Loss-of-function mutation in Pcsk1 increases serum APOA1 level and LCAT activity in mice

Background The convertase subtilisin/kexin family 1 gene (PCSK1) has been associated in various human genetics studies with a wide spectrum of metabolic phenotypes, including early-onset obesity, hyperphagia, diabetes insipidus, and others. Despite the evident influence of PCSK1 on obesity and the known functions of other PCSKs in lipid metabolism, the role of PCSK1 specifically in lipid and cholesterol metabolism remains unclear. This study evaluated the effect of loss of PCSK1 function on high-density lipoprotein (HDL) metabolism in mice. Results HDL cholesterol, apolipoprotein A1 (APOA1) levels in serum and liver, and the activities of two enzymes (lecithin-cholesterol acyltransferase, LCAT and phospholipid transfer protein, PLTP) were evaluated in 8-week-old mice with a non-synonymous single nucleotide mutation leading to an amino acid substitution in PCSK1, which results in a loss of protein’s function. Mutant mice had similar serum HDL cholesterol concentration but increased levels of serum total and mature APOA1, and LCAT activity in comparison to controls. Conclusions This study presents the first evaluation of the role of PCSK1 in HDL metabolism using a loss-of-function mutant mouse model. Further investigations will be needed to determine the underlying molecular mechanism.

Hepatocytic activating transcription factor 3 protects against steatohepatitis via hepatocyte nuclear factor 4

Activating transcription factor 3 (ATF3) has been shown to play an important role in HDL metabolism, yet the role of hepatocytic ATF3 in the development of steatohepatitis remains elusive. Here we show that adeno-associated virus-mediated over-expression of human ATF3 in hepatocytes prevents diet-induced steatohepatitis in C57BL/6 mice and reverses steatohepatitis in <i>db/db</i> mice. Conversely, global or hepatocyte-specific loss of ATF3 aggravates diet-induced steatohepatitis. Mechanistically, hepatocytic ATF3 induces hepatic lipolysis and fatty acid oxidation and inhibits inflammation and apoptosis. We further show that hepatocyte nuclear factor 4a (HNF4a) is required for ATF3 to improve steatohepatitis. Thus, the current study indicates that ATF3 protects against steatohepatitis through, at least in part, hepatic HNF4a. Targeting hepatic ATF3 may be useful for treatment of steatohepatitis.

Hepatocytic activating transcription factor 3 protects against steatohepatitis via hepatocyte nuclear factor 4

Activating transcription factor 3 (ATF3) has been shown to play an important role in HDL metabolism, yet the role of hepatocytic ATF3 in the development of steatohepatitis remains elusive. Here we show that adeno-associated virus-mediated over-expression of human ATF3 in hepatocytes prevents diet-induced steatohepatitis in C57BL/6 mice and reverses steatohepatitis in <i>db/db</i> mice. Conversely, global or hepatocyte-specific loss of ATF3 aggravates diet-induced steatohepatitis. Mechanistically, hepatocytic ATF3 induces hepatic lipolysis and fatty acid oxidation and inhibits inflammation and apoptosis. We further show that hepatocyte nuclear factor 4a (HNF4a) is required for ATF3 to improve steatohepatitis. Thus, the current study indicates that ATF3 protects against steatohepatitis through, at least in part, hepatic HNF4a. Targeting hepatic ATF3 may be useful for treatment of steatohepatitis.

Structural and Functional Impairments of Reconstituted High-Density Lipoprotein by Incorporation of Recombinant β-Amyloid42

Beta (β)-amyloid (Aβ) is a causative protein of Alzheimer’s disease (AD). In the pathogenesis of AD, the apolipoprotein (apo) A-I and high-density lipoprotein (HDL) metabolism is essential for the clearance of Aβ. In this study, recombinant Aβ42 was expressed and purified via the pET-30a expression vector and E.coli production system to elucidate the physiological effects of Aβ on HDL metabolism. The recombinant human Aβ protein (51 aa) was purified to at least 95% purity and characterized in either the lipid-free and lipid-bound states with apoA-I. Aβ was incorporated into the reconstituted HDL (rHDL) (molar ratio 95:5:1, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC):cholesterol:apoA-I) with various apoA-I:Aβ ratios from 1:0 to 1:0.5, 1:1 and 1:2. With an increasing molar ratio of Aβ, the α-helicity of apoA-I was decreased from 62% to 36% with a red shift of the Trp wavelength maximum fluorescence from 337 to 340 nm in apoA-I. The glycation reaction of apoA-I was accelerated further by the addition of Aβ. The treatment of fructose and Aβ caused more multimerization of apoA-I in the lipid-free state and in HDL. The phospholipid-binding ability of apoA-I was impaired severely by the addition of Aβ in a dose-dependent manner. The phagocytosis of LDL into macrophages was accelerated more by the presence of Aβ with the production of more oxidized species. Aβ severely impaired tissue regeneration, and a microinjection of Aβ enhanced embryotoxicity. In conclusion, the beneficial functions of apoA-I and HDL were severely impaired by the addition of Aβ via its detrimental effect on secondary structure. The impairment of HDL functionality occurred more synergistically by means of the co-addition of fructose and Aβ.

