High density lipoproteins (HDL) mediate cholesterol transport and protection from cardiovascular disease. Although synthetic HDLs have been studied for 30 years, the structure of human plasma-derived HDL, and its major protein apolipoprotein (apo)A-I, is unknown. We separated normal human HDL into 5 density subfractions and then further isolated those containing predominantly apoA-I (LpA-I). Using cross-linking chemistry and mass spectrometry, we found that apoA-I adopts a structural framework in these particles that closely mirrors that in synthetic HDL. We adapted established structural models for synthetic HDL to generate the first detailed models of authentic human plasma HDL in which apoA-I adopts a symmetrical cage-like structure. The models suggest that HDL particle size is modulated via a twisting motion of the resident apoA-I molecules. This understanding offers insights into how apoA-I structure modulates HDL function and its interactions with other apolipoproteins.

Spherical high density lipoproteins (HDL) † predominate in human plasma. However, little information exists on the structure of the most common HDL protein, apolipoprotein (apo) A-I, in spheres vs. better studied discoidal forms. We produced spherical HDL by incubating reconstituted discoidal HDL with physiological plasmaremodeling enzymes and compared apoA-I structure in discs and spheres of comparable diameter (79 -80 and 93-96 Å). Using cross-linking chemistry and mass spectrometry, we determined that the general structural organization of apoA-I was overall similar between discs and spheres, regardless of diameter. This was the case despite the fact that the 93 Å spheres contained three molecules of apoA-I per particle compared with only two in the discs. Thus, apoA-I adopts a consistent general structural framework in HDL particles-irrespective of shape, size and the number of apoA-Is present. Furthermore, a similar cross-linking pattern was demonstrated in HDL particles isolated from human serum. We propose the first experiment-based molecular model of apoA-I in spherical HDL particles. This model provides a new foundation for understanding how apoA-I structure modulates HDL function and metabolism.sphere ͉ disk G iven the inverse correlation between high density lipoprotein (HDL) levels and cardiovascular disease, a key question in vascular biology relates to how apolipoproteins modulate the metabolism and function of HDL. Significant evidence supports a role for HDL in the process of reverse cholesterol transport whereby lipids and cholesterol in the vessel wall are transported to the liver for catabolism. However, because of a lack of information on HDL structure and the molecular basis of its interactions with other proteins, our understanding of HDL metabolism and function is at a basic stage.The ''glue'' that holds most HDL particles together is apolipoprotein (apo)A-I, a highly ␣-helical, 28-kDa polypeptide. It comprises some 70% of HDL protein mass, making it the clear starting point for deriving a basic understanding of HDL structure. In humans, apoA-I is primarily present in two major spherical HDL species, HDL 2 (d ϭ 1.063-1.125 g/ml) and HDL 3 (d ϭ 1.125-1.210 g/ml) with diameters ranging from 70 to 120 Å. More minor, but clearly important, HDL subspecies include lipid-poor apoA-I and nascent discoidal particles (reviewed in ref. 1). Highly reactive but low abundance discoidal HDLs are critical intermediates between lipid-poor apoA-I and mature spherical HDL. Easily produced in vitro, they have been heavily used for structural studies (2). Despite some debates on details of certain regions of apoA-I in discs, the majority of recent theoretical and experimental data supports the so-called ''double belt'' model (3). In this scheme, each of two ring-shaped apoA-I molecules wrap around a leaflet of a disk-like patch of lipid bilayer in an anti-parallel orientation.Despite their abundance in plasma, much less is known about the structure of apoA-I in spherical particles. They contain a neutral lipid...

