Elevated Reactive Oxygen Species Production Research Articles

Angiotensin II Increases Oxidative Stress and Inflammation in Female, But Not Male, Endothelial Cells.

Women are at elevated risk for certain cardiovascular diseases, including pulmonary arterial hypertension, Alzheimer's disease, and vascular complications of diabetes. Angiotensin II (AngII), a circulating stress hormone, is elevated in cardiovascular disease; however, our knowledge of sex differences in the vascular effects of AngII are limited. We therefore analyzed sex differences in human endothelial cell response to AngII treatment. Male and female endothelial cells were treated with AngII for 24h and analyzed by RNA sequencing. We then used endothelial and mesenchymal markers, inflammation assays, and oxidative stress indicators to measure female and male endothelial cell functional changes in response to AngII. Our data show that female and male endothelial cells are transcriptomically distinct. Female endothelial cells treated with AngII had widespread gene expression changes related to inflammatory and oxidative stress pathways, while male endothelial cells had few gene expression changes. While both female and male endothelial cells maintained their endothelial phenotype with AngII treatment, female endothelial cells showed increased release of the inflammatory cytokine interleukin-6 and increased white blood cell adhesion following AngII treatment concurrent with a second inflammatory cytokine. Additionally, female endothelial cells had elevated reactive oxygen species production compared to male endothelial cells after AngII treatment, which may be partially due to nicotinamide adenine dinucleotide phosphate oxidase-2 (NOX2) escape from X-chromosome inactivation. These data suggest that endothelial cells have sexually dimorphic responses to AngII, which could contribute to increased prevalence of some cardiovascular diseases in women. The online version contains supplementary material available at 10.1007/s12195-023-00762-2.

Aging Hallmarks and the Role of Oxidative Stress

Aging is a complex biological process accompanied by a progressive decline in the physical function of the organism and an increased risk of age-related chronic diseases such as cardiovascular diseases, cancer, and neurodegenerative diseases. Studies have established that there exist nine hallmarks of the aging process, including (i) telomere shortening, (ii) genomic instability, (iii) epigenetic modifications, (iv) mitochondrial dysfunction, (v) loss of proteostasis, (vi) dysregulated nutrient sensing, (vii) stem cell exhaustion, (viii) cellular senescence, and (ix) altered cellular communication. All these alterations have been linked to sustained systemic inflammation, and these mechanisms contribute to the aging process in timing not clearly determined yet. Nevertheless, mitochondrial dysfunction is one of the most important mechanisms contributing to the aging process. Mitochondria is the primary endogenous source of reactive oxygen species (ROS). During the aging process, there is a decline in ATP production and elevated ROS production together with a decline in the antioxidant defense. Elevated ROS levels can cause oxidative stress and severe damage to the cell, organelle membranes, DNA, lipids, and proteins. This damage contributes to the aging phenotype. In this review, we summarize recent advances in the mechanisms of aging with an emphasis on mitochondrial dysfunction and ROS production.

Delivery of miR-3529-3p using MnO2 -SiO2 -APTES nanoparticles combined with phototherapy suppresses lung adenocarcinoma progression by targeting HIGD1A.

The present study aimed to investigate the function of miR-3529-3p in lung adenocarcinoma and MnO2 -SiO2 -APTES (MSA) as a promising multifunctional delivery agent for lung adenocarcinoma therapy. Expression levels of miR-3529-3p were evaluated in lung carcinoma cells and tissues by qRT-PCR. The effects of miR-3529-3p on apoptosis, proliferation, metastasis and neovascularization were assessed by CCK-8, FACS, transwell and wound healing assays, tube formation and xenografts experiments. Luciferase reporter assays, western blot, qRT-PCR and mitochondrial complex assay were used to determine the targeting relationship between miR-3529-3p and hypoxia-inducible gene domain family member 1A (HIGD1A). MSA was fabricated using MnO2 nanoflowers, and its heating curves, temperature curves, IC50, and delivery efficiency were examined. The hypoxia and reactive oxygen species (ROS) production was investigated by nitro reductase probing, DCFH-DA staining and FACS. MiR-3529-3p expression was reduced in lung carcinoma tissues and cells. Transfection of miR-3529-3p could promote apoptosis and suppress cell proliferation, migration and angiogenesis. As a target of miR-3529-3p, HIGD1A expression was downregulated, through which miR-3529-3p could disrupt the activities of complexes III and IV of the respiratory chain. The multifunctional nanoparticle MSA could not only efficiently deliver miR-3529-3p into cells, but also enhance the antitumor function of miR-3529-3p. The underlying mechanism may be that MSA alleviates hypoxia and has synergistic effects in cellular ROS promotion with miR-3529-3p. Our results establish the antioncogenic role of miR-3529-3p, and demonstrate that miR-3529-3p delivered by MSA has enhanced tumor suppressive effects, probably through elevating ROS production and thermogenesis.

