Bone marrow transplantation induces changes in the gut microbiota that chronically increase the cytokine response pattern of splenocytes.

Bone marrow transplantation (BMT) involves conditioning regimens which acutely induce side effects, including systemic inflammation, intestinal damage and shifts in the gut microbial composition, some of which may persist chronically. As the gut microbiota affect systemic immune responses, we aimed to investigate whether, post-BMT, the peripheral immune system is modulated as a direct consequence of alterations in the gut microbiota. We show that 24 weeks post-BMT, splenocytes but not peritoneal macrophages display increased cytokine response patterns upon ex-vivo stimulation with various pathogens as compared to untreated controls. The pattern of BMT-induced cytokine responses was transferred to splenocytes, and not to peritoneal macrophages, of healthy controls via co-housing and transferred to germfree mice via transplantation of cecum content. Thus, BMT induces changes in gut microbiota that in their turn increase cytokine responsiveness of splenocytes. Thus, BMT establishes a dominant microbiota that attenuates normalization of the immune-response.

Dual targeting of hepatic fibrosis and atherogenesis by icosabutate, an engineered eicosapentaenoic acid derivative.

BACKGROUND & AIMS: While fibrosis stage predicts liver-associated mortality, cardiovascular disease (CVD) is still the major overall cause of mortality in patients with NASH. Novel NASH drugs should thus ideally reduce both liver fibrosis and CVD. Icosabutate is a semi-synthetic, liver-targeted eicosapentaenoic acid (EPA) derivative in clinical development for NASH. The primary aims of the current studies were to establish both the anti-fibrotic and anti-atherogenic efficacy of icosabutate in conjunction with changes in lipotoxic and atherogenic lipids in liver and plasma respectively. METHODS: The effects of icosabutate on fibrosis progression and lipotoxicity were investigated in amylin liver NASH (AMLN) diet (high fat, cholesterol and fructose) fed ob/ob mice with biopsy-confirmed steatohepatitis and fibrosis and compared with the activity of obeticholic acid. APOE*3Leiden.CETP mice, a translational model for hyperlipidaemia and atherosclerosis, were used to evaluate the mechanisms underlying the lipid-lowering effect of icosabutate and its effect on atherosclerosis. RESULTS: In AMLN ob/ob mice, icosabutate significantly reduced hepatic fibrosis and myofibroblast content in association with downregulation of the arachidonic acid cascade and a reduction in both hepatic oxidised phospholipids and apoptosis. In APOE*3Leiden.CETP mice, icosabutate reduced plasma cholesterol and TAG levels via increased hepatic uptake, upregulated hepatic lipid metabolism and downregulated inflammation pathways, and effectively decreased atherosclerosis development. CONCLUSIONS: Icosabutate, a structurally engineered EPA derivative, effectively attenuates both hepatic fibrosis and atherogenesis and offers an attractive therapeutic approach to both liver- and CV-related morbidity and mortality in NASH patients.

Disruption of circadian rhythm by alternating light-dark cycles aggravates atherosclerosis development in APOE*3-Leiden.CETP mice.

Disruption of circadian rhythm by means of shift work has been associated with cardiovascular disease in humans. However, causality and underlying mechanisms have not yet been established. In this study, we exposed hyperlipidemic APOE*3-Leiden.CETP mice to either regular light-dark cycles, weekly 6 hours phase advances or delays, or weekly alternating light-dark cycles (12 hours shifts), as a well-established model for shift work. We found that mice exposed to 15 weeks of alternating light-dark cycles displayed a striking increase in atherosclerosis, with an approximately twofold increase in lesion size and severity, while mice exposed to phase advances and delays showed a milder circadian disruption and no significant effect on atherosclerosis development. We observed a higher lesion macrophage content in mice exposed to alternating light-dark cycles without obvious changes in plasma lipids, suggesting involvement of the immune system. Moreover, while no changes in the number or activation status of circulating monocytes and other immune cells were observed, we identified increased markers for inflammation, oxidative stress, and chemoattraction in the vessel wall. Altogether, this is the first study to show that circadian disruption by shifting light-dark cycles directly aggravates atherosclerosis development.

Deletion of hematopoietic Dectin-2 or CARD9 does not protect against atherosclerotic plaque formation in hyperlipidemic mice.