Metabolism of glutamate and glutamine in cultured mouse cerebral cortical astrocytes has been investigated using either radioactively labelled (¹⁴C) amino acids or ¹³C-labelled amino acids combined with NMR spectroscopy of cell extracts and lyophilyzed incubation media. Using [U-¹³C]glutamate it has been shown that in astrocytes exogenously supplied glutamate is primarily (70%) metabolized oxidatively through the tricarboxylic acid (TCA) cycle and to a lesser jextent (30%) directly to glutamine. Glutamate metabolized in the TCA cycle is to a large extent recovered as lactate showing that the astrocyte-specific enzyme, malic enzyme is functionally active. Incubation with [U-¹⁴C]glutamine led to a higher specific radioactivity in glutamate than in glutamine. It could also be shown that glutamate and glutamine were metabolized differently to aspartate and alanine. These results taken together strongly suggest that glutamate/glutamine metabolism in astrocytes is compartmentalized and a model with multiple cytoplasmic and mitochondrial compartments of these amino acids is proposed.

Diabetes mellitus is a strong risk factor for chronic kidney disease and end-stage renal disease. Whether sex differences in chronic kidney disease and end-stage renal disease incidence exist among diabetic patients remains unclear. This systematic review and meta-analysis was conducted to evaluate the relative effect of diabetes on chronic kidney disease and end-stage renal disease risk in women compared with men. We systematically searched Embase, PubMed, and the Cochrane Library for both cohort and case-control studies until October 2015. Studies were selected if they reported a sex-specific relationship between diabetes mellitus and chronic kidney disease or end-stage renal disease. We generated pooled estimates across studies using random-effects meta-analysis after log transformation with inverse variance weighting. Ten studies with data from more than 5 million participants were included. The pooled adjusted risk ratio of chronic kidney disease associated with diabetes mellitus was 3.34 (95 % CI 2.27, 4.93) in women and 2.84 (95 % CI 1.73, 4.68) in men. The data showed no difference in diabetes-related chronic kidney disease risk between the sexes (pooled adjusted women-to-men relative risk ratio was 1.14 [95 % CI 0.97, 1.34]) except for end-stage renal disease-the pooled adjusted women-to men relative risk ratio was 1.38 (95 % CI 1.22, 1.55; p = 0.114, I² = 38.1 %). The study found no evidence of a sex difference in the association between diabetes mellitus and chronic kidney disease. However, the excess risk for end-stage renal disease was higher in women with diabetes than in men with the same condition, from which we assume that the female gender could accelerate the disease progression. Further studies are needed to support this notion and elucidate the underlying mechanisms.

Transforming growth factor-beta (TGF-β), a key mediator of cardiac fibroblast activation, has a major influence on collagen type I production. However, the epigenetic mechanisms by which TGF-β induces collagen type I alpha 1 (COL1A1) expression are not fully understood. This study was designed to examine whether or not DNA methylation is involved in TGF-β-induced COL1A1 expression in cardiac fibroblasts. Cells isolated from neonatal Sprague-Dawley rats were cultured and stimulated with TGF-β1. The mRNA levels of COL1A1 and DNA methyltransferases (DNMTs) were determined via quantitative polymerase chain reaction and the protein levels of collagen type I were determined via Western blot as well as enzyme-linked immunosorbent assay. The quantitative methylation of the COL1A1 promoter region was analyzed using the MassARRAY platform of Sequenom. Results showed that TGF-β1 upregulated the mRNA expression of COL1A1 and induced the synthesis of cell-associated and secreted collagen type I in cardiac fibroblasts. DNMT1 and DNMT3a expressions were significantly downregulated and the global DNMT activity was inhibited when treated with 10 ng/mL of TGF-β1 for 48 h. TGF-β1 treatment resulted in a significant reduction of the DNA methylation percentage across multiple CpG sites in the rat COL1A1 promoter. Thus, TGF-β1 can induce collagen type I expression through the inhibition of DNMT1 and DNMT3a expressions as well as global DNMT activity, thereby resulting in DNA demethylation of the COL1A1 promoter. These findings suggested that the DNMT-mediated DNA methylation is an important mechanism in regulating the TGF-β1-induced COL1A1 gene expression.