Increased Reactive Oxygen Species Levels Research Articles

MiRNA-494 induces trophoblast senescence by targeting SIRT1

ABSTRACT Objective Although the mechanism underlying preeclampsia (PE) has been widely explored, the mechanisms related to senescence have not yet been fully revealed. Therefore, we investigated the role of the miR-494/longevity protein Sirtuin 1 (SIRT1) axis in PE. Methods Human placental tissue was obtained from severe preeclampsia (SPE) (n = 20) and gestational age-matched normotensive pregnancies (n = 20), and senescence-associated β-galactosidase (SAβG) and SIRT1 expression levels were measured. The TargetScan and miRDB databases predicted candidate miRNAs targeting SIRT1, and intersected with differentially expressed miRNAs in the GSE15789 dataset (p < 0.05, |log2FC|≥1.5). Subsequently, we showed that miRNA (miR)-494 expression was significantly elevated in SPE, revealing miR-494 as a candidate SIRT1-binding miRNA. A dual-luciferase assay confirmed the targeting relationship between miR-494 and SIRT1. The senescence phenotype, migration, cell viability, reactive oxygen species (ROS) production levels and inflammatory molecule expression levels were measured after miR-494 expression was altered. We conducted a rescue experiment using SIRT1 plasmids to further demonstrate the regulatory relationship. Results SIRT1 expression was lower(p < 0.01) and miR-494 expression was higher (p < 0.001) in SPE, and SaβG staining showed premature placental aging in SPE (p < 0.001). Dual-luciferase reporter assays revealed that miR-494 targeted SIRT1. Compared to control cells, HTR-8/SVneo cells with upregulation of miR-494 had remarkably downregulated SIRT1 expression (p < 0.001), more SAβG-positive cells (p < 0.001), cell cycle arrested (p < 0.05), and upregulated P21 and P16 expression (p < 0.01). miR-494 overexpression also decreased HTR-8/SVneo cell migration (p < 0.05) and ATP synthesis (p < 0.001), increased ROS levels (p < 0.001), and upregulated NLRP3 and IL-1β expression (p < 0.01). SIRT1-overexpressing plasmids partially reversed the effects of miR-494 overexpression in HTR-8/SVneo cells. Conclusion The miR-494/SIRT1 interaction plays a role in the mechanism of premature placental aging in PE patients.

Toxicological and Mutagenic Effects of Particulate Matter from Domestic Activities.

People spend most of their time indoors, particularly in their houses where daily activities are carried out, enhancing particulate matter (PM) emissions with consequent adverse health impacts. This study intended to appraise the toxicological and mutagenic responses of particulate matter with a diameter less than 10 μm (PM10) released from cooking and ironing activities under different conditions. The cytotoxicity of the PM10 total organic extracts was tested in A549 cells using the WST-8 and the lactate dehydrogenase (LDH) assays, while the interference in cell cycle dynamics and reactive oxygen species (ROS) production was analysed by flow cytometry. The S. typhimurium TA98 and TA100 Ames tester strains with and without metabolic activation were employed to determine the mutagenic potential of the PM10-bound polycyclic aromatic hydrocarbons (PAHs). PM10 organic extracts decreased the metabolic activity of A549 cells; however, no effects in the LDH release were observed. An increase in ROS levels was registered only for cells treated with PM10 at IC20 from steam ironing, in low ventilation conditions, while cell cycle dynamics was only affected by exposure to PM10 at IC20 from frying horse mackerel and grilling boneless pork strips. No mutagenic effects were observed for all the PM10-bound PAHs samples.

A Novel Strategy Based on Zn(II) Porphyrins and Silver Nanoparticles to Photoinactivate Candida albicans.

