L-精氨酸在诱导型一氧化氮合酶(iNOS)的作用下可产生一氧化氮(NO),而5-氟尿嘧啶(5-FU)可诱导iNOS表达的增加。本研究旨在探讨L-精氨酸联合5-FU通过iNOS/NO/AKT通路调节肝细胞癌(HCC)糖代谢的机制。L-精氨酸和5-FU的组合导致细胞存活率降低,并在HepG2和SMMC7721细胞中表现出协同的细胞毒性作用。同时,L-精氨酸通过增加NO的产生来增强5-FU对HepG2和SMMC7721细胞的抑制作用。用L-精氨酸和5-FU共处理导致G6PDH和LDH酶活性显著降低,以及与单独用L-精氨酸或5-FU治疗相比降低的ATP和LD水平。此外,L-精氨酸和5-FU的组合导致GLUT1,PKM2,LDHA的表达减少,p-PI3K和p-AKT。此外,该组合在下调HIF-1α和β-catenin的表达方面表现出协同作用,随着紫草素的加入,PKM2的特异性抑制剂。与组合组相比,LY294002处理进一步降低了由组合的L-精氨酸和5-FU处理诱导的GLUT1、PKM2和LDHA蛋白的表达。然而,p-PI3K的减少,p-AKT,在iNOS敲低的HepG2和SMMC7721细胞中,L-精氨酸和5-FU组合引起的GLUT1表达也被逆转,分别。此外,L-精氨酸和5-FU的组合导致ALT酶活性的更大降低,AST,G6PDH和LDH,以及肝指数显著降低,法新社,肝癌大鼠模型中AFP-L3、ATP和LD水平。此外,L-精氨酸和5-FU的同时给药显著改善了肝脏的总体形态,减少核异型,抑制癌细胞的增殖,并降低了p-PI3K的表达水平,p-AKT,GLUT1、PKM2和LDHA,而联合组iNOS表达增加。一起,L-精氨酸和5-FU组合通过iNOS/NO/AKT途径抑制有氧糖酵解中的酶,导致葡萄糖代谢的抑制和核转录因子的下调,从而阻碍肝癌细胞的增殖。
L-arginine can produce nitric oxide (NO) under the action of inducible nitric oxide synthase (iNOS), while 5-fluorouracil (5-FU) can induce the increase of iNOS expression. The present study was to investigate the mechanism of L-arginine combined with 5-FU regulating glucose metabolism of hepatocellular carcinoma (HCC) through iNOS/NO/AKT pathway. The combination of L-arginine and 5-FU resulted in decreased cell survival and exhibited synergistic cytotoxic effects in HepG2 and SMMC7721 cells. Meanwhile, L-arginine increased 5-FU inhibitory effect on HepG2 and SMMC7721 cells by increasing NO production. Co-treatment with L-arginine and 5-FU resulted in a significant decrease in both G6PDH and LDH enzymatic activities, as well as reduced levels of ATP and LD compared to treatment with L-arginine or 5-FU alone. Moreover, the combination of L-arginine and 5-FU resulted in a decrease in the expression of GLUT1, PKM2, LDHA, p-PI3K and p-AKT. Furthermore, the combination demonstrated a synergistic effect in downregulating the expression of HIF-1α and β-catenin, which were further diminished upon the addition of shikonin, a specific inhibitor of PKM2. LY294002 treatment further reduced the expression of GLUT1, PKM2, and LDHA proteins induced by combined L-arginine and 5-FU treatment compared to the combined group. However, the reduction in p-PI3K, p-AKT, and GLUT1 expression caused by L-arginine and 5-FU combination was also reversed in HepG2 and SMMC7721 cells with iNOS knockdown, respectively. Additionally, the combination of L-arginine and 5-FU led to a greater reduction in the enzymatic activity of ALT, AST, G6PDH and LDH, as well as a significant reduction in hepatic index, AFP, AFP-L3, ATP and LD levels in a rat model of HCC. Moreover, the simultaneous administration of L-arginine and 5-FU significantly improved the gross morphology of the liver, reduced nuclear atypia, inhibited the proliferation of cancer cells, and decreased the expression levels of p-PI3K, p-AKT, GLUT1, PKM2, and LDHA, while iNOS expression was increased in the combination group. Taking together, L-arginine and 5-FU combination resulted in the inhibition of enzymes in aerobic glycolysis via the iNOS/NO/AKT pathway, which led to the suppression of glucose metabolism and downregulation of nuclear transcription factors, thereby impeding the proliferation of hepatocellular carcinoma cells.