Matrix protein SPARC drives pro-inflammatory macrophage polarization through TLR4-MyD88/TRIF signaling in dairy cows with subclinical ketosis.

Abstract

Secreted protein acidic and rich in cysteine (SPARC) is a matricellular glycoprotein with important roles in extracellular matrix dynamics and immune-metabolic regulation. Although SPARC has been widely implicated in inflammatory and fibrotic disorders in humans and rodents, its involvement in metabolic diseases of ruminants remains unclear. In this study, we investigated the role of SPARC in macrophage-mediated inflammation in dairy cows with subclinical ketosis (SCK) and explored the underlying molecular mechanisms. SPARC levels were significantly elevated in the serum, liver, and adipose tissue of cows with SCK and were positively correlated with pro-inflammatory cytokines, including IL-1β, IL-18, and IFN-γ. In vitro, SPARC stimulation activated the TLR4/NF-κB signaling pathway in a concentration-dependent manner, enhanced iNOS expression, increased inflammatory mediator production, and promoted macrophage polarization toward a pro-inflammatory M1 phenotype, as evidenced by increased CD86 and decreased CD206 and IL-10 expression. Transcriptomic profiling revealed extensive remodeling of immune- and metabolism-related gene networks, with significant enrichment of pathways associated with Toll-like receptor signaling, TNF signaling, and cytokine-receptor interactions. Notably, TLR4 silencing abolished SPARC-induced signaling activation and macrophage polarization, indicating that SPARC functions upstream of the canonical TLR4-MyD88/TRIF axis. Collectively, these findings identify SPARC as a key immune-metabolic regulator that amplifies macrophage-driven inflammation through TLR4-dependent signaling, providing new mechanistic insight into inflammation-associated metabolic disorders in dairy cows.