Assessing the Potential Value and Mechanism of Kaji-Ichigoside F1 on Arsenite-Induced Skin Cell Senescence

Chronic exposure to inorganic arsenic is a major environmental public health issue worldwide affecting more than 220 million of people. Previous studies have shown the correlation between arsenic poisoning and cellular senescence; however, knowledge regarding the mechanism and effective prevention measures has not been fully studied. First, the associations among the ERK/CEBPB signaling pathway, oxidative stress, and arsenic-induced skin cell senescence were confirmed using the HaCaT cell model. In the arsenic-exposed group, the relative mRNA and protein expressions of ERK/CEBPB signaling pathway indicators (ERK1, ERK2, and CEBPB), cell cycle-related genes (p21, p16INK4a), and the secretion of SASP (IL-1α, IL-6, IL-8, TGF-β1, MMP-1, MMP-3, EGF, and VEGF) and the lipid peroxidation product (MDA) were significantly increased in cells ( P < 0.05 ), while the activity of antioxidant enzyme (SOD, GSH-Px, and CAT) was significantly decreased ( P < 0.05 ), and an increased number of cells accumulated in the G1 phase ( P < 0.05 ). Further Kaji-ichigoside F1 intervention experiments showed that compared to that in the arsenic-exposed group, the expression level of the activity of antioxidant enzyme was significantly increased in the Kaji-ichigoside F1 intervention group ( P < 0.05 ), but the indicators of ERK/CEBPB signaling pathway, cell cycle-related genes, and SASP were significantly decreased ( P < 0.05 ), and the cell cycle arrest relieved to a certain extent ( P < 0.05 ). Our study provides some limited evidence that the ERK/CEBPB signaling pathway is involved in low-dose arsenic-induced skin cell senescence, through regulating oxidative stress. The second major finding was that Kaji-ichigoside F1 can downregulate the ERK/CEBPB signaling pathway and regulate the balance between oxidation and antioxidation, alleviating arsenic-induced skin cell senescence. This study provides experimental evidence for further understanding of Kaji-ichigoside F1, a natural medicinal plant that may be more effective in preventing and controlling arsenic poisoning.

Type I Calreticulin Mutations Result in Hyperactivation of Its Acetyltransferase Function and Iron Metabolism, Inducing a Susceptibility to Ferroptosis

Approximately 20% of patients with myeloproliferative neoplasms (MPN) harbor mutations in the gene calreticulin (CALR). Of these, approximately half are classified as type 1 and 30% as type 2, characterized by a 52 bp deletion (CALRdel52) and a 5 bp insertion (CALRins5) respectively. Although both share identical mutant C-termini and are able to bind and activate MPL, type 1 and type 2 CALR mutations display different clinical and prognostic presentation: type 1 mutations are associated primarily with a fibrotic phenotype and increased proclivity towards fibrotic transformation, while type 2 mutations are more common in ET. Molecularly, type 1 and type 2 mutations result in differential C-domain amino acid sequences with the potential to affect the function of the protein. Various well known functions of CALR, including calcium binding ability and protein folding capacity, have begun to be explored in the context of CALR mutations; however, the impact of CALR mutations on its acetyltransferase ability, which was only discovered in 2006, remains unknown. Here, we show that in accordance with our structural models, mutant CALR not only retains its acyltransferase ability, but type 1 CALRdel52 mutations specifically lead to increased activation of its acetyltransferase ability, revealing a new gain of function phenotype for CALRdel52 mutations. As a result, type 1 CALR mutations lead to increased acetylation of CALR's acetyltransferase targets downstream, such as glutathione-S-transferase and cytochrome P450 reductase, which affects the outputs of these pathways downstream. Exploratory RNA-Seq on CALR-mutated cells revealed a concurrent upregulation of transferrin receptor mediated iron metabolism by CALRdel52. We subsequently validated this finding and show that CALRdel52 cells display differential iron metabolism. Given the upregulation of the transferrin receptor and the increased acetyltransferase ability affecting proteins involved in reactive oxygen species pathways (ROS), ferroptosis-an iron-dependent form of cell death characterized by the accumulation of lipid peroxides-emerged as a potential therapeutic target for CALRdel52 mutated cells. To test this, we first assessed basal proclivity to ferroptosis by measuring the lipid peroxidation product, classic ferroptotic marker 4-HNE (4-hydroxynonenal) as well as both ROS and global lipid peroxide levels in cells expressing wild-type CALR, CALRdel52, and CALRins5. We found that all of these ferroptotic markers were significantly increased in CALRdel52 cells. Therapeutic modulation of these pathways such as iron supplementation resulted in targeting of CALRdel52 cells and ferroptosis induction. This work is the first to examine the acetyltransferase ability of mutant CALR and reveal downstream phenotypic differences based on this ability that set the groundwork for a host of unexplored cellular consequences. Moreover, this study unites the novel understanding of the acetyltransferase function of mutant CALR with changes in transferrin receptor mediated iron metabolism to reveal not only how CALRdel52 induces a ferroptotic proclivity, but the potential of this sensitivity for therapeutic targeting. Disclosures No relevant conflicts of interest to declare.

