Over the past decade, the incidence of PCa has risen rapidly, reaching an annual growth of 12.07%. and incubated at 4C overnight. Then the membranes were washed, and incubated with secondary antibody. Blots were developed using Pierce Fast Western Blot Kit and exposed to film. Image_4.jpeg (53K) GUID:?2877735A-93C8-4CEE-9901-BCD2F44FC1A4 Data Availability StatementThe raw data supporting the conclusions of this article will GKT137831 be made available by the authors, without undue reservation, to any qualified researcher. Abstract Background The anticancer potential of pharmacologic ascorbic acid (AA) has been detected in a number of cancer cells. However, study suggested a strongly reduced cytotoxic activity of AA. It was known that pH could be a critical influencing factor for multiple anticancer treatments. In this study, we explored the influence of pH on the cytotoxicity of ascorbic acid. We employed castration-resistant prostate cancer (CRPC) cell lines PC3 and DU145 to observe the therapeutic effect of AA on PCa cells that were cultured with different pH GKT137831 studies demonstrate that acidic pH attenuates the cytotoxic activity of pharmacologic ascorbic acid by inhibiting AA uptake in PCa cells. Additionally, we found that the cancer cell-selective toxicity of AA depends on ROS. (Jacobs et al., 2015). Sodium AA (0C10?mM) decreases the viability of both androgen-independent (DU145) and androgen-dependent (LNCaP) human prostate cancer (PCa) cell lines (Maramag et al., 1997). However, these results were not confirmed in clinical trials following administration of AA infusion in castration-resistant prostate cancer (CRPC) patients and patients with advanced stages of other cancers (Creagan et al., 1979; Chen et al., 2005; Nielsen et al., 2017). So far there was no study investigating whether pH could play a role in the anticancer effect of AA on CRPC. Previous studies were conducted using commercially available cell culture media buffered to physiological pH ranging from 7.2 to 7.4 (Raghunand et al., 1999a). Metabolic reprogramming in cancer is often accompanied by acidification of extracellular matrix (Szatrowski and Nathan, 1991). Measurements of pH in tumor tissues, using microelectrodes, magnetic resonance, or fluorescence techniques, typically yield an extracellular pH range of 6.5 to 6.9 (Flavell et al., 2016). In most tumors, the pH is more acidic near the surface and less acidic in the tumor center (Stock et al., 2007). The pH at surfaces which consisted of highly metastatic cells was around 6.1 to 6.4. Whereas in non-metastatic tumors, the pH was at a range of 6.7 to 6.9, as measured by positioning a pH-sensitive fluorescent dye (Anderson et al., 2016). Furthermore, different results from preclinical research and clinical studies indicate that different conditions between tumor cells in a 2D cell culture and the microenvironment of human tumors might be the decisive factor for failure of AA in cancer treatment (Hickman et al., 2014). We proposed that the mild acidic microenvironment of human tumors might be an important factor for impairing the cytotoxicity of AA. However, the role of microenvironmental pH in the cytotoxicity of GKT137831 AA remains poorly understood. The cellular p44erk1 transportation of AA is mediated by two transport protein families (Liang et al., 2001), (i) the solute carrier gene family 23, which comprises the sodium-dependent vitamin C transporters (SVCTs) 1 and 2; and (ii) the solute carrier 2 family of glucose transporters (GLUTs). GLUTs transport the oxidized form of AA, dehydroascorbate (DHA) (Wohlrab et al., 2017). SVCT1 and SVCT2 cotransport sodium and ascorbate in a ratio of 2:1 down to an electrochemical sodium gradient, which is maintained by K/Na+ exchange mechanisms (Tsukaguchi et al., 1999). SVCTs transport is sensitive to pH changes and the optimum pH is 7.5 (Ormazabal et al., 2010). Acidic pH impairs SVCTs function through a mechanism involving the reversible protonation-deprotonation of five histidine residues in SVCTs (Tsukaguchi et al., 1999). The five histidine residues are central regulators of SVCTs function that modulate pH sensitivity, transporter kinetics, Na+ cooperativity,.