Hepatocytes direct the formation of a pro-metastatic niche in the liver.

The liver is the most common site of metastatic disease1. Although this metastatic tropism may reflect the mechanical trapping of circulating tumour cells, liver metastasis is also dependent, at least in part, on the formation of a 'pro-metastatic' niche that supports the spread of tumour cells to the liver2,3. The mechanisms that direct the formation of this niche are poorly understood. Here we show that hepatocytes coordinate myeloid cell accumulation and fibrosis within the liver and, in doing so, increase the susceptibility of the liver to metastatic seeding and outgrowth. During early pancreatic tumorigenesis in mice, hepatocytes show activation of signal transducer and activator of transcription 3 (STAT3) signalling and increased production of serum amyloid A1 and A2 (referred to collectively as SAA). Overexpression of SAA by hepatocytes also occurs in patients with pancreatic and colorectal cancers that have metastasized to the liver, and many patients with locally advanced and metastatic disease show increases in circulating SAA. Activation of STAT3 in hepatocytes and the subsequent production of SAA depend on the release of interleukin 6 (IL-6) into the circulation by non-malignant cells. Genetic ablation or blockade of components of IL-6-STAT3-SAA signalling prevents the establishment of a pro-metastatic niche and inhibits liver metastasis. Our data identify an intercellular network underpinned by hepatocytes that forms the basis of a pro-metastatic niche in the liver, and identify new therapeutic targets.

Serum amyloid A augments the atherogenic effects of cholesteryl ester transfer protein.

Serum amyloid A (SAA) is predictive of CVD in humans and causes atherosclerosis in mice. SAA has many proatherogenic effects in vitro. However, HDL, the major carrier of SAA in the circulation, masks these effects. The remodeling of HDL by cholesteryl ester transfer protein (CETP) liberates SAA restoring its proinflammatory activity. Here, we investigated whether deficiency of SAA suppresses the previously described proatherogenic effect of CETP. ApoE-/- mice and apoE-/- mice deficient in the three acute-phase isoforms of SAA (SAA1.1, SAA2.1, and SAA3; "apoE-/- SAA-TKO") with and without adeno-associated virus-mediated expression of CETP were studied. There was no effect of CETP expression or SAA genotype on plasma lipids or inflammatory markers. Atherosclerotic lesion area in the aortic arch of apoE-/- mice was 5.9 ± 1.2%; CETP expression significantly increased atherosclerosis in apoE-/- mice (13.1 ± 2.2%). However, atherosclerotic lesion area in the aortic arch of apoE-/- SAA-TKO mice (5.1 ± 1.1%) was not significantly increased by CETP expression (6.2 ± 0.9%). The increased atherosclerosis in apoE-/- mice expressing CETP was associated with markedly increased SAA immunostaining in aortic root sections. Thus, SAA augments the atherogenic effects of CETP, which suggests that inhibiting CETP may be of particular benefit in patients with high SAA.

Adipocyte-Derived Serum Amyloid A Promotes Angiotensin II-Induced Abdominal Aortic Aneurysms in Obese C57BL/6J Mice.

BACKGROUND: Obesity increases the risk for human abdominal aortic aneurysms (AAAs) and enhances Ang II (angiotensin II)-induced AAA formation in C57BL/6J mice. Obesity is also associated with increases in perivascular fat that expresses proinflammatory markers including SAA (serum amyloid A). We previously reported that deficiency of SAA significantly reduces Ang II-induced inflammation and AAA in hyperlipidemic apoE-deficient mice. In this study. we investigated whether adipose tissue-derived SAA plays a role in Ang II-induced AAA in obese C57BL/6J mice. METHODS: The development of AAA was compared between male C57BL/6J mice (wild type), C57BL/6J mice lacking SAA1.1, SAA2.1, and SAA3 (TKO); and TKO mice harboring a doxycycline-inducible, adipocyte-specific SAA1.1 transgene (TKO-Tgfat; SAA expressed only in fat). All mice were fed an obesogenic diet and doxycycline to induce SAA transgene expression and infused with Ang II to induce AAA. RESULTS: In response to Ang II infusion, SAA expression was significantly increased in perivascular fat of obese C57BL/6J mice. Maximal luminal diameters of the abdominal aorta were determined by ultrasound before and after Ang II infusion, which indicated a significant increase in aortic luminal diameters in wild type and TKO-TGfat mice but not in TKO mice. Adipocyte-specific SAA expression was associated with MMP (matrix metalloproteinase) activity and macrophage infiltration in abdominal aortas of Ang II-infused obese mice. CONCLUSIONS: We demonstrate for the first time that SAA deficiency protects obese C57BL/6J mice from Ang II-induced AAA. SAA expression only in adipocytes is sufficient to cause AAA in obese mice infused with Ang II.

