Calpastatin Counteracts Pathological Angiogenesis by Inhibiting Suppressor of Cytokine Signaling 3 Degradation in Vascular Endothelial Cells
Rationale: Janus kinase/signal transducer and activator of transcription (JAK/STAT) signals and their endogenous inhibitor suppressor of cytokine signalling 3 (SOCS3) in vascular endothelial cells (ECs) reportedly dominate the pathological angiogenesis. However, how these inflammatory signals are potentiated during pathological angiogenesis has not been fully elucidated. We suspected that an intracellular protease calpain, which compose the multifunctional proteolytic systems together with its endogenous inhibitor calpastatin (CAST), contributes to the JAK/STAT regulations.
Objective: To specify the impact of EC calpain/CAST systems on JAK/STAT signals and its relationship to pathological angiogenesis.
Methods and Results: The loss of CAST, which is ensured by several growth factor classes, was detectable in neovessels in murine allograft tumours, some human malignant tissues and oxygen-induced retinopathy (OIR) lesions in mice. EC-specific transgenic introduction of CAST caused down-regulation of JAK/STAT signals, up-regulation of SOCS3 expression and depletion of vascular endothelial growth factor (VEGF)-C, thereby counteracting unstable pathological neovessels and disease progression in tumours and OIR lesions in mice. Neutralizing antibody against VEGF-C ameliorated pathological angiogenesis in OIR lesions. Small-interfering RNA-based silencing of endogenous CAST in cultured ECs facilitated μ-calpain-induced proteolytic degradation of SOCS3, leading to VEGF-C production through amplified interleukin-6-driven STAT3 signals. Interleukin-6-induced angiogenic tube formation in cultured ECs was accelerated by CAST silencing, which is suppressible by pharmacological inhibition of JAK/STAT signals, antibody-based blockage of VEGF-C and transfection of calpain-resistant SOCS3, while transfection of wild-type SOCS3 exhibited modest angiostatic effects.
Conclusions: Loss of CAST in angiogenic ECs facilitates µ-calpain-induced SOCS3 degradation, which amplifies pathological angiogenesis through interleukin-6/STAT3/VEGF-C axis.
- Received September 30, 2014.
- Revision received February 2, 2015.
- Accepted February 3, 2015.