Inflammatory reactions activated by pattern recognition receptors (PRRs) on the membrane of innate immune cells play an important role in atherosclerosis. Whether the PRRs of the C-type lectin receptor (CLR) family including Dectin-2 may be involved in the pathogenesis of atherosclerosis remains largely unknown. Recently, the CLR-adaptor molecule caspase recruitment domain family member 9 (CARD9) has been suggested to play a role in cardiovascular pathologies as it provides the link between CLR activation and transcription of inflammatory cytokines as well as immune cell recruitment. We therefore evaluated whether hematopoietic deletion of Dectin-2 or CARD9 reduces inflammation and atherosclerosis development. Low-density lipoprotein receptor (Ldlr)-knockout mice were transplanted with bone marrow from wild-type, Dectin-2- or Card9-knockout mice and fed a Western-type diet containing 0.1% (w/w) cholesterol. After 10 weeks, lipid and inflammatory parameters were measured and atherosclerosis development was determined. Deletion of hematopoietic Dectin-2 or CARD9 did not influence plasma triglyceride and cholesterol levels. Deletion of hematopoietic Dectin-2 did not affect atherosclerotic lesion area, immune cell composition, ex vivo cytokine secretion by peritoneal cells or bone marrow derived macrophages. Unexpectedly, deletion of hematopoietic CARD9 increased atherosclerotic lesion formation and lesion severity. Deletion of hematopoietic CARD9 did also not influence circulating immune cell composition and peripheral cytokine secretion. Besides a tendency to a reduced macrophage content within these lesions, plasma MCP-1 levels decreased upon WTD feeding. Deletion of hematopoietic Dectin-2 did not influence atherosclerosis development in hyperlipidemic mice. The absence of CARD9 unexpectedly increased atherosclerotic lesion size and severity, suggesting that the presence of CARD9 may protect against initiation of atherosclerosis development.

Acute and chronic effects of treatment with mesenchymal stromal cells on LPS-induced pulmonary inflammation, emphysema and atherosclerosis development.

BACKGROUND: COPD is a pulmonary disorder often accompanied by cardiovascular disease (CVD), and current treatment of this comorbidity is suboptimal. Systemic inflammation in COPD triggered by smoke and microbial exposure is suggested to link COPD and CVD. Mesenchymal stromal cells (MSC) possess anti-inflammatory capacities and MSC treatment is considered an attractive treatment option for various chronic inflammatory diseases. Therefore, we investigated the immunomodulatory properties of MSC in an acute and chronic model of lipopolysaccharide (LPS)-induced inflammation, emphysema and atherosclerosis development in APOE*3-Leiden (E3L) mice. METHODS: Hyperlipidemic E3L mice were intranasally instilled with 10 µg LPS or vehicle twice in an acute 4-day study, or twice weekly during 20 weeks Western-type diet feeding in a chronic study. Mice received 0.5x106 MSC or vehicle intravenously twice after the first LPS instillation (acute study) or in week 14, 16, 18 and 20 (chronic study). Inflammatory parameters were measured in bronchoalveolar lavage (BAL) and lung tissue. Emphysema, pulmonary inflammation and atherosclerosis were assessed in the chronic study. RESULTS: In the acute study, intranasal LPS administration induced a marked systemic IL-6 response on day 3, which was inhibited after MSC treatment. Furthermore, MSC treatment reduced LPS-induced total cell count in BAL due to reduced neutrophil numbers. In the chronic study, LPS increased emphysema but did not aggravate atherosclerosis. Emphysema and atherosclerosis development were unaffected after MSC treatment. CONCLUSION: These data show that MSC inhibit LPS-induced pulmonary and systemic inflammation in the acute study, whereas MSC treatment had no effect on inflammation, emphysema and atherosclerosis development in the chronic study.

The Effects of Selective Hematopoietic Expression of Human IL-37 on Systemic Inflammation and Atherosclerosis in LDLr-Deficient Mice.

