Differential expression of LIGHT and its receptors in human placental villi and amniochorion membranes

Abstract

mRNA encoding LIGHT (homologous to lymphotoxins, exhibits inducible expression, competes with herpes simplex virus glycoprotein D for HVEM, a receptor expressed by T lymphocytes), a member of the tumor necrosis factor superfamily of ligands, as well as mRNAs encoding LIGHT receptors [HVEM, LTbetaR, and TR6 (DcR3)] are present in placentas and cytotrophoblast cells at term. To establish translation of these messages and determine directions for functional studies, term placentas, amniochorion membranes, and purified cytotrophoblast cells were evaluated by immunoblotting and immunohistochemistry. Ligand and receptor proteins were identified in lysates from all three sources although the soluble receptor, TR6, was scarce in placentas and all receptors were in low abundance in cytotrophoblast cells. These results were confirmed and cell type-specific expression was documented by immunohistochemistry. Ligand and receptor proteins were differentially expressed according to cell type. For example, HVEM was identified on syncytiotrophoblast but not in villous mesenchymal cells; amnion epithelial cells were positive for all proteins whereas chorion membrane cytotrophoblasts exhibited none. Because LIGHT is a powerful cytokine that can alter gene expression and promote apoptosis, these experiments suggest that ligand-receptor interactions may critically influence structural and functional aspects of human placentas through as yet undefined autocrine/paracrine pathways.

Figure 1.

A schematic illustration of LIGHT and its related TNF superfamily members. Arrows indicate receptor-ligand interactions.

Figure 2.

RT-PCR of whole placenta, purified cytotrophoblasts, and JAR cells. β-actin was used as a loading control. The sequence of the 470-bp product matches that of human LIGHT.

Figure 3.

Immunoblot of whole placenta, amniochorion and purified cytotrophoblast protein for LIGHT [predicted molecular weight (M.W.), 26.4 kd], TR6 (predicted M.W., 32.7 kd), HVEM (predicted M.W., 30.4 kd) and LTβR (predicted M.W., 46.7 kd). LIGHT migrates at approximately 33 kd, TR6 at 35 kd, HVEM at 50 kd and LTβR at 75 kd. Actual migration significantly differs from the predicted molecular weight for HVEM and LTβR, suggesting post-translational modifications.

Figure 4.

Immunolocalization of LIGHT, TR6, HVEM, and LTβR in term human placenta and amniochorion. A, C, E, and G show placenta and B, D, F, and H show amniochorion immunolocalized for LIGHT, TR6, HVEM, and LTβR and their controls, respectively. Annotations: a, amnion; m, mesenchymal layer; c, chorion; d, decidua. A: Arrow (vertical), syncytiotrophoblast layer; arrow (horizontal), mesenchymal cell. B: Arrow (vertical), amnion; arrow (horizontal), mesenchymal cell; arrow (blue), decidua capsularis. C: Arrow (vertical), mesenchymal cell; arrow (horizontal), syncytiotrophoblast layer. D: Arrow (horizontal), amnion; arrow (vertical), mesenchymal cell; arrow (blue), decidua capsularis. E: Arrow, syncytiotrophoblast layer. F: Arrow (horizontal), amnion; arrow (vertical), decidua capsularis. G: Arrow (vertical), mesenchymal cell; arrow (horizontal), syncytiotrophoblast layer. H: Arrow (vertical), amnion; arrow (horizontal), mesenchymal cell; arrow (blue), decidua capsularis. Table 1 ▶ has antibody specifications. Images captured at magnification, ×200.