Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 1. ABCG1 enhances amyloid-b protein (Ab) production in vitro. A, B: HEK-APPswe cells were transiently transfected with empty vector or ABCG1. Conditioned medium was collected at var- ious times, assayed for Ab40 (A) and Ab42 (B) by ELISA, and nor- malized to total cell protein. Data represent means 6 SD of two independent experiments for empty vector and ABCG1. Data were analyzed by unpaired Student’s t-test at each time point; * P , 0.05, ** P , 0.01. C: Western blot shows the detection of ABCG1 protein after transfection with ABCG1.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: In Vitro, Transfection, Plasmid Preparation, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 2. ABCG1 requires cholesterol efflux activity to affect Ab production. HEK-APPswe cells were trans- fected with vector, wild-type (WT) ABCG1, or ABCG1 containing a S220G mutation in the Walker A domain. A: Wild-type ABCG1, but not ABCG1 S220G, effluxes cholesterol to exogenous HDL above baseline levels. Data represent means 6 SEM from at least two independent cholesterol efflux assays over a 6 h period, each measured in triplicate. B, C: Wild-type ABCG1, but not ABCG1 S220G, augments Ab40 (B) and Ab42 (C) production above baseline levels. Data represent means 6 SEM of at least two independent experiments.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Activity Assay, Plasmid Preparation, Mutagenesis

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 3. ABCG1 promotes the release of secreted of b-amyloid pre- cursor protein (sAPPa and sAPPb). A: HEK-APPswe cells were trans- fected with ABCG1 or empty vector for 24 h, followed by a 6 h period of conditioning in fresh medium. sAPPa and sAPPb levels were eval- uated by Western blot analysis of medium and normalized to total cellular protein. B: The graph represents means 6 SEM of a rep- resentative experiment from three independent transfections. G1, ABCG1; V, vector.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Plasmid Preparation, Western Blot, Transfection

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 4. ABCG1 enhances a-secretase and b-secretase cleav- age of APP. A: Representative Western blot of total APP as well as C-terminal fragment a (CTFa) and CTFb from HEK-APPswe cells transfected with vector. B–E: The graphs represents means 6 SEM of three transfections. B, D: To- tal CTF levels corrected for GAPDH. C,E: CTF levels cor- rected for total APPs. A total of three independent rounds of transfections, each at least in triplicate, were performed.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Western Blot, Transfection, Plasmid Preparation

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 5. ABCG1 increases cell surface presentation of APP. A: Representative Western blot of HEK-APPswe cells transiently transfected with empty vector, murine ABCG1, or human ABCG1 and treated with sulfo- NHS-biotin followed by precipitation of biotinylated cell surface proteins using streptavidin agarose beads. The levels of total and cell surface APP are shown and normalized to actin levels as an internal control. B: Quantitation of APP distribution. The graphs represent means 6 SEM of a representative round from three independent transfections, analyzed by Student’s t-test. Three rounds of at least triplicate indepen- dent transfections were performed. Asterisks represent P , 0.05.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Western Blot, Transfection, Plasmid Preparation, Control, Quantitation Assay

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 6. ABCG1 does not increase the cellular adherence of exogenous sAPP. Conditioned medium and cell lysates were prepared from HEK-APPswe and HEK293 cells (input APP and HEK). HEK293 cells were transfected with vector or ABCG1 and exposed to conditioned medium from HEK293 cells (HEK CM) or conditioned medium from HEK-APPswe cells (APP CM) for 24 h, after which medium and cell lysates were immunoblotted for APP and GAPDH. The upper panel (Media sAPP) shows the input levels of sAPP in the conditioned medium at the beginning of the experiment and demonstrates no loss of input signal after 24 h of incubation on transfected cells. The middle panel (Cellular APP) shows the levels of cell-associated APP in HEK-APPswe and HEK293 cells (input lanes) and in HEK293 cells exposed to HEK293 CM or APP CM using 75 mg of protein per lane. The lower panel shows GAPDH as an internal loading control.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Transfection, Plasmid Preparation, Incubation, Control

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 7. ABCG1 is highly expressed in neurons. LacZ staining of ABCG1 heterozygous (A, C, E, G–I) and wild-type (WT; B, D, F) mice. Coronal sections are shown at 2.53 (A, B), 103 (C–F), and 403 (G–I) magnification. Strong LacZ staining, indicative of ABCG1 expression, is observed in hippocampus (A, C), all cortical layers (A, E, G–I), striatum, and thalamus (A).

