Apelin as a new therapeutic target for COVID-19 treatment

Abstract

Background: Apelin is an endogenous neuropeptide that binds to the G-protein-coupled receptor (APJ) and participates in a variety of physiological processes in the heart, lungs and other peripheral organs. Intriguingly, [Pyr1]-Apelin-13, a highly potent pyroglutamic form of apelin, has the potential to bind to and be degraded by angiotensin-converting enzyme 2 (ACE2). ACE2 is known to operate as a viral receptor in the early stages of severe acute respiratory coronavirus (SARS-CoV-2) infection.

Aim: This study aimed to determine if apelin protects against SARS-CoV-2 infection by inhibiting ACE2 binding to SARS-CoV-2 spike protein.

Design and methods: To determine whether [Pyr1]-Apelin-13 inhibits ACE2 binding to the SARS-CoV-2 spike protein (S protein), we performed a cell-to-cell fusion assay using ACE2-expressing cells and S protein-expressing cells and a pseudovirus-based inhibition assay. We then analyzed publicly available transcriptome data while focusing on the beneficial effects of apelin on the lungs.

Results: We found that [Pyr1]-Apelin-13 inhibits cell-to-cell fusion mediated by ACE2 binding to the S protein. In this experiment, [Pyr1]-Apelin-13 protected human bronchial epithelial cells, infected with pseudo-typed lentivirus-producing S protein, against viral infection. In the presence of [Pyr1]-Apelin-13, the level of viral spike protein expression was also reduced in a concentration-dependent manner. Transcriptome analysis revealed that apelin may control inflammatory responses to viral infection by inhibiting the nuclear factor kappa B pathway.

Conclusion: Apelin is a potential therapeutic candidate against SARS-CoV-2 infection.

Figure 1.

Cell-to-cell fusion assay. (A) Apelin-17 and apelin-13 effectively inhibited the binding of SARS-CoV-2 spike protein to angiotensin-converting enzyme 2 (ACE2) in human embryonic kidney 293 cells. (B) Apelin-13 effectively reduced the binding of SARS-CoV-2 spike protein to ACE2 in both human lung cell lines (A549, human lung cancer cells; 16HBE14o-, human bronchial epithelial cells). Apelin-13; [Pyr1]-Apelin-13. *P < 0.05; **P < 0.01.

Figure 2.

Co-localization of angiotensin-converting enzyme 2 and apelin. Representative images of co-localization of apelin and angiotensin-converting enzyme 2 in human cell human embryonic kidney 293 (HEK293) cells (A) and human bronchial epithelial cells (B). Apelin (red), GFP (green) and 4',6-diamidino-2-phenylindole (blue) signals can be seen in the images. The overlap of red and green displays as yellow (arrow). (C) Pseudovirus inhibition assay using lentivirus containing SARS-CoV-2 spike protein. [Pyr1]-Apelin-13 significantly suppressed SARS-CoV-2 entry into HEK293 cells.

Figure 3.

Profiling of SARS-CoV-2 infected human lung transcriptomes. (A) Apelin (APLN) expression in human lungs infected with SARS-CoV-2. Low viral: low SARS-CoV-2 virus and high viral: high SARS-CoV-2 virus content groups can be seen. (B) Volcano plot using differentially expressed genes (DEGs) between high APLN and low APLN expression groups. (C) Heatmap for the enriched pathway analysis using DEGs. (D) Heatmap for enriched gene-related extracellular matrix organization. *P < 0.05.

Figure 4.

Transcriptome analysis of apelin-null endothelial cells. (A) Volcano plot using differentially expressed genes (DEGs) between apelin knock out (KO) and wild type (WT) mouse endothelial transcriptome. (B) Heatmap for the enriched pathway analysis using DEGs. (C) Enrichment plot of nuclear factor kappa B (NF-κB) signaling. (D) Overview of proposed beneficial roles of apelin in the lungs in SARS-CoV-2 pathogenesis.