Beneficial effects of ApoA-I on LPS-induced acute lung injury and endotoxemia in mice

High density lipoprotein (HDL) binds lipopolysaccharide (LPS) and neutralizes its toxicity. The aim of our study was to investigate the effects of Apolipoprotein (ApoA-I), the major apolipoprotein of HDL, on LPS-induced acute lung injury (ALI) and endotoxemia. BALB/c mice were challenged with LPS, followed by ApoA-I or saline administration for 24h. The mice were then sacrificed and histopathological analysis of the lung was performed. We found that ApoA-I could attenuate LPS-induced acute lung injury and inflammation. To investigate the mechanisms, we measured tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) levels in the serum and bronchoalveolar lavage (BAL) fluid and found that ApoA-I could significantly inhibit LPS-induced increases in the IL-1beta and TNF-alpha levels in serum (P<0.05, respectively), as well as in the IL-1beta, TNF-alpha, and IL-6 levels in BAL fluid (P<0.01 and P<0.05, P<0.05, respectively). Moreover, we evaluated the effect of ApoA-I on the mortality of L-929 cells which were attacked by LPS-activated peritoneal macrophages. We found that ApoA-I could significantly inhibit the LPS-induced cell death in a dose-dependent fashion. Furthermore, we investigated in vivo the effects of ApoA-I on the mortality rate and survival time after LPS administration and found that ApoA-I significantly decreased the mortality (P<0.05) and increased the survival time (P<0.05). In summary, the results suggest that ApoA-I could effectively protect against LPS-induced endotoxemia and acute lung damage. The mechanism might be related to inhibition of inflammatory cytokine release from macrophages.     

Searching for a successful HDL-based treatment strategy

Despite strong evidence that HDL-cholesterol levels predict atherosclerotic events in a population, attempts at using an HDL-based treatment strategy have not yet been successful. Most of the efforts to date have focused on raising plasma HDL-cholesterol levels. This brief review focuses on a different strategy, which is based on the use of 18-amino acid apoA-I mimetic peptides. The story of these peptides spans decades and illustrates the remarkable complexity of HDL-based treatment strategies, but suggests that such a strategy may still be successful.     

A novel polymorphism of apolipoprotein A-IV is the result of an asparagine to serine substitution at residue 127

We have identified a hitherto genetic polymorphism of apolipoprotein A-IV (apo-IV). The molecular basis for this polymorphism is an A to G substitution at nucleotide 1687 resulting in an Asn to Ser change of amino acid 127. The frequencies of the two apoA-IV alleles (designated apoA-IV^127Asn and apoA-IV^127Ser), determined by Hinc II restriction analysis of PCR amplified exon three of the apoA-IV gene, were 0.788 and 0.212, respectively, in a Finnish population sample. Allele frequencies of another polymorphism due to a Thr to Ser substitution at amino acid 347 were determined using Hinf I restriction analysis. The allele frequencies were 0.823 for apoA-IV^347Thr and 0.177 for apoA-IV^247Ser. None of the apoA-IV polymorphisms (apoA-IV_{127:Asn→Ser}, apoA-IV_{347:Thr→Ser} and apoA-IV_{360:Gln→His}) had any effect of plasma lipid and lipoprotein concentrations in cohorts of dyslipidemic men and in a population sample of normolipidemic controls. There was also no association between the history of previous myocardial infarction and any of the apoA-IV alleles.     

Effect in vitro of apolipoprotein A-V on the structure and functions of recombinant high density lipoprotein

The neighboring position of apolipoprotein A-I (apoA-I) and apolipoprotein A-V (apoA-V) gene and the modulation of apoA-V on the concentrations, size and maturation of high density lipoprotein (HDL) may indicate a special relationship between apoA-V and HDL. To assess the effects of apoA-V on HDL structure and related functions in vitro, a series of recombinant HDL (rHDL) were synthesized in vitro with various mass ratios of recombinant apoA-I: apoA-V. An increase in apoA-V in rHDL resulted in enhanced lipid-binding ability, increased phospholipid content and larger particle size. Furthermore, the lipid-free and lipid-bound apoA-V in rHDL showed antioxidant capacity against low density lipoprotein (LDL) in vitro. In THP-1 derived macrophages, apoA-V of rHDL was shown to have no influence on the uptake of oxidized LDL (oxLDL) and intracellular lipid accumulation. Thus, the addition of apoA-V to rHDL resulted in changes in several rHDL properties, including increased lipid-binding ability, phospholipid content, particle size and antioxidant capacity. These alterations may explain the modulation of apoA-V on HDL in vivo and the beneficial functions of apoA-V on atherosclerosis.     

Apolipoprotein A-I induces tubulin phosphorylation in association with cholesterol release in fetal rat astrocytes

We previously identified cytosolic lipid–protein particles (CLPP) having size and density of HDL in rat astrocytes, to which apoA-I induces translocation of cholesterol, caveolin-1 and protein kinase Cα (PKCα) following its association with microtubules prior to cholesterol release/biogenesis of HDL (JBC 277: 7929, 2002; JLR 45: 2269, 2004). To further understand the physiological relevance of these findings, we investigated the CLPP/microtubule association and its role in intracellular cholesterol trafficking by using a technique of reconstituted microtubule-like filaments (rMT) in rat astrocyte cytosol. When the cells were pretreated with apoA-I, α-tubulin as a 52-kDa protein in rMT was found phosphorylated while α-tubulin in a soluble monomeric form was little phosphorylated. The phosphorylation took place coincidentally to apoA-I-induced association with rMT of CLPP, a complex containing PKCα, and was suppressed by a PKC inhibitor, Bis indolylmaleimide 1 (BIM). α-Tubulin dissociated from CLPP when phosphorylated, and it poorly bound to CLPP once dissociated. BIM did not influence association of PKCα with rMT but suppressed apoA-I-induced cholesterol translocation to the cytosol from the ER/Golgi apparatus and apoA-I-mediated cholesterol release. We thereby concluded that α-tubulin phosphorylation by PKCα on CLPP is involved in reversible CLPP association with the microtubules and intracellular cholesterol trafficking for apoA-I-dependent HDL biogenesis/cholesterol release in rat astrocytes.     