The human cytokine interleukin (IL)-37 has potent anti-inflammatory capacities, and hematopoietic cell-specific transgenic overexpression of IL-37 in mice protects against septic shock and colitis. In the present study we investigated the effect of hematopoietic expression of IL-37 on atherosclerosis development under low-grade inflammatory conditions. Low-density lipoprotein receptor (LDLr)-deficient mice were lethally irradiated and transplanted with bone marrow from IL-37-transgenic or control wild-type mice and fed a Western-type diet (WTD; 1% cholesterol) for eight weeks. Metabolic and inflammatory parameters were monitored and atherosclerosis was assessed in the aortic valve area. Hematopoietic IL-37 expression did not influence body weight, food intake and plasma cholesterol levels during the study. Plasma soluble E-selectin levels were increased with WTD-feeding as compared to chow-feeding, but were not influenced by IL-37 expression. IL-37 expression reduced the inflammatory state as indicated by reduced white blood cell counts and by reduced basal and lipopolysaccharide-induced cytokine response by peritoneal macrophages ex vivo. IL-37 expression did not influence the atherosclerotic lesion area. Lesion composition was marginally affected. Smooth muscle cell content was decreased, but macrophage and collagen content were not different. We conclude that under low-grade inflammatory conditions, hematopoietic IL-37 expression reduces the inflammatory state, but does not influence atherosclerosis development in hyperlipidemic LDLr-deficient mice.

Short-term cooling increases serum triglycerides and small high-density lipoprotein levels in humans.

BACKGROUND: Cold exposure and ß3-adrenergic receptor agonism, which both activate brown adipose tissue, markedly influence lipoprotein metabolism by enhancing lipoprotein lipase-mediated catabolism of triglyceride-rich lipoproteins and increasing plasma high-density lipoprotein (HDL) levels and functionality in mice. However, the effect of short-term cooling on human lipid and lipoprotein metabolism remained largely elusive. OBJECTIVE: The objective was to assess the effect of short-term cooling on the serum lipoprotein profile and HDL functionality in men. METHODS: Body mass index-matched young, lean men were exposed to a personalized cooling protocol for 2 hours. Before and after cooling, serum samples were collected for analysis of lipids and lipoprotein composition by 1H-nuclear magnetic resonance. Adenosine triphosphate-binding cassette A1 (ABCA1)-mediated cholesterol efflux capacity of HDL was measured using [3H]cholesterol-loaded ABCA1-transfected Chinese hamster ovary cells. RESULTS: Short-term cooling increased serum levels of free fatty acids, triglycerides, and cholesterol. Cooling increased the concentration of large very low-density lipoprotein (VLDL) particles accompanied by increased mean size of VLDL particles. In addition, cooling enhanced the concentration of small LDL and small HDL particles as well as the cholesterol levels within these particles. The increase in small HDL was accompanied by increased ABCA1-dependent cholesterol efflux in vitro. CONCLUSIONS: Our data show that short-term cooling increases the concentration of large VLDL particles and increases the generation of small LDL and HDL particles. We interpret that cooling increases VLDL production and turnover, which results in formation of surface remnants that form small HDL particles that attract cellular cholesterol.

BMT decreases HFD-induced weight gain associated with decreased preadipocyte number and insulin secretion.

Experimental bone marrow transplantation (BMT) in mice is commonly used to assess the role of immune cell-specific genes in various pathophysiological settings. The application of BMT in obesity research is hampered by the significant reduction in high-fat diet (HFD)-induced obesity. We set out to characterize metabolic tissues that may be affected by the BMT procedure and impair the HFD-induced response. Male C57BL/6 mice underwent syngeneic BMT using lethal irradiation. After a recovery period of 8 weeks they were fed a low-fat diet (LFD) or HFD for 16 weeks. HFD-induced obesity was reduced in mice after BMT as compared to HFD-fed control mice, characterized by both a reduced fat (-33%; p<0.01) and lean (-11%; p<0.01) mass, while food intake and energy expenditure were unaffected. As compared to control mice, BMT-treated mice had a reduced mature adipocyte volume (approx. -45%; p<0.05) and reduced numbers of preadipocytes (-38%; p<0.05) and macrophages (-62%; p<0.05) in subcutaneous, gonadal and visceral white adipose tissue. In BMT-treated mice, pancreas weight (-46%; p<0.01) was disproportionally decreased. This was associated with reduced plasma insulin (-68%; p<0.05) and C-peptide (-37%; p<0.01) levels and a delayed glucose clearance in BMT-treated mice on HFD as compared to control mice. In conclusion, the reduction in HFD-induced obesity after BMT in mice is at least partly due to alterations in the adipose tissue cell pool composition as well as to a decreased pancreatic secretion of the anabolic hormone insulin. These effects should be considered when interpreting results of experimental BMT in metabolic studies.