Peculiarities of changes in a functional condition of pro-oxidant and antioxidant systems in guinea pigs’ lungs in experimental allergic alveolitis under the conditions of immobilization stress

We have analyzed the results of investigation of alterations in indices of pro-oxidant (conjugated diene and malondialdehyde) and antioxidant (superoxide dismutase, ceruloplasmin, catalase) systems in guinea pigs’ lungs in experimental allergic alveolitis in the dynamics of EAA development under the conditions of immobilization stress. The investigation was conducted on 62 female guinea pigs weighing 180-220 g, divided into 4 groups: I – intact guinea pigs (n=20), II – guinea pigs (n=14) with EAA under the conditions of IS (1st day); III – guinea pigs (n=14) with EAA under the conditions of IS (2nd day); IV – guinea pigs (n=14) with EAA under the conditions of IS (64th day). The results of experimental investigation showed that a significant increase in conjugated diene level in animals’ lungs was observed at all stages of EAA development under the conditions of immobilization stress as compared with control group, indicating activation of this marker. The same changes occurred with MDA content, indicating excessive accumulation of this lipid peroxidation product in lung tissue. Intensive synthesis of free radical compounds caused activation of some components of enzymatic system of antioxidant defense. In particular, a moderate decrease in superoxide dismutase activity in lung tissue is observed in response to increased level of LOPs at early stages of EAA and immobilization stress development as compared with the indices in intact animals. The same situation is observed with catalase and ceruloplasmin activity in the lungs of guinea pigs with modeled AA and IS.

Association of Lipid Peroxidation Product 4-Hydroxynonenal with Post-Traumatic Stress Disorder

Repeated activation of the hypothalamic-pituitary-adrenal axis system, sleep disturbances, and other symptoms related to posttraumatic stress disorder (PTSD) elevate reactive oxygen species, increase inflammation, and accelerate cellular aging, leading to neuroprogression and cognitive decline. However, there is no information about possible involvement of 4-hydroxynonenal (4-HNE), the product of lipid peroxidation associated with stress-associated diseases, in the complex etiology of PTSD. Therefore, the aim of this study was to compare the plasma levels of 4-HNE between war veterans with PTSD (n = 62) and age-, sex- and ethnicity- matched healthy control subjects (n = 58) in order to evaluate the potential of HNE-modified proteins as blood-based biomarker of PTSD. The genuine 4-HNE-Enzyme-Linked Immunosorbent Assay (HNE-ELISA), based on monoclonal antibody specific for HNE-histidine (HNE-His) adducts, was used to determine plasma HNE-protein conjugates. Our results revealed significantly elevated levels of 4-HNE in patients with PTSD. Moreover, the accumulation of plasma 4-HNE seems to increase with aging but in a negative correlation with BMI, showing specific pattern of change for individuals diagnosed with PTSD. These findings suggest that oxidative stress and altered lipid metabolism reflected by increase of 4-HNE might be associated with PTSD. If confirmed with further studies, elevated 4-HNE plasma levels might serve as a potential biomarker of PTSD.