Photodynamic inactivation (PDI) is an attractive alternative to treat Candida albicans infections, especially considering the spread of resistant strains. The combination of the photophysical advantages of Zn(II) porphyrins (ZnPs) and the plasmonic effect of silver nanoparticles (AgNPs) has the potential to further improve PDI. Here, we propose the novel association of polyvinylpyrrolidone (PVP) coated AgNPs with the cationic ZnPs Zn(II) meso-tetrakis(N-ethylpyridinium-2-yl)porphyrin or Zn(II) meso-tetrakis(N-n-hexylpyridinium-2-yl)porphyrin to photoinactivate C. albicans. AgNPs stabilized with PVP were chosen to allow for (i) overlap between the NP extinction and absorption spectra of ZnPs and (ii) favor AgNPs-ZnPs interaction; prerequisites for exploring the plasmonic effect. Optical and zeta potential (ζ) characterizations were performed, and reactive oxygen species (ROS) generation was also evaluated. Yeasts were incubated with individual ZnPs or their respective AgNPs-ZnPs systems, at various ZnP concentrations and two proportions of AgNPs, then irradiated with a blue LED. Interactions between yeasts and the systems (ZnP alone or AgNPs-ZnPs) were evaluated by fluorescence microscopy. Subtle spectroscopic changes were observed for ZnPs after association with AgNPs, and the ζ analyses confirmed AgNPs-ZnPs interaction. PDI using ZnP-hexyl (0.8 µM) and ZnP-ethyl (5.0 µM) promoted a 3 and 2 log10 reduction of yeasts, respectively. On the other hand, AgNPs-ZnP-hexyl (0.2 µM) and AgNPs-ZnP-ethyl (0.6 µM) systems led to complete fungal eradication under the same PDI parameters and lower porphyrin concentrations. Increased ROS levels and enhanced interaction of yeasts with AgNPs-ZnPs were observed, when compared with ZnPs alone. We applied a facile synthesis of AgNPs which boosted ZnP efficiency. We hypothesize that the plasmonic effect combined with the greater interaction between cells and AgNPs-ZnPs systems resulted in an efficient and improved fungal inactivation. This study provides insight into the application of AgNPs in PDI and helps diversify our antifungal arsenal, encouraging further developments toward inactivation of resistant Candida spp.

FABP4 Regulates Cell Proliferation, Stemness, Apoptosis, and Glycolysis in Colorectal Cancer via Modulating ROS/ERK/mTOR Pathway.

Colorectal cancer is a common digestive tract malignancy. This study aimed to expound the functional role of fatty-acid-binding protein 4 (FABP4) and the potential underlying mechanisms in the development of colorectal cancer. Several techniques were utilized to investigate the role of FABP4 in colorectal cancer. FABP4 mRNA expression was quantified using Real time-quantitative PCR (RT-qPCR). Cell counting kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), sphere formation assays and flow cytometry evaluated cell growth, stemness, and apoptosis in SW480 and HT29 cells. Glycolysis was assessed via extracellular acidification rate (ECAR) , lactate production, glucose uptake, adenosine triphosphate (ATP)/adenosine 5'-diphosphate (ADP) ratio, and Glut1 and Elevated lactate dehydrogenase A (LDHA) protein expression. Reactive oxygen species (ROS) levels were analyzed by flow cytometry. Western blot measured the protein expression of FABP4, Proliferating cell nuclear antigen (PCNA), Bax, Bcl-2, Glut1, LDHA, stemness makers (Sox2, Oct4, and ALDHA1), and extracellular regulated protein kinase (ERK)/mammalian target of rapamycin (mTOR) pathway proteins. In vivo experiments, BALB/c nude mice (n = 12) were inoculated with 200 μL HT29 cells (5 × 106 cells) transfected with sh-FABP4 or short hairpin (sh)-negative control (NC), forming two groups with 6 mice each. The in vivo mice tumor model allowed for evaluating FABP4's impact on tumor growth. FABP4 was significantly upregulated in colorectal cancer tissues and cells (p < 0.05). FABP4 knockdown markedly inhibited cell proliferation, stemness, and glycolysis, while promoting apoptosis in these cells (p < 0.05). Additionally, FABP4 depletion led to a significant increase in ROS level (p < 0.05). However, N-acetyl-L-cysteine (NAC) (p < 0.05), a ROS scavenger, mitigates these effects. Furthermore, the effects of FABP4 depletion on cell growth, stemness, glycolysis, and apoptosis in colorectal cancer cells were also retarded by NAC (p < 0.05). Notably, FABP4 knockdown also suppressed the ERK/mTOR pathway, suggesting its regulation via ROS (p < 0.05). In vivo study results showed, FABP4 depletion significantly curbed tumor growth in colorectal cancer (p < 0.05). These results suggest that FABP4 depletion inhibits colorectal cancer progression by modulating cell growth, stemness, glycolysis and apoptosis. This regulation occurs through the ROS/ERK/mTOR pathway.

A novel ROS-Related chemiluminescent semiconducting polymer nanoplatform for acute pancreatitis early diagnosis and severity assessment

Acute pancreatitis (AP) is a common and potentially life-threatening inflammatory disease of the pancreas. Reactive oxygen species (ROS) play a key role in the occurrence and development of AP. With increasing ROS levels, the degree of oxidative stress and the severity of AP increase. However, diagnosing AP still has many drawbacks, including difficulties with early diagnosis and undesirable sensitivity and accuracy. Herein, we synthesized a semiconducting polymer nanoplatform (SPN) that can emit ROS-correlated chemiluminescence (CL) signals. The CL intensity increased in solution after optimization of the SPN. The biosafety of the SPN was verified in vitro and in vivo. The mechanism and sensitivity of the SPN for AP early diagnosis and severity assessment were evaluated in three groups of mice using CL intensity, serum marker evaluations and hematoxylin and eosin staining assessments. The synthetic SPN can be sensitively combined with different concentrations of ROS to produce different degrees of high-intensity CL in vitro and in vivo. Notably, the SPN shows an excellent correlation between CL intensity and AP severity. This nanoplatform represents a superior method to assess the severity of AP accurately and sensitively according to ROS related chemiluminescence signals. This research overcomes the shortcomings of AP diagnosis in clinical practice and provides a novel method for the clinical diagnosis of pancreatitis in the future.

