Journal: Journal of Lipid Research

Article Title: Dose-dependent effects of siRNA-mediated inhibition of SCAP on PCSK9, LDLR, and plasma lipids in mouse and rhesus monkey [S]

doi: 10.1194/jlr.M071498

Figure Lengend Snippet: KD of liver Scap mRNA and reduced SREBP activity in mice following Scap siRNA dosing. C57BL/6 mice were dosed with the indicated siRNAs. On day 10 after siRNA dosing, mice were injected with D 2 O and livers and plasma were collected after 4 h. Scap mRNA (A), Pcsk9 mRNA (B), and Ldlr mRNA (C) levels in liver. D: Direct comparison of Pcsk9 and Ldlr mRNA levels as a function of SCAP siRNA dose. De novo palmitate (E) and cholesterol synthesis (F) were measured as described in the Materials and Methods. Each bar represents mean ± SEM; n = 8 per group. *P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: TaqMan qPCR was done on an ABI 7900 real-time PCR system using the following primer pairs commercially obtained from ABI: Rh02621745_g1 (GAPDH), Rh02863945_m1 (SCAP), Rh03418189_m1 (PCSK9), Rh02828936_m1 (LDLR).

Techniques: Activity Assay, Injection, Clinical Proteomics, Comparison

Journal: Journal of Lipid Research

Article Title: Dose-dependent effects of siRNA-mediated inhibition of SCAP on PCSK9, LDLR, and plasma lipids in mouse and rhesus monkey [S]

doi: 10.1194/jlr.M071498

Figure Lengend Snippet: Effect of Scap siRNA on plasma PCSK9, LDL-C, TG production, and LDLR protein. C57BL/6 mice were dosed with the indicated siRNAs and plasma and livers were collected 10 days after dosing. PCSK9 (A) and LDL-C (C) were measured in EDTA-plasma. B: Quantification of LDLR protein as measured by Western blot. D: TG production rate was measured in plasma by LC-MS/MS after an intravenous dose of [ 13 C 18 ]oleic acid as described in the Materials and Methods. E: Representative Western blot of liver LDLR protein. Each bar represents mean ± SEM; n = 8 per group [except (B), n = 6 per group]. * P < 0.05 and *** P < 0.001.

Article Snippet: TaqMan qPCR was done on an ABI 7900 real-time PCR system using the following primer pairs commercially obtained from ABI: Rh02621745_g1 (GAPDH), Rh02863945_m1 (SCAP), Rh03418189_m1 (PCSK9), Rh02828936_m1 (LDLR).

Techniques: Clinical Proteomics, Western Blot, Liquid Chromatography with Mass Spectroscopy

Journal: Journal of Lipid Research

Article Title: Dose-dependent effects of siRNA-mediated inhibition of SCAP on PCSK9, LDLR, and plasma lipids in mouse and rhesus monkey [S]

doi: 10.1194/jlr.M071498

Figure Lengend Snippet: Scap siRNA KD adds to simvastatin LDL-C lowering in NFR-CETP mice. Mice were dosed orally daily with 100 mg/kg simvastatin or vehicle starting on day 0. On day 4, mice were dosed with nt control-1 siRNA, Pcsk9 siRNA (0.5 mg/kg), and m-Scap-1 siRNA at the indicated doses (milligrams per kilogram). Plasma and livers were collected 10 days later. Liver Scap mRNA (A), liver Hmgcr mRNA (B), plasma PCSK9 (C), and plasma LDL-C (D). Each bar in (A) and (B) represents mean ± SEM. Box and whiskers in (C) and (D) represent the 25th percentile, the median, the 75th percentile (boxes), and the minimum and maximum (whiskers); n = 8 per group. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: TaqMan qPCR was done on an ABI 7900 real-time PCR system using the following primer pairs commercially obtained from ABI: Rh02621745_g1 (GAPDH), Rh02863945_m1 (SCAP), Rh03418189_m1 (PCSK9), Rh02828936_m1 (LDLR).