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Staining, Expressing

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 8. ABCG1 mRNA levels are increased significantly in Down syndrome (DS) cortex. Quantitative RT-PCR was used to determine ABCG1 (A) and APP (B) mRNA levels from eight trisomy 21 pa- tients (black bars) and eight age- and sex-matched controls (Con; white bars) as well as eight Alzheimer’s disease (AD) patients (dark gray bars) and eight age- and sex-matched controls (light gray bars). Data represent means 6 SEM of each patient measured in duplicate for the DS patients and matched controls and in triplicate for the AD patients and matched controls, analyzed by Student’s unpaired t-test.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Quantitative RT-PCR

Journal: Journal of Lipid Research

Article Title: The cholesterol transporter ABCG1 modulates the subcellular distribution and proteolytic processing of β-amyloid precursor protein

doi: 10.1194/jlr.m600542-jlr200

Figure Lengend Snippet: Fig. 9. ABCG1 protein levels are increased significantly in DS cortex. A: Western blots of total membrane fractions from four age-matched controls and trisomy 21 patients. Blots were probed sequentially for ABCG1, APP, and NaK-ATPase as an internal load- ing control. B: Relative ABCG1/NaK-ATPase and APP/NaK-ATPase values are shown, with the values in control (Con) fractions set to 1. Data represent means 6 SEM of each patient assayed in duplicate and analyzed by Student’s unpaired t-test.

Article Snippet: SDS was added to a final concentration of 1% before SDS-PAGE and immunoblotting with antibodies against APP, ABCG1, and NaK-ATPase as an internal loading control (Novus).

Techniques: Western Blot, Membrane, Control

Journal: Arteriosclerosis, thrombosis, and vascular biology

Article Title: SREBF1/MicroRNA-33b Axis Exhibits Potent Effect on Unstable Atherosclerotic Plaque Formation In Vivo.

doi: 10.1161/ATVBAHA.118.311409

Figure Lengend Snippet: Figure 5. MicroRNA (miR)-33b alters macrophage functions and characteristics via repression of cholesterol efflux. A, Western blotting analysis of ATP-binding cas- sette transporter (ABC) A1 and ABCG1 expression in peritoneal macrophages (PEMs). PEMs were treated with 300 μg/mL acetylated low-density lipoprotein (AcLDL- C) for the indicated times. Representative images from several independent experiments are shown. β-actin was used as the loading control. B, Cholesterol efflux from PEMs in the presence or absence of 10 μg/mL ApoA-1 and 100 μg/mL high-density lipoprotein (HDL) cholesterol (n=8). C, Quantification of Annexin V–positive PEMs in the presence or absence of 300 μg/mL AcLDL-C and 10 ng/mL acyl-CoA: cholesterol acyl-transferase inhibitor (ACATi) for 24 h (total 8–9 fields each containing ≈1000 cells). D, Quantitative real-time polymerase chain reaction (PCR) analysis of Il-1β, Il-6, and Ccl2 in PEMs from miR-33b−/−Apoe−/− and miR-33b+/+Apoe−/− mice. Values from miR-33b−/−Apoe−/− mice were set at 1. E, PEMs were stained with fluorescein isothiocyanate (FITC)–conjugated cholera toxin subunit B (CTB) to assess the amount of lipid rafts on the cellular membrane. FITC-CTB was quantified using flow cytometry: a representative histogram (left) and a bar graph of mean fluores- cence intensity (MFI) from replicated samples (right; n=4). F, Expression levels of inflammatory genes in THP-1 macrophage transfected with miR-control and miR-33b overexpression vector. Values from miR-control vector transfected cells were set at 1 (n=9). G, MiR-control or miR-33b–overexpressed THP-1 macrophages with or without cholesterol depletion using methyl-β-cyclodextrin (MβCD) stained with FITC-CTB: a representative histogram (left) and a bar graph of MFI (the second from the left; n=3). Quantitative real-time PCR analysis of IL-1β and IL-6 from miR-control or miR-33b overexpressed THP-1 macrophage with or without cholesterol deple- tion using MβCD (the second from right and right). Values from miR-control vector transfected cells were set at 1 (n=9 each). Error bars indicate the SEM. *P<0.05, **P<0.01, and ****P<0.0001, by 1-way ANOVA with post hoc Tukey test (B, C, G) and Student t test (D, E, F). Apoe−/− indicates apolipoprotein E–deficient.

