Extracellular Superoxide Dismutase Induces Mouse Embryonic Fibroblast Proliferative Burst, Growth Arrest, Immortalization, and Consequent In Vivo Tumorigenesis

2014 ◽  
Vol 21 (10) ◽  
pp. 1460-1474 ◽  
Author(s):  
Maria Domenica Castellone ◽  
Angela Langella ◽  
Silvia Cantara ◽  
Juha P. Laurila ◽  
Lilja E. Laatikainen ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Growth Arrest ◽  
Mouse Embryonic Fibroblast ◽  
Extracellular Superoxide Dismutase ◽  
Embryonic Fibroblast

scholarly journals Proteolytic modification of the heparin-binding affinity of extracellular superoxide dismutase

1993 ◽  
Vol 290 (2) ◽  
pp. 623-626 ◽  
Author(s):  
K Karlsson ◽  
A Edlund ◽  
J Sandström ◽  
S L Marklund
Keyword(s):  
Superoxide Dismutase ◽  
Binding Affinity ◽  
Heparan Sulphate ◽  
Limited Proteolysis ◽  
Size Exclusion ◽  
Extracellular Superoxide Dismutase ◽  
Heparin Binding ◽  
Binding Domains ◽  
Heparin Affinity

The heparin-binding affinity of the tetrameric extracellular superoxide dismutase (EC-SOD) is a result of the cooperative effect of the heparin-binding domains of the subunits, located in the hydrophilic, strongly positively charged C-terminal ends. EC-SOD C, the high-heparin-affinity type, exposed to immobilized trypsin and plasmin was found to rapidly lose its affinity for heparin, without any loss of enzymic activity or major change in molecular mass as judged by size-exclusion chromatography. Heparin and dextran sulphate 5000 inhibited the proteolysis, suggesting that EC-SOD C sequestered by heparan sulphate proteoglycan in vivo is partially protected against proteolysis. The loss of heparin-affinity occurred with the stepwise formation of intermediates, and the pattern upon chromatography on heparin-Sepharose and subsequent immunoblotting was compatible with the notion that the changes are due to sequential truncations of heparin-binding domains from subunits composing the EC-SOD tetramers. A similar pattern with intermediates and apparent truncations has previously been found with EC-SOD of human plasma. The findings show that the unique design of the heparin-binding domain of EC-SOD allows easy modification of the heparin-affinity by means of limited proteolysis, and suggest that such proteolysis is a major contributor to the heterogeneity in heparin-affinity of EC-SOD in mammalian plasma.

Plasma extracellular superoxide dismutase and erythrocyte Cu, Zn-containing superoxide dismutase in alcoholics treated with disulfiram

1986 ◽  
Vol 70 (4) ◽  
pp. 365-369 ◽  
Author(s):  
Michael Öhman ◽  
Stefan L. Marklund
Keyword(s):  
Superoxide Dismutase ◽  
Chronic Alcoholism ◽  
Term Treatment ◽  
Extracellular Superoxide Dismutase ◽  
Healthy Controls ◽  
Efficient Inhibitor ◽  
Long Term Treatment

1. Disulfiram has long been used in the treatment of chronic alcoholism. It is in vivo partially reduced to diethyldithiocarbamate, which is an efficient inhibitor of Cu, Zn-containing superoxide dismutase both in vitro and in vivo. The recently described extracellular superoxide dismutase is even more sensitive to diethyldithiocarbamate than Cu, Zn-superoxide dismutase. 2. To test for the possibility that long term treatment with disulfiram leads to inhibition of the superoxide dismutases, plasma extracellular superoxide dismutase and erythrocyte Cu, Zn-superoxide dismutase were determined in 12 disulfiram-treated alcoholics, and compared with 11 non-treated alcoholics and 19 healthy controls. 3. Plasma extracellular superoxide dismutase was moderately reduced (about 20%) in the disulfiram-treated alcoholics as compared with the non-treated alcoholics and the healthy controls. No effect of disulfiram treatment on erythrocyte Cu, Zn-superoxide dismutase activity was demonstrated.

