*3.3.2 Epigenetic control of MDSC plasticity*

*Cells of the Immune System*

**116**

**Figure 1.**

*KLF4 regulates FSP-1 gene expression in fibrocyte generation. (A) Representative photographs of* 

*transfection and dual luciferase assays, \*P < 0.05, \*\*P < 0.01.*

*morphological fibrocyte generation from splenocytes in the absence and presence are indicated by red arrows. KLF4 deficiency was induced by 4-OH tamoxifen (TAM). (B) Quantification of the data from (A). (C) Relative levels of KLF4 and FSP-1 mRNA in fibrocyte generation as assessed by qRT-PCR. (D) Different MDSC subsets in mouse splenocytes measured by flow cytometry. (E) Relative levels of KLF4, FSP-1 and CCR2 mRNA in different MDSC subsets by qRT-PCR. (F) Potential of fibrocyte generation from MDSC subsets in mouse spleen. (G) Left—binding of KLF4 to the FSP-1 promoter as assessed by chromatin immunoprecipitation assay using two KLF4 antibodies (KLF4-1 and KLF4-2). IgG was used as a negative control. Right—the effect of KLF4 overexpression on FSP-1 promoter activities, as examined by transient* 

The studies of epigenetics, heritable changes to gene expression without changes to DNA, are significantly advancing our knowledge of the inflammatory conditions [86]. They include DNA modifications mainly methylation, histone tail modifications, and non-coding RNA-mediated gene regulation. Recent data revealed that epigenetic mechanisms could provide novel strategies for modulating wound healing [87–89].

Critical functions of KLF4 have been shown in the generation of induced pluripotent stem cells and in cancer development through epigenetic mechanisms [90, 91]. In addition, there are numerous reports showing that microRNAs regulate KLF4 [92– 94] or KLF4 regulate microRNAs [95, 96] in varied pathological conditions. KLF4 mediated DNA methylation have also been reported in hTert promoter [97] and methylation of KLF4 promoter is associated with urothelial cancer progression and early recurrence [98]. Moreover, the correlation of KLF4 and histone modifications has also been reported. For example, histone methyltransferase KMT2D, a frequently aberrant epigenetic modifier in various cancer, sustains prostate carcinogenesis and metastasis via epigenetically activating KLF4 [99]. From the perspective of MDSCs, epigenetic regulation of their differentiation and function is not completely understood. However, there is evidence to indicate the importance of epigenetic regulation. Shang et al. showed that long non-coding RNA retinal non-coding RNA3 (RNCR3) promotes C/EBP homologous protein (Chop) expression by sponging microRNA 185- 5p during MDSC differentiation [100]. In addition, although histone modifications related to myeloid differentiation have been extensively studied [101], currently there is no clear indication about epigenetic markers that can discriminate specific MDSC subsets. Given the role of KLF4 in epigenetic regulation and the importance of MDSC plasticity in cancer and wound healing, it will be very interesting to examine how KLF4 is involved in epigenetic control of MDSC subsets or plasticity.
