Inhibition of integrin β3, a binding partner of kallistatin, leads to reduced viability, invasion and proliferation in NCI-H446 cells

Abstract

BackgroundKallistatin is a serine proteinase inhibitor and heparin-binding protein. It is considered an endogenous angiogenic inhibitor. In addition, multiple studies demonstrated that kallistatin directly inhibits cancer cell growth. However, the molecular mechanisms underlying these effects remain unclear.MethodsPull-down, immunoprecipitation, and immunoblotting were used for binding experiments. To elucidate the mechanisms, integrin β3 knockdown (siRNA) or blockage (antibody treatment) on the cell surface of small the cell lung cancer NCI-H446 cell line was used.ResultsInterestingly, kallistatin was capable of binding integrin β3 on the cell surface of NCI-H446 cells. Meanwhile, integrin β3 knockdown or blockage resulted in loss of antitumor activities induced by kallistatin. Furthermore, kallistatin suppressed tyrosine phosphorylation of integrin β3 and its downstream signaling pathways, including FAK/-Src, AKT and Erk/MAPK. Viability, proliferation and migration of NCI-H446 cells were inhibited by kallistatin, with Bcl-2 and Grb2 downregulation, and Bax, cleaved caspase-9 and caspase 3 upregulation.ConclusionsThese findings reveal a novel role for kallistatin in preventing small cell lung cancer growth and mobility, by direct interaction with integrin β3, leading to blockade of the related signaling pathway.

Cell surface expression of nucleolin mediates the antiangiogenic and antitumor activities of kallistatin

“…Immunoprecipitation of the kallistatin complex with its binding protein(s) was conducted as previously described [ 12 ]. Whole-cell HUVEC lysates were pretreated with protein A-Sepharose beads for 1 h at 4°C with rotation.…”

“…It was later shown to exert a variety of biological effects in response to pathological conditions such as abnormal blood pressure regulation, tumor formation, and inflammation [ 6 , 8 , 9 ]. Previous studies conducted by our research group demonstrated that adenovirus-mediated kallistatin gene delivery inhibited human colorectal cancer, hepatocellular carcinomas, and small cell lung cancer mainly through inhibition of angiogenesis [ 10 – 12 ]. The anti-angiogenic functions of kallistatin are associated with its structure.…”

“…Although several kallistatin-binding proteins such as integrin β3, Krüppel-like factor 4 (KLF4), and lipoprotein receptor-related protein 6 have been reported as kallistatin receptors [ 12 , 15 , 16 ], the exact mechanisms underlying kallistatin effects remain unclear. Moreover, the involvement of these putative kallistatin receptors in the antitumoral action of kallistatin remains elusive.…”

“…We previously reported the binding of kallistatin to integrin β3 on the surface of human lung cancer NCI-H446 cells, an interaction that negatively affected tumor cell proliferation, migration, and survival [ 12 ]. Shen et al .…”

Deer thymosin beta 10 functions as a novel factor for angiogenesis and chondrogenesis during antler growth and regeneration

“…HUVEC migration was investigated using a modified Transwell assay (Costar, NY, USA; pore size 8 μm, 3422) in 24-well plates [ 29 ]. Deer eTMSB10 at 50, 100, and 150 ng/ml in 700 μl culture medium containing 5% FBS was placed in the bottom reservoir of each chamber.…”