Identification and characterization of two transcriptional regulators in Xanthomonas oryzae pv. oryzicola YM15

Bacterial leaf streak disease caused by Xanthomonas oryzae pv. Oryzicola (Xoc) is a serious threat to rice production worldwide. Like most bacterial phytopathogens, Xoc also contains multiple regulators involved in the complex regulatory networks to ensure its virulence. In this study, we found two genes, FE36_09860 and FE36_06255, through quantitative fluorescence PCR essential for Xoc virulence and might be regulated by the diffusible signal factor (DSF) and the global regulator (Clp). We evaluated the characteristics of the TetR family (FE36_09860) and Fur family (FE36_06255) transcriptional regulator genes in in-vitro and in-vivo experiments. Results demonstrated that the deletion of two transcriptional regulators genes, FE36_09860 and FE36_06255, reduced the virulence of Xoc. Further analysis revealed that the deletion of the FE36_06255 gene results in decreased EPS production, elevated biofilm formation and reactive oxygen species production, and maintained the growth rate of Xoc YM15. In contrast, the deletion of FE36_09860 showed an opposite trend. It is concluded that, as transcriptional regulators, the FE36_09860 and FE36_06255 genes played an important role in the biological processes of Xoc YM15 and were essential for its virulence.

Mitochondrial alternative NADH dehydrogenases NDA1 and NDA2 promote survival of reoxygenation stress in Arabidopsis by safeguarding photosynthesis and limiting ROS generation.

Plant submergence stress is a growing problem for global agriculture. During desubmergence, rising O2 concentrations meet a highly reduced mitochondrial electron transport chain (mETC) in the cells. This combination favors the generation of reactive oxygen species (ROS) by the mitochondria, which at excess can cause damage. The cellular mechanisms underpinning the management of reoxygenation stress are not fully understood. We investigated the role of alternative NADH dehydrogenases (NDs), as components of the alternative mETC in Arabidopsis, in anoxia-reoxygenation stress management. Simultaneous loss of the matrix-facing NDs, NDA1 and NDA2, decreased seedling survival after reoxygenation, while overexpression increased survival. The absence of NDAs led to reduced maximum potential quantum efficiency of photosystem II linking the alternative mETC to photosynthetic function in the chloroplast. NDA1 and NDA2 were induced upon reoxygenation, and transcriptional activation of NDA1 was controlled by the transcription factors ANAC016 and ANAC017 that bind to the mitochondrial dysfunction motif (MDM) in the NDA1 promoter. The absence of NDA1 and NDA2 did not alter recovery of cytosolic ATP levels and NADH : NAD+ ratio at reoxygenation. Rather, the absence of NDAs led to elevated ROS production, while their overexpression limited ROS. Our observations indicate that the control of ROS formation by the alternative mETC is important for photosynthetic recovery and for seedling survival of anoxia-reoxygenation stress.

