5'-CMP and 5'-UMP alleviate dexamethasone-induced muscular atrophy in C2C12 myotubes.

Atrogin-1 and muscle RING finger 1 (MuRF1) are ubiquitin ligases specifically expressed during skeletal muscle atrophy and mediate muscle protein degradation. In contrast, PGC-1α (peroxisome proliferator-activated receptor γ coactivator 1α), which is a master regulator of mitochondrial biosynthesis, protects skeletal muscle from atrophy. Pyrimidine nucleoside 5'-monophosphates, such as cytidine 5'-monophosphate (5'-CMP) and uridine 5'-monophosphate (5'-UMP), induce PGC-1α expression and promote myotube formation in mouse C2C12 cells. In this study, we determined the effect of 5'-CMP and 5'-UMP on muscular atrophy in C2C12 myotube cells. 5'-UMP decreased Atrogin-1 and MuRF1 mRNA levels that were upregulated by dexamethasone treatment. 5'-CMP and 5'-UMP ameliorated dexamethasone-mediated atrophy in C2C12 myotubes. Furthermore, the combination of 5'-CMP and 5'-UMP further alleviated dexamethasone-mediated atrophy. In addition, cytidine and uridine, the precursors of 5'-CMP and 5'-UMP, markedly ameliorated dexamethasone-mediated atrophy. Considering nucleotide metabolism and absorption, the active metabolites underlying the observed effects of 5'-CMP and 5'-UMP appear to be cytidine and uridine. Our results indicate that 5'-CMP alleviates muscle atrophy by activating PGC-1α and differentiation, and 5'-UMP alleviates muscle atrophy by suppressing the activation of the myolytic system, whereas the combined use of both enhances the muscle atrophy inhibitory effect. 5'-CMP and 5'-UMP may be an effective and safe treatment for muscular atrophy.

5'-CMP and 5'-UMP promote myogenic differentiation and mitochondrial biogenesis by activating myogenin and PGC-1α in a mouse myoblast C2C12 cell line.

Ribonucleotides are basic monomeric building blocks for RNA considered as conditionally essential nutrients. They are normally produced in sufficient quantity, but can become insufficient upon stressful challenges. The administration of pyrimidine nucleotides, such as cytidine-5'-monophosphate (5'-CMP) and uridine-5'-monophosphate (5'-UMP), enables rats to endure prolonged exercise. However, the underlying mechanisms have remained elusive. To investigate these mechanisms, we studied the effect of 5'-CMP and 5'-UMP on muscular differentiation and mitochondrial biogenesis in myoblast C2C12 cells. 5'-CMP and 5'-UMP were found to increase the mRNA levels of myogenin, which is a myogenic regulatory protein expressed during the final differentiation step and fusion of myoblasts into myotubes. 5'-CMP and 5'-UMP also promoted myoblast differentiation into myotube cells. 5'-CMP and 5'-UMP further increased the mRNA levels of PGC-1α which regulates mitochondrial biogenesis and skeletal muscle fiber type. In addition, 5'-CMP and 5'-UMP increased mitochondrial DNA copy number and enhanced mRNA levels of slow-muscle myosin heavy chains. Moreover, cytidine and uridine, nucleosides corresponding to 5'-CMP and 5'-UMP, markedly promoted myotube formation in C2C12 cells. Considering the metabolism and absorption of nucleotides, the active bodies underlying the effects observed with 5'-CMP and 5'-UMP could be cytidine and uridine. In conclusion, our results indicate that 5'-CMP and 5'-UMP can promote myogenic differentiation and mitochondrial biogenesis, as well as increase slow-twitch fiber via the activation of myogenin and PGC-1α. In addition, 5'-CMP and 5'-UMP may be considered as safe and effective agents to enhance muscle growth and improve the endurance in skeletal muscles.

Epigallocatechin-3-O-gallate induces acid sphingomyelinase activation through activation of phospholipase C.

Epigallocatechin-3-O-gallate (EGCG)-induced cyclic guanosine monophosphate (cGMP) plays a crucial role in EGCG-induced cell death in various types of cancer cells. However, little is known regarding the early molecular events after cGMP induction. In this study, we showed that cGMP induction is sufficient to induce the phosphorylation of protein kinase C delta (PKCδ) at Ser664, the crucial kinase for EGCG-induced activation of acid sphingomyelinase (ASM). Using a chemical inhibitor library, we revealed that the inhibitors of the negative regulators of diacylglycerol strongly increase the effect of EGCG. We also showed that EGCG treatment increased phospholipase C (PLC) activity, and the same results were obtained with cGMP inducer treatment. EGCG-induced ASM activation was completely suppressed by pharmacological inhibition of PLC. Collectively, EGCG-induced cGMP activated the cGMP/PLC/PKCδ/ASM signaling axis in multiple myeloma cells.

PDE3 inhibitor and EGCG combination treatment suppress cancer stem cell properties in pancreatic ductal adenocarcinoma.

Recurrence following chemotherapy is observed in the majority of patients with pancreatic ductal adenocarcinoma (PDAC). Recent studies suggest that cancer stem cells (CSCs) may be involved in PDAC recurrence and metastasis. However, an efficient approach to targeting pancreatic CSCs remains to be established. Here we show that in cancer cells overexpressing the 67-kDa laminin receptor (67LR)-dependent cyclic GMP (cGMP) inducer, epigallocatechin-3-O-gallate (EGCG) and a phosphodiesterase 3 (PDE3) inhibitor in combination significantly suppressed the Forkhead box O3 and CD44 axis, which is indispensable for the CSC properties of PDAC. We confirmed that the EGCG and PDE3 inhibitor in combination strongly suppressed tumour formation and liver metastasis in vivo. We also found that a synthesized EGCG analog capable of inducing strong cGMP production drastically suppressed the CSC properties of PDAC and extended the survival period in vivo. In conclusion, the combination treatment of EGCG and a PDE3 inhibitor as a strong cGMP inducer could be a potential treatment candidate for the eradication of CSCs of PDAC.

The FOXO3/PGC-1ß signaling axis is essential for cancer stem cell properties of pancreatic ductal adenocarcinoma.

In 95% of patients with pancreatic ductal adenocarcinoma, recurrence is observed following chemotherapy. Findings from several studies have indicated that cancer stem cells (CSCs) are resistant to anticancer agents and may be involved in cancer recurrence and metastasis. The CD44 protein is a major CSC marker, and CD44 also plays an indispensable role in the CSC properties in several cancers, including pancreatic cancer; however, no clinical approach exists to inhibit CD44 activity. Here, we have performed knock-in/knockdown experiments, and we demonstrate that the forkhead box O3 (FOXO3)/liver kinase B1 (LKB1)/AMP-activated protein kinase/peroxisome proliferator-activated receptor-γ co-activator-1ß (PGC-1ß)/pyruvate dehydrogenase-A1 pathway is essential for CD44 expression and CSC properties. We observed that patients exhibiting high pyruvate dehydrogenase-A1 expression have a poor prognosis. Systemic PGC-1ß knock-out mice are fertile and viable and do not exhibit an overt phenotype under normal conditions. This suggests that cGMP induction and PGC-1ß inhibition represent potential strategies for treating patients with pancreatic ductal adenocarcinoma.