Murine models of cardiovascular comorbidity in chronic obstructive pulmonary disease.

Patients with chronic obstructive pulmonary disease (COPD) have an increased risk for cardiovascular disease (CVD). Currently, COPD patients with atherosclerosis (i.e., the most important underlying cause of CVD) receive COPD therapy complemented with standard CVD therapy. This may, however, not be the most optimal treatment. To investigate the link between COPD and atherosclerosis and to develop specific therapeutic strategies for COPD patients with atherosclerosis, a substantial number of preclinical studies using murine models have been performed. In this review, we summarize the currently used murine models of COPD and atherosclerosis, both individually and combined, and discuss the relevance of these models for studying the pathogenesis and development of new treatments for COPD patients with atherosclerosis. Murine and clinical studies have provided complementary information showing a prominent role for systemic inflammation and oxidative stress in the link between COPD and atherosclerosis. These and other studies showed that murine models for COPD and atherosclerosis are useful tools and can provide important insights relevant to understanding the link between COPD and CVD. More importantly, murine studies provide good platforms for studying the potential of promising (new) therapeutic strategies for COPD patients with CVD.

Lack of LCAT reduces the LPS-neutralizing capacity of HDL and enhances LPS-induced inflammation in mice.

HDL has important immunomodulatory properties, including the attenuation of lipopolysaccharide (LPS)-induced inflammatory response. As lecithin-cholesterol acyltransferase (LCAT) is a critical enzyme in the maturation of HDL we investigated whether LCAT-deficient (Lcat(-/-)) mice present an increased LPS-induced inflammatory response. LPS (100µg/kg body weight)-induced cytokine response in Lcat(-/-) mice was markedly enhanced and prolonged compared to wild-type mice. Importantly, reintroducing LCAT expression using adenovirus-mediated gene transfer reverted their phenotype to that of wild-type mice. Ex vivo stimulation of whole blood with LPS (1-100ng/mL) showed a similar enhanced pro-inflammatory phenotype. Further characterization in RAW 264.7 macrophages in vitro showed that serum and HDL, but not chylomicrons, VLDL or the lipid-free protein fraction of Lcat(-/-) mice, had a reduced capacity to attenuate the LPS-induced TNFα response. Analysis of apolipoprotein composition revealed that LCAT-deficient HDL lacks significant amounts of ApoA-I and ApoA-II and is primarily composed of ApoE, while HDL from Apoa1(-/-) mice is highly enriched in ApoE and ApoA-II. ApoA-I-deficiency did not affect the capacity of HDL to neutralize LPS, though Apoa1(-/-) mice showed a pronounced LPS-induced cytokine response. Additional immunophenotyping showed that Lcat(-/-) , but not Apoa1(-/-) mice, have markedly increased circulating monocyte numbers as a result of increased Cd11b(+)Ly6C(med) monocytes, whereas 'pro-inflammatory' Cd11b(+)Ly6C(hi) monocytes were reduced. In line with this observation, peritoneal macrophages of Lcat(-/-) mice showed a markedly dampened LPS-induced TNFα response. We conclude that LCAT-deficiency increases LPS-induced inflammation in mice due to reduced LPS-neutralizing capacity of immature discoidal HDL and increased monocyte number.

Plasma cholesteryl ester transfer protein is predominantly derived from Kupffer cells.