SOD2 in platelets: with age comes responsibility

SOD2 in platelets: with age comes responsibility

Perfluorooctane sulfonate-induced oxidative stress contributes to pancreatic β-cell apoptosis by inhibiting cyclic adenosine monophosphate pathway: Prevention by pentoxifylline

Endocrine-disrupting chemical perfluorooctane sulfonate (PFOS) acute exposure stimulates insulin secretion from pancreatic β-cells. However, chronic exposure to PFOS on pancreatic β-cells, its role in insulin secretion, and the underlying mechanisms have not been studied. We used rat insulinoma INS-1 and human 1.1b4 islet cells to investigate the chronic effects of PFOS on glucose-stimulated insulin secretion and toxicity implicated in the downregulation of β-cell functionality. Chronic exposure of INS-1 cells or human pancreatic 1.1b4 β-cells to PFOS stimulated the small G-protein RAC1-guanosine triphosphate-dependent nicotinamide adenine dinucleotide phosphate oxidase (NOX2/gp91phox) subunit expression and activation. Upregulated NOX2/gp91phox activation led to elevated reactive oxygen species (ROS) production with a decrease in the cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) pathway in both cell types. Inhibition of cAMP/PKA signaling induces β-cell mitochondrial dysfunction and endoplasmic stress via the loss of PDX1-SERCA2B and glucose-stimulated insulin release. Inhibiting RAC1-NOX2/gp91phox activation or elevating cAMP by pentoxifylline, a Food and Drug Administration-approved phosphodiesterase inhibitor, significantly reduced PFOS-induced ROS production and restored insulin secretory function of pancreatic β-cells. Enhanced secretory function in pentoxifylline-treated cells was associated with increased stability of PDX1-SERCA2B protein levels. Intriguingly, inhibition of cAMP/PKA signaling impaired pentoxifylline-induced insulin secretion caused by the activation of ROS production and mitochondrial dysfunction. Overall, our findings show that PFOS has a new and first-ever direct chronic effect on pancreatic β-cell failure through increased RAC1-NOX2/gp91phox activation and pentoxifylline-induced cAMP/PKA signaling, which inhibits PFOS-mediated mitochondrial dysfunction.

Conditional Knockout of Hypoxia-Inducible Factor 1-Alpha in Tumor-Infiltrating Neutrophils Protects against Pancreatic Ductal Adenocarcinoma.

Large numbers of neutrophils infiltrate tumors and comprise a notable component of the inflammatory tumor microenvironment. While it is established that tumor cells exhibit the Warburg effect for energy production, the contribution of the neutrophil metabolic state to tumorigenesis is unknown. Here, we investigated whether neutrophil infiltration and metabolic status promotes tumor progression in an orthotopic mouse model of pancreatic ductal adenocarcinoma (PDAC). We observed a large increase in the proportion of neutrophils in the blood and tumor upon orthotopic transplantation. Intriguingly, these tumor-infiltrating neutrophils up-regulated glycolytic factors and hypoxia-inducible factor 1-alpha (HIF-1α) expression compared to neutrophils from the bone marrow and blood of the same mouse. This enhanced glycolytic signature was also observed in human PDAC tissue samples. Strikingly, neutrophil-specific deletion of HIF-1α (HIF-1αΔNφ) significantly reduced tumor burden and improved overall survival in orthotopic transplanted mice, by converting the pro-tumorigenic neutrophil phenotype to an anti-tumorigenic phenotype. This outcome was associated with elevated reactive oxygen species production and activated natural killer cells and CD8+ cytotoxic T cells compared to littermate control mice. These data suggest a role for HIF-1α in neutrophil metabolism, which could be exploited as a target for metabolic modulation in cancer.

25-Hydroxycholesterol-induced Osteoblast Oxiapoptophagy Is Involved in the Pathophysiological Process of Osteoporosis.

