Insulin sensitivity of the human glomerular podocyte: Its biological importance and the role of nephrin in this process.
Lead Research Organisation:
University of Bristol
Department Name: Academic Renal Unit
Abstract
The leading cause of kidney failure in the world is secondary to diabetes mellitus (DM). In DM there is either a lack of the production, or cellular action, of insulin. The first sign of kidney involvement in DM is the appearance of small amounts of protein in the urine which increases as the kidneys fail.
The glomerular filtration barrier (GFB) is the part of the kidney that acts as a sieve and prevents protein leaking into the urine from the blood. It is made up of two different cell types called podocytes and endothelial cells. We have developed a unique way of studying these cells in the laboratory by culturing them in dishes. Using these cells we have discovered that podocytes are rapidly insulin sensitive, suggesting that insulin sensitivity of the GFB is important in health.
Our research intends to study the precise mechanism that insulin exerts its actions on the GFB. We hope that by understanding the insulin responsiveness of the GFB that we may be able to develop new treatments that will treat and prevent urinary protein loss early in disease and ultimately prevent kidney failure in the setting of diabetes.
The glomerular filtration barrier (GFB) is the part of the kidney that acts as a sieve and prevents protein leaking into the urine from the blood. It is made up of two different cell types called podocytes and endothelial cells. We have developed a unique way of studying these cells in the laboratory by culturing them in dishes. Using these cells we have discovered that podocytes are rapidly insulin sensitive, suggesting that insulin sensitivity of the GFB is important in health.
Our research intends to study the precise mechanism that insulin exerts its actions on the GFB. We hope that by understanding the insulin responsiveness of the GFB that we may be able to develop new treatments that will treat and prevent urinary protein loss early in disease and ultimately prevent kidney failure in the setting of diabetes.
Technical Summary
The leading cause of albumin loss into the urine (albuminuria) and end stage renal failure in the Western world is due to abnormalities of the production of insulin (type-1 diabetes) or its cellular effects (type-2 diabetes). Furthermore, albuminuria occurs in non-diabetic patients who are insulin resistant and is importantly also an independent risk factor for cardiovascular morbidity. Therefore understanding the action of insulin on the filtration barrier of the kidney may lead to novel treatment strategies against these major health care problems.
The filtration barrier of the kidney is found in the glomerulus. It consists of 3 layers: the endothelium lining the blood vessel, the glomerular basement membrane and the podocytes that are adjacent to the urinary space. Intense interest has focused on the role of the podocyte in the prevention of albuminuria in recent years since the landmark discoveries of human single gene defects, that cause early onset massive albuminuria, code for proteins specific to the podocyte. The most severe form of this disease is caused by the mutation of a protein called nephrin. Using the only published immortalised human podocyte cell line in the world which expresses all the phenotypic markers of mature podocytes I have shown differentiated human podocytes are insulin sensitive doubling their glucose uptake within 15 minutes. This is specific to the podocyte in the glomerular filtration barrier as immortalised endothelial cells are insulin resistant. Furthermore by studying podocytes derived from children with mutations of the nephrin gene I have robust evidence that this protein is critical for this process. This project aims to examine the importance of nephrin in the action of insulin on podocytes in three major areas (1) insulin signalling pathways, (2) the reorganisation of the actin cytoskeleton targeting and (3) docking of glucose transporter rich vesicles with the plasma membrane of the cell. Studying the role of nephrin in these three areas will also be important in understanding other fundamental biological properties of the podocyte.
Finally the in vivo biological importance of the effect of insulin on the podocyte will be studied by selectively knocking out the insulin receptor in podocytes in an inducible manner. This will allow the podocyte to develop normally before switching the insulin receptor off, and rendering the podocyte insulin resistant. This will be achieved using a tetracycline inducible, podocyte specific, cre-lox mouse model.
The filtration barrier of the kidney is found in the glomerulus. It consists of 3 layers: the endothelium lining the blood vessel, the glomerular basement membrane and the podocytes that are adjacent to the urinary space. Intense interest has focused on the role of the podocyte in the prevention of albuminuria in recent years since the landmark discoveries of human single gene defects, that cause early onset massive albuminuria, code for proteins specific to the podocyte. The most severe form of this disease is caused by the mutation of a protein called nephrin. Using the only published immortalised human podocyte cell line in the world which expresses all the phenotypic markers of mature podocytes I have shown differentiated human podocytes are insulin sensitive doubling their glucose uptake within 15 minutes. This is specific to the podocyte in the glomerular filtration barrier as immortalised endothelial cells are insulin resistant. Furthermore by studying podocytes derived from children with mutations of the nephrin gene I have robust evidence that this protein is critical for this process. This project aims to examine the importance of nephrin in the action of insulin on podocytes in three major areas (1) insulin signalling pathways, (2) the reorganisation of the actin cytoskeleton targeting and (3) docking of glucose transporter rich vesicles with the plasma membrane of the cell. Studying the role of nephrin in these three areas will also be important in understanding other fundamental biological properties of the podocyte.
Finally the in vivo biological importance of the effect of insulin on the podocyte will be studied by selectively knocking out the insulin receptor in podocytes in an inducible manner. This will allow the podocyte to develop normally before switching the insulin receptor off, and rendering the podocyte insulin resistant. This will be achieved using a tetracycline inducible, podocyte specific, cre-lox mouse model.