Deficiency of Acute-Phase Serum Amyloid A Exacerbates Sepsis-Induced Mortality and Lung Injury in Mice.

Serum amyloid A (SAA) is a family of proteins, the plasma levels of which may increase >1000-fold in acute inflammatory states. We investigated the role of SAA in sepsis using mice deficient in all three acute-phase SAA isoforms (SAA-TKO). SAA deficiency significantly increased mortality rates in the three experimental sepsis mouse models: cecal ligation and puncture (CLP), cecal slurry (CS) injection, and lipopolysaccharide (LPS) treatments. SAA-TKO mice had exacerbated lung pathology compared to wild-type (WT) mice after CLP. A bulk RNA sequencing performed on lung tissues excised 24 h after CLP indicated significant enrichment in the expression of genes associated with chemokine production, chemokine and cytokine-mediated signaling, neutrophil chemotaxis, and neutrophil migration in SAA-TKO compared to WT mice. Consistently, myeloperoxidase activity and neutrophil counts were significantly increased in the lungs of septic SAA-TKO mice compared to WT mice. The in vitro treatment of HL-60, neutrophil-like cells, with SAA or SAA bound to a high-density lipoprotein (SAA-HDL), significantly decreased cellular transmigration through laminin-coated membranes compared to untreated cells. Thus, SAA potentially prevents neutrophil transmigration into injured lungs, thus reducing exacerbated tissue injury and mortality. In conclusion, we demonstrate for the first time that endogenous SAA plays a protective role in sepsis, including ameliorating lung injury.

The effect of diabetes on abdominal aortic aneurysm growth over 2 years.

OBJECTIVE: Abdominal aortic aneurysm (AAA) is a common progressive disease and a significant cause of morbidity and mortality. Prior investigations have shown that diabetes mellitus (DM) may be relatively protective of AAA incidence and growth. The Non-invasive Treatment of Aortic Aneurysm Clinical Trial (N-TA3CT) is a contemporary study of small AAA growth that provides a unique opportunity to validate and explore the effect of DM on AAA. Confirming the effect of DM on AAA growth in this study may present opportunities to explore for clues to potential biologic mechanisms as well as inform current patient management. METHODS: This is a secondary analysis examining the association of diabetes and aneurysm growth within N-TA3CT: a placebo-controlled multicenter trial of doxycycline in 261 patients with AAA maximum transverse diameters (MTDs) between 3.5 and 5 cm. The primary outcome is the change in the MTD from baseline as determined by computed tomography (CT) scans obtained during the trial. Secondary outcome is the growth pattern of the AAA. Baseline characteristics and growth patterns were assessed with t tests (continuous) or χ2 tests (categorical). Unadjusted and adjusted longitudinal analyses were performed with a repeated measures linear mixed model to compare AAA growth rates between patients with and without diabetes. RESULTS: Of 261 patients, 250 subjects had sufficient imaging and were included in this study. There were 56 patients (22.4%) with diabetes and 194 (77.6%) without. Diabetes was associated with higher body mass index and increased rates of hypercholesterolemia and coronary artery disease (P < .05). Diabetes was also associated with increased frequency of treatment for atherosclerosis and hypertension including treatment with statin, angiotensin-converting enzyme inhibitor, angiotensin II receptor blocker, anti-platelet, and diuretic therapy (P < .05). Baseline MTD was not significantly different between those with (4.32 cm) and without DM (4.30 cm). Median growth rate for patients with diabetes was 0.12 cm/y (interquartile range, 0.07-0.22 cm/y) and 0.19 cm/y (interquartile range, 0.12-0.27 cm/y) in patients without DM, which was significantly different on unadjusted analysis (P < .0001). Diabetes remained significantly associated with AAA growth after adjustment for other relevant clinical factors (coef, -0.057; P < .0001). CONCLUSIONS: Patients with diabetes have more than a 35% reduction in the median growth rates of AAA despite more severe concomitant vascular comorbidities and similar initial sizes of aneurysms. This effect persists and remains robust after adjusted analysis; and slower growth rates may delay the time to reach repair threshold. Rapid growth (>0.5 cm/y) is infrequent in patients with DM.

