Capmatinib suppresses LPS-induced interaction between HUVECs and THP-1 monocytes through suppression of inflammatory responses

Abstract

Background: Capmatinib (CAP) is a drug that has been used to treat non-small cell lung cancer (NSCLC) in adults. Presently, its novel effects on skeletal muscle insulin signaling, inflammation, and lipogenesis in adipocytes have been uncovered with a perspective of drug repositioning. However, the impact of CAP on LPS-mediated interaction between human umbilical vein endothelial cells (HUVECs) and THP-1 monocytes has yet to be investigated.

Methods: HUVECs and THP-1 monocytes were treated with LPS and CAP. The protein expression levels were determined using Western blotting. Target protein knockdown was conducted using small interfering (si) RNA transfection. Interactions between HUVECs and THP-1 cells were assayed using green fluorescent dye.

Results: This study found that CAP treatment ameliorated cell adhesion between THP-1 monocytes and HUVECs and the expression of adhesive molecules, such as intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin. Moreover, phosphorylation of inflammatory markers, such as NFκB and IκB as well as TNFα and monocyte chemoattractant protein-1 (MCP-1) released from HUVECs and THP-1 monocytes, was prevented by CAP treatment. Treatment with CAP augmented PPARδ and IL-10 expression. siRNA-associated suppression of PPARδ and IL-10 abolished the effects of CAP on cell interaction between HUVECs and THP-1 cells and inflammatory responses. Further, PPARδ siRNA mitigated CAP-mediated induction of IL-10 expression.

Conclusion: These findings imply that CAP improves inflamed endothelial-monocyte adhesion via a PPARδ/IL-10-dependent pathway. The current study provides in vitro evidence for a therapeutic approach for treating atherosclerosis.

Keywords: Capmatinib; HUVEC; IL-10; Inflammation; PPAR delta; THP-1.

Graphical abstract

Fig. 1

CAP reduces monocyte adhesion on endothelial cells as well as adhesion molecule expression in endothelial cells. (A) MTT assay in HUVECs and THP-1 cells treated with 0–10 nM CAP for 24 h. THP-1 cell adhesion assay (B) and Western blotting of ICAM-1, VCAM-1, and E-selectin (C) in HUVECs treated with LPS (200 ng/mL) and 0–5 nM CAP for 24 h ∗∗∗p < 0.001 when compared to control. ^!!!p < 0.001, ^!!p < 0.01, and ^!p < 0.05 when compared to LPS.

Fig. 2

CAP ameliorates inflammatory responses in LPS-treated HUVECs and THP-1 monocytes. (A) Western blot analysis of phosphorylated NFκB and IκB in HUVECs and THP-1 cells treated with LPS (200 ng/mL) and 0–5 nM CAP for 24 h. (B) ELISA of TNF and MCP-1 in culture media of HUVECs and THP-1 cells treated with LPS (200 ng/mL) and 0–5 nM CAP for 24 h ∗∗∗p < 0.001 when compared to control. ^!!!p < 0.001, ^!!p < 0.01, and ^!p < 0.05 when compared to LPS.

Fig. 3

CAP attenuates the adhesion between HUVECs and THP-1 monocytes as well as inflammation through PPARδ-dependent signaling. (A) Western blot analysis of PPARδ in HUVECs and THP-1 cells treated with 0–5 nM CAP for 24 h. THP-1 cell adhesion assay (B) and Western blotting of ICAM-1, VCAM-1, and E-selectin (C) in scrambled or PPARδ siRNA (20 nM)-transfected HUVECs treated with LPS (200 ng/mL) and CAP (5 nM) for 24 h. (D) Western blot analysis of phosphorylated NFκB and IκB in scrambled or PPARδ siRNA (20 nM)-transfected HUVECs and THP-1 cells treated with LPS (200 ng/mL) and CAP (5 nM) for 24 h. (E) ELISA of TNF and MCP-1 in culture media of scrambled or PPARδ siRNA (20 nM)-transfected HUVECs and THP-1 cells treated with LPS (200 ng/mL) and CAP (5 nM) for 24 h ∗∗∗p < 0.001 and ∗∗p < 0.01 when compared to control. ^!!!p < 0.001, ^!!p < 0.01, and ^!p < 0.05 when compared to LPS. ^###p < 0.001, ^##p < 0.01, and ^#p < 0.05 when compared to LPS plus CAP.

Fig. 4

PPARδ/IL-10 pathway contributes to the suppressive effects of CAP on inflammation and adhesion between HUVECs and THP-1 cells. (A) Western blot analysis of IL-10 in HUVECs and THP-1 cells treated with 0–5 nM CAP for 24 h. THP-1 cell adhesion assay (B) and Western blotting of ICAM-1, VCAM-1, and E-selectin (C) in scrambled or IL-10 siRNA (20 nM)-transfected HUVECs treated with LPS (200 ng/mL) and CAP (5 nM) for 24 h. (D) Western blot analysis of phosphorylated NFκB and IκB in scrambled or IL-10 siRNA (20 nM)-transfected HUVECs and THP-1 cells treated with LPS (200 ng/mL) and CAP (5 nM) for 24 h. (E) ELISA of TNFα and MCP-1 in culture media of scrambled or IL-10 siRNA (20 nM)-transfected HUVECs and THP-1 cells treated with LPS (200 ng/mL) and CAP (5 nM) for 24 h. (F) Western blotting of IL-10 in scrambled or PPARδ siRNA (20 nM)-transfected HUVECs and THP-1 cells treated with CAP (5 nM) for 24 h. Western blotting of PPARδ in scrambled or IL-10 siRNA (20 nM)-transfected HUVECs and THP-1 cells treated with CAP (5 nM) for 24 h ∗∗∗p < 0.001 and ∗∗p < 0.01 when compared to control. ^!!!p < 0.001, ^!!p < 0.01, and ^!p < 0.05 when compared to LPS or CAP. ^###p < 0.001, ^##p < 0.01, and ^#p < 0.05 when compared to LPS plus CAP.

Fig. 5

Schematic diagram of the effects of CAP on the adhesion between endothelial cells and monocytes, and its related pathways.

Suppl. Fig. 1

The cell viability assay in HUVECs and THP-1 monocytes treated with LPS. MTT assay in HUVECs and THP-1 cells treated with 0–500 ng/mL LPS for 24 h ∗∗∗P < 0.001 and ∗∗P < 0.01 when compared to control.