High throughput screening identifies microRNAs regulating PCSK9 and LDL receptor expression
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PROVIDER: PRJNA701607 | ENA |
REPOSITORIES: ENA
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Project description:Using a high throughput human microRNA library screen, we identified microRNAs targeting the 3’untranslated region of human PCSK9
2021-07-07 | GSE166680 | GEO
Project description:Circulating Proprotein Convertase Subtilisin/Kexin type 9 (PCSK9) levels, known for regulating plasma cholesterol by degrading LDL receptors, correlate with acute myocardial infarction (AMI) risk. Recent studies suggest that PCSK9 improves cardiac function beyond LDL cholesterol levels after cardiac ischemic injury, but the precise role of PCSK9 in this process is not clarified yet. Our study reveals that PCSK9 deficiency induces heterogeneous changes in myeloid cells and macrophages, potentially protecting the heart in AMI regardless of LDL cholesterol homeostasis. Single-cell RNA sequencing identifies PCSK9-dependent cardiac macrophages (PDCMs) as reparative macrophages enriched in activator protein-1 (AP-1) transcription factor (TF)-related pathways. PDCMs enhance VEGF-C secretion and activate Akt signalling in cardiac endothelial cells, improving cardiac remodelling. Indeed, PCSK9 inhibitor-treated coronary artery disease (CAD) patients show increased myeloid cells with PDCM-like features. In sum, targeting myeloid-PCSK9 may offer cardio-protective effects through increased AP-1 activity and VEGF-C expression, providing a novel approach to prevent cardiac dysfunction in AMI.
2025-07-01 | GSE285004 | GEO
Project description:Targeting myeloid-PCSK9 enhancing cardio-protection beyond LDL cholesterol-lowering
| PRJNA1200779 | ENA
Project description:The majority of known pathogenic point mutations in the human genome are C•G to T•A substitutions. Adenine base editors (ABEs), comprised of nuclease-impaired Cas9 fused to adenine deaminases, enable direct repair of these mutations, making them promising tools for precision in vivo genome editing therapies. However, prior to application in patients, thorough safety and efficacy studies in relevant model organisms are needed. Here, we apply adenine base editing in vivo in the liver of mice and cynomolgus macaques to install a splice site mutation in PCSK9 and reduce blood low-density lipoprotein (LDL) levels, a well-known risk factor for cardiovascular disease. Intravenous delivery of ABE-encoding mRNA and a locus-specific single guide (sg)RNA utilizing lipid nanoparticle (LNP) technology induce up to 67% editing in the liver of mice and up to 34% editing in the liver of macaques, leading to a reduction of plasma PCSK9 and LDL levels. We observed rapid clearance of ABE mRNA after LNP-mediated delivery, and neither sgRNA-dependent nor sgRNA-independent off-target mutations are detected in genomic DNA. Together, our findings support safety and feasibility of adenine base editing to treat patients with monogenetic liver diseases.
2021-04-13 | GSE168363 | GEO
Project description:The majority of known pathogenic point mutations in the human genome are C•G to T•A substitutions. Adenine base editors (ABEs), comprised of nuclease-impaired Cas9 fused to adenine deaminases, enable direct repair of these mutations, making them promising tools for precision in vivo genome editing therapies. However, prior to application in patients, thorough safety and efficacy studies in relevant model organisms are needed. Here, we apply adenine base editing in vivo in the liver of mice and cynomolgus macaques to install a splice site mutation in PCSK9 and reduce blood low-density lipoprotein (LDL) levels, a well-known risk factor for cardiovascular disease. Intravenous delivery of ABE-encoding mRNA and a locus-specific single guide (sg)RNA utilizing lipid nanoparticle (LNP) technology induce up to 67% editing in the liver of mice and up to 34% editing in the liver of macaques, leading to a reduction of plasma PCSK9 and LDL levels. We observed rapid clearance of ABE mRNA after LNP-mediated delivery, and neither sgRNA-dependent nor sgRNA-independent off-target mutations are detected in genomic DNA. Together, our findings support safety and feasibility of adenine base editing to treat patients with monogenetic liver diseases.