Intracellular cholesterol transporters and modulation of hepatic lipid metabolism: Implications for diabetic dyslipidaemia and steatosis

Aims/hypotheses

To examine hepatic expression of cholesterol-trafficking proteins, mitochondrial StarD1 and endosomal StarD3, and their relationship with dyslipidaemia and steatosis in Zucker (fa/fa) genetically obese rats, and to explore their functional role in lipid metabolism in rat McArdle RH-7777 hepatoma cells.

Methods

Expression of StarD1 and StarD3 in rat liver and hepatoma samples were determined by Q-PCR and/or immunoblotting; lipid mass by colorimetric assays; radiolabelled precursors were utilised to measure lipid synthesis and secretion, and lipidation of exogenous apolipoprotein A-I.

Results

Hepatic expression of StarD3 protein was repressed by genetic obesity in (fa/fa) Zucker rats, compared with lean (Fa/?) controls, suggesting a link with storage or export of lipids from the liver. Overexpression of StarD1 and StarD3, and knockdown of StarD3, in rat hepatoma cells, revealed differential effects on lipid metabolism. Overexpression of StarD1 increased utilisation of exogenous (preformed) fatty acids for triacylglycerol synthesis and secretion, but impacted minimally on cholesterol homeostasis. By contrast, overexpression of StarD3 increased lipidation of exogenous apoA-I, and facilitated de novo biosynthetic pathways for neutral lipids, potentiating triacylglycerol accumulation but possibly offering protection against lipotoxicity. Finally, StarD3 overexpression altered expression of genes which impact variously on hepatic insulin resistance, inducing Ppargcla, Cyp2e1, Nr1h4, G6pc and Irs1, and repressing expression of Scl2a1, Igfbp1, Casp3 and Serpine 1.

Conclusions/interpretation

Targeting StarD3 may increase circulating levels of HDL and protect the liver against lipotoxicity; loss of hepatic expression of this protein, induced by genetic obesity, may contribute to the pathogenesis of dyslipidaemia and steatosis.     

HDL subfraction distribution of paraoxonase-1 and its relevance to enzyme activity and resistance to oxidative stress

The subfraction distribution of HDL-associated peptides has implications for their functions and the impact of pathological modifications to lipoprotein metabolism on these functions. We have analyzed the subfraction distribution of paraoxonase-1 (PON1) and the consequences for enzyme activity and stability. HDL subfractions were defined by the presence (LpA-I,A-II) or absence (LpA-I) of apolipoprotein A-II (apoA-II). PON1 was present in both subfractions, although increased concentrations of HDL were associated with significantly increased PON1 in LpA-I. ApoA-II did not modify the capacity of native human HDL or reconstituted HDL to promote PON1 secretion from cells or to stabilize enzyme activity, nor did apoA-II decrease PON1 activity when added to rabbit serum normally devoid of the apolipoprotein. LpA-I,A-II particles isolated from human serum or reconstituted HDL (LpA-I,A-II) showed a significantly greater capacity than HDL(LpA-I) to stabilize secreted PON1 and purified recombinant PON1 added to such particles. PON1 associated with apoA-II-containing particles showed greater resistance to inactivation arising from oxidation. ApoA-I, apoA-II, and LpA-I,A-II, but not LpA-I, were independent determinants of serum PON1 concentration and activity in multivariate analyses. PON1 is at least equally distributed between LpA-I and LpA-II,A-II HDL particles. This dichotomous distribution has implications for PON1 activity and stability that may impact on the physiological role of the enzyme.     

The ABC transporters in lipid flux and atherosclerosis

Atherosclerotic cardiovascular disease is the leading cause of morbidity and mortality in the United States and in many other countries. Dysfunctional lipid homeostasis plays a central role in the initiation and progression of atherosclerotic lesions. The ATP-binding cassette (ABC) transporters are transmembrane proteins that hydrolyze ATP and use the energy to drive the transport of various molecules across cell membranes. Several ABC transporters play a pivotal role in lipid trafficking. They are critically involved in cholesterol and phospholipid efflux and reverse cholesterol transport (RCT), processes that maintain cellular cholesterol homeostasis and protect arteries from atherosclerosis. In this article we provide a review of the current literature on the biogenesis of ABC transporters and highlight their proposed functions in atheroprotection.     

Cholesterol and bile acid-mediated regulation of autophagy in fatty liver diseases and atherosclerosis

Liver is the major organ that regulates whole body cholesterol metabolism. Disrupted hepatic cholesterol homeostasis contributes to the pathogenesis of nonalcoholic steatohepatitis, dyslipidemia, atherosclerosis, and cardiovascular diseases. Hepatic bile acid synthesis is the major catabolic mechanism for cholesterol elimination from the body. Furthermore, bile acids are signaling molecules that regulate liver metabolism and inflammation. Autophagy is a highly-conserved lysosomal degradation mechanism, which plays an essential role in maintaining cellular integrity and energy homeostasis. In this review, we discuss emerging evidence linking hepatic cholesterol and bile acid metabolism to cellular autophagy activity in hepatocytes and macrophages, and how these interactions may be implicated in the pathogenesis and treatment of fatty liver disease and atherosclerosis.