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Plasminogen activator inhibitor type-1 synthesis and mRNA expression in HepG2 cells are regulated by VLDL
1Institute of Pharmacological Sciences and E. Grossi Paoletti Center, University of Milan, Italy.
Abstract:
The effect of VLDL on plasminogen activator inhibitor type 1 biosynthesis in HepG2 cells was investigated. Exposure of HepG2 cells to VLDL (range, 10 to 100 micrograms protein per milliliter) for 16 hours resulted in an enhanced release of PAI-1 antigen and PAI activity into conditioned medium, accompanied by the accumulation of intracellular triglycerides. By using a monoclonal antibody (IgG C7) specific to the LDL receptor, we showed that the effect of VLDL is mediated by its interaction with the LDL receptor. Enhanced PAI-1 release was due to increased biosynthesis: PAI-1 mRNA was doubled, mainly because of the effect on the 2.2-kb PAI-1 mRNA rather than the 3.2-kb transcript. Addition of insulin with the VLDL further enhanced PAI-1 antigen release and PAI-1 mRNA accumulation. The effect of VLDL on steady state levels of PAI-1 mRNA was apparently not due to an increase of gene transcription but to stabilization of both PAI-1 mRNA transcripts. The enhancing effect of VLDL on PAI-1 biosynthesis in HepG2 cells may raise PAI-1 antigen levels not only in hypertriglyceridemic states but also in those conditions in which both insulin and VLDL are elevated.
Insights
Very-low-density lipoprotein (VLDL) increases the production of plasminogen activator inhibitor-1 (PAI-1) in liver cells by interacting with LDL receptors. This VLDL-induced rise in PAI-1 may contribute to elevated levels in certain metabolic conditions.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Plasminogen activator inhibitor-1 (PAI-1) is a key regulator of fibrinolysis.
- Elevated PAI-1 levels are associated with thrombotic disorders and metabolic syndrome.
- Very-low-density lipoprotein (VLDL) is implicated in cardiovascular disease, but its direct effect on PAI-1 biosynthesis is not fully understood.
Purpose of the Study:
- To investigate the effect of VLDL on the biosynthesis of PAI-1 in HepG2 cells.
- To elucidate the mechanism by which VLDL influences PAI-1 production.
- To determine the role of the LDL receptor in mediating VLDL's effect on PAI-1.
Main Methods:
- HepG2 cells were exposed to varying concentrations of VLDL.
- PAI-1 antigen and activity in conditioned medium were measured.
- Intracellular triglyceride accumulation was assessed.
- Monoclonal antibody against the LDL receptor was used to block VLDL interaction.
- PAI-1 mRNA levels (2.2-kb and 3.2-kb transcripts) were quantified using RT-PCR.
- Gene transcription rates were evaluated.
Main Results:
- VLDL exposure significantly enhanced PAI-1 antigen release and activity in HepG2 cells.
- VLDL treatment led to intracellular triglyceride accumulation.
- The effect of VLDL on PAI-1 was mediated through interaction with the LDL receptor.
- VLDL doubled PAI-1 mRNA levels, primarily affecting the 2.2-kb transcript.
- Insulin co-administration with VLDL further amplified PAI-1 antigen and mRNA levels.
- VLDL's effect on PAI-1 mRNA was attributed to transcript stabilization, not increased gene transcription.
Conclusions:
- VLDL significantly stimulates PAI-1 biosynthesis in HepG2 cells via LDL receptor interaction.
- VLDL-induced PAI-1 production is linked to increased mRNA stability.
- Combined elevation of VLDL and insulin may contribute to higher PAI-1 levels in metabolic disorders.