Human airway smooth muscle cells secrete vascular endothelial growth factor: up‐regulation by bradykinin via a protein kinase C and prostanoid‐dependent mechanism

Abstract

Bronchial vascular remodeling is an important feature of the pathology of chronic asthma, but the responsible mechanisms and main sources of angiogenic factors are unclear. Here we report that human airway smooth muscle cells express vascular endothelial growth factor (VEGF)121, 165, 189, 206 splice variants and secrete VEGF protein constitutively. VEGF protein secretion was increased by the proinflammatory asthma mediator bradykinin through post-transcriptional mechanisms. Bradykinin-induced VEGF secretion was dependent on the B2 bradykinin receptor, activation of protein kinase C, and generation of endogenous prostanoids. This is the first report that bradykinin can increase VEGF secretion in any biological system and the first to show that airway smooth muscle cells produce VEGF. Our results suggest a novel role for human airway smooth muscle in contributing to bronchial mucosal angiogenesis in chronic asthma by secretion of VEGF and suggest a wider role for mesenchymal cell products in mediating angiogenesis in inflammatory and allergic diseases.

Vascular Endothelial Growth Factor Induction by Prostaglandin E2 in Human Airway Smooth Muscle Cells Is Mediated by E Prostanoid EP2/EP4 Receptors and SP-1 Transcription Factor Binding Sites

“…VEGF-A Assay-The enzyme-linked immunosorbent assay (ELISA) was used to measure VEGF-A (R&D Systems, Abingdon, Oxon, UK) according to the manufacturer's instructions and has been described by us in detail elsewhere (15).…”

“…Total RNA was isolated by using the RNeasy mini kit (Qiagen, West Sussex, UK) following the manufacturer's protocol with on-column DNase digestion. 1 g of total RNA was reverse transcribed in a total volume of 20 l including 200 units of Moloney murine leukemia virus reverse transcriptase (Promega, Madison, WI), 25 units of RNase inhibitor (Promega), 0.5 g of oligo(dT) 15 primer, 0.5 mM of each dNTPs, and 1ϫ first-strand buffer provided by Promega. The reaction was incubated at 42°C for 90 min.…”

“…Inflammatory cytokines such as interleukin-1␤ and transforming growth factor-␤ increased VEGF release in human cholangiocellular carcinoma cells, synovial fibroblasts, cardiac myocytes, and airway smooth muscle cells (11)(12)(13)(14). We have previously shown that the pro-inflammatory asthma mediator, bradykinin, increased VEGF production in human airway smooth muscle cells (HASMC) (15). A number of studies have shown that the products of COX-2, the inducible form of cyclooxygenase, may mediate the effect of cytokines and mediators on the release of chemokines and cytokines in an autocrine manner through a mechanism involving endogenous prostanoid production.…”

“…Recent work suggests that this is also true of VEGF. Autocrine PGE 2 increases VEGF release in response to interleukin-1␤ in synovial fibroblasts and in response to bradykinin in HASMC (12,15). Furthermore, exogenous PGE 2 increases VEGF expression in fibroblasts and osteoblasts (16 -18).…”

Asthma: where is it going?

“…ASM cells are directly involved in this cycle, perpetuating lung inflammation (40). In addition, human ASM cells in culture can also produce several compounds implicated in remodeling, including fibronectin, laminin, percelan, chondroitin, and vascular endothelial growth factor (42,43,45,46). This may be especially relevant in the presence of increased smooth muscle mass observed in asthmatic lungs (35).…”