25-hydroxycholesterol (25-HC) plays important roles in lipid metabolism, inflammatory responses, and apoptosis, but its pathophysiological association with osteoporosis (OP) has not been verified in osteoblasts. Hence, we studied the pathophysiological linkage and underlying cellular mechanisms of 25-HC in human osteoblast-like MG-63 cells and an ovariectomy-induced osteoporotic mouse model. To investigate the pathophysiological linkage between 25-HC-induced osteoblast oxiapoptophagy and OP, 25-HC ELISA assay, MTT assay, cell live/dead staining, hematoxylin and eosin staining, DAPI staining, flow cytometry analysis, western blot, caspase-3 staining, reactive oxygen species (ROS) assay, autophagy staining, immunocytochemistry, Micro-CT image analysis and immunocytochemistry were performed in MG-63 cells and ovariectomy-induced OP animals. The expression of cholesterol-25-hydroxylase (CH25H), an enzyme catalyzing the conversion of cholesterol to 25-HC, and the production of 25-HC were increased by lipopolysaccharide in MG-63 cells. Cytotoxicity was increased by 25-HC in MG-63 cells. Apoptosis with condensed chromatin and altered morphology was induced by 25-HC through cleavage of caspases-8, -9, and -3 in MG-63 cells. 25-HC induced oxidative stress in MG-63 cells via elevation of ROS production, cyclooxygenase-2, and inducible nitric oxide synthase. Furthermore, the expression of autophagy biomarkers, including beclin-1 and microtubule-associated protein 1A/1B-light chain 3, was elevated by 25-HC in MG-63 cells. In addition, p53 expression was increased, whereas Akt phosphorylation was suppressed in 25-HC-incubated MG-63 cells. The expression of CH25H, cleaved caspase-3, and beclin-1 were up-regulated in the femoral bone of ovariectomy-induced mouse osteoporotic animals. 25-HC plays a role in OP via the induction of oxiapoptophagic osteoblast death.

Metformin Alleviates Epirubicin-Induced Endothelial Impairment by Restoring Mitochondrial Homeostasis

Vascular endothelial injury is important in anthracycline-induced cardiotoxicity. Anthracyclines seriously damage the mitochondrial function and mitochondrial homeostasis. In this study, we investigated the damage of epirubicin to vascular endothelial cells and the protective role of metformin from the perspective of mitochondrial homeostasis. We found that epirubicin treatment resulted in DNA double-strand breaks (DSB), elevated reactive oxygen species (ROS) production, and excessive Angiotensin II release in HUVEC cells. Pretreatment with metformin significantly mitigated the injuries caused by epirubicin. In addition, inhibited expression of Mitochondrial transcription factor A (TFAM) and increased mitochondria fragmentation were observed in epirubicin-treated cells, which were partially resumed by metformin pretreatment. In epirubicin-treated cells, knockdown of TFAM counteracted the attenuated DSB formation due to metformin pretreatment, and inhibition of mitochondrial fragmentation with Mdivi-1 decreased DSB formation but increased TFAM expression. Furthermore, epirubicin treatment promoted mitochondrial fragmentation by stimulating the expression of Dynamin-1-like protein (DRP1) and inhibiting the expression of Optic atrophy-1(OPA1) and Mitofusin 1(MFN1), which could be partially prevented by metformin. Finally, we found metformin could increase TFAM expression and decrease DRP1 expression in epirubicin-treated HUVEC cells by upregulating the expression of calcineurin/Transcription factor EB (TFEB). Taken together, this study provided evidence that metformin treatment was an effective way to mitigate epirubicin-induced endothelial impairment by maintaining mitochondrial homeostasis.

Sunscreen exposure interferes with physiological processes while inducing oxidative stress in seagrass Posidonia oceanica (L.) Delile

The effects of a commercial sunscreen mixture on the Mediterranean seagrass Posidonia oceanica were investigated, evaluating its response in physiological processes and biochemical indicators of oxidative stress. Short-term laboratory experiments were conducted recreating summer conditions, and two sunscreen concentrations were tested in whole P. oceanica plants placed inside aquaria. Although primary productivity of leaf segments seemed to benefit from sunscreen addition, probably due to inorganic nutrients released, the rest of the biological parameters reflected possible impairments in the overall functioning of P. oceanica as a result of oxidative damages. Chlorophyll production and nitrogen fixation associated with old leaves were inhibited under high sunscreen concentrations, which concurred with elevated reactive oxygen species production, catalase activity and polyphenols content in the seagrass leaves. These results emphasize the importance of directing future investigations on determining which specific components of sunscreen products are likely threatening the wellbeing of critical species, such as P. oceanica.

Molecular basis of diseases induced by the mitochondrial DNA mutation m.9032T>C.