Unraveling the Complexity of HDL Remodeling: On the Hunt to Restore HDL Quality

Increasing evidence has cast doubt over the HDL-cholesterol hypothesis. The complexity of the HDL particle and its proven susceptibility to remodel has paved the way for intense molecular investigation. This state-of-the-art review discusses the molecular changes in HDL particles that help to explain the failure of large clinical trials intending to interfere with HDL metabolism, and details the chemical modifications and compositional changes in HDL-forming components, as well as miRNA cargo, that render HDL particles ineffective. Finally, the paper discusses the challenges that need to be overcome to shed a light of hope on HDL-targeted approaches.

Disturbances of the transfer of cholesterol to high-density lipoprotein (HDL) in patients with peripheral artery disease with or without type 2 diabetes mellitus

Introduction: Low high-density lipoprotein (HDL)-cholesterol is frequent in patients with peripheral artery disease (PAD) and also in type 2 diabetes mellitus (T2DM), the major risk factor for PAD. The transfer of cholesterol from the other lipoproteins to HDL is an important aspect of HDL metabolism and function, and may contribute to atherogenic mechanisms that lead to PAD development. Objective: The aim of this study was to investigate the status of cholesterol transfers in patients with PAD without or with T2DM. Methods: Patients with PAD ( n = 19), with PAD and T2DM (PAD + DM, n = 19), and healthy controls ( n = 20), all paired for age, gender, and BMI were studied. Transfer of both forms of cholesterol, unesterified (UC) and esterified (EC), was performed by incubating plasma with a donor nanoemulsion containing radioactive UC and EC, followed by chemical precipitation and HDL radioactive counting. Results: Low-density lipoprotein (LDL)-cholesterol and triglycerides were similar in the three groups. Compared to controls, HDL-C was lower in PAD + DM ( p < 0.05), but not in PAD. Transfer of UC was lower in PAD + DM than in PAD and controls (4.18 ± 1.17%, 5.13 ± 1.44%, 6.59 ± 1.25%, respectively, p < 0.001). EC transfer tended to be lower in PAD + DM than in controls (2.96 ± 0.60 vs 4.12 ± 0.89%, p = 0.05). Concentrations of cholesteryl ester transfer protein (CETP) and lecithin-cholesterol acyltransferase (LCAT), both involved in HDL metabolism, were not different among the three groups. Conclusion: Deficient cholesterol transfer to HDL may play a role in PAD pathogenesis. Since UC transfer to HDL was lower in PAD + DM compared to PAD alone, it is possible that defective HDL metabolism may contribute to the higher PAD incidence in patients with T2DM.Keywords

Lipid transfer to high-density lipoprotein is greater in elderly individuals with higher predictive cardiorespiratory fitness for age

Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): FAPESP and CNPq Background Elderly individuals have higher coronary artery disease (CAD) risk. Triggering factors include diminished peak of oxygen consumption (VO2peak), commonly referred as cardiorespiratory fitness (CF), and increase in atherogenic lipid profile: non high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C). The effect of aging on HDL-C is yet not clear, however, evaluating only the concentrations do not represent all HDL anti-atherosclerosis potential, aspects related to HDL metabolism and function can be more accurate to understand atherosclerosis process. Beyond that, elderly individuals also show reduced lipid transfer to HDL, an important assessment of HDL metabolism and function reported to be lower in CAD patients. Additionally, higher CF level is associated with longevity and lower hospitalizations rates and higher lipid transfer, independently of HDL-C. Data about different ages and absolute values of CF on lipids transfer are nonexistent. Purpose: To analyze the effect of paired CF absolute values and BMI in a healthy population of different ages on lipid transfer to HDL. Methods: 61 healthy individuals, enrolled in two groups: 30 young (27 ± 4 years, 21 females, BMI: 24.6 ± 2.6 kg/m², VO2peak: 33.9 ± 5.0 ml/kg/min) and 31 elderly (67 ± 5 years, 15 females, BMI: 23.9 ± 3.3 kg/m², VO2peak: 33.2 ± 4.5 ml/kg/min) that do not use any medication for chronic disease. Anthropometric data were collected. All participants performed treadmill cardiopulmonary test and the CF predictive value for age was calculated, according to each sex. Plasma lipids and cholesterol ester transfer protein (CETP) concentrations were determined. Lipid transfer was performed by incubating plasma with a donor lipoprotein-like nanoparticle containing radioactively labeled unesterified cholesterol (UC) and esterified cholesterol (EC), followed by chemical precipitation and radioactive count. Cholesterol transfers from the donor nanoparticle to HDL are expressed as %. For the statistical analysis Student"s t and Mann-Whitney tests were used according to data distribution. Results: Waist circumference (p = 0.0291) and waist-hip ratio (p &lt; 0.0001) were higher in elderly. LDL-C (p = 0.0005) and non HDL-C (p = 0.0003) were higher in elderly, as well apolipoprotein A-I, B, and CETP concentration. HDL-C and triglycerides were equal between groups. EC (p = 0.0268) and UC (p = 0.0348) transfers to HDL were higher in elderly. VO2peak predictive by age expressed in percentage was higher in elderly (90 ± 13 vs. 120 ±18; p &lt; 0.0001). Conclusion: Elderly individuals show higher anthropometric data and concentrations of atherogenic cholesterol lipoproteins; however, HDL-C was equal. When paired by absolute CF values and BMI with young adults, elderly subjects with high CF predicted by age, have higher cholesterol transfer to HDL which demonstrates greater HDL metabolism.

Effect of Menopause and Hypertriglyceridemia on Lipid Transfer and Paraoxonase Activity in Diabetic Women

Introduction. The high prevalence of type 2 diabetes mellitus (DM2) and cardiovascular atherosclerotic complications are increasing in women after menopause. Studies have shown that the in vitro transfer of lipids to HDL and the determination of paraoxonase activity are robust methods to HDL functional evaluation. Objective. This study aimed at observing the difference in the paraoxonase activity and the transfer of lipids to HDL in menopause DM2-women with hypertriglyceridemia. Material and Methods. Blood samples were collected from 63 female diabetic patients with hypertriglyceridemia, matched by age group and comorbidities. The artificial nanoparticle (LDE) determined the transfer to HDL by liquid scintillation, as well the activity of paraoxonase by spectrophotometry. Results. There was a difference in paraoxonase-1 activity between the groups due to the high hypertriglyceridemia, regardless of the menopausal or non-menopausal condition. And, there was also a difference in phospholipid transfer between groups. Discussion. This study showed the presence of alterations in the activity of paraoxonase and phospholipid and cholesterol ester transfer to HDL in menopausal women groups when compared to those non-menopausal groups. Conclusion. This study showed that both diabetic hypertriglyceridemia and menopause influence the HDL metabolism, and the action of paraoxonase.

Obesity Affects HDL Metabolism, Composition and Subclass Distribution

Background: Obesity increases the risk of coronary heart disease, partly due to its strong association with atherogenic dyslipidemia, characterized by high triglycerides and low high-density lipoprotein (HDL) cholesterol levels. Functional impairment of HDL may contribute to the increased cardiovascular mortality, but the effect of obesity on composition, structure, and function of HDL is not well understood. Design and Methods: We determined HDL composition, HDL subclass distribution, parameters of HDL function, and activities of most important enzymes involved in lipoprotein remodeling, including lecithin–cholesterol acyltransferase (LCAT) and cholesteryl ester transfer protein (CETP) in relatively young normal weight (n = 26), overweight (n = 22), and obese (n = 20) women. Results: Obesity (body mass index (BMI) ≥ 30) was associated with noticeable changes in LCAT and CETP activities and altered HDL composition, such as decreased apolipoprotein A-I, cholesterol, and phospholipid content, while pro-inflammatory HDL serum amyloid a content was increased. We observed a marked shift towards smaller HDL subclasses in obesity linked to lower anti-oxidative capacity of serum. LCAT activity, HDL subclass distribution, and HDL-cholesterol were associated with soluble leptin receptor, adiponectin, and liver enzyme activities. Of note, most of these alterations were only seen in obese women but not in overweight women. Conclusions: Obesity markedly affects HDL metabolism, composition, and subclass distribution linked to changes in liver and adipose tissue. HDL dysfunction may contribute to increased cardiovascular risk in obesity.