Reactive Oxygen Species-dependent Manner Research Articles

Arnicolide D Inhibits Oxidative Stress-induced Breast Cancer Cell Growth and Invasion through Apoptosis, Ferroptosis, and Parthanatos.

Breast cancer is the most common malignant tumor in women, and its pathogenesis is very complicated. More and more studies have found that Traditional Chinese Medicine plays an important role in tumor prevention. To investigate the mechanism of arnicolide D isolated from Centipeda minima in breast cancer. Cell Counting Kit-8 (CCK-8), western blot, RT-qPCR, ELISA, flow cytometry, and Transwell were used to detect the effect of arnicolide D on the biological function of breast cancer cells. Arnicolide D promoted reactive oxygen species (ROS) production and induced a decrease in mitochondrial membrane potential in breast cancer cells, thereby inhibiting cell viability and increasing lactate dehydrogenase (LDH) release. Arnicolide D activated the classical apoptosis pathway to induce cell apoptosis; it significantly promoted PARP-1 expression, enhanced the nuclear translocation of apoptosis-inducing factor (AIF), and reduced the expression of AIF in mitochondria, indicating that it can induce the occurrence of parthanatos in a ROS dependent manner. In addition, arnicolide D down-regulated glutathione peroxidase 4 (GPX4) expression and increased the accumulation of Fe2+ and malondialdehyde (MDA), thereby activating ferroptosis. Apoptosis inhibitor, ferroptosis inhibitor, PARP inhibitor, PARP-1 siRNA, AIF siRNA and GPX4 overexpression vector significantly attenuated the inhibitory effect of arnicolide D on cell viability and reduced LDH release, which indicates that arnicolide D inhibits breast cancer cell growth by inducing apoptosis, parthanatos and ferroptosis. Arnicolide D also reduced breast cancer cell invasion and inhibited the expression of matrix metallopeptidase (MMP)-2 and MMP-9. Arnicolide D can activate a variety of cell death modes by inducing oxidative stress, thereby inhibiting the growth and invasion of breast cancer cells, indicating that arnicolide D has a good anti-tumor effect.

STING1-accelerated vascular smooth muscle cell senescence-associated vascular calcification in diabetes is ameliorated by oleoylethanolamide via improved mitochondrial DNA oxidative damage

Vascular calcification is a prevalent hallmark of cardiovascular risk in elderly and diabetic individuals. Senescent vascular smooth muscle cells (VSMCs) participate in calcification; however, the associated underlying mechanisms remain unknown. Aberrant activation of the cytosolic DNA sensing adaptor stimulator of interferon gene 1 (STING1) caused by cytosolic DNA, particularly that leaked from damaged mitochondria, is a catalyst for aging-related diseases. Although oleoylethanolamide (OEA) is an endogenous bioactive lipid mediator with lipid overload-associated vasoprotective effects, its benefit in diabetic vascular calcification remains uncharacterized. This study focused on the role of STING1 in mitochondrial dysfunction-mediated calcification and premature VMSC senescence in diabetes and the effects of OEA on these pathological processes. In diabetic in vivo rat/mouse aorta calcification models and an in vitro VSMC calcification model induced by Nε-carboxymethyl-lysine (CML), senescence levels, STING1 signaling activation, and mitochondrial damage markers were significantly augmented; however, these alterations were markedly alleviated by OEA, partially in a nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent manner, and similar anti-calcification and senescence effects were observed in STING1-knockout mice and STING1-knockdown VSMCs. Mechanistically, mitochondrial DNA (mtDNA) damage was aggravated by CML in a reactive oxygen species-dependent manner, followed by mtDNA leakage into the cytosol, contributing to VSMC senescence-associated calcification via STING1 pathway activation. OEA treatment significantly attenuated the aforementioned cytotoxic effects of CML by enhancing cellular antioxidant capacity through the maintenance of Nrf2 translocation to the nucleus. Collectively, targeting STING1, a newly defined VSMC senescence regulator, contributes to anti-vascular calcification effects.

