ABSTRACT
In pancreatic ductal adenocarcinoma (PDAC), metabolic rewiring and resistance to standard therapy are closely associated. PDAC cells show enormous requirements for glucose-derived citrate, the first rate-limiting metabolite in the synthesis of new lipids. Both the expression and activity of citrate synthase (CS) are extraordinarily upregulated in PDAC. However, no previous relationship between gemcitabine response and citrate metabolism has been documented in pancreatic cancer. Here, we report for the first time that pharmacological doses of vitamin C are capable of exerting an inhibitory action on the activity of CS, reducing glucose-derived citrate levels. Moreover, ascorbate targets citrate metabolism towards the de novo lipogenesis pathway, impairing fatty acid synthase (FASN) and ATP citrate lyase (ACLY) expression. Lowered citrate availability was found to be directly associated with diminished proliferation and, remarkably, enhanced gemcitabine response. Moreover, the deregulated citrate-derived lipogenic pathway correlated with a remarkable decrease in extracellular pH through inhibition of lactate dehydrogenase (LDH) and overall reduced glycolytic metabolism. Modulation of citric acid metabolism in highly chemoresistant pancreatic adenocarcinoma, through molecules such as vitamin C, could be considered as a future clinical option to improve patient response to standard chemotherapy regimens.
ABSTRACT
Mitochondrial dysfunction associates with several pathological processes and contributes to chronic inflammatory and ageing-related diseases. Mitochondrial transcription factor A (TFAM) plays a critical role in maintaining mtDNA integrity and function. Taking advantage of Tfamfl/fl UBC-Cre/ERT2+/+ mice to investigate mitochondrial dysfunction in the stromal cell component, we describe an inducible in vitro model of mitochondrial dysfunction by stable depletion of TFAM in primary mouse skin fibroblasts (SK-FBs) after 4-hydroxytamoxifen (4-OHT) administration. Tfam gene deletion caused a sustained reduction in Tfam and mtDNA-encoded mRNA in Cre(+) SK-FBs cultured for low (LP) and high (HP) passages that translated into a loss of TFAM protein. TFAM depletion led to a substantial reduction in mitochondrial respiratory chain complexes that was exacerbated in HP SK-FB cultures. The assembly pattern showed that the respiratory complexes fail to reach the respirasome in 4-OHT-treated Cre(+) SK-FBs. Functionally, mito-stress and glycolysis-stress tests showed that mitochondrial dysfunction developed after long-term 4-OHT treatment in HP Cre(+) SK-FBs and was compensated by an increase in the glycolytic capacity. Finally, expression analysis revealed that 4-OHT-treated HP Cre(+) SK-FBs showed a senescent and pro-inflammatory phenotype.
Subject(s)
DNA, Mitochondrial , Mitochondrial Proteins , Animals , DNA, Mitochondrial/genetics , DNA-Binding Proteins/metabolism , Fibroblasts/metabolism , Gene Expression Regulation , Glycolysis , High Mobility Group Proteins/genetics , High Mobility Group Proteins/metabolism , Mice , Mitochondria/metabolism , Mitochondrial Proteins/metabolismABSTRACT
Peyronie and Dupuytren are pathologies characterized by the appearance of localized fibrotic lesions in an organ. These disorders originate from an excessive production of collagen in the tissue provoking dysfunction and functional limitations to the patients. Local administration of collagenase is the most used treatment for these fibrotic-type diseases, but a high lability of the enzyme limits its therapeutic efficacy. Herein, we present a novel methodology for the preparation of collagenase nanocapsules without affecting its enzymatic activity and capable of releasing the enzyme in response to an ultraviolet A (UVA) light stimulus. Polymeric coating around collagenase was formed by free-radical polymerization of acrylamide-type monomers. Their degradation capacity under UVA irradiation was provided by incorporating a novel photocleavable acrylamide-type crosslinker within the polymeric framework. This property allowed collagenase release to be triggered in a controlled manner by employing an easily focused stimulus. Additionally, UVA irradiation presents considerable benefits by itself due to its capacity to induce collagenase production in situ. An expected synergistic effect of collagenase nanocapsules in conjunction with UVA effect may present a promising treatment for these fibrotic diseases.