Effects of Lycopene on Sperm Parameter in Healthy Male: A Review

Infertility is an ability of couple to have a baby after one year of unprotected intercourse affects 48.5million couple in worldwide. Out one of four couple are affected by infertility. A minimum 30 million men world-wide are infertile and male factor are responsible for approximately 20-30% in all infertile cases. It is now understood that oxidative stress plays a significant role in the aetiology of male infertility. Reactive oxygen species (ROS) in excess lead to an oxidative stress condition that reduces sperm viability and motility by causing DNA damage, apoptosis, and lipid peroxidation of the sperm membrane. Increased ROS levels are a crucial role in idiopathic male factor infertility, which is a disease that is becoming more prevalent in society today. Due to its antioxidant qualities, lycopene, the most effective singlet oxygen quencher of all carotenoids, may be used to treat male infertility. Lycopene may minimise the occurrence of oxidative stress and, consequently, the harm that would otherwise be done to spermatozoa by interacting with and neutralising free radicals. We have analysed data from a number of sources and provided information regarding lycopene and its impact on male infertility in this review paper. The focus of this review is on how lycopene affects sperm parameters in healthy men.

Anti-leishmanial activity of sanguinarine and nano chitosan is modulated by increased ROS production and upregulated TNF-α and iNOS expression

The current study aimed to evaluate the anti-leishmanial activity of sanguinarine and nano chitosan (CS NPs) on Leishmania major, potentially mediated through gene expression of TNF-α and iNOS. To identify the potential targets for sanguinarine, the energy-minimized structure was docked using AutoDock 4.3 software into the active sites of selected peterdin reductase (PTR1) and arginase (ARG-L), which were modeled using homology modeling programs. The colorimetric MTT assay was used to determine the viability of L. major promastigotes subjected to different concentrations of sanguinarine and CS NPs in a concentration- and time-dependent manner. Reactive oxygen species (ROS) levels were measured in treated promastigotes. TNF-α and iNOS expression was determined in PBMCs infected at 24, 48, and 72 h after treatment with sanguinarine and CS NPs. The sanguinarine - PTR1 and ARG-L interactions and calculated energy of these enzymes indicated an excellent affinity for sanguinarine with both enzymes. The IC50 values of the sanguinarine in combination with 10% CS NPs) were 124.76 and 89.25 μM after 24 and 48 h, respectively. Sanguinarine-CS NPs enhanced the expression of TNF-α and iNOs in L. major-infected PBMCs after 24 h (p < 0.05). Sanguinarine-CS NPs could therefore reflect a promising anti-leishmanial agent. Sanguinarine and CS NPs in combination significantly inhibited the proliferation and viability of L.major promastigotes whilst concurrently regulating the expression of TNF-α and iNOS and increasing ROS levels. To better understand the effect of CS NPs and sanguinarine on leishmaniasis, further studies are required in an experimental murine model.

Enhanced mitochondrial fission inhibits triple-negative breast cancer cell migration through an ROS-dependent mechanism

Mitochondria produce reactive oxygen species (ROS), which function in signal transduction. Mitochondrial dynamics, encompassing morphological shifts between fission and fusion, can directly impact ROS levels in cancer cells. In this study, we identified an ROS-dependent mechanism for how enhanced mitochondrial fission inhibits triple negative breast cancer (TNBC) cell migration. We found that enforcing mitochondrial fission in TNBC resulted in an increase in intracellular ROS levels and reduced cell migration and the formation of actin-rich migratory structures. Consistent with mitochondrial fission, increasing ROS levels in cells inhibited cell migration. Conversely, reducing ROS levels with either a global ormitochondrially targeted scavenger overcame the inhibitory effects of mitochondrial fission. Mechanistically, we found that the ROS sensitive SHP-1/2 phosphatases partially regulate inhibitory effects of mitochondrial fission on TNBC migration. Overall, our work reveals the inhibitory effects of ROS in TNBC and supports mitochondrial dynamics as a potential therapeutic target for cancer.