Synergistic cytotoxicity effect of ultrasound and plasma microbubble in glioblastoma 3D tumor sphere model

AbstractThe combined application of ultrasound (US) and plasma microbubble (PMB) is a promising technology against glioblastoma (GBM). Here, we determined the effects of US and PMB (alone and combined) on U‐251 MG GBM and A431 epidermoid carcinoma spheroids under different conditions. The synergistic treatment of US‐ and PMB‐induced tumor sphere cell death in a time‐, dose‐, treatment frequency‐, and reactive oxygen species dependent manner, while multiple treatments augmented this effect. Synergistic treatments significantly reduced three‐dimensional GBM spheroid regrowth and cell proliferation while inducing cytotoxicity, spheroid shrinkage, and damage to the tumor microenvironment. PMB‐induced cytotoxicity throughout the tumor sphere, likely via long‐lived reactive oxygen and nitrogen species (H2O2, NO2−, and NO3−) and other reactive species.

Omeprazole induces profibrotic gene expression in rat kidney: implication of TGF-β/Smad signaling pathway

Proton pump inhibitors (PPIs) are one of the most commonly prescribed medications. However, PPI usage is linked to a higher risk of both acute and chronic renal damage by mechanisms not entirely known. The present study demonstrates that omeprazole (10 mg/kg body weight, i.p.) causes TGF-β/Smad signaling activation and subsequent expression of the profibrotic genes CTGF and TIMP-1 in rat kidney. Increased production of CTGF and TIMP-1 accompany activation of the TGF-β/Smad signaling cascade. However, simultaneous treatment of omeprazole and the TGF-β inhibitor, disitertide (P144) (1 mg/kg body weight i.p.) suppresses the TGF-β/Smad signaling pathway and subsequent production of CTGF and TIMP-1. Additionally, TGF-β level in rat kidney was highly reduced in animals treated with the ROS (reactive oxygen species) scavenger, N-acetyl cysteine (NAC) (100 mg/kg body weight i.p.) before omeprazole administration. Furthermore, the reduction in SOD activity brought by omeprazole was returned to the normal level in those animals. However, MDA level increased by omeprazole was highly reduced in the presence of NAC. Collectively, the current findings demonstrate that omeprazole has the ability to promote the expression of the profibrotic genes CTGF and TIMP-1 in a ROS and TGF-β dependent manner. The present study suggests the co-use of ROS scavenger to improve the therapeutic use of the PPI omeprazole.

Retracted: Isolation and purifications of an ambuic acid derivative compound from marine algal endophytic fungi Talaromyces flavus that induces apoptosis in MDA-MB-231 cancer cells.

In recent years, there has been a lot of buzz about the possibilities of marine microflora as a source of new therapeutic drugs. The strong anti-tumor potency of compounds found in marine resources reflects the ocean's enormous potential as a source of anticancer therapeutics. In this present investigation, an ambuic acid derivative anticancer compound was isolated from Talaromyces flavus, and its cytotoxicity and apoptosis induction potential were analyzed. T. flavus was identified through morphological and molecular analysis. The various organic solvent extracts of T. flavus grown on different growth mediums were evaluated for cytotoxicity on different cancer cell lines. The potent cytotoxicity was shown in the ethyl acetate extract of a fungal culture grown in the M1-D medium for 21 days. Furthermore, the anticancer compound was identified using preparative thin layer chromatography, followed by its purification in significant proportions using column chromatography. The spectroscopic and chromatographic analysis revealed that the structure of the purified molecules was an ambuic acid derivative. The ambuic acid derivative compound showed potent cytotoxicity on MDA-MB-231 (breast cancer cells) with an IC50 value of 26 μM and induced apoptosis in the MDA-MB-231 cells in a time-dependent and reactive oxygen species-independent manner.

Enhanced IRE1α Phosphorylation/Oligomerization-Triggered XBP1 Splicing Contributes to Parkin-Mediated Prevention of SH-SY5Y Cell Death under Nitrosative Stress.

