Macrophages are found in almost all tissues such as Kupffer cells in hepatocyte (1) and microglia in central nervous system (2). These macrophages engulf cellular debris, microbes, death cells and foreign substances by stretching filopodia (3, 4). Although the polarizations of macrophages are multiple, they are roughly polarized to two distinct subsets: classically activated (M1) phenotype and alternatively activated (M2) phenotype (5, 6). Macrophages polarize into M1 phenotype to perform their pathogen-scavenging function when exposed to T-helper 1 (Th 1) type cytokines or inflammatory mediators, such as interferon gamma (IFN-γ) and lipopolysaccharide (LPS) (7), or M2 phenotype to perform their anti-inflammatory effects, including wound healing and anti-tumor ability under conditions of exposure to Th 2 cytokines like IL-4 and IL-10 (8). Indeed, various contributors are related to the fate of macrophages. Notably, metabolism pathways and metabolites are the best examples for directing macrophage growth and survival by providing energy and substrates, and instructing functions of macrophages (9, 10). For example, altered amino acid metabolism [e.g., arginine metabolism (11)] is a well-accepted character to define macrophage polarization.

Traditionally, amino acids are simply divided into two categories: essential amino acids and non-essential amino acids (12). However, many traditionally considered non-essential amino acids are not only used as substrates for protein and peptide synthesis, but also involved in regulating metabolism, signal transduction and immune responses (13). Glycine consists of one carbon (C) atom, two hydrogen (H) atom, one carboxyl-group (COOH) and one amino-group (NH₂) (14). Of note, recent studies have shown that glycine affects functions of macrophage (15, 16). In this review, we will summarize glycinergic system in macrophages, discuss how glycine contributes to the polarization of macrophages, and list some examples that glycine mediates macrophage-associated diseases.

In mammals, glycine can be synthesized from serine, choline, threonine and hydroxyproline by different metabolic pathways (32). Since serine and glycine are biosynthetically linked (33), serine and its precursors can generate glycine. The conversion of serine to glycine catalyzed by serine hydroxymethyltransferase (SHMT) is the main way for glycine synthesis (34, 35). When glycine deficiency occurs, such as intrinsic glycine uptake capacity limitation or environmental glycine deprivation, SHMT can support glycine synthesis (36).

In addition to participating in protein synthesis, glycine is a precursor of peptides, nucleic acids as well as methyl donors. Upon LPS stimulation, the levels of intracellular glycine and glycine metabolites such as glutathione (GSH) and S-adenosylmethionine (SAM) increased (37–39). Interestingly, adding glycine to the serine-deprived medium failed to rescue IL-1β secretion in macrophages upon LPS stimulation (38). Besides this, lack of glycine cannot affect the polarization of macrophages (39). Thus, extracellular glycine may not influence macrophage metabolism. U-[13C]-labeling shows that glycine is mainly converted from glucose and serine, and it can be subsequently converted to ADP, ATP, GSH and SAM (38). Strikingly, U-[13C]-glycine revealed a remarkable attenuation of extracellular glycine-derived GSH compared to serine (synthesis from glycine)-derived GSH (38). Moreover, supplementary glycine in serine deprived medium failed to rescue intracellular GSH in macrophage. These phenomena indicate that glycine utilization in macrophages is mainly through intracellular conversion of serine, not via exogenous glycine supply.

The functions of macrophages are highly responsive to their micro-environmental stimuli. Upon the activation of Toll-like receptor (TLR) or interferon signaling, M1 macrophages arise in inflammatory to eliminate pathogens (40–42). Whereas M2 macrophages, usually found in Th2-dominated responses, can mediate helminth immunity, asthma, and allergy (43).

Among various signaling pathways regulating macrophage inflammation, NF-κB is a main contributor to orchestrate macrophage polarization (44). Glycine can prevent the activation of nuclear factor-κB (NF-κB) by inhibiting the degradation of inhibitor of NF-κB (IκB) in pro-inflammatory macrophages ( Figure 1A ) (45). Additionally, glycine affects inflammasome assembly in pro-inflammatory macrophages (46). However, given glycine treatment could induce IκB degradation in resting macrophages (45), we still could not exclude the possibility that glycine causes stress responses in resting macrophages. In addition, in the context of glycine treatment, the decreased phosphorylation of IκB kinase-α (IKK-α) and IκB kinase-β (IKK-β) is also observed (45, 46) ( Figure 1B ). Glycine reduces LPS-induced upregulation of nucleotide binding domain like receptor protein 3 (NLRP3) (47). This process can be achieved by up-regulating the expression of NRF2 and its down-stream signaling pathways to eliminate reactive oxygen species (ROS) (47) ( Figure 1C ).