Article Snippet: The antibodies used were a polyclonal anti–ATP-binding cassette transporter (ABC) A1 antibody (NB400-105), a polyclonal anti-ABCG1 antibody (NB400-132), a polyclonal anti–SR-BI (SCARB1) antibody (NB400-104; Novus Biologicals, Littleton, CO), an anti–single strand DNA antibody (no. 18731; IBL, Gunma, Japan), an anti-CPT1α antibody (ab128568), an anti-CROT antibody (ab103448), an anti–LDL receptor antibody (ab52818; Abcam, Cambridge, United Kingdom), an anti–β-actin antibody (AC-15; A5441, Sigma-Aldrich, St. Louis, MO), a PEconjugated anti-CD11b antibody (no. 12-0112; eBioscience, Santa Clara, CA), an APC-conjugated anti-F4/80 antibody (no. 123115; BioLegend, San Diego, CA), and an anti-CD68 antibody (FA-11, Serotec, Kidlington, United Kingdom).

Techniques: Western Blot, Binding Assay, Expressing, Control, Real-time Polymerase Chain Reaction, Staining, Membrane, Flow Cytometry, Transfection, Over Expression, Plasmid Preparation

Journal: Cell metabolism

Article Title: ABCA12 regulates ABCA1-dependent cholesterol efflux from macrophages and the development of atherosclerosis.

doi: 10.1016/j.cmet.2013.07.003

Figure Lengend Snippet: Figure 1. Cholesterol and Phospholipid Efflux from Abca12-Deficient Cells (A and B) Cholesterol efflux to apoA-I (A) or HDL (B) from liver macrophages isolated from the Abca12El12 strain. (C and D) Cholesterol efflux to apoA-I (C) or HDL (D) from liver macrophages isolated from the Abca12Lx12 strain. (E) Abundance of Abca12 in RAW 264.7 cells after knockdown of Abca12 with siRNA. (F) Cholesterol efflux to apoA-I from RAW 264.7 cells after knockdown of Abca12 with siRNA. (G) Phospholipid efflux to apoA-I from liver macrophages isolated from Abca12El12 strain. (H) Abundance of Abca1, Abcg1, and SR-B1 in RAW 264.7 cells after knockdown of Abca12 with siRNA. (I) Abca1 and Abcg1 abundance in liver macrophages isolated from the Abca12Lx12 strain. (legend continued on next page)

Article Snippet: The antibodies against ABCA1, ABCG1, SR-B1, LXRα, LXRβ, and GAPDH were from Abcam and Aviva Systems Biology.

