Molecular Cloning, Expression and Macrophage Activation of an Immunoregulatory Protein from Cordyceps militaris.

Qing Yang,Binmei Jia,Xiaomei Liu,Hui Sun,Lin Xu,Zhiyong Gong,Jialing Fang,Luyang Zhao,Min Fang

doi:10.3390/molecules26237107

Qing Yang, Binmei Jia + Show 7 more

Open Access

PDF Available

https://doi.org/10.3390/molecules26237107

Copy DOI

Export

Save

Cite

Abstract
Highlights/Summary
Full-Text PDF
Similar Papers

Abstract

Listen

Protein components of C. militaris have been reported to possess various biological activities. In our previous research, a Cordyceps militaris-derived immunoregulatory protein (CMIP) was naturally isolated and showed the activity of inhibiting the metastasis of breast cancer cells. This study aimed to obtain recombinant CMIP (rCMIP) using recombinant expression and elucidate its ability to activate macrophages. Recombinant CMIP showed one band at approximately 15 kDa or 30 kDa, or two bands at 15 kDa and 30 kDa, under different denaturation conditions of electrophoresis. The cell binding assay showed that rCMIP selectively binds to the surface of macrophages. After adhesion, it did not induce the apoptosis of RAW 264.7 cells, but promoted their proliferation. Moreover, rCMIP significantly induced the expression of M1 macrophage polarization-related molecules. The mean fluorescence intensity (MFI) of CD 86 was enhanced by 2.1-fold and 3.2-fold under 0.64 μM and 1.6 μM of rCMIP treatment, respectively. Cytokines typically expressed in M1 macrophages, such as TNF-α, iNOS, IL-6, CCL 4, CCL 5 and CXCL 10, were also considerably induced by rCMIP, while the expression of cytokines in typical M2 macrophages, like Arg-1, CCL17 and CCL22, were not changed or slightly decreased. Under rCMIP treatment, the release of NO was also appreciably induced. In the present study, we reported cloning, expression and functional characterization of rCMIP, which was naturally isolated from the fruiting body of C. militaris in our previous study. The data imply that rCMIP possesses immunomodulatory activity in macrophages.

Highlights

Mushrooms are considered nutritional foods and a precious source of health-promoting agents, especially in Asian countries [1]
We found that the mRNA expressions of the typical cytokines of M1 macrophages, including chemokines (CCL 2, CCL 4, CCL 5, CXCL 10 and CXCL 11), TNF-α, IL-6, IL-1β and iNOS were all remarkably up-regulated in recombinant CMIP (rCMIP)-treated RAW 264.7 cells
C. militaris is renowned as a functional food for its immuno-regulatory, anti-tumor and nutritional benefits [22]

Summary

Introduction

Mushrooms are considered nutritional foods and a precious source of health-promoting agents, especially in Asian countries [1]. One of the most important active ingredients in mushrooms, have been extensively studied for their pharmacological activities, including anti-tumor, antioxidant and immuno-regulatory activities [2,3,4]. Cordyceps militaris (C. militaris) is one of the most famous functional mushrooms as it contains multiple active ingredients, like polysaccharides, mannitol and proteins [7,8,9,10]. Various pharmacological activities, such as anti-cancer, anti-inflammatory and immunoregulatory activities, of Cordyceps militaris have been reported extensively [11]. Many of these pharmacological activities are attributed to the protein components from C. militaris [12]

Objectives

Results

Conclusion