The ability to suppress macrophage-mediated inflammation in orbital fat stem cells is controlled by miR-671-5p

Lien, Gi-Shih; Liu, Jen-Fang; Chien, Ming-Hsien; Hsu, Wei-Tse; Chang, Tzu-Hao; Ku, Chia-Chi; Ji, Andrea Tung-Qian; Tan, Peng; Hsieh, Ting-Lieh; Lee, Liang-Ming; Ho, Jennifer H

TGFβ, IL-10, IDO and IL-1RA are known as MSC-secreted factors in regulating innate immunity response to procytokines [12, 14, 15, 21] . Our previous in vivo study demonstrated that systemic OFSC transplantation enhanced the serum level of sTNFR type II in mice with LPS-induced ALI [23] . In the present study, OFSCs did express these immunomodulating factors when co-cultured with naïve macrophages (Figure 4) . Furthermore, gene expression of IL-10 ( Figure 4B ), IDO ( Figure 4C ), sTNFR type II ( Figure 4D ) and IL-1RA ( Figure 4E ) in OFSCs were highly upregulated by LPS-activated macrophages within 6 hours of noncontact culture. TGFβ was not affected upon coculture ( Figure 4A ). silencing mainly through miRNAs, deep-sequencing analysis and miRTar prediction were performed to identify the potential endogenous miRNAs in OFSCs, MSCs derived from subcutaneous tissue as well as MSCs derived from bone marrow targeting on inducible, secreted immunomodulating factors reported in MSCs (that is, IL-6, IL-10, IDO, hepatocyte growth factor, TGFβ, sTNFR type I, sTNFR type II, and IL-1RA). Seven miRNAs were found to potentially regulate the above genes by target prediction ( Table 2 ). For OFSCs, the expression levels of hsa-miR-28-5p, hsa-miR-503 and hsa-miR-769-5p (transcripts per million < 1,000) were too low to act as a regulator for immunomodulation. hsa-let-7c, hsa-miR-370, and hsa-miR-423-5p were strongly expressed in OFSCs (transcripts per million > 2,000) but each of them targeted only one gene, making them less possible as the key regulator in this study. Hsa-miR-671-5p had a strong expression level (transcripts per million = 5,140) in OFSCs and potentially regulated those genes upregulated in LPS-activated macrophages, including IL-10, sTNFR type II and IL-1RA (Figure 4) , indicating that miR-671-5p may be the key miRNA in OFSCs regulating inducible immunomodulating factors under procytokine stimulation. miR-671-5p regulates OFSC immunomodulation ability by targeting sTNFR type II and IL-1RA