Serum-based, diagnostic, biological assay to predict pregnancy disorders

Abstract

The invention provides serum-based, diagnostic, biological assays for predicting disorders of pregnancy resulting from poor trophoblast and/or placental ischemia, including preeclampsia. Serum samples from such subjects exhibit an ability to disrupt the architecture involving fetal trophoblasts and maternal endothelial cells in a three-dimensional, dual cell co-culture system provided herein, in contrast to normal pregnancy serum samples. Based on these distinctions, the assays are employed to predict pregnancy outcomes as early as first trimester.

1 . An assay for assessing whether a pregnant female is at risk of developing a pregnancy disorder characterized by either poor trophoblast invasion or placental ischemia, or both, comprising (a) incubating a co-culture of human endothelial cells and human trophoblast cells in the presence of serum obtained from a pregnant female for a period of time sufficient to permit vacuolization, and (b) after incubation, determining whether substantial vacuolization in the co-culture has occurred. 
     
     
         2 . The assay according to  claim 1  wherein the disorder is selected from preeclampsia, gestational diabetes, intrauterine growth restriction, and trophoblasitic diseases with hyper- or hypo invasive features. 
     
     
         3 . The assay according to  claim 2 , additionally comprising the steps of incubating a co-culture of human endothelial cells and human trophoblast cells under the same conditions as the co-culture in  claim 1 (a) in the absence of serum or plasma from a pregnant female for a period of time sufficient to permit vacuolization and after incubation determining whether substantial vacuolization in either co-culture has occurred. 
     
     
         4 . The assay according to  claims 1 - 3  wherein incubation time is at least 12 hours. 
     
     
         5 . The assay according to  claim 4  wherein the serum is from the first trimester of the pregnant female's pregnancy. 
     
     
         6 . The assay according to  claim 5  wherein the amount of serum used is 0.05-2 ml. 
     
     
         7 . The assay according to  claim 6  wherein the endothelial cells are selected from umbilical vein, uterine, myometrial, decidual, aorta, microvascular, dermal or other endothelial cells that express VE-cadherin, PECAM, and/or Aquaporin 1. 
     
     
         8 . The assay according to  claim 6  wherein the trophoblasts are selected from primary villous or extravillous trophoblasts, HTR8 and 3A trophoblasts or other invasive trophoblasts isolated from choriocarcinomas. 
     
     
         9 . The assay according to  claim 7 - 8  wherein the co-culture is performed on a natural or synthetic soluble matrix. 
     
     
         10 . The assay according to  claim 9  wherein the matrix is selected from matrigel, collagen, fibronectin, elastin and combinations thereof. 
     
     
         11 . The assay according to  claim 10  wherein the incubation step is performed at 37° C. in carbon dioxide. 
     
     
         12 . A method of screening for agents to treat a pregnancy disorder characterized by either poor trophoblast invasion or placental ischemia, or both, comprising (a) administering a potential endothelial-trophoblast cross-talk restoring agent to an IL 10 null animal that has previously been administered human PE serum and is exhibiting PE-associated symptoms, and (b) observing whether the PE-associated symptoms are alleviated thereby. 
     
     
         13 . The method according to  claim 12  wherein the animal is a mouse. 
     
     
         14 . The method according to  claim 12 - 13  wherein the disorder of preeclampsia. 
     
     
         15 . A method of treating a pregnant female human patient suffering from a placental ischemia disorder comprising administering to the patient a therapeutically effective amount of a compliment split C5a modulating agent. 
     
     
         16 . A method of screening for agents useful in the treatment pregnancy disorders characterized by either poor trophoblast invasion or placental ischemia, or both, comprising (a) treating co-cultured endothelial cells and trophoblast cells with a potential endothelial-trophoblast cross-talk restoring agent in the presence of preeclampsia serum and in the absence of preeclampsia serum, and (b) comparing the results to determine whether the preeclampsia serum induced disruption of endothelial-trophoblast cross-talk is reversed thereby. 
     
     
         17 . A method of screening for agents useful as contraceptives or abortificients, comprising (a) treating co-culture of endothelial cells and trophoblast cells with a potential contraceptive or abortificient agent in the presence of normal pregnancy serum and in the absence of normal pregnancy serum, and (b) comparing the results to determine whether the normal endothelial-trophoblast cross-talk is disrupted thereby.