Macrophages rely on extracellular serine to suppress aberrant cytokine production

Kento Kurita,Hiroshi Arima,Yasuyuki Kitaura,Takayoshi Suganami,Seiichiro Aoe,Ibuki Shirakawa,Hiroya Ohta,Hitoshi Shimano,Yoshihiro Ogawa,Yorihiro Iwasaki,Miyako Tanaka,Ayaka Ito,Takashi Matsuzaka

doi:10.1038/s41598-021-90086-w

Abstract

A growing body of evidence indicates that cellular metabolism is involved in immune cell functions, including cytokine production. Serine is a nutritionally non-essential amino acid that can be generated by de novo synthesis and conversion from glycine. Serine contributes to various cellular responses, but the role in inflammatory responses remains poorly understood. Here, we show that macrophages rely on extracellular serine to suppress aberrant cytokine production. Depleting serine from the culture media reduced the cellular serine content in macrophages markedly, suggesting that macrophages depend largely on extracellular serine rather than cellular synthesis. Under serine deprivation, macrophages stimulated with lipopolysaccharide showed aberrant cytokine expression patterns, including a marked reduction of anti-inflammatory interleukin-10 expression and sustained expression of interleukine-6. Transcriptomic and metabolomics analyses revealed that serine deprivation causes mitochondrial dysfunction: reduction in the pyruvate content, the NADH/NAD+ ratio, the oxygen consumption rate, and the mitochondrial production of reactive oxygen species (ROS). We also found the role of mitochondrial ROS in appropriate cytokine production. Thus, our results indicate that cytokine production in macrophages is tightly regulated by the nutritional microenvironment.

Highlights

Neutral amino acid transporters include ASCT1, ASCT2, and SNAT2. (b) Gene expression of transcription factor and enzymes for serine biosynthesis and neutral amino acid transporters in peripheral blood mononuclear cells (PBMC) of 10 patients infected with rotavirus compared to 8 age-matched healthy controls (GEO dataset: GSE2729). (c) Gene expression of transcription factor and enzymes for serine biosynthesis and neutral amino acid transporters in macrophages from synovial fluids of 5 patients with rheumatoid arthritis compared to PBMC of 3 healthy donors (GEO dataset: GSE10500). (d) Gene expression of Atf[4], Phgdh and Asct[1] in bone marrow-derived macrophages (BMDMs) at before and 4, 8, 12, 24, and 48 h after LPS stimulation
Because serine is considered to be produced mainly within each cell and the role of serine uptake is largely unknown, we first analyzed the expression of representative genes involved in serine metabolism, such as activating transcription factor 4 (ATF4, a transcription factor that promotes serine metabolism genes), phosphoglycerate dehydrogenase (PHGDH, a rate-limiting enzyme involved in serine biosynthesis), phosphoserine phosphatase (PSPH, a key enzyme involved in serine biogenesis), and neutral amino acid transporters, including alanine serine cysteine transporters (ASCT1 and ASCT2), using publicly accessible gene expression omnibus (GEO) datasets
Genes involved in serine metabolism were upregulated by lipopolysaccharide (LPS) stimulation in mouse bone marrow–derived macrophages (BMDMs) as well (Fig. 1d), whereas the gene expression pattern of PHGDH was distinct between the species (Fig. 1b–d), probably due to its complex regulatory mechanism[23]

Summary

Introduction

(b) Gene expression of transcription factor and enzymes for serine biosynthesis and neutral amino acid transporters in peripheral blood mononuclear cells (PBMC) of 10 patients infected with rotavirus compared to 8 age-matched healthy controls (GEO dataset: GSE2729). (c) Gene expression of transcription factor and enzymes for serine biosynthesis and neutral amino acid transporters in macrophages from synovial fluids of 5 patients with rheumatoid arthritis compared to PBMC of 3 healthy donors (GEO dataset: GSE10500). (e,f) Gene expression of Atf[4], Phgdh, and neutral amino acid transporters in BMDMs (e) and peritoneal macrophages (periMΦs) (f). BMDMs or periMΦs were cultured in control (Full) or serine/glycine-depleted (ΔSG) medium for 24 h, followed by stimulation with LPS (100 ng/ ml) for 8 h (n = 3). This study demonstrates that extracellular serine is required for LPS-induced IL10 production, thereby suppressing sustained proinflammatory cytokine expression

Methods

Results

Conclusion