Techniques: Isolation, Knockdown

Journal: The Journal of Experimental Medicine

Article Title: Normalization of cholesterol metabolism in spinal microglia alleviates neuropathic pain

doi: 10.1084/jem.20202059

Figure Lengend Snippet: ABCA1 and ABCG1 expression in microglia controls nociception and is required for AIBP-mediated reversal of allodynia in a mouse model of CIPN. (A and B) BV-2 cells were incubated for 30 min with AIBP (0.2 µg/ml) or vehicle in complete media, followed by a 5-min incubation with LPS (100 ng/ml). Colocalization of accessible cholesterol with ABCA1 (A) and APOA1 (B) in lipid rafts. Scale bar, 7 µm. Bar graphs show Manders’ tM1 coefficient. (C) Experimental design and timeline: Tamoxifen (TAM, 10 mg/ml, 200 µl/d), cisplatin (2.3 mg/kg), AIBP (0.5 µg/5 µl), or saline (5 µl). (D) Baseline (day 0) withdrawal thresholds before the start of cisplatin intervention. Data from three independent experiments ( n = 8 for vehicle-treated ABC-imKO mice; n = 16 for TAM-treated ABC-imKO mice, and n = 15 for littermates Abca1 fl/fl Abcg1 fl/fl no-Cre [WT] mice treated with TAM). (E) TLR4 surface expression and dimerization and lipid rafts (CTxB) in CD11b + TMEM119 + spinal microglia of naive WT and ABC-imKO mice at baseline (day 0; n = 5 for TLR4 surface expression and lipid raft content analysis for both groups, n = 8 for WT, and n = 9 for ABC-imKO for TLR4 dimerization). (F) Withdrawal thresholds after i.t. saline or AIBP (0.5 µg/5 µl), followed by i.t. LPS (0.1 µg/5 µl) in TAM-induced ABC-imKO mice ( n = 4 per group). (G and H) Withdrawal thresholds following i.p. cisplatin and i.t. saline or AIBP (0.5 µg/5 µl) injections in TAM-induced ABC-imKO (G) and noninduced (vehicle) ABC-imKO (H) mice ( n = 6 per group); data from two independent experiments. (I and J) TLR4 dimerization (I) and lipid rafts (J) in CD11b + TEMEM119 + spinal microglia at day 8 in the groups shown in G and H. Mean ± SEM ( n = 7 or 8) from two independent experiments. *, P < 0.05; **, P < 0.01; ***, P < 0.001. Two-way ANOVA with Bonferroni post hoc test for multiple comparisons in time course analysis; t test for two groups, and one-way ANOVA with Tukey post hoc test for multiple comparisons of more than two groups. sac, sacrificed.

Article Snippet: Separate sections were stained with 1:100 rabbit anti-ABCA1 or 1:100 rabbit anti-ABCG1 antibodies (Novus Biological; RRID:AB_10000630 and RRID:AB_10125717) overnight at 4°C.

Techniques: Expressing, Incubation, Saline

Journal: The Journal of Experimental Medicine

Article Title: Normalization of cholesterol metabolism in spinal microglia alleviates neuropathic pain

doi: 10.1084/jem.20202059

Figure Lengend Snippet: Validation of ABCA1 and ABCG1 KO in spinal microglia of tamoxifen-induced ABC-imKO mice. (A–D) Immunohistochemistry of spinal cord frozen sections from vehicle- and tamoxifen-induced ABC-imKO mice, showing colocalization (COLOC) of ABCA1 and ABCG1 staining with IBA1 (microglia), NeuN (neurons), and GFAP (astrocytes). Slides were mounted with Prolong Gold with DAPI. Confocal images were acquired with a 63× objective and analyzed with ImageJ software for colocalization. Colocalization masks and Pearson’s R values, Manders’ colocalization coefficients above threshold, and randomization Costes P values were calculated as described in Materials and methods for at least one slide for each animal in the experiment. Representative images and values shown correspond to one animal per condition. Scale bar, 50 µm.

Article Snippet: Separate sections were stained with 1:100 rabbit anti-ABCA1 or 1:100 rabbit anti-ABCG1 antibodies (Novus Biological; RRID:AB_10000630 and RRID:AB_10125717) overnight at 4°C.