2021-04-13 | GSE168364 | GEO
Project description:Proprotein convertase subtilisin kexin type 9 (PCSK9) is a critical modulator of cholesterol homeostasis. Whereas PCSK9 gain-of-function (GOF) mutations are associated with autosomal dominant hypercholesterolemia (ADH) and premature atherosclerosis, PCSK9 loss-of-function (LOF) mutations have a cardio-protective effect and in some cases can lead to familial hypobetalipoproteinemia (FHBL). However, limitations of the currently available cellular models preclude deciphering the consequences of PCSK9 mutation further. We aimed to validate urine-sample-derived human induced pluripotent stem cells (UhiPSCs) as an appropriate tool to model PCSK9-mediated ADH and FHBL. To achieve our goal, urine-sample-derived somatic cells were reprogrammed into hiPSCs by using episomal vectors. UhiPSC were efficiently differentiated into hepatocyte-like cells (HLCs). Compared to control cells, cells originally derived AQ3 from an individual with ADH (HLC-S127R) secreted less PCSK9 in the media (−38.5%; P=0.038) and had a 71% decrease (P<0.001) of low-density lipoprotein (LDL) uptake, whereas cells originally derived from an individual with FHBL (HLC-R104C/V114A) displayed a strong decrease in PCSK9 secretion (−89.7%; P<0.001) and had a 106% increase (P=0.0104) of LDL uptake. Pravastatin treatment significantly enhanced LDL receptor (LDLR) and PCSK9 mRNA gene expression, as well as PCSK9 secretion and LDL uptake in both control and S127R HLCs. Pravastatin treatment of multiple clones led to an average increase of LDL uptake of 2.19±0.77-fold in HLC-S127R compared to 1.38±0.49 fold in control HLCs (P<0.01), in line with the good response to statin treatment of individuals carrying the S127R mutation (mean LDL cholesterol reduction=60.4%, n=5). In conclusion, urine samples provide an attractive and convenient source of somatic cells for reprogramming and hepatocyte differentiation, but also a powerful tool to further decipher PCSK9 mutations and function.
2015-12-01 | GSE75545 | GEO
Project description:Investigations into the regulatory mechanisms controlling cholesterol homeostasis have proven fruitful in identifying low-density lipoprotein (LDL)-lowering therapies to reduce the risk of atherosclerotic cardiovascular disease. A major advance was the discovery of proprotein convertase subtilisin/kexin type 9 (PCSK9), a secreted protein that binds the LDL receptor (LDLR) on the cell surface and internalizes it for degradation, thereby blunting its ability to take up circulating LDL. The discovery that loss-of-function mutations in PCSK9 lead to lower plasma levels of LDL cholesterol and protection from cardiovascular disease led to the therapeutic development of PCSK9 inhibitors at an unprecedented pace. However, there remain many gaps in our understanding of PCSK9 regulation and biology, including its posttranscriptional control by microRNAs. Using a high-throughput region(3'-UTR) of human microRNA library screen, we identified microRNAs targeting the 3' untranslated region of human PCSK9. The top 35 hits were confirmed by large-format PCSK9 3'-UTR luciferase assays, and 10 microRNAs were then selected for further validation in hepatic cells, including effects on PCSK9 secretion and LDLR cell surface expression. These studies identified seven novel microRNAs that reduce PCSK9 expression, including miR-221-5p, miR-342-5p, miR-363-5p, miR-609, miR-765, and miR-3165. Interestingly, several of these microRNAs were also found to target other genes involved in LDLR regulation and potently upregulate LDLR cell surface expression in hepatic cells. Together, these data enhance our understanding of post-transcriptional regulators of PCSK9 and their potential for therapeutic manipulation of hepatic LDLR expression.
