Apoptosis of endothelial cells triggers a caspase‐dependent anti‐apoptotic paracrine loop active on vascular smooth muscle cells

Abstract

Increased endothelial apoptosis and decreased apoptosis of vascular smooth muscle cells (VSMC) are central to initiation of myo-intimal thickening. We hypothesized that apoptosis of endothelial cells (EC) induces the release of anti-apoptotic mediator(s) active on VSMC. We found that serum-free medium conditioned by apoptotic EC decreases apoptosis of VSMC compared with fresh serum-free medium. Inhibition of endothelial apoptosis during conditioning with a pan-caspase inhibitor ZVAD-FMK blocked the release of the anti-apoptotic factor(s) active on VSMC. VSMC exposed to serum-free medium conditioned by apoptotic EC showed increased ERK 1/2 phosphorylation, enhanced Bcl-xl expression, and inhibition of p53 expression. Fractionation of the conditioned medium followed by mass spectral analysis identified one bioactive component as a C-terminal fragment of the domain V of perlecan. Serum-free medium supplemented with either a synthetic peptide containing the EGF motif of the domain V of perlecan or chondroitin 4-sulfate, a glycosaminoglycan anchored on the domain V of perlecan, increased ERK 1/2 phosphorylation and Bcl-xl protein levels while inhibiting apoptosis of VSMC. These results suggest that a proteolytic activity developing downstream of activated caspases in apoptotic EC initiates degradation of pericellular proteoglycans and liberation of bioactive fragments with a robust impact on inhibition of VSMC apoptosis.

Caspase-3-dependent export of TCTP: a novel pathway for antiapoptotic intercellular communication

“…To study the secretome of apoptotic EC, human umbilical vein endothelial cells (HUVEC) were rendered apoptotic in vitro by serum starvation (SS) for 4 h, as reported previously. [3][4][5][6][7][8] In serum-starved EC, the percentage of cells with chromatin condensation in absence of cell membrane permeabilization increased over time, suggesting increased apoptosis ( Figure 1a). The absence of lactate dehydrogenase (LDH) release in serum-starved EC was also consistent with absence of cell membrane permeabilization ( Figure 1b Characterization of the secretome of apoptotic EC.…”

“…6 Concomitantly, SSC-Apo was fractionated by fast protein liquid chromatography (FPLC), and the antiapoptotic activity of each fraction was evaluated on VSMC. 3 The protein mediators present in fractions with significant antiapoptotic activity were further characterized by SDS-PAGE LC-MS/MS.…”

“…A C-terminal fragment of perlecan (LG3), released as a consequence of cathepsin L translocation from apoptotic endothelial cells (EC), induces Bcl-xl upregulation and resistance to apoptosis in fibroblasts, mesenchymal stem cells and smooth muscle cells, all of which are pivotal to tissue or vascular remodeling and repair. [3][4][5][6][7] Recently, we showed that caspase-3 activation in EC favors the release of connective tissue growth factor (CTGF), 8 which induces the differentiation of fibroblasts into myofibroblasts, a mandatory process in all forms of repair. To date, caspase-3 activation has been implicated in the release of most, if not all, mediators secreted by apoptotic cells, 1,2,6,8 suggesting that the paracrine response is embedded within the effector phase of apoptosis.…”

“…[11][12][13] Mediators produced by apoptotic EC, such as LG3 and CTGF, actively induce ERK 1/2-and/or Bcl-xl-dependent antiapoptotic phenotypes in vascular smooth muscle cells (VSMC), mesenchymal stem cells and myofibroblasts. [3][4][5][6][7][8] This raises the possibility that a chronic increase in EC apoptosis, resulting in a sustained paracrine response, could contribute to maladaptive vascular repair.…”

In Vitro and Pilot In Vivo Evaluation of a Bioactive Coating for Stent Grafts Based on Chondroitin Sulfate and Epidermal Growth Factor

“…Two different bioactive molecules were selected to create a bioactive coating: (i) chondroitin sulfate (CS), a glycosaminoglycan of the extracellular matrix that participates in the integrity of the arterial wall in normal aortas [9] and exhibits anti-apoptotic properties [10]; and (ii) epidermal growth factor (EGF), a major actor of wound-healing [11] presenting both anti-apoptotic and pro-proliferative properties [12,13]. This choice is based on previous studies that have shown the superiority of the combination of CS and EGF [14].…”