The mitochondrial DNA mutation m.9032T>C was previously identified in patients presenting with NARP (Neuropathy Ataxia Retinitis Pigmentosa). Their clinical features had a maternal transmission and patient's cells showed a reduced oxidative phosphorylation capacity, elevated reactive oxygen species (ROS) production and hyperpolarization of the mitochondrial inner membrane, providing evidence that m.9032T>C is truly pathogenic. This mutation leads to replacement of a highly conserved leucine residue with proline at position 169 of ATP synthase subunit a (L169P). This protein and a ring of identical c-subunits (c-ring) move protons through the mitochondrial inner membrane coupled to ATP synthesis. We herein investigated the consequences of m.9032T>C on ATP synthase in a strain of Saccharomyces cerevisiae with an equivalent mutation (L186P). The mutant enzyme assembled correctly but was mostly inactive as evidenced by a > 95% drop in the rate of mitochondrial ATP synthesis and absence of significant ATP-driven proton pumping across the mitochondrial membrane. Intragenic suppressors selected from L186P yeast restoring ATP synthase function to varying degrees (30-70%) were identified at the original mutation site (L186S) or in another position of the subunit a (H114Q, I118T). In light of atomic structures of yeast ATP synthase recently described, we conclude from these results that m.9032T>C disrupts proton conduction between the external side of the membrane and the c-ring, and that H114Q and I118T enable protons to access the c-ring through a modified pathway.

Copper sulfide anchored MXene improving photo-responsive self-healing polyurethane with enhanced mechanical and antibacterial properties

Diseases caused by bacterial infection are becoming a major threat to human health. Therefore, developing efficient antibacterial materials is of great significance in improving medical care and protecting people's health. In this work, an accordion-like structural Ti3C2@CuS was synthesized by copper sulfide (CuS) nanospheres anchored firmly on the surface of Ti3C2Tx via the hydrothermal method. The multilayer Ti3C2@CuS becomes few-layered nanosheets after ultrasonic treatment, which have an enjoyable dispersion in the polyurethane (PU) matrix. PU and the released Cu2+ from Ti3C2@CuS are firmly linked by a coordination bond, which improves the mechanical properties and thermal stability of Ti3C2@CuS-PU and reduces the heavy metal ion pollution by blocking the Cu2+ released by forming coordination bonds. Moreover, Ti3C2@CuS-PU exhibits an excellent self-healing performance after 30 tensile cycles. Additionally, Ti3C2Tx and CuS could improve the separation efficiency of the electron-hole pairs of CuS to produce more reactive oxygen species (ROS) to kill bacteria. Ti3C2@CuS-PU maintains a highly long-term sterilization ability of more than 90 % in 30 days because of the synergistic effect of the sustained release of copper ions, the elevated ROS production ability, and the excellent dispersion of Ti3C2@CuS in PU. This work demonstrates a simple and promising route for designing multifunctional antibacterial self-healing materials.

Basal Gp78-dependent mitophagy promotes mitochondrial health and limits mitochondrial ROS

Mitochondria are major sources of cytotoxic reactive oxygen species (ROS), such as superoxide and hydrogen peroxide, that when uncontrolled contribute to cancer progression. Maintaining a finely tuned, healthy mitochondrial population is essential for cellular homeostasis and survival. Mitophagy, the selective elimination of mitochondria by autophagy, monitors and maintains mitochondrial health and integrity, eliminating damaged ROS-producing mitochondria. However, mechanisms underlying mitophagic control of mitochondrial homeostasis under basal conditions remain poorly understood. E3 ubiquitin ligase Gp78 is an endoplasmic reticulum membrane protein that induces mitochondrial fission and mitophagy of depolarized mitochondria. Here, we report that CRISPR/Cas9 knockout of Gp78 in HT-1080 fibrosarcoma cells increased mitochondrial volume, elevated ROS production and rendered cells resistant to carbonyl cyanide m-chlorophenyl hydrazone (CCCP)-induced mitophagy. These effects were phenocopied by knockdown of the essential autophagy protein ATG5 in wild-type HT-1080 cells. Use of the mito-Keima mitophagy probe confirmed that Gp78 promoted both basal and damage-induced mitophagy. Application of a spot detection algorithm (SPECHT) to GFP-mRFP tandem fluorescent-tagged LC3 (tfLC3)-positive autophagosomes reported elevated autophagosomal maturation in wild-type HT-1080 cells relative to Gp78 knockout cells, predominantly in proximity to mitochondria. Mitophagy inhibition by either Gp78 knockout or ATG5 knockdown reduced mitochondrial potential and increased mitochondrial ROS. Live cell analysis of tfLC3 in HT-1080 cells showed the preferential association of autophagosomes with mitochondria of reduced potential. Xenograft tumors of HT-1080 knockout cells show increased labeling for mitochondria and the cell proliferation marker Ki67 and reduced labeling for the TUNEL cell death reporter. Basal Gp78-dependent mitophagic flux is, therefore, selectively associated with reduced potential mitochondria promoting maintenance of a healthy mitochondrial population, limiting ROS production and tumor cell proliferation.

