Overexpression of nucleotide metabolic enzyme DUT in hepatocellular carcinoma potentiates a therapeutic opportunity through targeting its dUTPase activity.

Uracil misincorporation during DNA replication is a major cell toxic event, of which cancer cells overcome by activating the dUTPase enzyme. The DUT gene is the only known dUTPase in human. Despite reports on common upregulations in cancers, the role of DUT in human hepatocellular carcinoma (HCC) remains largely undetermined. In this study, we investigated the mechanism underlying DUT biology in HCC and tumor susceptibility to drug targeting dUTPase. Overexpression of DUT was found in 42% of HCC tumors and correlated with advanced stage HCC. Knockout of DUT in HCC cell lines showed suppressed proliferation through cell cycle arrest and a spontaneous induction of DNA damage. A protective effect from oxidative stress was also demonstrated in both knockout and overexpression DUT assays. Transcriptome analysis highlighted the NF-κB survival signaling as the downstream effector pathway of DUT in overriding oxidative stress-induced cell death. Interestingly, stably expressed DUT in liver progenitor organoids conferred drug resistance to TKI Sorafenib. Targeting dUTPase activity by TAS-114, could potentiate suppression of HCC growth that synergized with Sorafenib for better treatment sensitivity. In conclusion, upregulated DUT represents a nucleotide metabolic weakness and therapeutic opportunity in HCC.

Medium Chain Triglycerides Promote the Uptake of ß-Carotene in O/W Emulsions via Intestinal Transporter SR-B1 in Caco-2 Cells.

This study aimed to elucidate the impacts of carrier oil types (long chain triglycerides (LCT), medium chain triglycerides (MCT), and orange oil (indigestible oil)) on the micellization and cellular uptake of ß-carotene (BC) formulated in O/W emulsions, with an emphasis on the role of intestinal transporters. The micellization and cellular uptake of BC in the gastrointestinal tract were evaluated via an in vitro digestion model and a Caco-2 cell monolayer. And the interactions between lipids and intestinal transporters were monitored by nontargeted lipidomics, RT-PCR, and Western blot. The BC micellization rates followed a decreasing trend in emulsions: corn oil (69.47 ± 4.19%) > MCT (22.22 ± 0.89%) > orange oil (11.01 ± 2.86%), whereas the cellular uptake rate of BC was significantly higher in MCT emulsion (56.30 ± 20.13%) than in corn oil emulsion (14.01 ± 1.04%, p < 0.05). The knockdown of SR-B1 led to a 31.63% loss of BC cellular uptake from MCT micelles but had no effect on corn oil micelles. Lipidomics and transporter analysis revealed that TG (10:0/10:0/12:0) and TG (10:0/12:0/12:0) might be the fingerprint lipids that promoted the cellular absorption of BC-MCT micelles via stimulating the mRNA expression of SR-B1.

Calycosin-7-O-ß-D-glucoside regulates nitric oxide /caveolin-1/matrix metalloproteinases pathway and protects blood-brain barrier integrity in experimental cerebral ischemia-reperfusion injury.

ETHNOPHARMACOLOGY RELEVANCE: Astragali Radix (AR) has been used for thousands years to treat ischemic stroke. Calycosin and its glycoside form calycosin-7-O-ß-D-glucoside (CG) are two representative isoflavones in Astragali Radix. However, its neurological effects and related molecular mechanisms are largely unknown. The present study aims to evaluate the neuroprotective effects of CG on blood-brain barrier (BBB) integrity of ischemic brain tissue and explore the relevant signaling mechanisms. MATERIAL AND METHOD: Male adult Sprague-Daweley rats were subjected to 2 h of middle cerebral artery occlusion (MCAO) plus 24 h or 14 days of reperfusion. CG (26.8 mg/kg) was intraperitoneally administered into the rats at 15 min before onset of ischemia. The neuroprotective effects of CG were evaluated by measuring infarct volume, histological damage and BBB permeability. Furthermore, the effects of CG on scavenging nitric oxide (NO), and modulating matrix metalloproteinases (MMPs) and caveolin-1 (cav-1) were investigated with in vitro cultured brain microvascular endothelial cells treated with NO donor or oxygen-glucose deprivation (OGD) and/or in vivo rat model of MCAO cerebral ischemia-reperfusion injury. RESULTS: CG treatment significantly reduced infarct volume, histological damage and BBB permeability in the in vivo MCAO ischemia-reperfusion rat model. CG treatment remarkably inhibited the expression and activities of MMPs, and secured the expression of cav-1 and tight junction proteins in the microvessels isolated from ischemic rat cortex. Furthermore, CG was revealed to scavenge NO, inhibit the activities of MMP-2 and MMP-9, and attenuate cell death in the in vitro cultured brain microvascular endothelial cells under OGD condition. CONCLUSION: CG could protect BBB integrity in experimental cerebral ischemia-reperfusion injury via regulating NO/cav-1/MMPs pathway.

Baicalin can scavenge peroxynitrite and ameliorate endogenous peroxynitrite-mediated neurotoxicity in cerebral ischemia-reperfusion injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Baicalin is one of the principal flavonoids isolated from the dried root of Scutellaria baicalensis Georgi that has long been used to treat ischemic stroke. However, its neuroprotective mechanisms against cerebral ischemia injury are poorly understood. AIM OF THE STUDY: To explore the neuroprotective mechanisms of baicalin against cerebral ischemia reperfusion injury. MATERIAL AND METHODS: In chemical systems, we conducted electron paramagnetic resonance (EPR) spin trapping experiments to evaluate the scavenging effects of baicalin on superoxide and nitric oxide, and mass spectrometry (MS) studies on the reaction of baicalin and peroxynitrite. In cellular experiments, we investigated the effects of baicalin against extraneous and endogenous peroxynitrite mediated neurotoxicity in SH-SY5Y cells treated with peroxynitrite donor, synthesized peroxynitrite and exposed to oxygen glucose deprivation and reoxygenation (OGD/RO) in vitro. Moreover, we studied the neuroprotective effects of baicalin by using a rat model of middle cerebral artery occlusion in vivo. FeTMPyP, a peroxynitrite decomposition catalyst, was used as positive control. Cell viability and apoptotic cell death was accessed by MTT assay and TUNEL assay respectively; 3-nitrotyrosine formation and infarction volume were detected by immunostaining experiments and TTC staining respectively. RESULTS: Baicalin revealed strong antioxidant ability by directly scavenging superoxide and reacting with peroxynitrite. Baicalin protected the neuronal cells from extraneous and endogenous peroxynitrite-induced neurotoxicity. In ischemia-reperfused brains, baicalin inhibited the formation of 3-nitrotyrosine, reduced infarct size and attenuated apoptotic cell death, whose effects were similar to FeTMPyP. CONCLUSIONS: Baicalin can directly scavenge peroxynitrite and the peroxynitrite-scavenging ability contributes to its neuroprotective mechanisms against cerebral ischemia reperfusion injury.