An Yue Tu

Neutralization and Transfer of Lipopolysaccharide by Phospholipid Transfer Protein

Phospholipid transfer protein (PLTP) and lipopolysaccharide-binding protein (LBP) are lipid transfer proteins found in human plasma. PLTP shares 24% sequence similarity with LBP. PLTP mediates the transfer and exchange of phospholipids between lipoprotein particles, whereas LBP transfers bacterial lipopolysaccharide (LPS) either to lipoprotein particles or to CD14, a soluble and cell-surface receptor for LPS. We asked whether PLTP could interact with LPS and mediate the transfer of LPS to lipoproteins or to CD14. PLTP was able to bind and neutralize LPS: incubation of LPS with purified recombinant PLTP (rPLTP) resulted in the inhibition of the ability of LPS to stimulate adhesive responses of neutrophils, and addition of rPLTP to blood inhibited cytokine production in response to LPS. Transfer of LPS by rPLTP was examined using fluorescence dequenching experiments and native gel electrophoresis. The results suggested that rPLTP was able to mediate the exchange of LPS between micelles and the transfer of LPS to reconstituted HDL particles, but it did not transfer LPS to CD14. Consonant with these findings, rPLTP did not mediate CD14-dependent adhesive responses of neutrophils to LPS. These results suggest that while PLTP and LBP both bind and transfer LPS, PLTP is unable to transfer LPS to CD14 and thus does not mediate responses of cells to LPS.

Functional expression of human and mouse plasma phospholipid transfer protein: effect of recombinant and plasma PLTP on HDL subspecies

The molecular cloning of mouse plasma phospholipid transfer protein (PLTP) and the eukaryotic cell expression of complementary DNA for mouse and human PLTP are described. Mouse PLTP was found to share 83% amino acid sequence identity with human PLTP. PLTP was produced in baby hamster kidney cells. Conditioned medium from BHK cells expressing PLTP possessed both phospholipid transfer activity and high density lipoprotein (HDL) conversion activity. PLTP mRNA was detected in all 16 human tissues examined by Northern blot analysis with ovary, thymus, and placenta having the highest levels. PLTP mRNA was also examined in eight mouse tissues with the highest PLTP mRNA levels found in the lung, brain, and heart. The effect of purified human plasma-derived PLTP and human recombinant PLTP (rPLTP) on the two human plasma HDL subspecies Lp(A-I) and Lp(A-I/A-II) was evaluated. Plasma PLTP or rPLTP converted the two distinct size subspecies of Lp(A-I) into a larger species, an intermediate species, and a smaller species. Lp(A-I/A-II) particles containing multiple size subspecies were significantly altered by incubation with either plasma or rPLTP with the largest but less prominent subspecies becoming the predominant one, and the smallest subspecies increasing in concentration. Thus, PLTP promoted the conversion of both Lp(A-I) and Lp(A-I/A-II) to populations of larger and smaller particles. Also, both human PLTP and mouse rPLTP were able to convert human or mouse HDL into larger and smaller particles. These observations suggest that PLTP may play a key role in extracellular phospholipid transport and modulation of HDL particles.

Transgenic mice expressing human phospholipid transfer protein have increased HDL/non-HDL cholesterol ratio

The role of plasma phospholipid transfer protein (PLTP) in lipoprotein metabolism is poorly understood. In vitro studies suggest that PLTP influences HDL size and composition and transfers phospholipids among lipoproteins. To provide an in vivo model for studies of PLTP physiology, transgenic mice that express human PLTP were generated. Human PLTP transcripts were detected in total RNA from adipose tissue, lung, heart, and spleen of the two distinct lines (A and C) of transgenic mice. Despite minimal expression of human PLTP in the liver of these transgenic mice and similar plasma phospholipid transfer activity in transgenic and non-transgenic mice (19.1±3.1 vs 18.9±2.7 μmol/ml/h), differences in lipoprotein levels were observed between transgenic and control mice receiving the same chow diet. Male transgenic mice of line C had significantly higher HDL cholesterol than control mice (76.4±4.6 vs 71.9±7.0 mg/dl, p<0.05) and the male transgenic mice of lines A and C had a significantly lower non-HDL cholesterol (15.1±4.1 and 15.6 ±4.7 vs 20.9±5.5 mg/dl,P<0.01 andP<0.02) and a significantly higher HDL cholesterol/non-HDL cholesterol ratio than the control mice (5.3±1.3 and 5.5±2.2 vs 3.9 ±1.9 mg/dl,P<0.01 andP<0.02). Female mice from transgenic line C had higher HDL cholesterol than control mice (64.6±4.8 vs 57.4±5.1 mg/dl,P<0.01) while female mice from line A tended to have higher HDL cholesterol/non-HDL cholesterol ratio than control mice (5.5±3.7 vs 3.8±1.4). These observations suggest that expression of PLTP in peripheral tissues play an important role in lipoprotein metabolism. Expression of human PLTP produced a more favorable lipoprotein profile and thus, enhanced expression of PLTP could potentially retard atherosclerosis.

