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Revascularization is an effective therapy for rescuing myocardial tissue after ischemic events. However, the process of reperfusion can lead to more severe cardiomyocyte damage, called myocardial ischemia-reperfusion (I/R) injury (MIRI). We have previously shown that vitexin (VT) (a flavonoid compound derived from natural products) protects against MIRI; however, the exact mechanisms underpinning this effect require further elucidation. This study is aimed at elucidating the protective mechanism of VT in inhibiting ischemic myocardial mitochondrial dysfunction and reducing cardiomyocyte apoptosis by regulating Epac1-Rap1 signaling. Isolated rat hearts were subjected to MIRI in a Langendorff perfusion system, and H9c2 cells were subjected to hypoxia/reoxygenation (H/R) in vitro. Our analyses show that during I/R, Epac1 expression was upregulated, left ventricular dysfunction deteriorated, mitochondrial dynamics were disrupted, and both myocardial cells and tissues exhibited apoptosis. Furthermore, administration of 8-CPT (an Epac agonist) exacerbated cardiomyocyte injury and mitochondrial dysfunction. Interestingly, suppressing the function of Epac1 through VT or ESI-09 (an Epac inhibitor) treatment during I/R reduced the myocardial infarct size, cardiomyocyte apoptosis, and reactive oxygen species production; alleviated mitochondrial dysfunction by increasing mitochondrial membrane potential; elevated MFN2 expression; and inhibited Drp1 expression. To our knowledge, our results reveal, for the first time, the mechanisms underlying the protective effect of VT in the myocardium of rats with MIRI. Moreover, we provide a new target and theoretical basis for VT in the treatment of ischemic heart disease.

T cell hyporesponsiveness is crucial for functional immune system and prevent the damage induced by alloreactive T cells in autoimmune pathology and transplantation. As one of the most important regulators during B cell development, PRDM1 (also known as BLIMP-1) has been demonstrated its essential role for maintaining T cell hyporesponsiveness and homeostasis, evidenced by Prdm1 deficient mice accumulating activated T cells and developing multiorgan inflammatory disease. However, the mechanism of PRDM1 regulating T cell hyporesponsiveness is still ambiguous. In this study, we took advantage of multiomics technologies and systemic report the central role of PRDM1 in inducing human primary T cell hyporesponsiveness. Firstly, we overexpressed PRDM1 in human primary T cells and found increased ratio of CD4 +CD25 +FOXP3 + Treg cell subsets and increased IL-4 secretion. In parallel, inhibited PRDM1 expression level in human T cells decreased ratio of Treg cells and secretion of IL-4. Meanwhile, transcriptome analyses revealed that overexpressed PRDM1 enriched negative regulation of cell proliferation signaling pathway and resulted in a global reduction in IL-2 and inflammatory response signaling pathways. Furthermore, overexpressed PRDM1 in primary T cells upregulated several negative regulators of T cell function like EOMES, KLF2, LILRB1, KLRB1 and CD244, indicating a pioneer role of PRDM1 in inducing T cell hyporesponsive. To further investigate the regulation role of T cell hyporesponsiveness of PRDM1, we performed CUT&Tag and ATAC-seq in PRDM1 overexpressed primary T cells. CUT&Tag analysis showed PRDM1 could directly upregulated T cell hyporesponsiveness related gene such as KLF2, CD244 and KLRD1. Importantly, we observed consistent changes of IL-2, central regulator of T cell activation, in PRDM1 overexpressed T cell from ATAC-seq, CUT&Tag and RNA-seq data. We found PRDM1 could binding to IL-2 locus and decreased the chromatin accessibility of IL-2, consequently downregulated the expression level of IL-2 in human primary T cells. Moreover, altered open chromatin regions (OCRs) in PRDM1 overexpressed T cells enriched the similar transcription factors (TFs) with PRDM1 binding sites, indicating PRDM1 might be a pioneer TF in T cell hyporesponsiveness. These results demonstrated PRDM1 is sufficient for inducing T cell hyporesponsiveness in human primary T cells. To further validate the coexpression relationship between PRDM1 and Treg cell central TF FOXP3, we upregulated PRDM1 expression level on Jurkat T cells lines. The results also showed that elevated FOXP3 both in mRNA and protein level accompanied with upregulated PRDM1 expression level. To analyze the mechanism of PRDM1 regulating FOXP3 expression level, CUT&Tag data analyses showed that PRDM1 might upregulated FOXP3 by directly binding to the enhancer region of upstream of FOXP3 locus. Meanwhile, PRDM1 indirectly upregulated FOXP3 by upregulated KLF2, evidenced by inhibiting KLF2 in PRDM1 overexpressed primary T cells downregulated FOXP3 expression level. To further investigate the clinical implication of PRDM1 inducing T cell hyporesponsiveness, we detected the relationship of PRDM1 expression level and acute graft-versus-host disease(aGVHD) occurrence after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Our results showed that patients with aGVHD (n=7) exhibited lower PRDM1 expression level than those without aGVHD in the same period after HSCT (n=7). Furthermore, we detected the expression level of PRDM1 in CD4 + T cell and CD8 + T cells from bone marrow allografts (BM) or peripheral blood allografts (PB) and followed up the occurrence of GVHD after HSCT for 2 years (n=18). There are low expression levels of PRDM1 in CD4 + T cells both from BM or PB grafts corelated with aGVHD occurrence in patients after allo-HSCT compared with those without aGVHD occurrence. In conclusion, our study provides the global regulatory model of PRDM1 in human primary T cell. We introduced PRDM1 as a sufficient regulator in T cell hyporesponsiveness induction, which is altering the chromatin accessibility and directly upregulated T cell inhibitory signals and downregulated T cell activated signals. The negative relationship between PRDM1 expression level with GVHD occurrence indicated it might be a potential biomarker for indicating HSCT prognosis. Disclosures No relevant conflicts of interest to declare.

The aim of this study was to investigate the prevention of diet-induced obesity by a high safflower oil diet and adipocytic gene expression in mice. Forty 3-week-old C57BL/6 mice were randomly divided into three groups: control group (CON, 5% lard 1 5% safflower oil), high lard group (LAR, 45% lard 1 5% safflower oil), and high safflower oil group (SAF, 45% safflower oil 1 5% lard). After 10 weeks, 10 mice of the LAR group were switched to high safflower oil diet (LAR–SAF). Ten weeks later, glucose tolerance tests were performed by intraperitoneal injection of glucose. Circulating levels of lipid and insulin were measured and white adipose tissues were taken for gene chip and reverse transcriptase–polymerase chain reaction analysis. The LAR group showed higher body weight, adiposity index, insulin, and lipids than the CON group (P < 0.05). The body weight in the LAR–SAF group decreased after dietary reversal. The plasma biochemical profiles decreased in the LAR–SAF and SAF groups (P < 0.05) compared with those of the LAR group. The blood glucose level of the LAR–SAF group was reduced during intraperitoneal glucose tolerance test compared with that of the LAR group. The LAR–SAF group had lower levels of Orexin and Ghrelin gene expression, whereas the level of PPARa gene expression was significantly enhanced compared with that of the LAR group. So, the SAF diet can alter adipocytic adiposity-related gene expression and result in effective amelioration of diet-induced obesity.