Anti-Malarial and Anti-Lipid Peroxidation Activities of Deferiprone-Resveratrol Hybrid in Plasmodium berghei-Infected Mice

Iron is essential for all organisms including fast-dividing malarial parasites. Inversely, iron chelators can inhibit parasite growth through the inhibition of DNA synthesis and can ameliorate oxidative cell damage. Deferiprone (DFP)-resveratrol (RVT) hybrid (DFP-RVT) is a lipophilic anti-oxidative, iron-chelating agent that has displayed potent neuroprotective and anti-plasmodium activities in vitro. The goal of this work was to investigate the inhibitory effects of DFP-RVT on parasite growth and oxidative stress levels during malaria infections. Mice were intraperitoneally infected with P. berghei and orally administered with DFP, DFP-RVT and pyrimethamine for 4 d. The percentage of parasitemia was determined using Giemsa’s staining/microscopic examination. Amounts of the lipid-peroxidation product, thiobarbituric acid-reactive substance (TBARS), were determined in both plasma and liver tissue. In our findings, DFP-RVT exhibited a greater potent inhibitory effect and revealed an improvement in anemia and liver damage in infected mice than DFP. To this point, the anti-malarial activity was found to be associated with anti-RBC hemolysis and the liver weight index. In addition, plasma and liver TBARS levels in the DFP-RVT-treated mice were lower than those in DFP-treated mice. Thus, DFP-RVT could exert anti-plasmodium, anti-hemolysis and anti-lipid peroxidation activities to a better degree than DFP in P. berghei-infected mice.

Combretum dolichopentalum extract normalized biochemical and haematological parameters in carbon tetrachloride (CCL4) intoxicated rats

Background: The ethanol extract of Combretum dolichopentalum (EECD) is employed in Nigeria to stabilize the uterus after parturition. The ability of EECD to confer protection on rats destabilized by moderate concentrations of carbon tetrachloride (CCl4) was evaluated. Methods: Fifty rats were assigned to 5 groups of 10 rats each. The experimental animals after acclimatization were handled accordingly: Groups 1 and 2 respectively were maintained on food and water only throughout the study. Group 3, 4, and group 5 were pre-treated with 250 mg/kg and 500 mg/kg body weight of EECD and 50 mg/kg of silymarin for 28 respectively. All groups except group 1 were intoxicated to 0.2 ml/kg body weight of CCl4, administered via an intraperitoneal route on day 29. Serum pipetted from the blood of the rats after cardiac puncture was assayed for antioxidant enzymes, lipid peroxidation product and serum iron, zinc and biocarbonate. Haematological analysis was also conducted. Results: Administration of CCl4 at 0.2 ml/kg b.w slightly increased the oxidizing species as indicated in the concentration of malondialdehyde in the rats while reducing the antioxidant enzymes; it increased the Iron and zinc concentrations and also the haematological parameters except for the white blood cells. However, this was corrected by pre-treatment with the EECD dosedependently. Conclusion: These characteristics portends that the crude ethanol extract of C. dolichopentalum could be employed to correct minor oxidative perturbation induced by CCl4 intoxication

Short-Term and Long-Term Carcinogenic Effects of Food Contaminants (4-Hydroxynonenal and Pesticides) on Colorectal Human Cells: Involvement of Genotoxic and Non-Genomic Mechanisms

To investigate environmental impacts upon colorectal carcinogenesis (CRC) by diet, we assessed two western diet food contaminants: 4-hydroxynonenal (HNE), a major lipid peroxidation product neoformed during digestion, and a mixture of pesticides. We used human colonic cell lines ectopically eliciting varied genetic susceptibilities to CRC: the non-transformed human epithelial colonic cells (HCECs) and their five isogenic cell lines with the loss of APC (Adenomatous polyposis coli) and TP53 (Tumor protein 53) and/or ectopic expression of mutated KRAS (Kristen-ras). These cell lines have been exposed for either for a short time (2–24 h) or for a long period (3 weeks) to 1 µM HNE and/or 10 µM pesticides. After acute exposure, we did not observe any cytotoxicity or major DNA damage. However, long-term exposure to pesticides alone and in mixture with HNE induced clonogenic transformation in normal HCECs, as well as in cells representing later stages of carcinogenesis. It was associated with genotoxic and non-genomic mechanisms (cell growth, metabolic reprogramming, cell mobility and epithelial-mesenchymal transition) depending on genetic susceptibility. This study demonstrated a potential initiating and promoting effect of food contaminants on CRC after long-term exposure. It supports that these contaminants can accelerate carcinogenesis when mutations in oncogenes or tumor suppressor genes occur.