Techniques: Control, Clinical Proteomics

Journal: Journal of Lipid Research

Article Title: Dose-dependent effects of siRNA-mediated inhibition of SCAP on PCSK9, LDLR, and plasma lipids in mouse and rhesus monkey [S]

doi: 10.1194/jlr.M071498

Figure Lengend Snippet: Effect of single dose SCAP siRNA on SCAP mRNA and SREBP activity in rhesus macaque monkeys. Lean rhesus monkeys were dosed intravenously with the indicated siRNAs. Liver biopsy samples were obtained on day −7 (prior) and days 7, 14, 28, and 35 after siRNA dosing for measurement of liver SCAP mRNA (A), liver PCSK9 mRNA (B), and liver LDLR mRNA. shows a direct comparison of liver PCSK9 and LDLR mRNA. D: On day 16, animals were administered D 2 O and blood samples were collected at −0.5, +4, +24, and +48 h for the determination of palmitate and cholesterol synthesis. Each symbol or bar represents mean ± SEM; n = 4–10 per group. Liver SCAP mRNA was significantly lower in the NHP SCAP-1 group relative to nt control-2 at all time points ( P < 0.05). Liver PCSK9 mRNA was lower in the PCSK9 and NHP SCAP-1 groups relative to nt control-2 group at all time points ( P < 0.05). * P < 0.05 and ** P < 0.01.

Article Snippet: TaqMan qPCR was done on an ABI 7900 real-time PCR system using the following primer pairs commercially obtained from ABI: Rh02621745_g1 (GAPDH), Rh02863945_m1 (SCAP), Rh03418189_m1 (PCSK9), Rh02828936_m1 (LDLR).

Techniques: Activity Assay, Comparison, Control

Journal: Journal of Lipid Research

Article Title: Dose-dependent effects of siRNA-mediated inhibition of SCAP on PCSK9, LDLR, and plasma lipids in mouse and rhesus monkey [S]

doi: 10.1194/jlr.M071498

Figure Lengend Snippet: Reduction in plasma PCSK9 and lipids in rhesus monkeys after treatment with SCAP siRNA. Lean rhesus macaque monkeys were dosed intravenously with the indicated siRNAs on day 0 and blood was collected for EDTA plasma on several days for analysis as indicated in the graphs. Plasma PCSK9 (A), plasma LDL-C (B), and plasma HDL-C (C). PCSK9 was significantly lower in the NHP SCAP-1 siRNA group relative to nt control-2 on days 3–21 and significantly lower in the PCSK9 siRNA group relative to nt control-2 on days 3–35 ( P < 0.05). LDL-C was significantly lower in the NHP SCAP-1 siRNA group relative to nt control-2 on days 14 and 18–35, and significantly lower in the PCSK9 siRNA group relative to nt control-2 on days 3–35 ( P < 0.05). HDL-C was significantly lower in the NHP SCAP-1 siRNA group relative to nt control-2 on days 10–35 ( P < 0.05). Each symbol represents mean ± SEM; n = 4–10 per group.

Article Snippet: TaqMan qPCR was done on an ABI 7900 real-time PCR system using the following primer pairs commercially obtained from ABI: Rh02621745_g1 (GAPDH), Rh02863945_m1 (SCAP), Rh03418189_m1 (PCSK9), Rh02828936_m1 (LDLR).

Techniques: Clinical Proteomics, Control

Journal: Journal of Lipid Research

Article Title: Dose-dependent effects of siRNA-mediated inhibition of SCAP on PCSK9, LDLR, and plasma lipids in mouse and rhesus monkey [S]

doi: 10.1194/jlr.M071498

Figure Lengend Snippet: Dose-dependent reduction in liver SCAP, PCSK9 mRNA, and plasma PCSK9 in rhesus monkeys following treatment with SCAP siRNA. Lean rhesus macaque monkeys were dosed intravenously with the indicated siRNAs on day 0 and blood was collected for analysis on the days indicated in the graphs. Liver biopsy samples were obtained on day −2 (prior) and days 3, 12, and 26 after siRNA dosing for measurement of liver SCAP mRNA (A) and liver PCSK9 mRNA (B). C: Comparison of liver PCSK9 and LDLR mRNA on day 3 for the indicated NHP SCAP-1 siRNA doses. D: Plasma PCSK9 levels. Each symbol represents mean ± SEM. Box and whiskers in (C) represent the 25th percentile, the median, the 75th percentile (boxes), and the minimum and maximum (whiskers). Liver SCAP mRNA was significantly reduced ( P < 0.05) for all doses and time points relative to nt control-2. Liver PCSK9 mRNA was significantly lower in the 26.7 mg/m 2 dose group relative to nt control-2 at all time points and significantly lower in the 13.35 mg/m 2 dose group relative to nt control-2 on day 12 ( P < 0.05). Plasma PCSK9 was significantly reduced for all SCAP siRNA doses relative to nt control-2 siRNA on days 3–28 ( P < 0.05) (n = 4 for the 26.7 mg/m 2 dose group; n = 8 for all other groups).