UNLABELLED: The role of Kupffer cells (KCs) in the pathophysiology of the liver has been firmly established. Nevertheless, KCs have been underexplored as a target for diagnosis and treatment of liver diseases owing to the lack of noninvasive diagnostic tests. We addressed the hypothesis that cholesteryl ester transfer protein (CETP) is mainly derived from KCs and may predict KC content. Microarray analysis of liver and adipose tissue biopsies, obtained from 93 obese subjects who underwent elective bariatric surgery, showed that expression of CETP is markedly higher in liver than adipose tissue. Hepatic expression of CETP correlated strongly with that of KC markers, and CETP messenger RNA and protein colocalized specifically with KCs in human liver sections. Hepatic KC content as well as hepatic CETP expression correlated strongly with plasma CETP concentration. Mechanistic and intervention studies on the role of KCs in determining the plasma CETP concentration were performed in a transgenic (Tg) mouse model expressing human CETP. Selective elimination of KCs from the liver in CETP Tg mice virtually abolished hepatic CETP expression and largely reduced plasma CETP concentration, consequently improving the lipoprotein profile. Conversely, augmentation of KCs after Bacille-Calemette-Guérin vaccination largely increased hepatic CETP expression and plasma CETP. Also, lipid-lowering drugs fenofibrate and niacin reduced liver KC content, accompanied by reduced plasma CETP concentration. CONCLUSIONS: Plasma CETP is predominantly derived from KCs, and plasma CETP level predicts hepatic KC content in humans.

Splenic autonomic denervation increases inflammatory status but does not aggravate atherosclerotic lesion development.

UNLABELLED: The brain plays a prominent role in the regulation of inflammation. Immune cells are under control of the so-called cholinergic anti-inflammatory reflex, mainly acting via autonomic innervation of the spleen. Activation of this reflex inhibits the secretion of proinflammatory cytokines and may reduce the development of atherosclerosis. Therefore, the aim of this study was to evaluate the effects of selective parasympathetic (Px) and sympathetic (Sx) denervation of the spleen on inflammatory status and atherosclerotic lesion development. Female APOE*3-Leiden.CETP mice, a well-established model for human-like lipid metabolism and atherosclerosis, were fed a cholesterol-containing Western-type diet for 4 wk after which they were subdivided into three groups receiving either splenic Px, splenic Sx, or sham surgery. The mice were subsequently challenged with the same diet for an additional 15 wk. Selective Px increased leukocyte counts (i.e., dendritic cells, B cells, and T cells) in the spleen and increased gene expression of proinflammatory cytokines in the liver and peritoneal leukocytes compared with Sx and sham surgery. Both Px and Sx increased circulating proinflammatory cytokines IL-1ß and IL-6. However, the increased proinflammatory status in denervated mice did not affect atherosclerotic lesion size or lesion composition. CONCLUSION: Predominantly selective Px of the spleen enhances the inflammatory status, which, however, does not aggravate diet-induced atherosclerotic lesion development.

Apolipoprotein A5 deficiency aggravates high-fat diet-induced obesity due to impaired central regulation of food intake.

Mutations in apolipoprotein A5 (APOA5) have been associated with hypertriglyceridemia in humans and mice. This has been attributed to a stimulating role for APOA5 in lipoprotein lipase-mediated triglyceride hydrolysis and hepatic clearance of lipoprotein remnant particles. However, because of the low APOA5 plasma abundance, we investigated an additional signaling role for APOA5 in high-fat diet (HFD)-induced obesity. Wild-type (WT) and Apoa5(-/-) mice fed a chow diet showed no difference in body weight or 24-h food intake (Apoa5(-/-), 4.5±0.6 g; WT, 4.2±0.5 g), while Apoa5(-/-) mice fed an HFD ate more in 24 h (Apoa5(-/-), 2.8±0.4 g; WT, 2.5±0.3 g, P<0.05) and became more obese than WT mice. Also, intravenous injection of APOA5-loaded VLDL-like particles lowered food intake (VLDL control, 0.26±0.04 g; VLDL+APOA5, 0.11±0.07 g, P<0.01). In addition, the HFD-induced hyperphagia of Apoa5(-/-) mice was prevented by adenovirus-mediated hepatic overexpression of APOA5. Finally, intracerebroventricular injection of APOA5 reduced food intake compared to injection of the same mouse with artificial cerebral spinal fluid (0.40±0.11 g; APOA5, 0.23±0.08 g, P<0.01). These data indicate that the increased HFD-induced obesity of Apoa5(-/-) mice as compared to WT mice is at least partly explained by hyperphagia and that APOA5 plays a role in the central regulation of food intake.

The apolipoprotein m-sphingosine-1-phosphate axis: biological relevance in lipoprotein metabolism, lipid disorders and atherosclerosis.