PI3K (phosphatidylinositol 3-kinase) and Akt (protein kinase B) pathways regulate tremendous signaling pathways, including NF-κB and mitogen-activated protein kinase (MAPK) signaling (48) related to macrophage polarization (49). Glycine can up-regulate Akt by blocking phosphatase and tensin homolog deleted on chromosome ten (PTEN), then inhibit NF-κB and hypoxia induced factor-1 α (HIF1-α) in microglia (50) in the context of ischemia-reperfusion injury. Except for macrophages, glycine also inhibits PTEN and activates Akt in other tissues or cells (51, 52) ( Figure 1D ). Unfortunately, there is still no direct evidence showing whether glycine can affect proinflammatory macrophage polarization induced by canonical stimuli (e.g., LPS and/or IFN-γ) through PTEN-Akt pathway. Notably, Akt kinases have distinct effects in macrophage polarization, with Akt1 ablation leading to an M1 phenotype and Akt2 ablation resulting in an M2 phenotype (53). It has not been studied which subunit of Akt is regulated by glycine. Therefore, it is necessary to further explore the connection between glycine and the Akt signaling pathway in guiding macrophages polarization.

MicroRNAs (miRNAs) play vital roles in a great deal of biological processes (54) and could function as crucial regulators that support macrophage polarization (54, 55). It has been reported that some miRNAs which associated with macrophages are related with glycine. For example, glycine alleviates subarachnoid-hemorrhage (SAH) induced neuron inflammation, which is mediated by miRNA-26b/PTEN/Akt signaling pathway in microglia (56) ( Figure 2A ). Inhibition of miRNA-26b or activation of PTEN expression suppressed the protective function of glycine (56). MiR-301a is abundantly expressed in hypoxic pancreatic cancer cell-derived exosomes (57, 58), which can promote M2 macrophage polarization through activating PTEN/PI3K signaling pathway (57). Interestingly, glycine has been reported to enhance the expression of miR-301a in the cortical neurons (59). Thus, miR-301a might be a potential target for glycine to regulate M2 macrophage functions ( Figure 2B ).

MiR-19a-3p can suppress LPS/IFN-γ-induced M1 macrophage polarization via inhibiting STAT1 (signal transducer and activator of transcription-1) (60). In addition, glycine regulates miR-19a-3p/AMPK pathway to alleviate ischemic stroke injury (61). Therefore, glycine may promote M1 macrophage polarization by regulating miR-19a-3p ( Figure 2C ). Besides influencing M1 macrophages polarization, miR-19a-3p is capable of suppressing M2 macrophage polarization by inhibiting STAT3 when overexpressed (62) ( Figure 2D ).

Notably, miRNAs can regulate GlyTs function. Human GlyT1 possesses several miRNAs targeting sites within the 3’UTR (miR-7, miR-30, miR-96, miR-137, miR-141). Among them, miR-96 and miR-137 negatively regulate GlyT1 under physiological conditions (63) ( Figure 2E ). It is intriguing to investigate whether microRNAs mediate the regulation of glycinergic system in macrophage polarization.

In this review, we introduced glycinergic system in macrophages, and summarized how glycine shapes macrophages polarization. For glycinergic system, GlyRs could be found in macrophages, and the subunits of GlyRs are varied in macrophages with different origins. Though it has been already noted that NAATs exist in macrophages, it is not clear which type of NAATs is expressed in macrophages. Glycine is supposed to affect macrophage through different contributors. Mechanistically, glycine alters macrophage signaling pathways (e.g., NF-κB, NRF2, and Akt) and miRNAs. Interestingly, other signaling pathways [e.g., ERK (109)] might also mediate the functions of glycine. Therefore, it is not surprising that glycine could influence the progresses of several macrophage-associated diseases (e.g., colitis and NAFLD).

Indeed, the influences of glycine in macrophage activation are still worth further investigation. Firstly, it is not clear whether glycine can affect methylation reaction in macrophages. In one-carbon metabolism, glycine partly provides the carbon backbones required for the generation of SAM (110), which is the main methyl donor for cellular methylation reaction (39, 111). Recent studies have shown that the methylation of histone (39), DNA (112) or mRNA (113, 114) is closely related to macrophage polarization. Therefore, glycine is likely to affect macrophage polarization through methylation modification. Secondly, there are few studies on the effect of glycine on the metabolism of macrophages. Macrophage metabolism is highly related with the function output of macrophages (54). Considering glycine could impact HIF-1α and mTORC1 that are related to cellular metabolism (e.g., glycolysis), thus studying the effect of glycine on macrophage metabolism is meaningful to reveal the working mechanism of glycine on macrophages function. Finally, studying the effect of glycine on macrophages in the tumor microenvironment may reveal a potential target for cancer therapy. Therefore, it is necessary to find out the relationship between glycine, macrophage function and cancer progression.

ZG wrote the review article. ZG, SH, and XW revised the review article. CH, JF, and LF helped with designing figures and finding relevant literatures. MZ and DX reviewed and revised the grammar error in the manuscript. All authors contributed to the article and approved the submitted version.

This study was supported by the Guangdong Basic and Applied Basic Research Foundation (2019B1515210002) and National Natural Science Foundation of China (31922079).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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