Mutations in parkin, a neuroprotective protein, are the predominant cause of autosomal recessive juvenile Parkinson's disease. Neuroinflammation-derived nitrosative stress has been implicated in the etiology of the chronic neurodegeneration. However, the interactions between genetic predisposition and nitrosative stress contributing to the degeneration of dopaminergic (DA) neurons remain incompletely understood. Here, we used the SH-SY5Y neuroblastoma cells to investigate the function of parkin and its pathogenic mutants in relation to cell survival under nitric oxide (NO) exposure. The results showed that overexpression of wild-type parkin protected SH-SY5Y cells from NO-induced apoptosis in a reactive oxygen species-dependent manner. Under nitrosative stress conditions, parkin selectively upregulated the inositol-requiring enzyme 1α/X-box binding protein 1 (IRE1α/XBP1) signaling axis, an unfolded protein response signal through the sensor IRE1α, which controls the splicing of XBP1 mRNA. Inhibition of XBP1 mRNA splicing either by pharmacologically inhibiting IRE1α endoribonuclease activity or by genetically knocking down XBP1 interfered with the protective activity of parkin. Furthermore, pathogenic parkin mutants with a defective protective capacity showed a lower ability to activate the IRE1α/XBP1 signaling. Finally, we demonstrated that IRE1α activity augmented by parkin was possibly mediated through interacting with IRE1α to regulate its phosphorylation/oligomerization processes, whereas mutant parkin diminished its binding to and activation of IRE1α. Thus, these results support a direct link between the protective activity of parkin and the IRE1α/XBP1 pathway in response to nitrosative stress, and mutant parkin disrupts this function.

Immunoregulatory CD71+ Erythroid Cells (CECs) Expand in Multiple Myeloma and Impair Control of L. Monocytogenes Infection

Introduction Although the clinical management of multiple myeloma (MM) patients has significantly improved over recent years, MM remains a largely incurable disease. One of the most common complication of MM, which correlates with the worse clinical outcome, is anemia. Moreover, MM patients are prone to develop life-threatening infections mainly due to the disease-related impairment of the immune system function. Recently, CD71+ erythroid progenitor and precursor cells (CECs) arose as a relevant immunomodulatory population. CECs have been shown to create an immunosuppressive microenvironment via multiple mechanisms including arginase 2 (ARG2), PDL-1, and TGF-beta. It was recently observed in our laboratory that mice bearing late-stage tumors are characterized by the presence of an increased percentage of CECs in their blood and spleens. Moreover, the tumor-associated CECs show elevated levels of arginases. The abovementioned facts may suggest an important role of CECs and their related mechanisms in compromising the immune response in MM. Methods Vk*MYC MM model Recipient C57BL/6 (wild type or ARG2 KO) mice were injected via tail vein with 1 × 106 Vk*MYC cells. M-spike levels were evaluated with serum protein electrophoresis. Five weeks later they were challenged with L. monocytogenes. L.monocytogenes infection and bacterial burden evaluation Mice were injected intraperitoneally with 1 x 103L. monocytogenes cells (strain EGDe). Bacterial burden was analyzed 72h and 7 days post infection. Briefly, spleens and livers of sacrificed mice were isolated, weighed, and homogenized. The homogenates were serially diluted and plated on culture plates. Bacterial burden was calculated using the number of colonies and the dilution factor. Flow cytometry analysis Flow cytometry analyses (immunophenotyping of spleen and bone marrow (BM) cells as well as cytokine beads arrays, Legenplex, BioLegend) were performed using BD Fortessa X-20 flow cytometer operated by BD FACSDiva Software ver. 8.0. The acquired data were re-analyzed using FlowJo™ ver. 10 software. Arginase inhibitor OAT-1746 arginase inhibitor (Molecure SA) was administered intraperitoneally at doses of 5 or 20 mg/kg twice daily, starting from the second or the fifth day before the bacterial challenge, respectively. The treatment was continued until the end of the experiment. Results In Vk*MYC-bearing mice we observed an accumulation of CECs in the blood and spleen in time, correlating with the development of anemia. At the late stage of the disease MM-associated CECs showed less differentiated phenotype and increased expression of arginase-1 (ARG1) and ARG2. MM-associated CECs inhibited T cells activation and proliferation in an ARG and reactive oxygen species-dependent manner. Interestingly, MM development was delayed in ARG2 KO vs WT mice. In the BM aspirates of MM patients we observed increased percentage of CECs expressing high levels of both ARG1 and ARG2 comparing with healthy controls. Moreover, MM impaired control of L. monocytogenes infection as we observed an elevated bacterial burden in the livers and spleens of Vk*MYC-bearing vs control mice. Infected MM-bearing mice had fewer NK cells and activated CD4+ and CD8+ T cells in the spleen than infected control mice. Furthermore, MM altered the dynamics of cytokines involved in the control of intracellular infection. 24h post Listeria challenge we observed their significant decrease followed by an increase at 72h comparing to infected non-MM mice. These results indicate that MM-bearing mice have delayed initial immune response leading to increased bacterial burden followed by excessive cytokines response resulting in sepsis-like condition leading to increased mice mortality. Finally, ARG inhibitor decreased L. monocytogenes burden in MM-bearing mice. Altogether, our results show that accumulation of CECs during MM progression leads to impaired bacterial infection control, at least partially dependent on ARG2 activity. Funding: National Science Centre in Poland [grants No 2017/25/B/NZ6/01139 and 2019/35/B/NZ6/00540]; Polish Ministry of Education and Science [grant No 013/RID/2018/19 (Regional Initiative for Excellence, project budget 12,000,000 PLN)].