Apolipoprotein M (apoM) is a plasma apolipoprotein that mainly associates with high-density lipoproteins. Hence, most studies on apoM so far have investigated its effect on and association with lipid metabolism and atherosclerosis. The insight into apoM biology recently took a major turn. ApoM was identified as a carrier of the bioactive lipid sphingosine-1-phosphate (S1P). S1P activates five different G-protein-coupled receptors, known as the S1P-receptors 1-5 and, hence, affects a wide range of biological processes, such as lymphocyte trafficking, angiogenesis, wound repair and even virus suppression and cancer. The ability of apoM to bind S1P is due to a lipophilic binding pocket within the lipocalin structure of the apoM molecule. Mice overexpressing apoM have increased plasma S1P concentrations, whereas apoM-deficient mice have decreased S1P levels. ApoM-S1P is able to activate the S1P-receptor-1, affecting the function of endothelial cells, and apoM-deficient mice display impaired endothelial permeability in the lung. This review will focus on the putative biological roles of the new apoM-S1P axis in relation to lipoprotein metabolism, lipid disorders and atherosclerosis.

Apolipoprotein isoform E4 does not increase coronary heart disease risk in carriers of low-density lipoprotein receptor mutations.

BACKGROUND: In humans, the E4 allele of the apolipoprotein E gene is associated with increased coronary heart disease risk. Surprisingly, in rodents, apolipoprotein E4 only accelerates the atherosclerotic process when transgenic for the human low-density lipoprotein receptor (LDLR) protein. We therefore investigated whether the LDLR locus interacted with the apolipoprotein E gene genotype on coronary heart disease risk in patients clinically diagnosed with familial hypercholesterolemia with and without LDLR mutation. We investigated whether the presence of an LDLR mutation diminishing LDLR function was protective in E4/E4 carriers. METHODS AND RESULTS: In a cohort of 2400 patients clinically diagnosed with familial hypercholesterolemia, we found an LDLR gene mutation in 1383 patients, whereas in 1013 patients, such mutation was not present. In 92 patients homozygous for the apolipoprotein E4, the presence of an LDLR mutation conferred lower coronary heart disease risk (hazard ratio, 0.16; 95% CI, 0.05-0.58; P=0.005). Mirroring these results, the apolipoprotein E4/E4 genotype was also associated with lower coronary heart disease risk in patients with familial hypercholesterolemia with an LDLR mutation (hazard ratio, 0.26; hazard ratio, 0.08-0.80; P=0.02). CONCLUSIONS: LDLR function is key to the detrimental effects of apolipoprotein E4 in humans. Kinetic studies in humans are now required to study the consequences of our observation for prevention of both coronary heart disease and Alzheimer disease.

Apolipoprotein CI knock-out mice display impaired memory functions.

The ε4 allele of apolipoprotein E (APOE4), which is a well established genetic risk factor for development of Alzheimer's disease (AD), is in genetic disequilibrium with the H2 allele of apolipoprotein C1 (APOC1), giving rise to increased expression of apoC-I. This raises the possibility that the H2 allele of APOC1, either alone or in combination with APOE4, provides a major risk factor for AD. In line herewith, we previously showed that mice overexpressing human APOC1 display impaired learning and memory functions. Here, we tested the hypothesis that the absence of Apoc1 expression in mice may improve memory functions. In contrast with our expectations, Apoc1(-/-) mice showed impaired hippocampal-dependent memory functions, as judged from their performance in the object recognition task (p < 0.001) as compared to their wild-type littermates. No gross changes in brain morphology or in brain sterol concentrations were detected in knockout mice compared to wild-type littermates. Apoc1 deficiency reduced the expression of ApoE mRNA (-25%, p < 0.05), but not ApoE protein levels. In line with a role for apoC-I in inflammatory processes, we observed significantly increased mRNA concentrations of the proinflammatory marker tumor necrosis factor α and oxidative stress related heme oxygenase 1 (Hmox1) in the absence of glial activation. In conclusion, the absence of ApoC-I results in impaired memory functions, which is, together with previous data, suggestive of an important, bell-shaped gene-dose dependent role for ApoC-I in appropriate brain functioning. The relative contributions of the H2 allele of APOC1 and/or APOE4 in the risk assessment in AD remain to be determined.