High-Density Lipoproteins and Serum Amyloid A (SAA).

PURPOSE OF REVIEW: Serum amyloid A (SAA) is a highly sensitive acute phase reactant that has been linked to a number of chronic inflammatory diseases. During a systemic inflammatory response, liver-derived SAA is primarily found on high-density lipoprotein (HDL). The purpose of this review is to discuss recent literature addressing the pathophysiological functions of SAA and the significance of its association with HDL. RECENT FINDINGS: Studies in gene-targeted mice establish that SAA contributes to atherosclerosis and some metastatic cancers. Accumulating evidence indicates that the lipidation state of SAA profoundly affects its bioactivities, with lipid-poor, but not HDL-associated, SAA capable of inducing inflammatory responses in vitro and in vivo. Factors that modulate the equilibrium between lipid-free and HDL-associated SAA have been identified. HDL may serve to limit SAA's bioactivities in vivo. Understanding the factors leading to the release of systemic SAA from HDL may provide insights into chronic disease mechanisms.

Aortic Aneurysms and Dissections Series: Part II: Dynamic Signaling Responses in Aortic Aneurysms and Dissections.

Aortic structure and function are controlled by the coordinated actions of different aortic cells and the extracellular matrix. Several pathways have been identified that control the aortic wall in a cell-type-specific manner and play diverse roles in various phases of aortic injury, repair, and remodeling. This complexity of signaling in the aortic wall poses challenges to the development of therapeutic strategies for treating aortic aneurysms and dissections. Here, in part II of this Recent Highlights series on aortic aneurysms and dissections, we will summarize recent studies published in Arteriosclerosis, Thrombosis, and Vascular Biology that have contributed to our knowledge of the signaling pathway-related mechanisms of aortic aneurysms and dissections.

Aortic Aneurysms and Dissections Series.

The aortic wall is composed of highly dynamic cell populations and extracellular matrix. In response to changes in the biomechanical environment, aortic cells and extracellular matrix modulate their structure and functions to increase aortic wall strength and meet the hemodynamic demand. Compromise in the structural and functional integrity of aortic components leads to aortic degeneration, biomechanical failure, and the development of aortic aneurysms and dissections (AAD). A better understanding of the molecular pathogenesis of AAD will facilitate the development of effective medications to treat these conditions. Here, we summarize recent findings on AAD published in ATVB. In this issue, we focus on the dynamics of aortic cells and extracellular matrix in AAD; in the next issue, we will focus on the role of signaling pathways in AAD.

Serum amyloid A is not incorporated into HDL during HDL biogenesis.

Liver-derived serum amyloid A (SAA) is present in plasma where it is mainly associated with HDL and from which it is cleared more rapidly than are the other major HDL-associated apolipoproteins. Although evidence suggests that lipid-free and HDL-associated forms of SAA have different activities, the pathways by which SAA associates and disassociates with HDL are poorly understood. In this study, we investigated SAA lipidation by hepatocytes and how this lipidation relates to the formation of nascent HDL particles. We also examined hepatocyte-mediated clearance of lipid-free and HDL-associated SAA. We prepared hepatocytes from mice injected with lipopolysaccharide or an SAA-expressing adenoviral vector. Alternatively, we incubated primary hepatocytes from SAA-deficient mice with purified SAA. We analyzed conditioned media to determine the lipidation status of endogenously produced and exogenously added SAA. Examining the migration of lipidated species, we found that SAA is lipidated and forms nascent particles that are distinct from apoA-I-containing particles and that apoA-I lipidation is unaltered when SAA is overexpressed or added to the cells, indicating that SAA is not incorporated into apoA-I-containing HDL during HDL biogenesis. Like apoA-I formation, generation of SAA-containing particles was dependent on ABCA1, but not on scavenger receptor class B type I. Hepatocytes degraded significantly more SAA than apoA-I. Taken together, our results indicate that SAA's lipidation and metabolism by the liver is independent of apoA-I and that SAA is not incorporated into HDL during HDL biogenesis.