Drosophila hemocytes recognize lymph gland tumors of mxc mutants and activate the innate immune pathway in a reactive oxygen species-dependent manner.

Mechanisms of cancer cell recognition and elimination by the innate immune system remains unclear. The immune signaling pathways are activated in the fat body to suppress the tumor growth in mxcmbn1 hematopoietic tumor mutants in Drosophila by inducing antimicrobial peptides (AMP). Here, we investigated the regulatory mechanism underlying the activation in the mutant. Firstly, we found that reactive oxygen species (ROS) accumulated in the hemocytes due to induction of dual oxidase and one of its activators. This was required for the AMP induction and the tumor growth suppression. Next, more hemocytes transplanted from normal larvae were associated with the mutant tumor than normal lymph glands (LGs). Matrix metalloproteinase 1 and 2 (MMP2) were highly expressed in the tumors. The basement membrane components in the tumors were reduced and ultimately lost inside. Depletion of the MMP2 rather than MMP1 resulted in a significantly reduced AMP expression in the mutant larvae. The hemocytes may recognize the disassembly of basement membrane in the tumors and activate the ROS production. Our findings highlight the mechanism via which macrophage-like hemocytes recognize tumor cells and subsequently convey the information to induce AMPs in the fat body. They contribute to uncover the role of innate immune system against cancer.

Methamphetamine signals transcription of IL1β and TNFα in a reactive oxygen species-dependent manner and interacts with HIV-1 Tat to decrease antioxidant defense mechanisms.

Methamphetamine (Meth) abuse is a common HIV co-morbidity that is linked to aggravated Central Nervous System (CNS) inflammation, which accentuates HIV- associated neurological disorders, triggered both directly or indirectly by the drug. We used the well-established human innate immune macrophage cell line system (THP1) to demonstrate that Reactive Oxygen Species (ROS) immediately induced by Meth play a role in the increased transcription of inflammatory genes, in interaction with HIV-1 Tat peptide. Meth and Tat, alone and together, affect early events of transcriptional activity, as indicated by changes in RNA polymerase (RNAPol) recruitment patterns throughout the genome, via ROS-dependent and -independent mechanisms. IL1β (IL1β) and TNF α (TNFα), two genes with defining roles in the inflammatory response, were both activated in a ROS-dependent manner. We found that this effect occurred via the activation of the activator protein 1 (AP-1) comprising cFOS and cJUN transcription factors and regulated by the SRC kinase. HIV-1 Tat, which was also able to induce the production of ROS, did not further impact the effects of ROS in the context of Meth, but promoted gene activity independently from ROS, via additional transcription factors. For instance, HIV-1 Tat increased NFkB activation and activated gene clusters regulated by Tata box binding peptide, ING4 and IRF2. Importantly, HIV-1 Tat decreased the expression of anti-oxidant genes, where its suppression of the detoxifying machinery may contribute to the aggravation of oxidative stress induced by ROS in the context of Meth. Our results provide evidence of effects of Meth via ROS and interactions with HIV Tat that promote the transcription of inflammatory genes such as IL1β and TNFα.

Impaired Mitochondrial Transcription Factor A Expression Promotes Mitochondrial Damage to Drive Fibroblast Activation and Fibrosis in Systemic Sclerosis.

