Effects of TNF-α and L-Arginine Ex Vivo Treatment on Left Internal Mammary Artery (LIMA) Grafts

Abstract

Background: Coronary artery bypass grafting (CABG) is the standard revascularization procedure, and the left internal mammary artery (LIMA) is the most widely used conduit owing to its long-term patency and favorable impact on survival. Although fibrosis and calcification are comparatively infrequent in internal mammary artery (IMA) grafts, progressive remodeling may still compromise patency. Tumor necrosis factor-alpha (TNF-alpha) is a pro-inflammatory cytokine implicated in vascular remodeling and calcification, whereas L-arginine, a precursor of nitric oxide, may exert vasoprotective effects. Aim: To investigate the effects of ex vivo TNF-alpha and combined TNF-alpha plus L-arginine treatment on the morphology, calcification, elemental composition, and biomechanical properties of human LIMA grafts. Patients and Methods: LIMA graft segments from 18 male CABG patients were allocated to one control group (G0) and two ex vivo treatment groups. Grafts were assessed using hematoxylin and eosin and Masson's Trichrome staining under brightfield microscopy, using Alizarin Red S fluorescence staining to measure calcification, and by scanning electron microscopy with energy-dispersive spectroscopy (SEM/EDS) to assess ultrastructure and elemental composition. Scanning acoustic microscopy (SAM) was used to derive acoustic impedance as a label-free index of tissue stiffness. Results: TNF-alpha treatment was associated with increased acoustic impedance, consistent with graft stiffening, together with greater calcification signatures, whereas combined TNF-alpha and L-arginine treatment shifted impedance toward control-like values and showed more organized tissue architectures with reduced calcification. Conclusions: In this preliminary ex vivo model, L-arginine attenuated TNF-alpha-associated remodeling and stiffening of LIMA grafts, suggesting a potential role in supporting graft durability; larger, statistically powered studies with molecular validation are required to confirm these findings.