Ubiquitin activation is essential for schizont maturation in Plasmodium falciparum blood-stage development

Ubiquitylation is a common post translational modification of eukaryotic proteins and in the human malaria parasite, Plasmodium falciparum (Pf) overall ubiquitylation increases in the transition from intracellular schizont to extracellular merozoite stages in the asexual blood stage cycle. Here, we identify specific ubiquitylation sites of protein substrates in three intraerythrocytic parasite stages and extracellular merozoites; a total of 1464 sites in 546 proteins were identified (data available via ProteomeXchange with identifier PXD014998). 469 ubiquitylated proteins were identified in merozoites compared with only 160 in the preceding intracellular schizont stage, suggesting a large increase in protein ubiquitylation associated with merozoite maturation. Following merozoite invasion of erythrocytes, few ubiquitylated proteins were detected in the first intracellular ring stage but as parasites matured through trophozoite to schizont stages the apparent extent of ubiquitylation increased. We identified commonly used ubiquitylation motifs and groups of ubiquitylated proteins in specific areas of cellular function, for example merozoite pellicle proteins involved in erythrocyte invasion, exported proteins, and histones. To investigate the importance of ubiquitylation we screened ubiquitin pathway inhibitors in a parasite growth assay and identified the ubiquitin activating enzyme (UBA1 or E1) inhibitor MLN7243 (TAK-243) to be particularly effective. This small molecule was shown to be a potent inhibitor of recombinant PfUBA1, and a structural homology model of MLN7243 bound to the parasite enzyme highlights avenues for the development of P. falciparum specific inhibitors. We created a genetically modified parasite with a rapamycin-inducible functional deletion of uba1; addition of either MLN7243 or rapamycin to the recombinant parasite line resulted in the same phenotype, with parasite development blocked at the schizont stage. Nuclear division and formation of intracellular structures was interrupted. These results indicate that the intracellular target of MLN7243 is UBA1, and this activity is essential for the final differentiation of schizonts to merozoites

Protection of Iron-Induced Oxidative Damage in Neuroblastoma (SH-SY5Y) Cells by Combination of 1-(N-Acetyl-6-aminohexyl)-3-hydroxy-2-methylpyridin-4-one and Green Tea Extract

Iron is a crucial trace element and essential for many cellular processes; however, excessive iron accumulation can induce oxidative stress and cell damage. Neurodegenerative disorders, such as Alzheimer’s disease and Parkinson’s disease, have been associated with altered iron homoeostasis causing altered iron distribution and accumulation in brain tissue. This study aims to investigate the protective effect of 1-(N-acetyl-6-aminohexyl)-3-hydroxy-2-methylpyridin-4-one (CM1) in combination with green tea extract (GTE) on iron-induced oxidative stress in neuroblastoma (SH-SY5Y) cells. Cells were cultured in medium with or without ferric chloride loading. Their viability and mitochondrial activity were assessed using MTT and JC-1 staining methods. Levels of the cellular labile iron pool (LIP), reactive oxygen species (ROS), and lipid-peroxidation products were determined using calcein acetoxymethyl ester, 2′,7′-dichlorohydrofluorescein diacetate, and TBARS-based assays, respectively. The viability of iron-loaded cells was found to be significantly increased after treatment with CM1 (10 µM) for 24 h. CM1 co-treatment with GTE resulted in a greater protective effect than their monotherapy. Combination of CM1 and GTE also reduced mitochondrial disruption and LIP content and ROS and TBARS production. In conclusion, the combination of CM1 and GTE exhibits protection against iron-induced oxidative stress in neuroblastoma cells

Impact of Green Extraction on Curcuminoid Content, Antioxidant Activities and Anti-Cancer Efficiency (In Vitro) from Turmeric Rhizomes (<i>Curcuma longa</i> L.)

Turmeric (Curcuma longa L.) powder is widely used as a spice and seasoning in Asian countries. This study investigated the effect of turmeric extracts on the anticancer activity of Huh7 and HCT 116 cells. The curcumin bioactive compounds were extracted using various methods such as microwave-assisted extraction (MAE), ultrasound-assisted extraction (UAE) and traditional extraction (TDE). The yield of dried extracts from MAE was found to be the highest at 17.89%, followed by UAE and TDE, with 11.34% and 5.54%, respectively. Antioxidant activities such as TPC, DPPH and FRAP from MAE were higher than those of UAE and TDE. The total curcuminoid contents from the novel extractions were higher than those from traditional extraction methods. For instance, curcuminoid contents from MAE, UAE and TDE were 326.79, 241.17 and 215.83 mg/g, respectively. Due to having the highest bioactive compounds and extraction yield, turmeric extract from MAE was used to investigate the potential anticancer properties. The extract showed significant cytotoxic potential against the human liver (Huh7) and human colon (HCT116) cell lines, in concentrations ranging from 31.25 to 1000.00 µg/mL. Turmeric extracts using MAE have potential anticancer effects on Huh7 and HCT116 cells. This study serves as scientific data for the chemotherapeutic properties of turmeric extracts and their use as functional ingredients

Antioxidant Potential and Cytotoxic Effect of Isoflavones Extract from Thai Fermented Soybean (Thua-Nao)

Thua-nao, or Thai fermented soybeans, is a traditional Lanna fermented food in Northern Thailand. It is produced by using a specific bacterial species called Bacillus subtilis var. Thua-nao. We investigated the antioxidant activity and cytotoxic effect of isoflavones from Thua-nao. The phenolic compound contents and total flavonoid contents were determined by spectrophotometry. The antioxidant activity was examined using the ABTS, FRAP, and DPPH assays. The isoflavone contents and phenolic compositions were examined by the high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) techniques. The ability of isoflavones to inhibit human cancer cell growth was assessed by the MTT assay. The total phenolic content, total flavonoid content, and antioxidant activities of the isoflavones were 49.00 &plusmn; 0.51 mg GAE/g of dry extract (DE), 10.76 &plusmn; 0.82 mg QE/g of DE, 61.03 &plusmn; 0.97 &micro;mol Trolox/g of DE, 66.54 &plusmn; 3.97 &micro;M FeSO4/g of DE, and 22.47 &plusmn; 1.92% of DPPH inhibition, respectively. Additionally, the isoflavone extracts from Thua-nao had high isoflavone contents and polyphenolic compound compositions, especially daidzein and genistein. The isoflavone demonstrated a weak inhibition of MCF-7 and HEK293 cancer cell growth. It has a high antioxidant component, which is beneficial and can be developed for new therapeutic uses. However, further studies on the benefits of Thua-nao should be performed for realizing better and more effective uses soon