High-density lipoprotein inhibits serum amyloid A-mediated reactive oxygen species generation and NLRP3 inflammasome activation.

Serum amyloid A (SAA) is a high-density apolipoprotein whose plasma levels can increase more than 1000-fold during a severe acute-phase inflammatory response and are more modestly elevated in chronic inflammation. SAA is thought to play important roles in innate immunity, but its biological activities have not been completely delineated. We previously reported that SAA deficiency protects mice from developing abdominal aortic aneurysms (AAAs) induced by chronic angiotensin II (AngII) infusion. Here, we report that SAA is required for AngII-induced increases in interleukin-1ß (IL-1ß), a potent proinflammatory cytokine that is tightly controlled by the Nod-like receptor protein 3 (NLRP3) inflammasome and caspase-1 and has been implicated in both human and mouse AAAs. We determined that purified SAA stimulates IL-1ß secretion in murine J774 and bone marrow-derived macrophages through a mechanism that depends on NLRP3 expression and caspase-1 activity, but is independent of P2X7 nucleotide receptor (P2X7R) activation. Inhibiting reactive oxygen species (ROS) by N-acetyl-l-cysteine or mito-TEMPO and inhibiting activation of cathepsin B by CA-074 blocked SAA-mediated inflammasome activation and IL-1ß secretion. Moreover, inhibiting cellular potassium efflux with glyburide or increasing extracellular potassium also significantly reduced SAA-mediated IL-1ß secretion. Of note, incorporating SAA into high-density lipoprotein (HDL) prior to its use in cell treatments completely abolished its ability to stimulate ROS generation and inflammasome activation. These results provide detailed insights into SAA-mediated IL-1ß production and highlight HDL's role in regulating SAA's proinflammatory effects.

Serum Amyloid A Is an Exchangeable Apolipoprotein.

Objective- SAA (serum amyloid A) is a family of acute-phase reactants that have proinflammatory and proatherogenic activities. SAA is more lipophilic than apoA-I (apolipoprotein A-I), and during an acute-phase response, <10% of plasma SAA is found lipid-free. In most reports, SAA is found exclusively associated with high-density lipoprotein; however, we and others have reported SAA on apoB (apolipoprotein B)-containing lipoproteins in both mice and humans. The goal of this study was to determine whether SAA is an exchangeable apolipoprotein. Approach and Results- Delipidated human SAA was incubated with SAA-free human lipoproteins; then, samples were reisolated by fast protein liquid chromatography, and SAA analyzed by ELISA and immunoblot. Both in vitro and in vivo, we show that SAA associates with any lipoprotein and does not remain in a lipid-free form. Although SAA is preferentially found on high-density lipoprotein, it can exchange between lipoproteins. In the presence of CETP (cholesterol ester transfer protein), there is greater exchange of SAA between lipoproteins. Subjects with diabetes mellitus, but not those with metabolic syndrome, showed altered SAA lipoprotein distribution postprandially. Proteoglycan-mediated lipoprotein retention is thought to be an underlying mechanism for atherosclerosis development. SAA has a proteoglycan-binding domain. Lipoproteins containing SAA had increased proteoglycan binding compared with SAA-free lipoproteins. Conclusions- Thus, SAA is an exchangeable apolipoprotein and increases apoB-containing lipoproteins' proteoglycan binding. We and others have previously reported the presence of SAA on low-density lipoprotein in individuals with obesity, diabetes mellitus, and metabolic syndrome. We propose that the presence of SAA on apoB-containing lipoproteins may contribute to cardiovascular disease development in these populations.