Mitochondrial transcription factor A (TFAM) controls the transcription of core proteins required for mitochondrial homeostasis. This study was undertaken to investigate changes in TFAM expression in systemic sclerosis (SSc), to analyze mitochondrial function, and to evaluate the consequences for fibroblast activation. TFAM expression was analyzed by immunofluorescence and Western blotting. The effects of TFAM knockout were investigated in cultured fibroblasts and in murine models of bleomycin-induced skin fibrosis, bleomycin-induced lung fibrosis, and skin fibrosis induced by overexpression of constitutively active transforming growth factor β type I receptor (TGFβRΙ). TFAM expression was down-regulated in fibroblasts in SSc skin and in cultured SSc fibroblasts. The down-regulation of TFAM was associated with decreased mitochondrial number and accumulation of damaged mitochondria with release of mitochondrial DNA (mtDNA), accumulation of deletions in mtDNA, metabolic alterations with impaired oxidative phosphorylation, and release of the mitokine GDF15. Normal fibroblasts subjected to long-term, but not acute, exposure to TGFβ mimicked SSc fibroblasts, with down-regulation of TFAM and accumulation of mitochondrial damage. Down-regulation of TFAM promoted fibroblast activation with up-regulation of fibrosis-relevant Gene Ontology terms in RNA-Seq, partially in a reactive oxygen species-dependent manner. Mice with fibroblast-specific knockout of Tfam were prone to fibrotic tissue remodeling, with fibrotic responses even to NaCl instillation and enhanced sensitivity to bleomycin injection and overexpression of constitutively active TGFβRI. TFAM knockout fostered Smad3 signaling to promote fibroblast activation. Alterations in the key mitochondrial transcription factor TFAM in response to prolonged activation of TGFβ and associated mitochondrial damage induce transcriptional programs that promote fibroblast-to-myofibroblast transition and drive tissue fibrosis.

Neutrophil extracellular traps induce tumor metastasis through dual effects on cancer and endothelial cells

ABSTRACT Neutrophils constitute a major component in human hepatocellular carcinoma (HCC) and can facilitate disease progression via poorly understood mechanisms. Here, we show that neutrophil extracellular traps (NETs) formation was increased in human HCC tumor tissues than in paired non-tumor liver tissues. Mechanism study revealed that tumor-induced metabolic switch toward glycolysis and pentose phosphate pathway in tumor infiltrating neutrophils promoted NETs formation in a reactive oxygen species dependent-manner. NETs subsequently induced the migration of cancer cells and down-regulation of tight junction molecules on adjacent endothelial cells, thus facilitating tumor intravasation and metastasis. Accordingly, NETs depletion could inhibit tumor metastasis in mice in vivo, and the infiltration levels of NETs-releasing neutrophils were negatively associated with patient survival and positively correlated with tumor metastasis potential of HCC patients. Our results unveiled a pro-metastatic role of NETs in the milieu of human HCC, and pointed to the importance of metabolic reprogramming in shaping their characteristics, thus providing an applicable efficient target for anti-cancer therapies.

H2 Inhibits the Formation of Neutrophil Extracellular Traps

Neutrophil extracellular traps (NETs) contribute to inflammatory pathogenesis in numerous conditions, including infectious and cardiovascular diseases, and have attracted attention as potential therapeutic targets. H2 acts as an antioxidant and has been clinically and experimentally proven to ameliorate inflammation. This study was performed to investigate whether H2 could inhibit NET formation and excessive neutrophil activation. Neutrophils isolated from the blood of healthy volunteers were stimulated with phorbol-12-myristate-13-acetate (PMA) or the calcium ionophore A23187 in H2-exposed or control media. Compared with control neutrophils, PMA- or A23187-stimulated human neutrophils exposed to H2 exhibited reduced neutrophil aggregation, citrullination of histones, membrane disruption by chromatin complexes, and release of NET components. CXCR4high neutrophils are highly prone to NETs, and H2 suppressed Ser-139 phosphorylation in H2AX, a marker of DNA damage, thereby suppressing the induction of CXCR4 expression. H2 suppressed both myeloperoxidase chlorination activity and production of reactive oxygen species to the same degree as N-acetylcysteine and ascorbic acid, while showing a more potent ability to inhibit NET formation than these antioxidants do in PMA-stimulated neutrophils. Although A23187 formed NETs in a reactive oxygen species-independent manner, H2 inhibited A23187-induced NET formation, probably via direct inhibition of peptidyl arginine deiminase 4-mediated histone citrullination. Inhalation of H2 inhibited the formation and release of NET components in the blood and bronchoalveolar lavage fluid in animal models of lipopolysaccharide-induced sepsis (mice and aged mini pigs). Thus, H2 therapy can be a novel therapeutic strategy for NETs associated with excessive neutrophil activation.

