Nitric Oxide (NO) in Apoptotic versus Necrotic RAW 264.7 Macrophage Cell Death: The Role of NO-Donor Exposure, NAD+Content, and p53 Accumulation

Initiation of nitric oxide (NO•)-mediated apoptotic cell death in RAW 264.7 macrophages is associated with up-regulation of mitochondrial manganese superoxide dismutase (MnSOD; SOD2) and down-regulation of cytosolic copper zinc superoxide dismutase (CuZnSOD; SOD1) at their individual mRNA and protein levels. To evaluate the decreased CuZnSOD expression and the initiation of apoptosis we stably transfected macrophages to overexpress human CuZnSOD. Individual clones revealed a 2-fold increase in CuZnSOD activity. Expression of a functional and thus protective CuZnSOD was verified by attenuated superoxide (O2•-)-mediated apoptotic as well as necrotic cell death. In this study we showed that SOD-overexpressing macrophages (R-SOD1-12) were also protected against NO•-initiated programmed cell death. Protection was substantial towards NO• derived from exogenously added NO donors or when NO• was generated by inducible NO synthase activation, and was evident at the level of p53 accumulation, caspase activation and DNA fragmentation. Stimulation of parent and SOD-overexpressing cells with a combination of lipopolysaccharide and murine interferon γ produced equivalent amounts of nitrite/nitrate, which ruled out attenuated inducible NO• synthase activity during protection. Because protection by a O2•--scavenging system during NO•-intoxication implies a role of NO• and O2•- in the progression of cell damage, we used uric acid to delineate the role of peroxynitrite during NO•-elicited apoptosis. The peroxynitrite scavenger uric acid left S-nitrosoglutathione or spermine-NO-elicited apoptosis unaltered, blocking only 3-morpholinosydnonimine-mediated cell death. As a result we exclude peroxynitrite from contributing, to any major extent, to NO•-mediated apoptosis. Therefore protection observed with CuZnSOD overexpression is unlikely to stem from interference with peroxynitrite formation and/or action. Unequivocally, the down-regulation of CuZnSOD is associated with NO• cytotoxicity, whereas CuZnSOD overexpression protects macrophages from apoptosis.

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Chromatographic Analysis and In Vitro Cytotoxic Properties of Different Root Extracts of Hermannia geniculata Eckl. & Zeyh on Vero, HepG2 And RAW 264.7 Macrophage

Nigerian Veterinary Journal ◽

10.4314/nvj.v41i2.4 ◽

2021 ◽

Vol 41 (2) ◽

pp. 117-124

Author(s):

L.A. Adeniran ◽

A.O.T. Ashafa

Keyword(s):

Nitric Oxide ◽

Cell Lines ◽

Chromatographic Analysis ◽

Ethanol Extract ◽

Raw 264.7 ◽

Macrophage Cell ◽

Root Extracts ◽

Lc50 Value ◽

Raw 264.7 Macrophage

Hermannia geniculata is widely used in the management of several illnesses in South African traditional medicine. Chromatographic analysis, in vitro cytotoxicity, and biological activities of secondary metabolites present in Hermannia geniculata root extracts were investigated. Vero monkey kidney cells, human hepatocellular carcinoma (HepG2) cells, and RAW 264.7 macrophage cell lines were used to determine the cytotoxicity of the extracts using MTT assay. The capabilities of the plant extracts to inhibit 5-lipoxygenase enzyme activities, the overproduction of nitric oxide (NO) following lipopolysaccharide (LPS)-activated RAW 264.7 macrophages by the ethanol extract was evaluated. Results showed selective toxicity of the extracts with LC50 values of Vero cells ranging from (0.40-0.57 mg/mL) while the LC50 value of HepG2 cells varies between (0.016-0.136 mg/mL). The selectivity indexes (SI) were recorded (31.87, 18.87, 33.33, and 13.52) for ethanol, hydro-ethanol, decoction, and aqueous extracts respectively. The ethanol extract inhibited NO production in a concentration-dependent manner showing a decrease of 82% at a concentration of 0.1 mg/mL. Its LC50 value (3.64 mg/mL) is lower and significantly different (p<0.05) compared to quercetin (standard) with an LC50 (8.28 mg/mL). Similarly, the ethanol extract is a potent inhibitor of 5-lipoxygenase enzyme with the lowest IC50 value of 0.14 mg/mL which is significantly different (p<0.05) from other extracts and indomethacin (standard). The GC-MS chromatograms of the ethanol extract revealed five principal compounds that have been reported to have antioxidant, anti-inflammatory, and antifungal properties. This result indicated that Hermannia geniculata root extracts is not toxic to Vero and RAW 264.7 macrophage cell lines and toxic to HepG2 cell lines used in this experiment, it may also possesses antiiflammatory and antiploriferative activities which could be exploited in the development of new, safer, and efficacious drugs. Keywords: Cytotoxicity; Nitric oxide; 5-lipoxygenase; Hermannia geniculata, Antioxidant GCMS.

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α2-Macroglobulin Functions as a Cytokine Carrier to Induce Nitric Oxide Synthesis and Cause Nitric Oxide-dependent Cytotoxicity in the RAW 264.7 Macrophage Cell Line

Journal of Biological Chemistry ◽

10.1074/jbc.270.37.21919 ◽

1995 ◽

Vol 270 (37) ◽

pp. 21919-21927 ◽

Cited By ~ 28

Author(s):

Jeffrey J. Lysiak ◽

Isa M. Hussaini ◽

Donna J. Webb ◽

William F. Glass ◽

Margaretta Allietta ◽

...

