Expression of lymphatic vascular endothelial hyaluronan receptor-1 (LYVE-1) in the human placenta.

BACKGROUND: Lymphatic vascular endothelial hyaluronan receptor-1 (LYVE-1) is a selective marker for lymphatic endothelium and a homolog of CD44, the hyaluronan (HA) receptor. HA in the extracellular matrix plays roles in tissue remodeling, development, and homeostasis, and as an HA receptor, LYVE-1 mediated HA metabolism might regulate these events. Currently, little is known about the lymphatic character within the human placenta. This study therefore determined LYVE-1 and other lymphatic markers in human placentas. METHODS AND RESULTS: Placentas and villous tissue were fixed and immunostained for human LYVE-1 and CD44 and examined by RT-PCR. LYVE-1 was expressed at both protein and mRNA levels in trophoblast cells (TC) and in villous core endothelium (VCE). Predominant protein expression for LYVE-1 was observed in syncytiotrophoblast cells (TCs) of preterm placentas. Neither mRNA or protein for CD44 was expressed. Other blood and lymphatic-lineage molecules (VEGF-A, -C, and -D, Flt-1, KDR, Flt-4, and Prox-1) were examined by RT-PCR. VEGF-A, VEGF-D, and Flt-1 mRNA were observed in TCs and VCEs, while mRNA for VEGF-C, KDR, and Flt-4 was mainly observed in VCEs. Prox-1 was found at the mRNA, but not protein level in TCs and VCEs. Our findings indicate (1) the importance of LYVE-1, but not CD44, in regulation of HA metabolism in the maternal-fetal interface and fetal circulation, and (2) possible dual blood and lymphatic phenotypic characteristics in fetal endothelium. These results provide new insights into HA metabolism and lymphatic-lineage molecule expression in the human placenta.

Down-regulation of TIMP3 leads to increase in TACE expression and TNFα production by placental trophoblast cells.

PROBLEM: To determine whether down-regulation of TIMP3 expression promotes TACE expression and increases in TNFα production by placental trophoblast cells. METHOD OF STUDY: Placental expression of TIMP3 and TACE was examined by immunostaining and Western blot. Effects of TIMP3 on TACE expression and TNFα production were assessed by transfection of TIMP3 siRNA into trophoblasts isolated from normal placentas. Effects of oxidative stress on trophoblast TIMP3 expression and TNFα production were also determined. Trophoblast production of TIMP3, TACE and TNFα were measured by ELISA. RESULTS: TIMP3 expression was markedly reduced in preeclamptic placentas compared with normal placentas; oxidative stress down-regulated trophoblast TIMP3 expression and production, P < 0.01. Down-regulation of TIMP3 expression by TIMP3 siRNA resulted in significant increases in TACE expression and TNFα production, P < 0.01. CONCLUSION: As TIMP3 is an endogenous TACE inhibitor, down-regulation of trophoblast TIMP3 expression/activity could result in increased TACE expression and subsequently lead to increased TNFα production in preeclamptic placentas.

Activation of vitamin D receptor promotes VEGF and CuZn-SOD expression in endothelial cells.

Endothelial dysfunction associated with vitamin D deficiency has been linked to many chronic vascular diseases. Vitamin D elicits its bioactive actions by binding to its receptor, vitamin D receptor (VDR), on target cells and organs. In the present study, we investigated the role of VDR in response to 1,25(OH)2D3 stimulation and oxidative stress challenge in endothelial cells. We found that 1,25(OH)2D3 not only induced a dose- and time-dependent increase in VDR expression, but also induced up-regulation of vascular endothelial growth factor (VEGF) and its receptors (Flt-1 and KDR), as well as antioxidant CuZn-superoxide dismutase (CuZn-SOD) expression in endothelial cells. We demonstrated that inhibition of VDR by VDR siRNA blocked 1,25(OH)2D3 induced increased VEGF and KDR expression and prevented 1,25(OH)2D3 induced endothelial proliferation/migration. Using CoCl2, a hypoxic mimicking agent, we found that hypoxia/oxidative stress not only reduced CuZn-SOD expression, but also down-regulated VDR expression in endothelial cells, which could be prevented by addition of 1,25(OH)2D3 in culture. These findings are important indicating that VDR expression is inducible in endothelial cells and oxidative stress down-regulates VDR expression in endothelial cells. We conclude that sufficient vitamin D levels and proper VDR expression are fundamental for angiogenic and oxidative defense function in endothelial cells.

Limited density of an antigen presented by RMA-S cells requires B7-1/CD28 signaling to enhance T-cell immunity at the effector phase.

The association of B7-1/CD28 between antigen presenting cells (APCs) and T-cells provides a second signal to proliferate and activate T-cell immunity at the induction phase. Many reports indicate that tumor cells transfected with B7-1 induced augmented antitumor immunity at the induction phase by mimicking APC function; however, the function of B7-1 on antitumor immunity at the effector phase is unknown. Here, we report direct evidence of enhanced T-cell antitumor immunity at the effector phase by the B7-1 molecule. Our experiments in vivo and in vitro indicated that reactivity of antigen-specific monoclonal and polyclonal T-cell effectors against a Lass5 epitope presented by RMA-S cells is increased when the cells expressed B7-1. Use of either anti-B7-1 or anti-CD28 antibodies to block the B7-1/CD28 association reduced reactivity of the T effectors against B7-1 positive RMA-S cells. Transfection of Lass5 cDNA into or pulse of Lass5 peptide onto B7-1 positive RMA-S cells overcomes the requirement of the B7-1/CD28 signal for T effector response. To our knowledge, the data offers, for the first time, strong evidence that supports the requirement of B7-1/CD28 secondary signal at the effector phase of antitumor T-cell immunity being dependent on the density of an antigenic peptide.