Abstract
Alpha-momorcharin (α-MMC), a member of the ribosome-inactivating protein (RIP) family, has been found in the seeds of Momordica charantia (bitter melon). α-MMC contributes a number of pharmacological activities; however, its inflammatory properties have not been well studied. Here, we aim to determine the inflammatory responses induced by recombinant α-MMC and identify the underlying mechanisms using cell culture and animal models. Recombinant α-MMC was generated in Rosetta™(DE3)pLysS and purified by the way of nitrilotriacetic acid (NTA) chromatography. Treatment of recombinant α-MMC at 40 μg/mL exerted sub-lethal cytotoxic effect on THP-1 monocytic cells. Transcriptional profiling revealed that various genes coding for cytokines and other proinflammatory proteins were upregulated upon recombinant α-MMC treatment in THP-1 cells, including MCP-1, IL-8, IL-1β, and TNF-α. Recombinant α-MMC was shown to activate IKK/NF-κB and JNK pathways and the α-MMC-induced inflammatory gene expression could be blocked by IKKβ and JNK inhibitors. Furthermore, murine inflammatory models further demonstrated that α-MMC induced inflammatory responses in vivo. We conclude that α-MMC stimulates inflammatory responses in human monocytes by activating of IKK/NF-κB and JNK pathways, raising the possibility that consumption of α-MMC-containing food may lead to inflammatory-related diseases.
Highlights
Ribosome-inactivating proteins (RIPs) are highly potent protein toxins which restrain protein synthesis by directly targeting the ribosomes [1]
Toxins 2019, 11, 694 types: (1) a polypeptide chain of about 30 kDa (Type I) and (2) a more toxic heterodimer composed of an A chain which has identical function to the Type I RIP, attached with a B subunit that can bind to lectin (Type II) [1]
The isolation of recombinant His-tagged α-MMC protein was achieved by Ni-nitrilotriacetic acid (NTA) affinity chromatography and the purity was shown in 12% SDS-PAGE electrophoresis (Figure 1A)
Summary
Ribosome-inactivating proteins (RIPs) are highly potent protein toxins which restrain protein synthesis by directly targeting the ribosomes [1]. Plant-derived RIPs can be classified into two main. Toxins 2019, 11, 694 types: (1) a polypeptide chain of about 30 kDa (Type I) and (2) a more toxic heterodimer composed of an A chain which has identical function to the Type I RIP, attached with a B subunit that can bind to lectin (Type II) [1]. RIPs gain biologists’ attention in the biomedical research area because of their diverse activities toward plant and animal cells, including their anticancer, antiviral, antifungal, abortifacient, and antiparasitic effects [2,3,4,5]. RIP-related biotechnological research is focused on obtaining a deeper understanding of the mechanism of cell entry and finding ways to increase specificity, reduce antigenicity, and prolong plasma half-life [1]
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