Imaging flow cytometry to characterize the relationship between abnormal sperm morphologies and reactive oxygen species in stallion sperm.

Low levels of intracellular reactive oxygen species (ROS) are essential for normal sperm function and are produced by sperm mitochondria as a byproduct of metabolism, but in excess, ROS can cause catastrophic cellular damage and has been correlated with infertility, poor sperm motility and abnormal morphology in humans. Stallion sperm motility is fueled predominantly by oxidative phosphorylation-produced ATP, requiring high basal rates of mitochondrial function. Consequently, whether elevated ROS production by stallion sperm is an indicator of dysfunctional or highly motile cells has been debated by researchers over the last decade. The objective of this study was to evaluate the relationship between various sperm morphologies and ROS production in fresh and cooled stallion semen by employing the novel method of imaging flow cytometry for stallion semen assessment. For evaluation of fresh semen, single ejaculates (n=5) were collected from four resident stallions at the University of California, Davis. For the evaluation of 24-h cool-stored semen, single ejaculates were collected from stallions at Texas A&M University (n=5) and shipped to the University of California, Davis overnight for evaluation. Ejaculate volume, sperm concentration and motility parameters were recorded. Samples were co-stained for viability and ROS detection with SytoxGreen™ and dihydroethidium (DHE), respectively, and evaluated with the Amnis® ImageStream® system (Luminex Corporation). Antimycin, an electron transport chain inhibitor that triggers ROS production (1μM), was used as a positive control for DHE, while dead cells (2× snap frozen in liquid nitrogen) served as a positive control for SytoxGreen™. Unstained samples were also evaluated as controls. Imaging flow cytometric analysis was performed with the ideas® software (Luminex Corporation). Evaluated morphologies included abnormal head (AH), abnormal midpiece (AM), abnormal tail (AT), proximal cytoplasmic droplet (PD), or distal cytoplasmic droplet (DD), and morphologically normal (MN) cells. For fresh semen, an additional abnormality, coiled tail and midpiece (CTM) was assessed; 24-h cool-stored semen did not contain enough viable CTM cells for analysis. Only cells with obvious, single abnormalities were selected for the first portion of analysis to minimize subjectivity. Mixed effects modelling was used to evaluate the relationship between each morphologic classification and the corresponding DHE fluorescence intensity. Compared to the MN population, ROS production was significantly higher in viable cells with AH, PD and AM (p < .0001) in both fresh and cooled semen. CTM cells had significantly higher levels of ROS production compared to MN cells in fresh semen (p < .0001). There was no significant difference in ROS levels between MN cells and AT and DD cells in either fresh or cooled semen (p > .05). These results suggest that ROS generation is indicative of abnormal cell morphology and function and confirm that imaging flow cytometry is a valuable tool for the assessment of stallion semen.