Acetaminophen causes mitochondrial dysfunction in heart cardiomyocytes

Acetaminophen (APAP) is a common over-the-counter medication used to treat pain and fever. It can be taken orally, topically, or intravenously and is considered a safe and effective drug when taken at therapeutic doses. However, recent research suggests that taking APAP regularly can result in increased blood pressure and cardiovascular risk. Previous publications found that Non-Steroidal-Anti-Inflammatory-Drugs (NSAIDs) increased Reactive Oxygen Species (ROS) in cardiomyocytes, resulting in cardiac dysfunction. Although APAP is not a NSAID, APAP likely has similar effects. We hypothesized that regular APAP usage resulted in increased ROS levels, leading to potential dysfunction in hearts and other organs. We used the embryonic rat heart cell line H9C2 to test our hypothesis. We conducted a ROS Assay using H9C2 cells treated with dichlorodihydrofluorescein diacetate (DFCDA). The cells were then treated with vehicle, various concentrations of APAP (25μM, 50μM, 100μM, and 200μM), or 100μM hydrogen peroxide for 1.5 hours before measuring the ROS levels of the cells. We also measured the cell viability of H9C2 cells. Cells were treated with 10μM Alamar Blue, then treated with vehicle, various concentrations of APAP (25μM, 50μM, 100μM, and 200μM), or 100μM or 200μM hydrogen peroxide for 48 hours before detecting the percentage of viable cells. The mitochondrial membrane potential (MMP) of cells treated with APAP was also measured. Cells were treated with vehicle, various concentrations of APAP (25μM, 50μM, 100μM, and 200μM), or 250μM p-triflouromethoxyphenylhydrazone (FCCP) for 24 hours before being incubated with JC-10 for one hour. The cells were measured using a fluorescent reader at excitation/emission 490/525 nm and 540/590 nm. For the ROS assay, the cells treated with 100μM APAP, 200μM APAP, and 100μM Hydrogen Peroxide significantly increased total ROS compared to the control. The cell viability assay resulted in a significant decrease in percent viability in cells treated with 200μM APAP, 100μM hydrogen peroxide, and 200μM hydrogen peroxide compared to the control. The MMP Assay resulted in a significant decrease in MMP at all concentrations of APAP and 250μM FCCP compared to the control. These results suggest that treatment of H9C2 cardiac cells with physiological concentrations of APAP causes an increase in intracellular ROS levels, decreased cell viability, and decreased MMP. Overall, APAP causes mitochondrial dysfunction, which is likely to cause cardiomyocyte dysfunction. This research is funded by the NIEHS/Superfund Research Program (P42 ES004699). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

LKB1 Regulates Inflammation of Fibroblast-like Synoviocytes from Patients with Rheumatoid Arthritis via AMPK-Dependent SLC7A11-NOX4-ROS Signaling.

Fibroblast-like synoviocytes (FLS) in rheumatoid arthritis (RA) patients have increased reactive oxygen species (ROS) levels and an impaired redox balance compared with FLS from control patients. Liver kinase B1 (LKB1) plays a key role in ROS scavenging and cellular metabolism in various cancers. Here, we aimed to determine the specific mechanism of LKB1 in RA pathogenesis. FLS were obtained from RA patients (n = 10). siRNA-induced LKB1 deficiency in RA FLS increased ROS levels via NADPH oxidase 4 (NOX4) upregulation. RA FLS migration and expression of inflammatory factors, including interleukin (IL)-1β, IL-6, IL-8, tumor necrosis factor-alpha (TNF-α), and vascular endothelial growth factor (VEGF), were enhanced by LKB1 deficiency. LKB1-deficient RA FLS showed increased sensitivity to oxidative stress damage caused by hydrogen peroxidase exposure. siRNA-induced solute carrier family 7 member 11 (SLC7A11) deficiency in RA FLS enhanced NOX4 and ROS expression and increased cell migration. When LKB1-deficient RA FLS were stimulated with an AMP-activated protein kinase (AMPK) activator, the LKB1-inhibition-induced cell migration significantly decreased through the restoration of SLC7A11/NOX4 expression. LKB1 regulates the AMPK-mediated SLC7A11-NOX4-ROS pathway to control cell migration and inflammation. Our data indicate that LKB1 is a key regulator of redox homeostasis in RA FLS.

Effect of Nisin on the Quality and Antioxidant Activity of Fresh-Cut Pumpkins (Cucurbita moschata Duch.)