Techniques: Biomarker Discovery, Immunohistochemistry, Staining, Software

Journal: The Journal of Experimental Medicine

Article Title: Normalization of cholesterol metabolism in spinal microglia alleviates neuropathic pain

doi: 10.1084/jem.20202059

Figure Lengend Snippet: Gene expression in spinal microglia of ABC-imKO mice. Microglia (CD11b + TEMEM119 + ) were FACS-sorted from three groups of ABC-imKO mice: naive or injected with cisplatin (days 1 and 3), followed on day 7 by i.t. saline (5 µl) or AIBP (0.5 µg/5 µl), and terminated on day 8; n = 3 biological replicates (each biological replicate collapsed from three technical replicates). RNA-seq datasets from ABC-imKO and WT (not littermates) mice were acquired in the same experiment. (A) Top: Overlapping genes (purple lines) and pathways (blue lines) induced in naive ABC-imKO microglia and shared with WT microglia in mice treated with cisplatin showed in purple lines connecting overlapping genes and in blue lines connecting the overlapping enriched pathways. Bottom: Venn diagram of up-regulated genes in spinal microglia from WT cisplatin and ABC-imKO naive mice. (B) Enrichment pathway analysis of up- and down-regulated genes induced by ABCA1 and ABCG1 knockdown in microglia, using cutoff P < 0.05, enrichment >1.5, and a minimum overlap of three genes in the pathway. (C) DEGs in naive spinal microglia of TAM-induced ABC-imKO mice. Adjusted P < 0.05 and Benjamini–Hochberg FDR <5%. (D) Overlapping genes and pathways induced by cisplatin treatment in ABC-imKO microglia and shared with WT microglia in mice treated with cisplatin. (E) DEGs in spinal microglia of cisplatin-treated, TAM-induced ABC-imKO mice compared with cisplatin-treated WT mice. Adjusted P < 0.05 and Benjamini–Hochberg FDR <5%. (F and G) Heatmap of DEGs up-regulated (F) or down-regulated (G) in ABC-imKO microglia either in naive or cisplatin condition. cisp, cisplatin.

Article Snippet: Separate sections were stained with 1:100 rabbit anti-ABCA1 or 1:100 rabbit anti-ABCG1 antibodies (Novus Biological; RRID:AB_10000630 and RRID:AB_10125717) overnight at 4°C.

Techniques: Gene Expression, Injection, Saline, RNA Sequencing, Knockdown

Journal: The Journal of Experimental Medicine

Article Title: Normalization of cholesterol metabolism in spinal microglia alleviates neuropathic pain

doi: 10.1084/jem.20202059

Figure Lengend Snippet: Microglial reprogramming by AIBP is dependent on ABCA1/ABCG1 expression. (A) Venn diagram comparing the effect of AIBP treatment on gene expression in WT and ABC-imKO mice in which CIPN was induced by cisplatin. (B) Volcano plot representation of up- and down-regulated genes by AIBP treatment in CIPN comparing AIBP effect on ABC-imKO versus WT mice. Cutoff of adjusted P < 0.05 and absolute fold change >1.5 shown in light green dots. (C) Heatmap of log 2 normalized gene counts of inflammatory genes altered by AIBP in an ABC-dependent manner (down-regulated by AIBP in WT microglia but up-regulated by AIBP in ABC-imKO. (D) Heatmap of cholesterol synthesis and LXR-related genes comparing cisplatin and AIBP effect in WT and ABC-imKO. (E) Heatmap of noninflammatory genes regulated by AIBP in an ABC-dependent manner. (F) Enrichment pathway analysis of up-regulated genes by AIBP in ABC-imKO microglia, using cutoff P < 0.05, enrichment >1.5, and a minimum overlap of three genes in the pathway. Black arrows highlight cholesterol-related pathways and signaling pathway related to neurotransmitter regulation. cisp, cisplatin.

Article Snippet: Separate sections were stained with 1:100 rabbit anti-ABCA1 or 1:100 rabbit anti-ABCG1 antibodies (Novus Biological; RRID:AB_10000630 and RRID:AB_10125717) overnight at 4°C.