Article Snippet: TaqMan qPCR was done on an ABI 7900 real-time PCR system using the following primer pairs commercially obtained from ABI: Rh02621745_g1 (GAPDH), Rh02863945_m1 (SCAP), Rh03418189_m1 (PCSK9), Rh02828936_m1 (LDLR).

Techniques: Clinical Proteomics, Comparison, Control

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 4. Angptl2 knockout improved renal UA excretion in HUA mice model (A) Angptl2 knockout and identification protocol (left panel), and genotype identification results by DNA agarose gel electrophoresis (right panel); (B) HUA mice model flow chart. (C) Fasting body weight before sacrifice. (D–G) Serum UA, CREA, BUN and XOD levels in WT, HUA, Angptl2 ko-HUA groups. (H) Urinary UA level in WT, HUA, Angptl2 ko-HUA groups. (I) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in the kidney. (J) Representative data of ABCG2 and GLUT9 in the kidney by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; CREA, creatinine; BUN, blood urea nitrogen; UA, uric acid; HUA, hyperuricemia; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Knock-Out, Agarose Gel Electrophoresis, Western Blot, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 5. Adipocyte-specific overexpression of Angptl2 significantly increased serum UA levels in mice induced by adenine and potassium oxonate (A) HUA mice model flow chart. (B) Fasting body weight before sacrifice. (C) Serum ANGPTL2 levels after adipocyte-specific Angptl2 overexpression. (D–F) Serum CREA, BUN and UA levels after adipocyte-specific Angptl2 overexpression. (G) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in the kidney. (H) Representative data of ABCG2 and GLUT9 in the kidney by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; CREA, creatinine; BUN, blood urea nitrogen; HUA, hyperuricemia; UA, uric acid; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Over Expression, Western Blot, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 6. ANGPTL2 decreased ABCG2 by suppressing AKT activity in HK2 cells (A) Flow chart of HK2 treatment groups. (B) UA levels in HK2 cell supernatants. (C) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in HK2 cells. (D) Representative data of ABCG2 and GLUT9 in HK2 cells by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; rhA2, human recombinant proteins ANGPTL2; HUA, hyperuricemia; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Activity Assay, Western Blot, Recombinant, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 7. ANGPTL2 decreased ABCG2 by suppressing AKT activity in primary renal tubular epithelial cells. (A) Flow chart of RTECs treatment groups. (B) UA levels in RTECs supernatants. (C) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in RTECs. (D) Representative data of ABCG2 and GLUT9 in RTECs by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; rhA2, human recombinant proteins ANGPTL2; HUA, hyperuricemia; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Activity Assay, Western Blot, Recombinant, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 9. Adipocyte-secreted ANGPTL2 promotes UA level through AKT/ABCG2 signaling in HK2 cells. (A) Flow diagram of C3H10 induction and treatment with FFA, as well as oil red O staining. (B) Representative data of ANGPTL2 after FFA treated C3H10 cells differentiated adipocytes by Western blotting (left panel), and statistic data (right panel). (C) ANGPTL2 levels in FFA-treated C3H10 cells differentiated adipocytes. (D) Flow chart of conditioned medium collected from C3H10 differ entiated into adipocytes treated with HK2 cells. (E) UA levels in HK2 cell supernatants after conditioned medium treatment. (F) Representative data of ABCG2 in HK2 cells by Western blotting (left panel), and statistic data (right panel). (G) Graphic summary of the study. Adipocyte-secreted ANGPTL2 decreased ABCG2 through inhibited AKT activation, then increased UA levels and led to HUA. ANGPTL2, angiopoietin-like protein 2; FFA, free fatty acid; HUA, hyperuricemia; ABCG2, ATP binding cassette G member 2; GLUT9, glucose transporter 9. *p < 0.05, **p < 0.01, ****p < 0.0001. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Staining, Western Blot, Activation Assay, Binding Assay