Apolipoprotein CI enhances the biological response to LPS via the CD14/TLR4 pathway by LPS-binding elements in both its N- and C-terminal helix.

Timely sensing of lipopolysaccharide (LPS) is critical for the host to fight invading Gram-negative bacteria. We recently showed that apolipoprotein CI (apoCI) (apoCI1-57) avidly binds to LPS, involving an LPS-binding motif (apoCI48-54), and thereby enhances the LPS-induced inflammatory response. Our current aim was to further elucidate the structure and function relationship of apoCI with respect to its LPS-modulating characteristics and to unravel the mechanism by which apoCI enhances the biological activity of LPS. We designed and generated N- and C-terminal apoCI-derived peptides containing varying numbers of alternating cationic/hydrophobic motifs. ApoCI1-38, apoCI1-30, and apoCI35-57 were able to bind LPS, whereas apoCI1-23 and apoCI46-57 did not bind LPS. In line with their LPS-binding characteristics, apoCI1-38, apoCI1-30, and apoCI35-57 prolonged the serum residence of 125I-LPS by reducing its association with the liver. Accordingly, both apoCI1-30 and apoCI35-57 enhanced the LPS-induced TNFalpha response in vitro (RAW 264.7 macrophages) and in vivo (C57Bl/6 mice). Additional in vitro studies showed that the stimulating effect of apoCI on the LPS response resembles that of LPS-binding protein (LBP) and depends on CD14/ Toll-like receptor 4 signaling. We conclude that apoCI contains structural elements in both its N-terminal and C-terminal helix to bind LPS and to enhance the proinflammatory response toward LPS via a mechanism similar to LBP.

Apolipoprotein CI inhibits scavenger receptor BI and increases plasma HDL levels in vivo.

Apolipoprotein CI (apoCI) has been suggested to influence HDL metabolism by activation of LCAT and inhibition of HL and CETP. However, the effect of apoCI on scavenger receptor BI (SR-BI)-mediated uptake of HDL-cholesteryl esters (CE), as well as the net effect of apoCI on HDL metabolism in vivo is unknown. Therefore, we evaluated the effect of apoCI on the SR-BI-mediated uptake of HDL-CE in vitro and determined the net effect of apoCI on HDL metabolism in mice. Enrichment of HDL with apoCI dose-dependently decreased the SR-BI-dependent association of [(3)H]CE-labeled HDL with primary murine hepatocytes, similar to the established SR-BI-inhibitors apoCIII and oxLDL. ApoCI deficiency in mice gene dose-dependently decreased HDL-cholesterol levels. Adenovirus-mediated expression of human apoCI in mice increased HDL levels at a low dose and increased the HDL particle size at higher doses. We conclude that apoCI is a novel inhibitor of SR-BI in vitro and increases HDL levels in vivo.

Preoperative apolipoprotein CI levels correlate positively with the proinflammatory response in patients experiencing endotoxemia following elective cardiac surgery.

OBJECTIVE: Experimental models show that apolipoprotein CI (apoCI) binds and enhances the inflammatory response to endotoxin. We studied in patients undergoing cardiopulmonary bypass surgery (CPB) and experiencing endotoxemia during reperfusion whether plasma apoCI levels correlate with the inflammatory response and perioperative cytokine release. DESIGN: Prospective, observational, clinical cohort study. SETTING: Operating room (OR) and intensive care unit (ICU) of a university hospital. PATIENTS: One hundred fifty-nine consecutive patients > 18 years of age (66% males =105), median age 65 and 67 years for males and females, respectively) undergoing elective cardiothoracic surgery with cardiopulmonary bypass. INTERVENTIONS: None. MEASUREMENTS: Baseline apoCI, apoCIII, total cholesterol and triglyceride levels, and perioperative endotoxin and TNF-alpha levels were determined. RESULTS: High preoperative plasma apoCI, but not apoCIII, levels were associated (p < 0.05) with increased perioperative levels of TNF-alpha in patients experiencing endotoxemia. This association was not observed in patients without endotoxemia. CONCLUSION: High plasma apoCI is positively related to proinflammatory response in patients experiencing endotoxemia and confirms the observations in animal models.