Techniques: Expressing, Gene Expression

Journal: Circulation Research

Article Title: Control of Cholesterol Metabolism and Plasma High-Density Lipoprotein Levels by microRNA-144

doi: 10.1161/circresaha.112.300626

Figure Lengend Snippet: Figure 3. miR-144 levels regulate adenosine triphosphate–binding cassette transporter A1 (ABCA1) expression. A, Quantitative real-time polymerase chain reaction analysis of ABCA1, and ABCG1 expression in mouse peritoneal macrophages transfected with miR- 144 mimics. Data are expressed as relative expression and correspond to the mean±SEM from 3 independent experiments. *Significantly different from cells transfected with control mimic (Con-miR), *P≤0.05. B and C, Western blot analysis of ABCA1, adenosine triphosphate– binding cassette transporter G1 (ABCG1), and HSP90 in mouse peritoneal macrophages and THP-1 cells transfected with (B) Con-miR or miR-144, or (C) control inhibitor (Con-inh) or anti–miR-144 in the presence or in the absence of acetylated low-density lipoprotein (Ac-LDL) or T0901317 (T090). Data correspond to a representative experiment among 3 that gave similar results. Values of the band densitometry analysis are shown. D, Luciferase reporter activity in COS-7 cells transfected with Con-miR or miR-144 (40 nmol/L) mimic and human Abca1 3′-untranslated region (UTR) containing the indicated point mutations (PM) in the miR-144 target sites. Data are expressed as mean percent of 3′UTR activity of Con-miR±SEM and are representative of ≥3 experiments. *Significantly different from cells cotransfected with Con-miR and wild-type (WT) 3′UTR. P≤0.05. E, Human Abca1 3′UTR containing the indicated PM in the miR-144 target sites.

Article Snippet: Rabbit polyclonal antibodies against ABCG1 and SR-B1 were obtained from Novus.

Techniques: Binding Assay, Expressing, Real-time Polymerase Chain Reaction, Transfection, Control, Western Blot, Luciferase, Activity Assay

Journal: Circulation Research

Article Title: Control of Cholesterol Metabolism and Plasma High-Density Lipoprotein Levels by microRNA-144

doi: 10.1161/circresaha.112.300626

Figure Lengend Snippet: Figure 6. Delivery of miR-144 oligonucleotides significantly reduces liver adenosine triphosphate–binding cassette transporter A1 (ABCA1) expression and high-density lipoprotein (HDL) levels in mice. A, Quantitative real-time polymerase chain reaction analysis of ABCA1 and ABCG1 expression from mouse livers treated with miR-144 mimics. Data are the mean±SEM from 6 mice per group. *Significantly different from mice injected with nontargeting control mimic (Con-miR) conjugated particles. P≤0.05. B, Analysis of hepatic gene expression 6 days after injection of Con-miR or miR-144. Western blot analysis of liver tissue from 3 representative mice per group. Quantification analysis is shown in the right. Data are the mean±SEM from 6 mice per group. *Significantly different from mice injected with Con-miR conjugated particles. P≤0.05. C Total cholesterol (left), HDL cholesterol (center), and triglyceride (TG; right) levels in mice treated with Con-miR or miR-144 mimics (n=6). *Significantly different from mice injected with nontargeting Con-miR conjugated particles. P≤0.05. D Lipoprotein profile from mice treated with Con-miR or miR-144 mimics (n=6). Cholesterol distribution across plasma fractions was analyzed by fast protein liquid chromatography. ABCG1 indicates adenosine triphosphate–binding cassette transporter G1.

Article Snippet: Rabbit polyclonal antibodies against ABCG1 and SR-B1 were obtained from Novus.

Techniques: Binding Assay, Expressing, Real-time Polymerase Chain Reaction, Injection, Control, Gene Expression, Western Blot, Clinical Proteomics, Fast Protein Liquid Chromatography