Serum amyloid A3 is a high density lipoprotein-associated acute-phase protein.

Serum amyloid A (SAA) is a family of acute-phase reactants. Plasma levels of human SAA1/SAA2 (mouse SAA1.1/2.1) can increase ≥1,000-fold during an acute-phase response. Mice, but not humans, express a third relatively understudied SAA isoform, SAA3. We investigated whether mouse SAA3 is an HDL-associated acute-phase SAA. Quantitative RT-PCR with isoform-specific primers indicated that SAA3 and SAA1.1/2.1 are induced similarly in livers (∼2,500-fold vs. ∼6,000-fold, respectively) and fat (∼400-fold vs. ∼100-fold, respectively) of lipopolysaccharide (LPS)-injected mice. In situ hybridization demonstrated that all three SAAs are produced by hepatocytes. All three SAA isoforms were detected in plasma of LPS-injected mice, although SAA3 levels were ∼20% of SAA1.1/2.1 levels. Fast protein LC analyses indicated that virtually all of SAA1.1/2.1 eluted with HDL, whereas ∼15% of SAA3 was lipid poor/free. After density gradient ultracentrifugation, isoelectric focusing demonstrated that ∼100% of plasma SAA1.1 was recovered in HDL compared with only ∼50% of SAA2.1 and ∼10% of SAA3. Thus, SAA3 appears to be more loosely associated with HDL, resulting in lipid-poor/free SAA3. We conclude that SAA3 is a major hepatic acute-phase SAA in mice that may produce systemic effects during inflammation.

Impact of individual acute phase serum amyloid A isoforms on HDL metabolism in mice.

The acute phase (AP) reactant serum amyloid A (SAA), an HDL apolipoprotein, exhibits pro-inflammatory activities, but its physiological function(s) are poorly understood. Functional differences between SAA1.1 and SAA2.1, the two major SAA isoforms, are unclear. Mice deficient in either isoform were used to investigate plasma isoform effects on HDL structure, composition, and apolipoprotein catabolism. Lack of either isoform did not affect the size of HDL, normally enlarged in the AP, and did not significantly change HDL composition. Plasma clearance rates of HDL apolipoproteins were determined using native HDL particles. The fractional clearance rates (FCRs) of apoA-I, apoA-II, and SAA were distinct, indicating that HDL is not cleared as intact particles. The FCRs of SAA1.1 and SAA2.1 in AP mice were similar, suggesting that the selective deposition of SAA1.1 in amyloid plaques is not associated with a difference in the rates of plasma clearance of the isoforms. Although the clearance rate of SAA was reduced in the absence of the HDL receptor, scavenger receptor class B type I (SR-BI), it remained significantly faster compared with that of apoA-I and apoA-II, indicating a relatively minor role of SR-BI in SAA's rapid clearance. These studies enhance our understanding of SAA metabolism and SAA's effects on AP-HDL composition and catabolism.

Ectopically expressed pro-group X secretory phospholipase A2 is proteolytically activated in mouse adrenal cells by furin-like proprotein convertases: implications for the regulation of adrenal steroidogenesis.

Group X secretory phospholipase A2 (GX sPLA2) hydrolyzes mammalian cell membranes, liberating free fatty acids and lysophospholipids. GX sPLA2 is produced as a pro-enzyme (pro-GX sPLA2) that contains an N-terminal 11-amino acid propeptide ending in a dibasic motif, suggesting cleavage by a furin-like proprotein convertase (PC). Although propeptide cleavage is clearly required for enzymatic activity, the protease(s) responsible for pro-GX sPLA2 activation have not been identified. We previously reported that GX sPLA2 negatively regulates adrenal glucocorticoid production, likely by suppressing liver X receptor-mediated activation of steroidogenic acute regulatory protein expression. In this study, using a FLAG epitope-tagged pro-GX sPLA2 expression construct (FLAG-pro-GX sPLA2), we determined that adrenocorticotropic hormone (ACTH) enhanced FLAG-pro-GX sPLA2 processing and phospholipase activity secreted by Y1 adrenal cells. ACTH increased the expression of furin and PCSK6, but not other members of the PC family, in Y1 cells. Overexpression of furin and PCSK6 in HEK 293 cells significantly enhanced FLAG-pro-GX sPLA2 processing, whereas siRNA-mediated knockdown of both PCs almost completely abolished FLAG-pro-GX sPLA2 processing in Y1 cells. Expression of either furin or PCSK6 enhanced the ability of GX sPLA2 to suppress liver X receptor reporter activity. The PC inhibitor decanoyl-Arg-Val-Lys-Arg-chloromethyl ketone significantly suppressed FLAG-pro-GX sPLA2 processing and sPLA2 activity in Y1 cells, and it significantly attenuated GX sPLA2-dependent inhibition of steroidogenic acute regulatory protein expression and progesterone production. These findings provide strong evidence that pro-GX sPLA2 is a substrate for furin and PCSK6 proteolytic processing and define a novel mechanism for regulating corticosteroid production in adrenal cells.