Preliminary Findings on the Association of the Lipid Peroxidation Product 4-Hydroxynonenal with the Lethal Outcome of Aggressive COVID-19

Major findings of the pilot study involving 21 critically ill patients during the week after admission to the critical care unit specialized for COVID-19 are presented. Fourteen patients have recovered, while seven passed away. There were no differences between them in respect to clinical or laboratory parameters monitored. However, protein adducts of the lipid peroxidation product 4-hydroxynonenal (HNE) were higher in the plasma of the deceased patients, while total antioxidant capacity was below the detection limit for the majority of sera samples in both groups. Moreover, levels of the HNE-protein adducts were constant in the plasma of the deceased patients, while in survivors, they have shown prominent and dynamic variations, suggesting that survivors had active oxidative stress response mechanisms reacting to COVID-19 aggression, which were not efficient in patients who died. Immunohistochemistry revealed the abundant presence of HNE-protein adducts in the lungs of deceased patients indicating that HNE is associated with the lethal outcome. It seems that HNE was spreading from the blood vessels more than being a consequence of pneumonia. Due to the limitations of the relatively small number of patients involved in this study, further research on HNE and antioxidants is needed. This might allow a better understanding of COVID-19 and options for utilizing antioxidants by personalized, integrative biomedicine approach to prevent the onset of HNE-mediated vitious circle of lipid peroxidation in patients with aggressive inflammatory diseases.

Effect of the Extracts of Sargassum fusiforme on Red Tide Microalgae in East China Sea

This study examined the effects of extracts of hijiki (Sargassum fusiforme) on the growth and physiology of three species of red tide microalgae (Prorocentrum donghaiensis, Skeletonema costatum, and Heterosigma akashiwo) that commonly grow in the East China Sea. The red tide algae were cultivated with the hijiki extracts at different concentrations to investigate the effects of the extracts on cells growth, chlorophyll a content, maximum quantum yield of PSII (Fv/Fm), the activities of four oxidoreductases including peroxidase (SOD), glutathione S-transferase (GST), glutathione peroxidase (GSH-Px), glutathione reductase (GR), and the level of the membrane lipid peroxidation product, malondialdehyde (MDA). The sensitivity of red tide algae to the extracts varied among the strains, with P. donghaiensis being the most sensitive, followed by S. costatum, and then H. akashiwo. Furthermore, the extracts had a rapid lethal effect on P. donghaiensis at over 1.6 g/L and on S. costatum and P. donghaiensis at over 8 g/L. From that concentration, increasing amounts of the extracts in cultures of S. costatum promoted a reduction in Chla contents and Fv/Fm values. In addition, the oxidoreductase activity of S. costatum was reduced at 4 and 6 g/L, as shown by the reduced activity of SOD, GR, GSH-Px, GST and MDA content in the cells. The results presented herein will be useful to the development and utilization of hijiki on red tide control, and marine environmental protection.

Selenium Biofortification of Soybean Seeds Influences Physiological Responses of Seedlings to Osmotic Stress

Climate change poses a serious threat to agricultural production. Water deficit in agricultural soils is one of the consequences of climate change that has a negative impact on crop growth and yield. Selenium (Se) is known to be involved in plant defense against biotic and abiotic stress through metabolic, structural, and physiological activity in higher plants. The aim of this study was to investigate the physiological response of Se-biofortified soybean (Glycine max (L.) Merrill) seedlings under osmotic stress. For this research, we used biofortified soybean grain obtained after foliar Se biofortification in 2020. The experiment was conducted in a growth chamber with two cultivars (Lucija and Sonja) grown on filter paper in three replicates. The experiment was carried out with two watering treatments: distilled water (PEG-0) and 2.5% polyethylene glycol 6000 (PEG-2.5) on Se-biofortified seeds (Se) and nonbiofortified seeds (wSe). Contents of lipid peroxidation product (LP), free proline (PRO), total phenolic content (TP), ferric reducing antioxidant power (FRAP), and ascorbic acid (AA) were analyzed in 7-days-old seedlings. Significant differences were detected in the Se content of soybean grains between the two cultivars. A milder reaction to PEG-2.5 was observed in cultivar Lucija in both Se and wSe treatments, which might represent the mitigating effects of Se on osmotic stress in this cultivar. Contrarily, in cultivar Sonja, Se adversely affected all analyzed traits in the PEG-2.5 treatment. Ultimately, Se is a pro-oxidant in Sonja, whereas it represents an anti-oxidant in Lucija. In conclusion, different soybean cultivars show contrasting physiological reactions to both osmotic stress and Se. However, the activation of antioxidant pathways in Sonja can also be interpreted as added value in soybean seedlings as a functional food.