Mechanisms underlying citrinin-induced toxicity via oxidative stress and apoptosis-mediated by mitochondrial-dependent pathway in SH-SY5Y cells

Citrinin (CIT) is a mycotoxin produced as a secondary product by the genera Aspergillus, Penicillium, Monascus, and other strains. CIT has the potential for contaminating animal feed and human food such as maize, wheat, rye, barley, oats, rice, cheese, and sake. Although CIT is primarily known as a nephrotoxic mycotoxin, it also affects other organs, including the liver and bone marrow, and its mechanisms of toxicity have not been clearly elucidated. There is a further lack of studies investigating the potential for CIT-induced neurotoxicity and its mechanisms. In the current study, SH-SY5Y human neuroblastoma cell line was treated with CIT for 24 h to evaluate various toxicological endpoints, such as reactive oxygen species (ROS) production and apoptosis induction. Results indicate that CIT has an IC50 value of 250.90 μM and cell proliferation decreased significantly at 50 and 100 μM CIT concentrations. These same concentrations also caused elevated ROS production (≥34.76%), apoptosis (≥9.43-fold) and calcium ion mobilization (≥36.52%) in the cells. Results show a significant decrease in the mitochondrial membrane potential (≥86.8%). We also found that CIT significantly upregulated the expression of some genes related to oxidative stress and apoptosis, while downregulating others. These results suggest that apoptosis and oxidative stress may be involved in the mechanisms underlying CIT-induced neurotoxicity.

Noncanonical Role of Telomerase in Regulation of Microvascular Redox Environment With Implications for Coronary Artery Disease.

Telomerase reverse transcriptase (TERT) (catalytic subunit of telomerase) is linked to the development of coronary artery disease (CAD); however, whether the role of nuclear vs. mitchondrial actions of TERT is involved is not determined. Dominant-negative TERT splice variants contribute to decreased mitochondrial integrity and promote elevated reactive oxygen species production. We hypothesize that a decrease in mitochondrial TERT would increase mtDNA damage, promoting a pro-oxidative redox environment. The goal of this study is to define whether mitochondrial TERT is sufficient to maintain nitric oxide as the underlying mechanism of flow-mediated dilation by preserving mtDNA integrity.Immunoblots and quantitative polymerase chain reaction were used to show elevated levels of splice variants α- and β-deletion TERT tissue from subjects with and without CAD. Genetic, pharmacological, and molecular tools were used to manipulate TERT localization. Isolated vessel preparations and fluorescence-based quantification of mtH2O2 and NO showed that reduction of TERT in the nucleus increased flow induced NO and decreased mtH2O2 levels, while prevention of mitochondrial import of TERT augmented pathological effects. Further elevated mtDNA damage was observed in tissue from subjects with CAD and initiation of mtDNA repair mechanisms was sufficient to restore NO-mediated dilation in vessels from patients with CAD. The work presented is the first evidence that catalytically active mitochondrial TERT, independent of its nuclear functions, plays a critical physiological role in preserving NO-mediated vasodilation and the balance of mitochondrial to nuclear TERT is fundamentally altered in states of human disease that are driven by increased expression of dominant negative splice variants.

New brefeldin A-cinnamic acid ester derivatives as potential antitumor agents: Design, synthesis and biological evaluation

Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and ranks third in mortality rate worldwide. Brefeldin A (BFA, 1), a natural Arf1 inhibitor, qualifies extremely superior antitumor activity against HCC while its low aqueous solubility, poor bioavailability, and high toxicity have greatly hindered its translation to the clinic. Herein, a series of BFA-cinnamic acid ester derivatives was rationally designed and synthesized via introducing active cinnamic acid and its analogues into the structure of 1. Their in vitro cytotoxic activities on five cancer cell lines, including HepG2, BEL-7402, HeLa, Eca-109 and PANC-1, were evaluated using MTT assay. As expected, favorable cytotoxic activity was observed on majority of the mono-substituted derivatives. Especially, the most potent brefeldin A 4-O-(4)-dimethylaminocinnamate (CHNQD-01269, 33) with improved aqueous solubility, demonstrated the strong cytotoxic activity against HepG2 and BEL-7402 cell lines with IC50 values of 0.29 and 0.84 μM, respectively. More importantly, 33 performed low toxicity on normal liver cell line L-02 with the selectivity index (SI) of 9.69, which was more than 17-fold higher than that of 1. Results from mechanistic studies represented that 33 blocked the cell cycle in the G1 phase, and induced apoptosis via elevating reactive oxygen species (ROS) production and increasing expression of apoptosis-related proteins of HepG2 cells. Docking experiment also suggested 33 a promising Arf1 inhibitor, which was confirmed by the cellular thermal shift assay that 33 displayed a significant effect on the stability of Arf1 protein. Furthermore, 33 possessed high safety profile (MTD >100 mg/kg, ip) and favorable pharmacokinetic properties. Notably, the superior antiproliferative activity was verified in HepG2 tumor-bearing xenograft model in which 33 markedly suppressed the tumor growth (TGI = 46.17%) in nude mice at a dose of 10 mg/kg once a day for 16 d. The present study provided evidence of exploiting this series of highly efficacious derivatives, especially 33, for the treatment of HCC.