Fresh-cut pumpkins refer to fresh pumpkin that has been graded, cleaned, peeled, sliced, preserved, and packaged. It has the qualities of freshness, nutrition, convenience, and being 100% edible. However, mechanical damages during the cutting processing can accelerate the quality deterioration, aging, and loss of nutritional values of fresh-cut pumpkins. Nisin, a natural preservative, has been widely used in fruits and vegetables with good preservation effects. To investigate the effect of different concentrations (0, 0.2, 0.4, and 0.6 g/L) of nisin on the quality of fresh-cut pumpkins, the critical indexes involved in weight loss, firmness, color, respiration intensity, reactive oxygen species (ROS) metabolism, ascorbate (AsA)—glutathione (GSH) cycle, and antioxidant capacity were monitored for fresh-cut pumpkins during storage at 4 °C for 10 days. The results showed that 0.4 g/L nisin was the best preservation concentration. Compared with 0 g/L nisin, 0.4 g/L nisin reduced the weight loss rate and whitening rate of fresh-cut pumpkins by 13.53% and 13.61%, inhibited respiration rate by 45.83%, and maintained hardness by 1.18 times. Meanwhile, 0.4 g/L nisin increased the activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) and maintained higher contents of GSH and AsA. It prevented the rapid increase in ROS levels by improving antioxidant capacity, including DPPH, ABTS free radical scavenging rate, and T-AOC (total antioxidant capacity). The collected results showed that nisin has an obvious influence on the quality by regulating physiological and antioxidant activity metabolism. It is envisaged that the combination of nisin and physical and chemical preservation technology will further enhance the quality of fresh-cut pumpkins during storage in the future.

Concurrent induction of apoptosis and necroptosis inapigenin‑treated malignant mesothelioma cells: Reversal ofWarburg effect through Akt inhibition and p53 upregulation.

A high dependence on aerobic glycolysis, known as the Warburg effect, is one of the metabolic features exhibited by tumor cells. Therefore, targeting glycolysis is becoming a very promising strategy for the development of anticancer drugs. In the present study, it was investigated whether pre‑adaptation of malignant mesothelioma (MM) cells to an acidic environment was associated with a metabolic shift to the Warburg phenotype in energy production, and whether apigenin targets acidosis‑driven metabolic reprogramming. Cell viability, glycolytic activity, Annexin V‑PE binding activity, reactive oxygen species (ROS) levels, mitochondrial membrane potential, ATP content, western blot analysis and spheroid viability were assessed in the present study. MM cells pre‑adapted to lactic acid were resistant to the anticancer drug gemcitabine, increased Akt activation, downregulated p53 expression, and upregulated rate‑limiting enzymes in glucose metabolism compared with their parental cells. Apigenin treatment increased cytotoxicity, Akt inactivation and p53 upregulation. Apigenin also reduced glucose uptake along with downregulation of key regulatory enzymes in glycolysis, increased ROS levels with loss of mitochondrial membrane potential, and downregulated the levels of complexes I, III and IV in the mitochondrial electron transport chain with intracellular ATP depletion, resulting in upregulation of molecules mediating apoptosis and necroptosis. Apigenin‑induced alterations of cellular responses were similar to those of Akt inactivation by Ly294002. Overall, the present results provide mechanistic evidence supporting the anti‑glycolytic and cytotoxic role of apigenin via inhibition of the PI3K/Akt signaling pathway and p53 upregulation.

Discovery and Mechanism of Azatryptanthrin Derivatives as Novel Anti-Phytopathogenic Bacterial Agents for Potent Bactericide Candidates.

The natural alkaloids of tryptanthrin and their derivatives have a wide range of biological activities. In this research, four series of azatryptanthrin derivatives containing 4-aza/3-aza/2-aza/1-aza tryptanthrin were prepared by condensation cyclization reaction against plant pathogens to develop a new natural product-based bacterial pesticide. Compound 4Aza-8 displayed a remarkable growth inhibitory effect on pathogenic bacteria of Xanthomonas axonopodis pv. citri (Xac), Xanthomonas oryzae pv. Oryzae (Xoo), and Pseudomonas syringae pv. actinidiae (Psa) with the final corrected EC50 values of 0.312, 1.91, and 18.0 μg/mL, respectively, which were greatly superior than that of tryptanthrin (Tryp). Moreover, 4Aza-8 also showed effective therapeutic and protective activities in vivo on citrus canker. Further mechanism studies on Xac elucidated that compound 4Aza-8 was able to affect the growth curve of Xac and the formation of biofilm, cause severe shrinkage in bacterial morphology, increase reactive oxygen species levels, and induce apoptosis in bacterial cells. Quantitative analysis of differential protein profiles found that the major differences were mainly concentrated on the endometrial protein in the bacterial secretion system pathway, which blocked the membrane transport and affected the transfer of DNA to the host cell. In summary, these research results suggest that 4Aza-8 represents a promising anti-phytopathogenic-bacteria agent, which is worth being further investigated as a bactericide candidate.

Zoledronic acid induces ferroptosis by upregulating POR in osteosarcoma.

