https://orcid.org/0000-0002-2757-304X
Figures & data
Cichoric acid (CA) extract of Dandelion advanced PAQR3 expression to ameliorate ferroptosis in DN by activating the PI3K/AKT signaling pathways. Importantly, CA advanced PAQR3 ubiquitination to restore PI3K/AKT signaling through the relieving the interaction between PAQR3 and P110α pathway.
Figure 1. CA improved DN in mice model.
Structural formula of CA (a), blood glucose (b), body weight (c), water intake (d), Kidney/body weight (e), serum creatinine (f), urea nitrogen (g); urinary albumin levels (h), glomerulus injury (HE staining, i). Sham, sham control mice; Model, mice model of DN; Med or High, 15 or 30 mg/kg/d of CA; ME, 195 mg/kg metformin. **P < .01 sham group; ##P < .01 versus Model group. Data were expressed as mean ± SEM.
Figure 2. CA reduced acute kidney injury (AKI) and inflammation in mice model or in vitro model.
E-cadherin and periostin mRNA expression (a), glomerulus injury (MASS staining, b/PAS, c), podocyte ultrastructure changes (electron microscope, d), IL-1β/IL-6/INF-γ/TNF-α in mice model (e) or in vitro model (f). Sham, sham control mice; Model, mice model of DN; Med or High, 15 or 30 mg/kg/d of CA; ME, 195 mg/kg of metformin. Control, control; Model, vitro model; Med or High, 10 and 20 μM of CA; ME, 250 μM of metformin. **P < .01 sham group; ##P < .01 versus Model group. Data were expressed as mean ± SEM.
Figure 3. CA reduced high glucose-induced ferroptosis of NRK-52E through the induction ferroptosis of GSH/GPX4 axle.
Cell viability (a), LDH activity level (b), cell apoptosis (c), proportions of PI positive cells (d), GSH activity (e), lron concentration (f), GPX4 protein expression (g) in vitro model; ROS levels (live imaging, h), GSH activity (i), GPX4 protein expression in kidney tissue (j); JC-1 disaggregation (k), calcein levels (l), CA increased calcein levels and reduced mitochondrial damage in high glucose-treated NRK-52E cells (). in mice model. Sham, sham control mice; Model, mice model of DN; Low, Med or High, 7.5 or 15 or 30 mg/kg/d of CA. Control, control; Model, vitro model; Low, Med or High, 5 or 10 and 20 μM of CA. **P < .01 sham group; ##P < .01 versus Model group. Data were expressed as mean ± SEM.
Figure 4. Ferroptosis activator reduced the effects of CA on DN in mice model or in vitro model.
Blood glucose (a), body weight (b), serum creatinine (c), urea nitrogen (d), urinary albumin levels (e), water intake (f), Kidney/body weight (g), E-cadherin and periostin mRNA expression (h and i), glomerulus injury (HE staining, j) in mice model; GPX4 protein expression in vitro model or mice model (k or l). Sham, sham control mice; Model, mice model of DN; Med or High, 15 or 30 mg/kg/d of CA; Ferroptosis, 50 mg/kg of Sorafenib Tosylate. Control, control; Model, vitro model; Med or High, 10 and 20 μM of CA; Ferroptosis, 5 μM of CIL56. **P < .01 sham group; ##P < .01 versus Model group; $P < .01. Data were expressed as mean ± SEM.
Figure 5. CA suppressed PAQR3 expression in model of DN to promote PI3K/AKT axle.
PAQR3 mRNA expression and PAQR3/PI3K/AKT protein expression in mice model (a) or in vitro model (c), PAQR3 expression in kidney tissue (Immunohistochemistry, b/Bioluminescent imaging, d), PAQR3 expression in vitro model (immunofluorescence, e). Sham, sham control mice; Model, mice model of DN; Low, Med or High, 7.5 or 15 or 30 mg/kg/d of CA. Control, control; Model, vitro model; Low, Med or High, 5 or 10 and 20 μM of CA. **P < .01 sham group; ##P < .01 versus Model group. Data were expressed as mean ± SEM.
Figure 6. PAQR3 activator reduced the effects of CA on DN in vitro model.
PAQR3/PI3K/AKT/GPX4 protein expression in vitro model (a and b), GSH activity (c), lron concentration (d), Cell viability (e), LDH activity level (f), cell apoptosis (g), proportions of PI positive cells (h), IL-1β/IL-6/INF-γ/TNF-α (i, j, k, l) in vitro model. Control, control; Model, vitro model; Med or High, 10 and 20 μM of CA; PAQR3, PAQR3 up-regulation. **P < .01 sham group; ##P < .01 versus Model group; $P < .01. Data were expressed as mean ± SEM.
Figure 7. CA directly targeted the PAQR3 protein to advance ubiquitination of PAQR3 protein.
3D image revealed that CA bond to the binding pocket and formed with PAQR3 (a); PAQR3 protein expression (b); Microscale thermophoresis (MST) of PAQR3 incubated with CA (c); TSA results in the presence or absence of CA (d); the thermal stability of WT PDPK1 and Mut PAQR3 plasmid after treatment with CA using CETSA (e and f); PAQR3 Ubiquitination (g).
Figure 8. PI3K inhibitor the effects of CA on DN in mice model.
PI3K/AKT/GPX4 protein expression (a and b), glomerulus injury (HE staining, c), Blood glucose (d), Kidney/body weight (e), water intake (f), serum creatinine (g), urea nitrogen (h), urinary albumin levels (i), GSH/TNF-α/IL-1β/IL-6/INF-γ(j, k, l, m, n). Sham, sham control mice; Model, mice model of DN; Med or High, 15 or 30 mg/kg/d of CA; Inhibitor, PI3K inhibitor. **p < .01 sham group; ##P < .01 versus Model group; $P < .01. Data were expressed as mean ± SEM.
Figure 9. CA reduced the interaction between PAQR3 and P110α pathway to induce PI3K/AKT signaling pathway.
LY294002 reversed the effects of CA on p-PI3K and p-AKT protein expression (a), the expression of PAQR3 and p110α protein expressions (b), the combination of PAQR3 and p110α (c), PAQR3 and P110α expression (Immunofluorescence staining, d), the combination of p110α and p85α (e), the expression of PAQR3 and p110α protein (f), the interaction between P110α and P85α protein expressions (g), PAQR3 up-regulation reversed the indcution of p-PI3K and p-AKT (h). **p < .01. Data were expressed as mean ± SEM.
Figure 10. Chicoric acid advanced PAQR3 ubiquitination to ameliorate Ferroptosis in diabetes nephropathy through the relieving of the interaction between PAQR3 and P110α pathway.
