Cardiovascular Inflammation and Lesion Cell Apoptosis

Abstract

Objective Increasing evidence suggests that chronic inflammation contributes to atherogenesis, and that acute inflammatory events cause plaque rupture, thrombosis, and myocardial infarction. The present studies examined how inflammatory factors, such as interferon-γ (IFNγ), cause increased sensitivity to apoptosis in vascular lesion cells. Methods and Results Cells from the fibrous cap of human atherosclerotic lesions were sensitized by interferon-γ (IFNγ) to Fas-induced apoptosis, in a Bcl-XL reversible manner. Microarray profiling identified 72 INFγ-induced transcripts with potential relevance to apoptosis. Half could be excluded because they were induced by IRF-1 overexpression, which did not sensitize to apoptosis. IFNγ treatment strongly reduced Mcl-1, phospho-Bcl-2 (ser70), and phospho-Bcl-XL (ser62) protein levels. Candidate transcripts were modulated by siRNA, overexpression, or inhibitors to assess the effect on IFNγ-induced Fas sensitivity. Surprisingly, siRNA knockdown of PSMB8 (LMP7), an ‘immunoproteasome’ component, reversed IFNγ-induced sensitivity to Fas ligation and prevented Fas/IFNγ-induced degradation of Mcl-1, but did not protect p-Bcl-2 or p-Bcl-XL. Proteasome inhibition markedly increased Mcl-1, p-Bcl-2, and p-Bcl-XL levels after IFNγ treatment. Conclusions While critical for antigen presentation, the immunoproteasome appears to be a key link between inflammatory factors and the control of vascular cell apoptosis, and thus may be an important factor in plaque rupture and myocardial infarction.

Apolipoprotein L6, Induced in Atherosclerotic Lesions, Promotes Apoptosis and Blocks Beclin 1-dependent Autophagy in Atherosclerotic Cells

“…The next morning, cells were treated with recombinant human IFN␥ (R&D Systems, 5 ng/50 IU/ml) in 1% FBS media, and 8 h later the sensitivity to apoptosis was examined by determining survival after challenge with a Fas-activating IgM (clone CH11; Upstate Biotechnology) (10 or 25 ng/ml). 20 h later, MTT was added for 4 h, the stained cells were dissolved in DMSO, and the level of reduced MTT was measured by absorbance at 540 nm in a plate reader (34). The efficiency of target silencing was assessed by quantitative RT-PCR (qPCR) mRNA expression level and/or Western blot analysis with anti-ApoL6 antibody.…”

“…Moreover, in the same pulldown experiment, we did not find ApoL6 physically interacted with Bcl-2 or Mcl-1 (data not shown). Importantly, we have previously documented that LDCs possessed elevated Bcl-X L mRNA and protein levels that inhibited caspase activation and apoptosis (34 (15,43,44); 2) P62 (also known as SQSTM1/ sequestome 1), an adaptor protein targeting protein aggregates and damaged organelles for autophagic degradation (39,(43)(44)(45). In so functioning, p62 is selectively incorporated into autophagosomes through binding to LC3-II, degraded by autophagy (46) and a good marker for efficient autophagic activity; 3) LC3/Atg8, activation and translocation during autophagy (14,39,43,44); and 4) Autophagic vesicles (AVs)/ compartments: autophagosomes, amphisomes and autolysosomes (13,43,44).…”

“…Utilizing microarray profiling, quantitative RT-PCR and immunoblot analysis, we recently showed that ApoL6, a BH3-only pro-apoptotic member of the Bcl-2 family, was one of the 36 major downstream targets of INF␥-sensitized, Fas-induced apoptosis in LDCs (34). These cell lines were originally isolated from the fibrous cap of human atherosclerotic lesions, possessing characteristics of myofibroblasts and were resistant to the antiproliferative effects of transforming growth factor-␤ and the pro-apoptotic effects of Fas ligation, similar to activated, apoptosis-resistant SMCs in neointima (35,36).…”

Andrographolide Enhances Nuclear Factor-κB Subunit p65 Ser536 Dephosphorylation through Activation of Protein Phosphatase 2A in Vascular Smooth Muscle Cells

“…3 Persistent increases in inflammatory cytokines derived from immune cells, endothelial cells, and VSMCs have been implicated in vascular dysfunction and vascular diseases, such as atherosclerosis, abdominal aortic aneurysms, and hypertension. These cytokines, including tumor necrosis factors (TNFs), interleukins (ILs), and interferons (IFNs), interact with specific receptors and activate signaling cascades leading to a variety of inflammatory responses involving matrix metalloproteinase (MMP) expressions, nitric oxide (NO) and reactive oxygen species production, and subsequent cell growth, adhesion, and migration (1,2). In addition, there is increasing evidence from animal models that pattern recognition receptors (i.e.…”

“…1 Both authors contributed equally to this work. 2 ptosis (11). Andrographolide was also shown to interfere with T-cell activation and dendritic cell maturation (12).…”