Osteosarcoma, usually originating in the stroma, is the most common primary bone cancer in adolescents, and its prognosis is poor. Surgery,adjuvant and neoadjuvant chemotherapy andradiationtherapyare not satisfactoryatthepresenttime. Therefore, itiscriticalto develop novel therapeutic strategies to improve the quality of life and long-term survival rate of osteosarcoma patients. In this study, we discovered that zoledronic acid (ZOL) dramatically increased cell deathin osteosarcoma cells, and this cytotoxicity was greatly reversed byliproxstatin-1 (a ferroptosis inhibitor). ZOL alsohadanobviouseffectonlipid peroxidation and reactive oxygen species(ROS), which suggested that ZOL most certainly induces ferroptosis in osteosarcoma cells. In addition, we further found thatZOLincreasescytochrome P450 oxidoreductase (POR) expression dose dependently in osteosarcoma cell lines. Knockdown of POR attenuated ZOL-induced cytotoxicity and attenuated the effect of ferroptosis in osteosarcoma cells, which indicated that POR plays an important role in ferroptosis. Moreover, we also found that ZOL inhibits osteosarcoma growth in vivo. Our findings suggest that ZOL induces ferroptosis by upregulating POR expression to increase ROS levels and upregulate lipid peroxidation levels in osteosarcoma cells. PORmay be used asa therapeutictarget to inhibit osteosarcoma.

Nanoplastics impacts on Thiobacillus denitrificans: Effects of size and dissolved organic matter

The widespread distribution of nanoplastics and dissolved organic matter (DOM) in sewage raises concerns about the potential impact of DOM on the bioavailability of nanoplastics. In this study, the effects of different sizes (100 nm and 350 nm) of polystyrene nanoplastics (PS-NPs, 50 mg/L) and combined with 10 mg/L or 50 mg/L DOMs (fulvic acid, humic acid and sodium alginate) on the growth and denitrification ability of Thiobacillus denitrificans were investigated. Results showed that 100 nm PS-NPs (50 mg/L) cause a longer delay in the nitrate reduction (3 days) of T. denitrificans than 350 nm PS-NPs (2 days). Furthermore, the presence of DOM exacerbated the adverse effect of 100 nm PS-NPs on denitrification, resulting in a delay of 1–4 days to complete denitrification. Fulvic acid (50 mg/L) and humic acid (50 mg/L) had the most significant adverse effect on increasing 100 nm PS-NPs (50 mg/L), causing a reduction of 20 mmol/L nitrate by T. denitrificans in nearly 7 days. It is noteworthy that the presence of DOM did not modify the adverse effect of 350 nm PS-NPs on denitrification. Further analysis of toxicity mechanism of PS-NPs revealed that they could induce reactive oxygen species (ROS) and suppressed denitrification gene expression. The results suggested that DOM may assist in the cellular internalization of PS-NPs by inhibiting PS-NPs aggregation, leading to the increased ROS levels and accelerated T. denitrificans death. This study highlights the potential risk of nanoplastics to autotrophic denitrifying bacteria in the presence of DOM and provides new insights for the treatment of nitrogen-containing wastewater by T. denitrificans.

Abstract 271: MYC mediates enhanced lactate reutilization and resistance to anti-angiogenesis therapy in preclinical models of LKB1-deficient NSCLC