Novel Mechanism by a Bis-Pyridinium Fullerene Derivative to Induce Apoptosis by Enhancing the MEK-ERK Pathway in a Reactive Oxygen Species-Independent Manner in BCR-ABL-Positive Chronic Myeloid Leukemia-Derived K562 Cells.

In the treatment of breakpoint cluster region-Abelson (BCR-ABL)-positive chronic myeloid leukemia (CML) using BCR-ABL inhibitors, the appearance of a gatekeeper mutation (T315I) in BCR-ABL is a serious issue. Therefore, the development of novel drugs that overcome acquired resistance to BCR-ABL inhibitors by CML cells is required. We previously demonstrated that a bis-pyridinium fullerene derivative (BPF) induced apoptosis in human chronic myeloid leukemia (CML)-derived K562 cells partially through the generation of reactive oxygen species (ROS). We herein show that BPF enhanced the activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase-extracellular signal-regulated kinase (MEK-ERK) pathway in a ROS-independent manner. BPF-induced apoptosis was attenuated by trametinib, suggesting the functional involvement of the MEK-ERK pathway in apoptosis in K562 cells. In addition, the constitutive activation of the MEK-ERK pathway by the enforced expression of the BRAFV600E mutant significantly increased the sensitivity of K562 cells to BPF. These results confirmed for the first time that BPF induces apoptosis in K562 cells through dual pathways—ROS production and the activation of the MEK-ERK pathway. Furthermore, BPF induced cell death in transformed Ba/F3 cells expressing not only BCR-ABL but also T315I mutant through the activation of the MEK-ERK pathway. These results indicate that BPF is as an effective CML drug that overcomes resistance to BCR-ABL inhibitors.

The GPCR adaptor protein norbin suppresses the neutrophil-mediated immunity of mice to pneumococcal infection

AbstractStreptococcal pneumonia is a worldwide health problem that kills ∼2 million people each year, particularly young children, the elderly, and immunosuppressed individuals. Alveolar macrophages and neutrophils provide the early innate immune response to clear pneumococcus from infected lungs. However, the level of neutrophil involvement is context dependent, both in humans and in mouse models of the disease, influenced by factors such as bacterial load, age, and coinfections. Here, we show that the G protein–coupled receptor (GPCR) adaptor protein norbin (neurochondrin, NCDN), which was hitherto known as a regulator of neuronal function, is a suppressor of neutrophil-mediated innate immunity. Myeloid norbin deficiency improved the immunity of mice to pneumococcal infection by increasing the involvement of neutrophils in clearing the bacteria, without affecting neutrophil recruitment or causing autoinflammation. It also improved immunity during Escherichia coli–induced septic peritonitis. It increased the responsiveness of neutrophils to a range of stimuli, promoting their ability to kill bacteria in a reactive oxygen species–dependent manner, enhancing degranulation, phagocytosis, and the production of reactive oxygen species and neutrophil extracellular traps, raising the cell surface levels of selected GPCRs, and increasing GPCR-dependent Rac and Erk signaling. The Rac guanine-nucleotide exchange factor Prex1, a known effector of norbin, was dispensable for most of these effects, which suggested that norbin controls additional downstream targets. We identified the Rac guanine-nucleotide exchange factor Vav as one of these effectors. In summary, our study presents the GPCR adaptor protein norbin as an immune suppressor that limits the ability of neutrophils to clear bacterial infections.

The quinazoline derivative, 04NB-03, induces cell cycle arrest and apoptosis in hepatocellular carcinoma cells in a reactive oxygen species-dependent manner

Hepatocellular carcinoma (HCC) is one of the most deadly malignancies worldwide. However, current therapeutic drugs for HCC are far from satisfactory. Thus, the development of new drugs is urgently needed. In this study, we identified a novel quinazoline derivative, 04NB-03, with potent anti-HCC activities both in vitro and in vivo. 04NB-03 effectively suppressed the viability and proliferation of HCC cells. It induced both cell cycle arrest at the G2/M phase and apoptosis in concentration- and time-dependent manners. Moreover, 04NB-03 treatment significantly reduced xenograft tumor growth without notable toxic effects. Mechanistically, 04NB-03 induced endogenous reactive oxygen species (ROS) accumulation in concentration- and time-dependent manners. Scavenging the ROS reversed 04NB-03-induced cell cycle arrest and apoptosis. Taken together, these results indicate that the quinazoline derivative, 04NB-03, inhibits the growth of HCC cells through the induction of cell cycle arrest and apoptosis in an ROS-dependent manner. 04NB-03 is, therefore, a potential small molecule candidate for the development of antitumor drugs targeting HCC.

