The role of uterine NK cell subsets in mediating placental implantation
Lead Research Organisation:
Imperial College London
Department Name: Metabolism, Digestion and Reproduction
Abstract
The placenta implants into the lining of the womb at the beginning of pregnancy. Sometimes this does not happen properly and the pregnancy fails, for example by an early pregnancy loss. However, sometimes the placenta does implant, but incompletely. In this case, we tend to see problems at the end of pregnancy, such as pre-eclampsia, the baby not growing properly, and preterm labour. All of these can result in preterm birth, which affects 15 million babies every year, with 1 million of them dying. Of those babies that survive, one in ten will have a disability. Therefore, research into how the placenta implants has the potential not just to help women conceive, but also to improve the health of babies by avoiding them being born too soon.
The lining of the womb is rich in immune cells called "uterine natural killer cells", or "uNK". These cells are thought to help the placenta to implant, but we don't yet know how. We have also recently discovered that there are three types of uNK: "uNK1", "uNK2" and "uNK3". Each of these is likely to have different roles, and perhaps only one of them is crucial for placental implantation.
I want to find out which uterine NK cell subset mediates placental implantation, and how it does it.
To do this, I will look at samples of the lining of the uterus from fertile women who are coming to the hospital to have a contraceptive coil fitted, compared to women attending our fertility clinic either because they have suffered more than two miscarriages, or more than two failures of implantation following IVF. I will use a technique called "flow cytometry" to look at cells from my participants one at a time and find out how many of each cell there are, and how active they are. If one particular subset is less numerous or active in women who have problems with placental implantation, that will suggest that these cells are important for implantation.
To back up my conclusions from the clinic, I will also do some more intensive laboratory work. I will develop two models of placental implantation using real placental cells, one in a microchip and one in a dish. I will then see how adding pure preparations of each of the uNK subsets changes the way placental cells behave in these two models: when I add the subset that is important for implantation, it will improve the growth and migration of the placental cells. I will use these models of implantation to help me understand how the uNK help placental implantation. For example, I can block the action of certain molecules that uNK make: if these molecules are important for implantation, blocking them will make the placental cells grow and move less well. Conversely, if I add more of those molecules, and they are important for implantation, they will improve the growth and migration of the placental cells.
Doing these experiments will allow me to identify which uNK subset is not working properly in clinical conditions characterised by poor placental implantation, and confirm my finding in two laboratory models of implantation. I will also use these models to begin to identify the molecules uNK make to help promote implantation. This will be an essential first step in developing diagnostic tools to identify women who are likely to suffer from disorders of pregnancy caused by insufficient implantation, and perhaps even promoting interventions that can promote placental implantation, ultimately reducing the burden of preterm birth associated with insufficient implantation.
The lining of the womb is rich in immune cells called "uterine natural killer cells", or "uNK". These cells are thought to help the placenta to implant, but we don't yet know how. We have also recently discovered that there are three types of uNK: "uNK1", "uNK2" and "uNK3". Each of these is likely to have different roles, and perhaps only one of them is crucial for placental implantation.
I want to find out which uterine NK cell subset mediates placental implantation, and how it does it.
To do this, I will look at samples of the lining of the uterus from fertile women who are coming to the hospital to have a contraceptive coil fitted, compared to women attending our fertility clinic either because they have suffered more than two miscarriages, or more than two failures of implantation following IVF. I will use a technique called "flow cytometry" to look at cells from my participants one at a time and find out how many of each cell there are, and how active they are. If one particular subset is less numerous or active in women who have problems with placental implantation, that will suggest that these cells are important for implantation.