Abstract Deletion or loss of function mutations of the STK11/LKB1 tumor suppressor are associated with primary resistance to immunotherapy in KRAS-mutant lung adenocarcinoma (LUAD) and drive metabolic reprogramming of tumor cells. We observed that LKB1-deficient tumors were resistant to anti-angiogenic therapy in the hypoxic and nutrient-depleted or acidic tumor microenvironment (TME). We determined that MYC which is elevated in LKB1-deficient cells, regulated the expression of the lactate transporter, MCT4. Moreover, knockdown of MYC decreases glycolysis and cell proliferation. Therefore, we hypothesize that metabolic changes in LKB1-deficient tumor cells is at least partially driven by MYC upregulation, and that depletion of MYC or targeting of key enzymes or transporters downstream of MYC such as MCT4 may abrogate lactate reutilization and sensitize LKB1-deficient tumors to anti-angiogenic therapy. To explore the impact of MYC on lactate metabolism in LKB1-deficient NSCLC cells, we performed isotope tracing in KRAS mutant (K) and KRAS mutant LKB1 knockout (KL) syngeneic murine lung cancer cells. 13C3-lactate tracing revealed that isotopologues were significantly enriched in TCA components such as pyruvate, citrate and a-ketoglutarate in KL cells. Meanwhile, lactate-treated KL cells shows less reactive oxygen species (ROS), suggesting that LKB1-deficient tumor cells reutilize lactate as an energy source more efficiently than LKB1-intact cells. MCT4 KO significantly abrogated lactate incorporation into TCA cycle. Similarly, MYC knockdown or MCT4 KO decreased lactate-induced oxygen consumption, but increased ROS levels. In contrast, exhausted T cells were inefficient at lactate reutilization, and high levels of lactate increased ROS in T cells. Hence, LKB1-deficient tumors have a survival advantage over T cells in the lactate-rich TME. We queried scRNAseq data from K, KL and KL MCT4KO syngeneic models and similarly observed that KL tumor cells exhibited elevated hypoxia and angiogenesis gene expression signatures, which was reversed by MCT4 KO. However, ROS detoxification was decreased in T cells from KL tumors but increased in MCT4 KO tumors. Finally, we injected KL murine tumor cells into immunocompetent mice, and randomly treated them with vehicle or the VEGF blocking antibody, DC101. MYC knockdown or MCT4 KO sensitized KL tumors to VEGF inhibition, and significantly increased overall survival. Collectively, our data indicates that in LKB1-deficient tumors, upregulation of MYC promotes tumor cell metabolic reprogramming and that targeting MYC or MCT4 can inhibit lactate reutilization and enhance the efficacy of anti-angiogenic agents. These findings provide insight into the mechanisms driving the aggressive phenotype of KRAS-mutant LKB1-deficient tumors and identify a novel therapeutic strategy for targeting this patient population. Citation Format: Yu Qian, David Molkentine, Chendong Yang, Ana Galan Cobo, Irene Guijarro, Minghao Dang, Alissa Poteete, Peixin Jiang, Ferdinandos Skoulidis, Linghua Wang, Alexandre Reuben, John D. Minna, Ralph J. DeBerardinis, John V. Heymach. MYC mediates enhanced lactate reutilization and resistance to anti-angiogenesis therapy in preclinical models of LKB1-deficient NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 271.

Dietary iodine attenuates allergic rhinitis by inducing ferroptosis in activated B cells

Iodine-containing formulations have been widely used to treat iodine deficiency and as antiseptics. Lecithin-bound iodine (LBI) has been approved to treat allergic diseases in Japan; however, its underlying mechanism remains unknown. In this study, we show that LBI ameliorated disease symptoms in an ovalbumin (OVA)-induced allergic rhinitis mouse model. LBI suppressed OVA-specific IgE production by attenuating germinal center (GC) reaction in the draining lymph nodes. The antiallergic effect of LBI is most likely attributed to increased serum iodine levels but not thyroid hormone levels. In vitro treatment of activated B cells with potassium iodide induced ferroptosis by increasing intracellular reactive oxygen species (ROS) and ferrous iron in a concentration-dependent manner. Accordingly, LBI diets increased ROS levels in GC B cells of the draining lymph nodes. This study suggests that iodine directly promotes ferroptosis in activated B cells and attenuates GC reactions, leading to the alleviation of allergic symptoms.

Ethanolic extract of Ocimum sanctum leaf modulates oxidative stress, cell cycle and apoptosis in head and neck cancer cell lines

Ocimum sanctum Linn. is a medicinal herb that has cytotoxic effects by inducing oxidative stress in some carcinomas. This study aimed to examine the impact of O. sanctum leaf extract on oxidative stress, cell cycle progression, and apoptosis in cell lines of head and neck squamous cell carcinoma (HNSCC). Isogenic primary (HN18/HN30) and metastatic (HN17/HN31) HNSCC cell lines were used. Preparation of the ethanolic extract of O.sanctum leaf (EEOS) was carried out. HNSCC cell lines were exposed to varying concentrations (0.1–0.8 mg/ml) of EEOS for a duration of 72 h, and the MTT assay was utilized to determine the cytotoxic doses. To assess the impact of EEOS on HNSCC cells, the levels of reactive oxygen species (ROS) and malondialdehyde were measured using a fluorometric method. Flow cytometry was utilized to evaluate effects of EEOS on the cell cycle, DNA damage, and apoptosis in HNSCC cells. Caspase-3 and -9 levels in the EEOS-treated HNSCC cells were measured by ELISA. The chemical components in EEOS were detected using high-performance liquid chromatography-electrospray ionization-time of flight-mass spectrometry. EEOS exhibited cytotoxicity against the HN18, HN17, HN30 and HN31 cells at minimum concentrations of 0.1, 0.3, 0.2 and 0.2 mg/ml, respectively. Treatment with EEOS resulted in a significant increase in ROS levels in HN18 and HN17 cells. Additionally, EEOS significantly induced the levels of malondialdehyde in HN18 and HN31 cells. Moreover, EEOS arrested the cell cycle in HN30 and HN31 cells, and significantly induced DNA damage and apoptosis in the HN18, HN30, and HN31 cells. EEOS selectively increased caspase-9 in the HN18 cells. However, caspase-3 was activated without apoptosis in the EEOS-treated HN17 cells. The constituents of EEOS were identified as rosmarinic acid, caffeic acid, and apigenin. In conclusion, EEOS exhibits various prooxidative and apoptotic effects between HNSCC cells.