| S-EPMC8310920 | biostudies-literature
Project description:PCSK9 promotes the lysosomal degradation of cell surface LDL receptor (LDLR). We analyzed how excess LDLR generated by PCSK9 deficiency is differently handled in male and female mice to possibly unveil the mechanism leading to the lower efficacy of PCSK9 mAb on LDL-cholesterol levels in women. Analysis of intact or ovariectomized PCSK9 knockout (KO) mice supplemented with placebo or 17β-estradiol (E2) demonstrated that female, but not male mice massively shed the soluble ectodomain of the LDLR in the plasma. Liver-specific PCSK9 KO or alirocumab-treated WT mice exhibit the same pattern. This shedding is distinct from the basal one and is inhibited by ZLDI-8, a metalloprotease inhibitor pointing at ADAM10/ADAM17. In PCSK9 KO female mice, ZLDI- 8 raises by 80 % the LDLR liver content in a few hours. This specific shedding is likely cholesterol-dependent: it is prevented in PCSK9 KO male mice that exhibit low intra-hepatic cholesterol levels without activating SREBP-2, and enhanced by mevalonate or high cholesterol feeding, or by E2 known to stimulate cholesterol synthesis via the estrogen receptor-α. Liver transcriptomics demonstrates that critically low liver cholesterol in ovariectomized female or knockout male mice also hampers the cholesterol-dependent G2/M transition of the cell cycle. Finally, higher levels of shed LDLR were measured in the plasma of women treated with PCSK9 mAb. PCSK9 knockout female mice hormonally sustain cholesterol synthesis and shed excess LDLR, seemingly like women. In contrast, male mice rely on high surface LDLR to replenish their stocks, despite 80 % lower circulating LDL.
2022-09-22 | GSE205008 | GEO
Project description:Purpose: PCSK9, which regulates hepatic lipid homeostasis by modulating LDL levels, is expressed in small quantities in the heart, but its role is unknown. Here, we sought to determine its cardiac function and identify the mechanisms. Mice with heart-specific deletion of Pcsk9 had reduced contractile capacity, heart failure with preserved ejection fraction, and left ventricular dilatation at 28 weeks of age and died prematurely. Results: Transcriptomic analyses revealed alterations of signaling pathways linked to cardiomyopathy and energy metabolism. Mitochondrial membrane lipid composition and cristae morphology were also altered, impairing the assembly and activity of mitochondrial respiratory complexes. Mitochondrial oxidation was reduced and compensatory glycolytic energy production was insufficient to support demand. Conclusions: Thus, PCSK9 is a key factor in cardiac metabolic function and PCSK9 deficiency is linked to cardiomyopathy, impaired heart function, and compromised energy production.
2023-07-12 | GSE178389 | GEO
Project description:Proprotein convertase subtilisin kexin type 9 (PCSK9) is a key regulator of LDL cholesterol metabolism and the target of lipid-lowering drugs. PCSK9 is mainly expressed in hepatocyte. Here, we show that PCSK9 is highly expressed in undifferentiated hiPSCs. PCSK9 inhibition in hiPSCs with the use of shRNA, CRISPR/cas9-mediated knockout or endogenous PCSK9 loss-of-function mutation R104C/V114A unveiled its new role as a potential cell cycle regulator through the NODAL signaling pathway. Indeed, PCSK9 inhibition leads to a decrease of SMAD2 phosphorylation and hiPSCs proliferation. Conversely, PCSK9 overexpression stimulates hiPSCs proliferation. PCSK9 can interfere with the NODAL pathway by regulating the expression of its endogenous inhibitor DACT2, which is involved in the TGFß-R1 lysosomal degradation. Using different PCSK9 constructs we show that PCSK9 interacts with DACT2 through its CHRD domain. Altogether these data highlight a new role of PCSK9 in cellular proliferation and development, beyond its canonical effect on lipid metabolism.
2021-08-05 | GSE181481 | GEO