To back up my conclusions from the clinic, I will also do some more intensive laboratory work. I will develop two models of placental implantation using real placental cells, one in a microchip and one in a dish. I will then see how adding pure preparations of each of the uNK subsets changes the way placental cells behave in these two models: when I add the subset that is important for implantation, it will improve the growth and migration of the placental cells. I will use these models of implantation to help me understand how the uNK help placental implantation. For example, I can block the action of certain molecules that uNK make: if these molecules are important for implantation, blocking them will make the placental cells grow and move less well. Conversely, if I add more of those molecules, and they are important for implantation, they will improve the growth and migration of the placental cells.
Doing these experiments will allow me to identify which uNK subset is not working properly in clinical conditions characterised by poor placental implantation, and confirm my finding in two laboratory models of implantation. I will also use these models to begin to identify the molecules uNK make to help promote implantation. This will be an essential first step in developing diagnostic tools to identify women who are likely to suffer from disorders of pregnancy caused by insufficient implantation, and perhaps even promoting interventions that can promote placental implantation, ultimately reducing the burden of preterm birth associated with insufficient implantation.
Technical Summary
Insufficient placental implantation is associated with disorders of pregnancy that can lead to preterm birth, including pre-eclampsia, intrauterine growth restriction and preterm labour. Uterine NK cells (uNK) are thought to promote placental implantation, but which of the three recently-defined cell subsets achieves this, and how have not yet been defined.
To address this question in a clinical setting, I will recruit recurrent pregnancy loss (RPL) patients and recurrent implantation failure (RIF) patients, since these patients are suffering a clear failure of implantation. Fertile women attending for coil fitting will be recruited as controls. I will collect endometrial biopsies at LH+7, isolate immune cells and determine uNK subset frequency (CD56, CD49a, CD39, CD103) and function (CD107a, IL-8, XCL1, GM-CSF) by flow cytometry: if any subset is subfunctional in fertility patients, this will suggest it plays a role in implantation. I will also collect information about the outcomes of the cycles of fertility patients, to determine whether any measurement of uNK subset frequency or function is predictive of outcome.
To confirm and extend my findings from the clinical samples, I will also examine the ability of the subsets to promote trophoblast invasion in two in vitro models of implantation: a microfluidics model in which primary trophoblast cells are loaded into a central Matrigel channel and allowed to migrate towards cells/molecules in the side channels, and a trophoblast explant outgrowth model. In both cases, sorted uNK subsets will be added to the coculture and the effect on trophoblast invasion/outgrowth observed. I will also use these models to test the effects of blocking specific soluble molecules made by uNK (eg. M-CSF, XCL1, IL-8) and trophoblast/uNK interactions (eg. CD2, CD7, CD44) to begin to identify the molecular mechanisms by which uNK promote trophoblast invasion/outgrowth.
To address this question in a clinical setting, I will recruit recurrent pregnancy loss (RPL) patients and recurrent implantation failure (RIF) patients, since these patients are suffering a clear failure of implantation. Fertile women attending for coil fitting will be recruited as controls. I will collect endometrial biopsies at LH+7, isolate immune cells and determine uNK subset frequency (CD56, CD49a, CD39, CD103) and function (CD107a, IL-8, XCL1, GM-CSF) by flow cytometry: if any subset is subfunctional in fertility patients, this will suggest it plays a role in implantation. I will also collect information about the outcomes of the cycles of fertility patients, to determine whether any measurement of uNK subset frequency or function is predictive of outcome.
To confirm and extend my findings from the clinical samples, I will also examine the ability of the subsets to promote trophoblast invasion in two in vitro models of implantation: a microfluidics model in which primary trophoblast cells are loaded into a central Matrigel channel and allowed to migrate towards cells/molecules in the side channels, and a trophoblast explant outgrowth model. In both cases, sorted uNK subsets will be added to the coculture and the effect on trophoblast invasion/outgrowth observed. I will also use these models to test the effects of blocking specific soluble molecules made by uNK (eg. M-CSF, XCL1, IL-8) and trophoblast/uNK interactions (eg. CD2, CD7, CD44) to begin to identify the molecular mechanisms by which uNK promote trophoblast invasion/outgrowth.