Keyword(s):

Nitric Oxide ◽

Cell Line ◽

Raw 264.7 ◽

Nitric Oxide Synthesis ◽

Macrophage Cell Line ◽

Macrophage Cell ◽

Raw 264.7 Macrophage ◽

Α2 Macroglobulin

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Nanotechnologies. 5-(and 6)-Chloromethyl-2',7' Dichlorodihydrofluorescein diacetate (CM-H2DCF- DA) assay for evaluating nanoparticle-induced intracellular reactive oxygen species (ROS) production in RAW 264.7 macrophage cell line

10.3403/30284328u ◽

2016 ◽

Keyword(s):

Reactive Oxygen Species ◽

Cell Line ◽

Reactive Oxygen ◽

Intracellular Reactive Oxygen Species ◽

Raw 264.7 ◽

Ros Production ◽

Oxygen Species ◽

Macrophage Cell Line ◽

Macrophage Cell ◽

Raw 264.7 Macrophage

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Role of Nitric Oxide in the Nickel and Cobalt Induced Cytotoxicity in RAW 264.7 cell

Korean Journal of Occupational and Environmental Medicine ◽

10.35371/kjoem.2001.13.3.274 ◽

2001 ◽

Vol 13 (3) ◽

pp. 274

Author(s):

Jung Ho Youm ◽

Gyung Jae Oh ◽

Young Cheun Yoo

Keyword(s):

Nitric Oxide ◽

Raw 264.7 ◽

Raw 264.7 Cell

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Immune Enhancing Effect of Medicinal Herb Extracts on a RAW 264.7 Macrophage Cell Line

Journal of the Korean Society of Food Science and Nutrition ◽

10.3746/jkfn.2012.41.11.1521 ◽

2012 ◽

Vol 41 (11) ◽

pp. 1521-1527 ◽

Cited By ~ 10

Author(s):

A-Reum Yu ◽

Ho-Young Park ◽

In-Wook Choi ◽

Yong-Kon Park ◽

Hee-Do Hong ◽

...

Keyword(s):

Cell Line ◽

Medicinal Herb ◽

Raw 264.7 ◽

Macrophage Cell Line ◽

Macrophage Cell ◽

Enhancing Effect ◽

Raw 264.7 Macrophage ◽

Herb Extracts

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Florigen governs shoot regeneration

Scientific Reports ◽

10.1038/s41598-021-93180-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yaarit Kutsher ◽

Michal Fisler ◽

Adi Faigenboim ◽

Moshe Reuveni

Keyword(s):

Nitric Oxide ◽

Shoot Regeneration ◽

Ectopic Expression ◽

Flowering Plants ◽

Regeneration Capacity ◽

No Donor ◽

Tobacco Leaves ◽

Age Related ◽

Delayed Flowering

AbstractIt is widely known that during the reproductive stage (flowering), plants do not root well. Most protocols of shoot regeneration in plants utilize juvenile tissue. Adding these two realities together encouraged us to study the role of florigen in shoot regeneration. Mature tobacco tissue that expresses the endogenous tobacco florigen mRNA regenerates poorly, while juvenile tissue that does not express the florigen regenerates shoots well. Inhibition of Nitric Oxide (NO) synthesis reduced shoot regeneration as well as promoted flowering and increased tobacco florigen level. In contrast, the addition of NO (by way of NO donor) to the tissue increased regeneration, delayed flowering, reduced tobacco florigen mRNA. Ectopic expression of florigen genes in tobacco or tomato decreased regeneration capacity significantly. Overexpression pear PcFT2 gene increased regeneration capacity. During regeneration, florigen mRNA was not changed. We conclude that florigen presence in mature tobacco leaves reduces roots and shoots regeneration and is the possible reason for the age-related decrease in regeneration capacity.

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Phaseolin Attenuates Lipopolysaccharide-Induced Inflammation in RAW 264.7 Cells and Zebrafish

Biomedicines ◽

10.3390/biomedicines9040420 ◽

2021 ◽

Vol 9 (4) ◽

pp. 420

Author(s):

Su-Jung Hwang ◽

Ye-Seul Song ◽

Hyo-Jong Lee

Keyword(s):

Nitric Oxide ◽

Nuclear Translocation ◽

Raw 264.7 Cells ◽

Network Pharmacology ◽

Raw 264.7 ◽

Dependent Manner ◽

Bioactive Constituents

Kushen (Radix Sophorae flavescentis) is used to treat ulcerative colitis, tumors, and pruritus. Recently, phaseolin, formononetin, matrine, luteolin, and quercetin, through a network pharmacology approach, were tentatively identified as five bioactive constituents responsible for the anti-inflammatory effects of S. flavescentis. However, the role of phaseolin (one of the primary components of S. flavescentis) in the direct regulation of inflammation and inflammatory processes is not well known. In this study, the beneficial role of phaseolin against inflammation was explored in lipopolysaccharide (LPS)-induced inflammation models of RAW 264.7 macrophages and zebrafish larvae. Phaseolin inhibited LPS-mediated production of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS), without affecting cell viability. In addition, phaseolin suppressed pro-inflammatory mediators such as cyclooxygenase 2 (COX-2), interleukin-1β (IL-1β), tumor necrosis factor α (TNF-α), monocyte chemoattractant protein-1 (MCP-1), and interleukin-6 (IL-6) in a dose-dependent manner. Furthermore, phaseolin reduced matrix metalloproteinase (MMP) activity as well as macrophage adhesion in vitro and the recruitment of leukocytes in vivo by downregulating Ninjurin 1 (Ninj1), an adhesion molecule. Finally, phaseolin inhibited the nuclear translocation of nuclear factor-kappa B (NF-κB). In view of the above, our results suggest that phaseolin could be a potential therapeutic candidate for the management of inflammation.

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