MOLECULAR BIOLOGY OF PHOSPHOLIPID TRANSFER PROTEIN

Lipid transfer proteins play an essential role in the intravascular dynamics of lipids among lipoproteins and between lipoproteins and cell membranes. Phospholipid transfer protein has been known for over a decade but, unlike cholesteryl ester transfer protein, has been investigated relatively little with regard to its physiological importance. The recent determination of the phospholipid transfer protein complementary DNA sequence as well as the further characterization of its gene structure will direct future studies toward the understanding of its structure-function correlations, physiological regulation, and clinical assessment at the molecular level. As a member of the lipid-transfer lipopolysaccharide-binding protein gene family, phospholipid transfer protein will attract investigators to studying its possible involvement in lipopolysaccharide or endotoxin interactions in addition to its phospholipid transfer activity.

Quantitative Trait Locus Mapping of Genes That Regulate Phospholipid Transfer Activity in SM/J and NZB/BlNJ Inbred Mice

ObjectivePhospholipid transfer protein (PLTP), an important protein in the transfer of phospholipids between lipoprotein particles and in the remodeling of HDL, is regulated at both the transcriptional and the protein level. We performed quantitative trait locus (QTL) analysis to identify genomic loci regulating PLTP activity in mice. Methods and ResultsPlasma PLTP activity was measured in 217 male F2 progeny from a SM/J × NZB/B1NJ intercross. Two QTL for plasma PLTP activity in mice fed chow (Pltpq1 and Pltpq2) were found on chromosomes 3 (34 cM, logarithm of odds [LOD] 3.5) and 10 (66 cM, LOD 4.1); two additional QTL in mice fed atherogenic diet (Pltpq3 and Pltpq4) were found on chromosomes 9 (56 cM, LOD 4.5) and 15 (34 cM, LOD 5.0); and one QTL (Pltiq1) for the inducibility of PLTP activity was found on chromosome 4 (70 cM, LOD 3.7). Several candidate genes for these 5 QTL were tested by sequence comparison and expression studies. ConclusionsWe identified five significant loci involved in PLTP activity in the mouse and provided supporting evidence for the candidacy of Nr1h4 and Apof as the genes underlying Pltpq2.

Low-Density Lipoprotein Inhibits Secretion of Phospholipid Transfer Protein in Human Trophoblastic BeWo Cells

Human plasma phospholipid transfer protein (PLTP) plays an important role in lipoprotein metabolism. In this study, we investigated the effects of lipoproteins on the secretion of PLTP in cultured BeWo choriocarcinoma cells. Low-density lipoproteins (LDLs) decreased PLTP secretion in a dose- and time-dependent manner, whereas very low density lipoproteins and high-density lipoproteins (HDLs) had little effect. LDL suppression of PLTP secretion was not altered by the inhibition of both LDL receptor and LDL receptor–related protein with receptor-associated protein. Mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor, U0126, could abolish the LDL-mediated inhibition of PLTP secretion. Furthermore, LDL, but not HDL, could stimulate the expression of MAPK phosphatase-1 (MKP-1) in BeWo cells that resulted in the inactivation of p44/p42 extracellular signal-regulated kinase (ERK) 1 and 2, the family members of MAPKs. These results support the conclusion that LDL-mediated suppression of PLTP secretion in BeWo cells is through a LDL receptor-independent MAPK signaling pathway.