Flavonoids regulate cell death-related cellular signaling via ROS in human colon cancer cells

Although several studies investigated effects of flavonoids on proliferation and apoptosis, yet no study has correlated the cellular damage caused by reactive oxygen species (ROS) production, cell death related pathways and cell adhesion molecules (CAM) expression with cell survival. Here, we investigate cytotoxic effect of catechin, epicatechin and naringenin on colon cancer cells. While especially naringenin demonstrated most significant inhibition of colon cancer cell viability, high concentrations treatment did not exhibit more pronounced effect in colon epithelial cells. In addition, these flavonoids caused excessive ROS generation resulting in the impairment of lipid and protein, followed by the induction of apoptosis and autophagy. In details, mechanism studies revealed that elevated ROS production leads to caspase activation and pretreated NAC, an antioxidant, blocks catechins and naringenin induced apoptosis and autophagy. PKC activity assay data also showed that catechin and naringenin treatment decreases PKC activity that leads to cell death. Moreover, studied flavonoids, especially naringenin, induced G2/M cell cycle arrest in a ROS-independent manner. Furthermore, the studied flavonoids suppressed or decreased the expression of the cancer progression and metastasis-related cell adhesion molecules. Taken together, our results indicate that studied flavonoids suppress colon cancer cell growth via inducing cell death in ROS dependent manner.

Pan-cancer analysis reveals synergistic effects of CDK4/6i and PARPi combination treatment in RB-proficient and RB-deficient breast cancer cells

DNA damage results in mutations and plays critical roles in cancer development, progression, and treatment. Targeting DNA damage response in cancers by inhibiting poly-(ADP-ribose) polymerases (PARPs) offers an important therapeutic strategy. However, the failure of PARP inhibitors to markedly benefit patients suggests the necessity for developing new strategies to improve their efficacy. Here, we show that the expression of cyclin-dependent kinase 4/6 (CDK4/6) complex members significantly correlates with mutations (as proxies of DNA damages), and that the combination of CDK4/6 and PARP inhibitors shows synergy in both RB-proficient and RB-deficient breast cancer cells. As PARPs constitute sensors of DNA damage and are broadly involved in multiple DNA repair pathways, we hypothesized that the combined inhibition of PARPs and DNA repair (or repair-related) pathways critical for cancer (DRPCC) should show synergy. To identify druggable candidate DRPCC(s), we analyzed the correlation between the genome-wide expression of individual genes and the mutations for 27 different cancer types, assessing 7146 exomes and over 1,500,000 somatic mutations. Pathway enrichment analyses of the top-ranked genes correlated with mutations indicated “cell cycle pathway” as the top candidate DRPCC. Additionally, among functional cell-cycle complexes, the CDK4/6 complex showed the most significant negative correlation with mutations, also suggesting that combined CDK4/6 and PARP inhibition might exhibit synergy. Furthermore, combination treatment showed synergy in not only RB-proficient but also RB-deficient breast cancer cells in a reactive oxygen species-dependent manner. These findings suggest a potential therapeutic strategy to improve the efficacy of PARP and CDK4/6 inhibitors in cancer treatment.

Human mast cells exhibit an individualized pattern of antimicrobial responses.

