Study of the Protective Effect and Mechanism of Inclisiran on Renal Tissue in a Type 2 Diabetes Mouse Model via the Transforming Growth Factor-β Pathway

Abstract

Objective 

To investigate whether the novel proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor Inclisiran exerts protective effects on the kidneys under high-glucose conditions, and to predict whether its mechanism involves the transforming growth factor-β (TGF-β) pathway using proteomic techniques, while constructing its regulatory network.

Methods 

Healthy male C57BL/6J mice were randomly assigned to four groups: Group A (control, n = 9), Group B (diabetes model, n = 9), Group C (diabetes + low-dose Inclisiran, n = 9), and Group D (diabetes + high-dose Inclisiran, n = 9). Groups B, C, and D were induced with type 2 diabetes using a high-fat diet combined with streptozotocin (STZ). Diabetes was confirmed by three consecutive days of fasting blood glucose levels > 16.7 mmol/L after modeling. The experiment ended 8 weeks after modeling. Renal tissue changes were evaluated using hematoxylin-eosin (HE) and periodic acid-Schiff (PAS) staining. Serum creatinine, low-density lipoprotein (LDL), cholesterol, and PCSK9 levels were measured, along with 24 h urinary protein-to-creatinine ratios. Renal tissue samples from Groups A, B, and D (4 mice per group) underwent transcriptomic sequencing to identify differentially expressed proteins. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were performed to assess the potential of Inclisiran to protect the kidneys via the TGF-β pathway.

Results 

After modeling, blood glucose, urine protein/creatinine ratio, blood creatinine, cholesterol, and other indicators in groups B, C, and D were significantly higher than those in group A, with group B showing the highest values (P < 0.05). In the renal tissues of groups B, C, and D, focal tubular cell degeneration and mesangial proliferation were observed. The glomerular proliferation index in group B was significantly higher than in the other groups. Proteomics identified 1096 differentially expressed proteins (579 upregulated and 517 downregulated) between groups A and B, and 911 differentially expressed proteins (475 upregulated and 436 downregulated) between groups B and D. KEGG enrichment analysis showed that the TGF-β pathway was enriched in both the A-B and B-D group comparisons. There were 11 downregulated differentially expressed proteins (P45481: Crebbp, P70387: Hfe, Q61502: E2f5, Q62312: Tgfbr2, Q62432: Smad2, Q8BSK8: Rps6kb1, Q8BUN5: Smad3, Q8CG19: Ltbp1, Q9CUN6: Smurf1, Q9JKX3: Tfr2, Q9Z1M4: Rps6kb2) related to this pathway between groups B and D.

Conclusion 

Inclisiran may improve the lipid profile of type 2 diabetic mice and reduce the activity of the TGF-β pathway. Its mechanism of action may be related to effects such as extracellular matrix proliferation.

Keywords: Proprotein convertase subtilisin/kexin type 9, Inclisiran, Diabetic kidney disease, Transforming growth factor-β, Proteomics