritis (GN) means inflammation of the glomeruli, which are the filtering units of the kidney made up of clusters of tiny blood vessels ("capillaries"). In this disease, white blood cells, which are part of the immune system and usually defend the body against infection, accumulate in the glomeruli where they "stick" to the walls of the capillaries and become inappropriately activated, releasing chemicals which cause inflammation and damage to these filtering units, causing kidney dysfunction. GN is a common cause of dialysis-dependent kidney failure, which is associated with long-term health problems and increased mortality. Crescentic GN (CrGN) is a particularly severe form of inflammation in the glomeruli which is potentially life-threatening and, untreated, leads to rapidly progressive kidney failure.GN is currently treated using powerful drugs that suppress the entire immune system, rather than therapy targeted specifically at the disease itself, because the mechanisms by which circulating white blood cells are recruited to and retained in the glomeruli are not well understood. These drugs are not always effective and can cause severe side effects because the immune system is needed to protect the body against infection and cancer. The goal of this research is to understand the mechanisms by which circulating white blood cells are recruited to the glomerulus to cause inflammation and kidney failure so that more targeted, safer therapy can be developed.I will study the mechanisms by which white blood cells "stick" to the glomerular capillary walls and cause inflammation and damage using a technique called intravital microscopy to directly visualise (using a high-powered microscope) their trafficking and behaviour as they travel through the glomerular capillaries in a rat model of CrGN. I will examine the hypothesis that different subpopulations of white blood cells are recruited to particular areas of the glomerulus during the disease process and that this is important in causing the glomerular damage that leads to progressive kidney failure. I will study the mechanisms that recruit these cells to the glomerulus and cause them to be retained there and, importantly, I will study their fate to determine whether they can be detected in urine, which may represent a useful clinical tool to assess disease activity in response to treatment. Importantly, I will use a more clinically relevant model of CrGN than has been previously described to carry out this work. The rat is biologically similar to humans, and experimental CrGN in the WKY rat strain produces changes in the kidney that look strikingly similar to those seen in human CrGN. This is an established model of CrGN which has been used successfully by our research group for decades to study the disease. I will be using the newer but increasingly popular technique of "intravital microscopy" to study the trafficking of white blood cells in "real time" as they travel through the glomeruli of rats that are alive but under general anaesthetic and therefore not in any distress. This much more closely resembles the environment of the human kidney and is therefore more translatable to human disease than other experimental techniques studying the behaviour and function of white blood cells in cell culture systems outside the body ("in vitro" studies). I will undertake the proposed research at Imperial College (IC) where scientists in the kidney unit have an excellent track-record conducting high quality research in the field of GN. Through collaborative work with Prof Aitman's genetics group at IC, our research group has identified a number of genes responsible for susceptibility to CrGN and have developed strains of rats which lack these responsible genes. This will provide me with the tools needed to study mechanisms of white blood cell recruitment and retention in the glomerulus in more detail than has been previously possible in rat models of CrGN.