Catalase regulates the homeostasis of hemolymph microbiota and autophagy of the hemocytes in mud crab (Scylla paramamosain)

Reactive oxygen species (ROS), a group of unstable and highly reactive molecules or free radicals, are productions of molecular oxygen through cell metabolism. In invertebrates, especially in crustaceans, hemolymph plays an extremely important role in innate immune regulation and inflammatory responses, in which various microbes were modified by ROS. Even so, the information about ROS in relation to hemolymph microbiota and innate immunity is still limited in mud crab. To investigate the underlying mechanism, the ROS-excessive mud crabs (Scylla paramamosain) caused by knocking down catalase (SpCAT) were used to demonstrate the effects of abnormal levels of ROS on the changes in the hemolymph microbiota. Our results showed that SpCAT silence significantly increase ROS production and decrease the load of the bacteria in the hemolymph in mud crab. In the siCAT group, the dominant phyla were Proteobacteria, Actinobacteria, Acidonacteria, Bacteroidetes, Firmicutes, Gemmatimonadetes, Chloroflexi, Planctomycetes, Rokubacteria and Verrucommicrobia. Among them, the relative abundance of Firmicutes and Cyanobacteria was significantly increased upon the elevated ROS production. Some bacterial genera such as Bacteroides, Parabacteroides, Faecalibacterium and Romboutsia also increased with higher level of ROS. Moreover, the results showed that excessive ROS could promote the evident autophagy of the hemocytes. The result of this study demonstrated that ROS plays a vital role in modulating the homeostasis of the microbiota, and SpCAT is also crucially important in the ROS-mediated hemolymph microbiota homeostasis and the autophagy of the hemocytes in mud crab.

Deuterated Linoleic Acid Attenuates the RBC Storage Lesion in a Mouse Model of Poor RBC Storage.

Background: Long-chain polyunsaturated fatty acids (PUFAs) are important modulators of red blood cell (RBC) rheology. Dietary PUFAs are readily incorporated into the RBC membrane, improving RBC deformability, fluidity, and hydration. However, enriching the lipid membrane with PUFAs increases the potential for peroxidation in oxidative environments (e.g., refrigerated storage), resulting in membrane damage. Substitution of bis-allylic hydrogens with deuterium ions in PUFAs decreases hydrogen abstraction, thereby inhibiting peroxidation. If lipid peroxidation is a causal factor in the RBC storage lesion, incorporation of deuterated linoleic acid (DLA) into the RBC membrane should decrease lipid peroxidation, thereby improving RBC lifespan, deformability, filterability, and post-transfusion recovery (PTR) after cold storage. Study Design and Methods: Mice associated with good (C57BL/6J) and poor (FVB) RBC storage quality received diets containing 11,11-D2-LA Ethyl Ester (1.0 g/100 g diet; deuterated linoleic acid) or non-deuterated LA Ethyl Ester (control) for 8 weeks. Deformability, filterability, lipidomics, and lipid peroxidation markers were evaluated in fresh and stored RBCs. Results: DLA was incorporated into RBC membranes in both mouse strains. DLA diet decreased lipid peroxidation (malondialdehyde) by 25.4 and 31% percent in C57 mice and 12.9 and 79.9% in FVB mice before and after cold storage, respectively. In FVB, but not C57 mice, deformability filterability, and post-transfusion recovery were significantly improved. Discussion: In a mouse model of poor RBC storage, with elevated reactive oxygen species production, DLA attenuated lipid peroxidation and significantly improved RBC storage quality.