Deficiency of Endogenous Acute-Phase Serum Amyloid A Protects apoE-/- Mice From Angiotensin II-Induced Abdominal Aortic Aneurysm Formation.

OBJECTIVE: Rupture of abdominal aortic aneurysm (AAA), a major cause of death in the aged population, is characterized by vascular inflammation and matrix degradation. Serum amyloid A (SAA), an acute-phase reactant linked to inflammation and matrix metalloproteinase induction, correlates with aortic dimensions before aneurysm formation in humans. We investigated whether SAA deficiency in mice affects AAA formation during angiotensin II (Ang II) infusion. APPROACH AND RESULTS: Plasma SAA increased ≈60-fold in apoE(-/-) mice 24 hours after intraperitoneal Ang II injection (100 µg/kg; n=4) and ≈15-fold after chronic 28-day Ang II infusion (1000 ng/kg per minute; n=9). AAA incidence and severity after 28-day Ang II infusion was significantly reduced in apoE(-/-) mice lacking both acute-phase SAA isoforms (SAAKO; n=20) compared with apoE(-/-) mice (SAAWT; n=20) as assessed by in vivo ultrasound and ex vivo morphometric analyses, despite a significant increase in systolic blood pressure in SAAKO mice compared with SAAWT mice after Ang II infusion. Atherosclerotic lesion area of the aortic arch was similar in SAAKO and SAAWT mice after 28-day Ang II infusion. Immunostaining detected SAA in AAA tissues of Ang II-infused SAAWT mice that colocalized with macrophages, elastin breaks, and enhanced matrix metalloproteinase activity. Matrix metalloproteinase-2 activity was significantly lower in aortas of SAAKO mice compared with SAAWT mice after 10-day Ang II infusion. CONCLUSIONS: Lack of endogenous acute-phase SAA protects against experimental AAA through a mechanism that may involve reduced matrix metalloproteinase-2 activity.

Group X secretory phospholipase A2 regulates insulin secretion through a cyclooxygenase-2-dependent mechanism.

Group X secretory phospholipase A2 (GX sPLA2) potently hydrolyzes membrane phospholipids to release arachidonic acid (AA). While AA is an activator of glucose-stimulated insulin secretion (GSIS), its metabolite prostaglandin E2 (PGE2) is a known inhibitor. In this study, we determined that GX sPLA2 is expressed in insulin-producing cells of mouse pancreatic islets and investigated its role in beta cell function. GSIS was measured in vivo in wild-type (WT) and GX sPLA2-deficient (GX KO) mice and ex vivo using pancreatic islets isolated from WT and GX KO mice. GSIS was also assessed in vitro using mouse MIN6 pancreatic beta cells with or without GX sPLA2 overexpression or exogenous addition. GSIS was significantly higher in islets isolated from GX KO mice compared with islets from WT mice. Conversely, GSIS was lower in MIN6 cells overexpressing GX sPLA2 (MIN6-GX) compared with control (MIN6-C) cells. PGE2 production was significantly higher in MIN6-GX cells compared with MIN6-C cells and this was associated with significantly reduced cellular cAMP. The effect of GX sPLA2 on GSIS was abolished when cells were treated with NS398 (a COX-2 inhibitor) or L-798,106 (a PGE2-EP3 receptor antagonist). Consistent with enhanced beta cell function, GX KO mice showed significantly increased plasma insulin levels following glucose challenge and were protected from age-related reductions in GSIS and glucose tolerance compared with WT mice. We conclude that GX sPLA2 plays a previously unrecognized role in negatively regulating pancreatic insulin secretion by augmenting COX-2-dependent PGE2 production.

