Fig 2: PRRSV infection upregulated AMPK phosphorylation levels. (A) PK-15CD163 or (B) PAMs were infected with PRRSV (MOI = 0.5) for 12, 24 or 36 h. Lysates were collected and analyzed by western blot assay for the levels of total AMPK, phosphorylated-AMPK (p-AMPK) and PRRSV-N protein. ß-actin was measured as a loading control.

Fig 3: Model of PRRSV-mediated manipulation of the AMPK–ACC1 pathway. Acetyl-CoA is metabolized to malonyl-CoA via ACC1, which is then transformed into palmitates by FA synthase (FASN). Palmitates are prolonged through FA elongase to form saturated FAs, which are subsequently metabolized to unsaturated FAs by desaturase. PRRSV infection activated AMPK, thus resulting in an increased level of phosphorylated ACC1 (inactive form) to block FA synthesis. Pharmacological agents targeting various steps of FA synthesis pathway, such as C75 (inhibitor of FASN), CP-640186 (inhibitor of ACC1), and A769662 (activator of AMPK), suppressed FA biosynthesis and subsequently inhibited the replication of PRRSV. In summary, host cells limit PRRSV infection via the AMPK–ACC1 pathway.

Fig 4: Knockdown of AMPK-enhanced PRRSV infection. (A) PK-15CD163 cells were transfected with AMPK-specific siRNAs (siAMPKs) or NC siRNA. At 36 h post transfection, cells were harvested to determine the knockdown efficiency through western blot assay. (B,C) PK-15CD163 were transfected with siAMPKs or NC siRNA. After 24 h, cells were infected with PRRSV (MOI of 0.5) and processed to detect the mRNA levels of the PRRSV NSP9 gene by qRT-PCR assay (B) and virus titers by plaque assay (C) at 24 hpi. The results represent the means and standard deviations from three independent experiments. ***, p < 0.001.

Fig 5: A proposed model of the potential mechanisms involved in the protective effect of THSG against gentamicin-induced ototoxicity. THSG, 2,3,4',5-tetrahydroxystilbene-2-O-ß-D-glucoside; AMPK, AMP-activated protein kinase.