P20BAP31 induces cell apoptosis via both AIF caspase-independent and the ROS/JNK mitochondrial pathway in colorectal cancer

BackgroundDuring cell apoptosis, the C-terminus of BAP31 is cleaved by caspase-8 and generates p20BAP31, which has been shown to induce an apoptotic pathway between the endoplasmic reticulum (ER) and mitochondria. However, the underlying mechanisms of p20BAP31 in cell apoptosis remains unclear.MethodsWe compared the effects of p20BAP31 on cell apoptosis in six cell lines and selected the most sensitive cells. Functional experiments were conducted, including Cell Counting Kit 8 (CCK-8), reactive oxygen species (ROS), and mitochondrial membrane potential (MMP) assay. Then, cell cycle and apoptosis were investigated by flow cytometry and verified by immunoblotting. Next, NOX inhibitors (ML171 and apocynin), ROS scavenger (NAC), JNK inhibitor (SP600125), and caspase inhibitor (Z-VAD-FMK) were used to further investigate the underlying mechanisms of p20BAP31 on cell apoptosis. Finally, apoptosis-inducing factor (AIF) translocation from the mitochondria to the nuclei was verified by immunoblotting and immunofluorescence assay.ResultsWe found that overexpression of p20BAP31 indeed induced apoptosis and had a much greater sensitivity in HCT116 cells. Furthermore, the overexpression of p20BAP31 inhibited cell proliferation by causing S phase arrest. Further study revealed that p20BAP31 reduced MMP, with a significant increase in ROS levels, accompanied by the activation of the MAPK signaling pathway. Importantly, the mechanistic investigation indicated that p20BAP31 induces mitochondrial-dependent apoptosis by activating the ROS/JNK signaling pathway and induces caspase-independent apoptosis by promoting the nuclear translocation of AIF.Conclusionsp20BAP31 induced cell apoptosis via both the ROS/JNK mitochondrial pathway and AIF caspase-independent pathway. Compared with antitumor drugs that are susceptible to drug resistance, p20BAP31 has unique advantages for tumor therapy.

Eriocitrin suppresses proliferation and migration of hepatocellular carcinoma SMMC-7721 cells by promoting ROS production and activating the MAPK pathway

To investigate the role of the ROS/MAPK signaling axis in mediating the inhibitory effect of eriocitrin on proliferation and migration of hepatocellular carcinoma SMMC-7721 cells. SMMC-7721 cells were treated with different concentrations of eriocitrin for 24 h, and the changes in cell viability were detected with CCK-8 assay. The migration and invasion abilities of the treated cells were evaluated using Transwell and scratch healing assays, the cell proliferation was assessed with colony-forming assay, and changes in nuclear morphology were observed with DAPI staining. Western blotting was performed to examine the changes in the expressions of E-cadherin, N-cadherin, MMP-2, MMP-9, PARP, Pro-caspase 3, pJNK, p-P38, and p-ERK. The effect of eriocitrin on PARP cleavage in SMMC-7721 cells pretreated with ERK, JNK and P38 inhibitors (U0126, SB203580 and SP600125, respectively) was detected using Western blotting. The effect of treatment with Nacetyl-cysteine (NAC, 30 μmol/L) and eriocitrin (100, 200, and 300 μg/mL), alone or in combination, on reactive oxygen species (ROS) levels in the cells was examined using a DCFH-DA fluorescent probe. Eriocitrin below 50 μg/mL did not produce significant effect on the viability of SMMC-7721 cells (P>0.05). Treatment with eriocitrin significantly inhibited scratch healing, migration, and colony formation of the cells (P < 0.01), reduced the protein expressions of N-cadherin, MMP-2, and MMP-9 (P < 0.01), and up-regulated E-cadherin protein expression (P < 0.05). Eriocitrin-treated SMMC-7721 cells showed obvious apoptotic morphologies with decreased Procaspase 3 expression and increased PARP cleavage (P < 0.01) and phosphorylation levels of JNK, P38, and ERK (P < 0.01); Eriocitrin-induced PAPR cleavage was obviously enhanced by U0126 and SB203580 but attenuated by SP600125. Treatment with 300 μg/mL eriocitrin for 30 min significantly increased ROS level in the cells, and this effect was obviously suppressed by NAC. Eriocitrin can suppress the proliferation and migration and promote apoptosis of hepatocellular carcinoma SMMC-7721 cells by promoting ROS production and activating the MAPKs signaling pathway.