Six2 Is a Coordinator of LiCl-Induced Cell Proliferation and Apoptosis.

The metanephric mesenchyme (MM) cells are a subset of kidney progenitor cells and play an essential role in mesenchymal-epithelial transition (MET), the key step of nephron generation. Six2, a biological marker related to Wnt signaling pathway, promotes the proliferation, inhibits the apoptosis and maintains the un-differentiation of MM cells. Besides, LiCl is an activator of Wnt signaling pathway. However, the role of LiCl in cellular regulation of MM cells remains unclear, and the relationship between LiCl and Six2 in this process is also little known. Here, we performed EdU assay and flow cytometry assay to, respectively, detect the proliferation and apoptosis of MM cells treated with LiCl of increasing dosages. In addition, reverse transcription-PCR (RT-PCR) and Western-blot were conducted to measure the expression of Six2 and some maker genes of Wnt and bone-morphogenetic-protein (BMP) signaling pathway. Furthermore, luciferase assay was also carried out to detect the transcriptional regulation of Six2. Then we found LiCl promoted MM cell proliferation at low-concentration (10, 20, 30, and 40 mM). The expression of Six2 was dose-dependently increased in low-concentration (10, 20, 30, and 40 mM) at both mRNA and protein level. In addition, both of cell proliferation and Six2 expression in MM cells declined when dosage reached high-concentration (50 mM). However, Six2 knock-down converted the proliferation reduction at 50 mM. Furthermore, Six2 deficiency increased the apoptosis of MM cells, compared with negative control cells at relative LiCl concentration. However, the abnormal rise of apoptosis at 30 mM of LiCl concentration implies that it might be the reduction of GSK3ß that increased cell apoptosis. Together, these demonstrate that LiCl can induce the proliferation and apoptosis of MM cells coordinating with Six2.

Glucocorticoid and glycolysis inhibitors cooperatively abrogate acute graft-versus-host disease.

Although glucorticosteroids (GCs) are the standard first-line therapy for acute graft-versus-host disease (aGvHD), nearly 50% of aGvHD patients have no response to GCs. The role of T cell metabolism in murine aGvHD was recently reported. However, whether GCs and metabolism regulators could cooperatively suppress T cell alloreactivity and ameliorate aGvHD remains to be elucidated. Increased glycolysis, characterized by elevated 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), and higher rates of glucose consumption and lactate production were found in T cells from aGvHD patients. Genetic upregulation of PFKFB3 induced T cell proliferation and differentiation into proinflammatory cells. In a humanized mouse model, PFKFB3-overexpressing or PFKFB3-silenced T cells aggravated or prevented aGvHD, respectively. Importantly, our integrated data from patient samples in vitro, in a humanized xenogeneic murine model of aGvHD and graft-versus-leukaemia (GVL) demonstrate that GCs combined with a glycolysis inhibitor could cooperatively reduce the alloreactivity of T cells and ameliorate aGvHD without loss of GVL effects. Together, the current study indicated that glycolysis is critical for T cell activation and induction of human aGvHD. Therefore, the regulation of glycolysis offers a potential pathogenesis-oriented therapeutic strategy for aGvHD patients. GCs combined with glycolysis inhibitors promises to be a novel first-line combination therapy for aGvHD patients.