Impact of phospholipid transfer protein on nascent high-density lipoprotein formation and remodeling.

OBJECTIVE: Phospholipid transfer protein (PLTP), which binds phospholipids and facilitates their transfer between lipoproteins in plasma, plays a key role in lipoprotein remodeling, but its influence on nascent high-density lipoprotein (HDL) formation is not known. The effect of PLTP overexpression on apolipoprotein A-I (apoA-I) lipidation by primary mouse hepatocytes was investigated. APPROACH AND RESULTS: Overexpression of PLTP through an adenoviral vector markedly affected the amount and size of lipidated apoA-I species that were produced in hepatocytes in a dose-dependent manner, ultimately generating particles that were <7.1 nm but larger than lipid-free apoA-I. These <7.1-nm small particles generated in the presence of overexpressed PLTP were incorporated into mature HDL particles more rapidly than apoA-I both in vivo and in vitro and were less rapidly cleared from mouse plasma than lipid-free apoA-I. The <7.1-nm particles promoted both cellular cholesterol and phospholipid efflux in an ATP-binding cassette transporter A1-dependent manner, similar to apoA-I in the presence of PLTP. Lipid-free apoA-I had a greater efflux capacity in the presence of PLTP than in the absence of PLTP, suggesting that PLTP may promote ATP-binding cassette transporter A1-mediated cholesterol and phospholipid efflux. These results indicate that PLTP alters nascent HDL formation by modulating the lipidated species and by promoting the initial process of apoA-I lipidation. CONCLUSIONS: Our findings suggest that PLTP exerts significant effects on apoA-I lipidation and nascent HDL biogenesis in hepatocytes by promoting ATP-binding cassette transporter A1-mediated lipid efflux and the remodeling of nascent HDL particles.

Deficiency of endogenous acute phase serum amyloid A does not affect atherosclerotic lesions in apolipoprotein E-deficient mice.

OBJECTIVE: Although elevated plasma concentrations of serum amyloid A (SAA) are associated strongly with increased risk for atherosclerotic cardiovascular disease in humans, the role of SAA in the pathogenesis of lesion formation remains obscure. Our goal was to determine the impact of SAA deficiency on atherosclerosis in hypercholesterolemic mice. APPROACH AND RESULTS: Apolipoprotein E-deficient (apoE(-/-)) mice, either wild type or deficient in both major acute phase SAA isoforms, SAA1.1 and SAA2.1, were fed a normal rodent diet for 50 weeks. Female mice, but not male apoE-/- mice deficient in SAA1.1 and SAA2.1, had a modest increase (22%; P≤0.05) in plasma cholesterol concentrations and a 53% increase in adipose mass compared with apoE-/- mice expressing SAA1.1 and SAA2.1 that did not affect the plasma cytokine levels or the expression of adipose tissue inflammatory markers. SAA deficiency did not affect lipoprotein cholesterol distributions or plasma triglyceride concentrations in either male or female mice. Atherosclerotic lesion areas measured on the intimal surfaces of the arch, thoracic, and abdominal regions were not significantly different between apoE-/- mice deficient in SAA1.1 and SAA2.1 and apoE-/- mice expressing SAA1.1 and SAA2.1 in either sex. To accelerate lesion formation, mice were fed a Western diet for 12 weeks. SAA deficiency had effect neither on diet-induced alterations in plasma cholesterol, triglyceride, or cytokine concentrations nor on aortic atherosclerotic lesion areas in either male or female mice. In addition, SAA deficiency in male mice had no effect on lesion areas or macrophage accumulation in the aortic roots. CONCLUSIONS: The absence of endogenous SAA1.1 and 2.1 does not affect atherosclerotic lipid deposition in apolipoprotein E-deficient mice fed either normal or Western diets.