IntroductionMast cells (MCs) are tissue‐resident immune cells implicated in antibacterial responses. These include chemokine secretion, degranulation, and the release of mast cell‐extracellular traps, which are primarily dependent on reactive oxygen species (ROS) production. Our study investigated whether human mast cells (hMCs) develop individual response patterns to bacteria located at different tissue sites: Escherichia coli (gut commensal), Listeria monocytogenes (foodborne intracellular pathogen), Staphylococcus aureus (skin commensal and opportunistic pathogen), and Streptococcus pneumoniae (upper respiratory tract commensal and lung pathogen).MethodsAfter live bacteria exposure, hMCs were analyzed by a combined flow cytometry assay for degranulation, ROS production, DNA externalization, and for β‐hexosaminidase, chemokine, and prostaglandin release.Results L. monocytogenes induced hMC degranulation, IL‐8 and MCP‐1 release coupled with DNA externalization in a novel hMC ROS independent manner. In contrast, S. pneumoniae caused ROS production without DNA release and degranulation. E. coli induced low levels of hMC degranulation combined with interleukin 8 and MCP‐1 secretion and in the absence of ROS and DNA externalization. Finally, S. aureus induced hMCs prostaglandin D2 release and DNA release selectively. Our findings demonstrate a novel hMC phenomenon of DNA externalization independent of ROS production. We also showed that ROS production, degranulation, DNA externalization, and mediator secretion occur as independent immune reactions in hMCs upon bacterial encounter and that hMCs contribute to bacterial clearance.ConclusionsThus, hMCs exhibit a highly individualized pattern of immune response possibly to meet tissue requirements and regulate bacteria coexistence vs defense.

Selenium nanoparticles reduce glucose metabolism and promote apoptosis of glioma cells through reactive oxygen species-dependent manner.

Gliomas are the most common, malignant, and lethal tumors in adults. Furthermore, gliomas are highly resistant to current chemotherapeutic drugs. Thus, new effective anticancer drugs for glioma are urgently needed. Selenium nanoparticles have been reported to have potent anti-tumor activity, although the specific mechanism is not fully understood. This study aimed to test the anti-tumor effect of selenium nanoparticles and its mechanism. We used selenium nanoparticles to treat commercial glioma cell lines, and patient-derived glioma cells, and then used the MTT assay to determine selenium nanoparticles effect against these. Apoptotic cell death was determined by annexin V-Fluos staining kit. Glucose uptake, lactate, and adenosine triphosphate production, together with hexokinase 2 and pyruvate kinase activities were measured to determine the glucose metabolism level. Reactive oxygen species production was tested using 2',7'-dichlorodihydrofluorescein diacetate. Our results showed that selenium nanoparticles had a potent cytotoxic effect in glioma cells, regardless of whether they were drug-resistant or not, whereas it showed less toxic effect in normal healthy cells. Further tests showed that selenium nanoparticles treatment leads to apoptotic cell death enhancement and glucose metabolism reduction, and this process was in a reactive oxygen species pathway-dependent manner. These results may provide a novel direction for glioma therapy in the future.

Formulation of a triple combination gemcitabine plus romidepsin + cisplatin regimen to efficaciously and safely control triple-negative breast cancer tumor development.

Triple-negative breast cancer (TNBC) is an aggressive, lethal, and heterogeneous subtype of breast cancers, tending to have lower 5-year survival rates than other BC subtypes in response to conventional chemotherapies. This study's aim was to identify advanced regimens to effectively control TNBC tumor development. We investigated the combination of the DNA synthesis inhibitor gemcitabine, the DNA-damaging agent cisplatin, and the histone deacetylase inhibitor romidepsin to control a variety of breast cells in vitro. We studied the toxicity of drug doses and administration schedules to determine tolerable combination regimens in immune-deficient nude and -competent BALB/c mice. We then studied the efficacy of tolerable regimens in controlling TNBC cell-derived xenograft development in nude mice. By reducing clinically equivalent doses of each agent in combination, we formulated tolerable regimens in animals. We verified that the tolerable triple combination gemcitabine plus romidepsin + cisplatin regimen more efficacious than double combination regimens in controlling xenograft tumor development in nude mice. A triple combination of gemcitabine + romidepsin + cisplatin synergistically induced death of the TNBC M.D. Anderson-Metastatic Breast cancer (MDA-MB) 231 and MDA-MB468, as well as Michigan Cancer Foundation (MCF) 7, MCF10A, and MCF10A-Ras cells. Cell death induced by gemcitabine + romidepsin + cisplatin was in a reactive oxygen species-dependent manner. Considering the high costs for developing a new anticancer agent, we used the FDA-approved drugs gemcitabine, romidepsin (is approved for T-cell lymphoma and is under clinical trial for TNBC), and cisplatin to economically formulate an efficacious and safe combination regimen. The highly efficacious gemcitabine plus romidepsin + cisplatin regimen should be poised for efficient translation into clinical trials, ultimately contributing to reduced mortality and improved quality of life for TNBC patients.