scholarly journals Phytochemical Characterization and Chemotherapeutic Potential of Cinnamomum verum Extracts on the Multiplication of Protozoan Parasites In Vitro and In Vivo

Molecules ◽  
2020 ◽  
Vol 25 (4) ◽  
pp. 996 ◽  
Author(s):  
Gaber El-Saber Batiha ◽  
Amany Magdy Beshbishy ◽  
Azirwan Guswanto ◽  
Arifin Nugraha ◽  
Tserendorj Munkhjargal ◽  
...  
Keyword(s):  
Drug Exposure ◽  
Inhibitory Effect ◽  
Mouse Embryonic Fibroblast ◽  
Bioactive Constituents ◽  
Viability Assay ◽  
Nih 3T3 ◽  
Embryonic Fibroblast ◽  
Herbal Plant

Cinnamomum verum is a commonly used herbal plant that has several documented properties against various diseases. The existing study evaluated the inhibitory effect of acetonic extract of C. verum (AECV) and ethyl acetate extract of C. verum (EAECV) against piroplasm parasites in vitro and in vivo. The drug-exposure viability assay was tested on Madin-Darby bovine kidney (MDBK), mouse embryonic fibroblast (NIH/3T3) and human foreskin fibroblast (HFF) cells. Qualitative phytochemical estimation revealed that AECV and EAECV containing multiple bioactive constituents namely alkaloids, tannins, saponins, terpenoids and remarkable amounts of polyphenols and flavonoids. AECV and EAECV inhibited B. bovis, B. bigemina, B. divergens, B. caballi, and T. equi multiplication at half-maximal inhibitory concentrations (IC50) of 23.1 ± 1.4, 56.6 ± 9.1, 33.4 ± 2.1, 40.3 ± 7.5, 18.8 ± 1.6 µg/mL, and 40.1 ± 8.5, 55.6 ± 1.1, 45.7 ± 1.9, 50.2 ± 6.2, and 61.5 ± 5.2 µg/mL, respectively. In the cytotoxicity assay, AECV and EAECV affected the viability of MDBK, NIH/3T3 and HFF cells with half-maximum effective concentrations (EC50) of 440 ± 10.6, 816 ± 12.7 and 914 ± 12.2 µg/mL and 376 ± 11.2, 610 ± 7.7 and 790 ± 12.4 µg/mL, respectively. The in vivo experiment showed that AECV and EAECV were effective against B. microti in mice at 150 mg/kg. These results showed that C. verum extracts are potential antipiroplasm drugs after further studies in some clinical cases.

scholarly journals Therapeutic Effects of Atranorin towards the Proliferation of Babesia and Theileria Parasites

Pathogens ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 127 ◽  
Author(s):  
Amany Magdy Beshbishy ◽  
Gaber El-Saber Batiha ◽  
Luay Alkazmi ◽  
Eman Nadwa ◽  
Eman Rashwan ◽  
...  
Keyword(s):  
Mouse Embryonic Fibroblast ◽  
Drug Candidate ◽  
Therapeutic Effects ◽  
Diminazene Aceturate ◽  
Human Foreskin Fibroblasts ◽  
Ic50 Values ◽  
Nih 3T3 ◽  
Embryonic Fibroblast

Atranorin (ATR), is a compound with multidirectional biological activity under different in vitro and in vivo conditions and it is effective as an antibacterial, antiviral, antiprotozoal and anti-inflammatory agent. In the current study, the in vitro as well as in vivo chemotherapeutic effect of ATR as well as its combined efficacy with the existing antibabesial drugs (diminazene aceturate (DA), atovaquone (AV) and clofazimine (CF)) were investigated on six species of piroplasm parasites. ATR suppressed B. bovis, B. bigemina, B. divergens, B. caballi and T. equi multiplication in vitro with IC50 values of 98.4 ± 4.2, 64.5 ± 3.9, 45.2 ± 5.9, 46.6 ± 2.5, and 71.3 ± 2.7 µM, respectively. The CCK test was used to examine ATR’s cytotoxicity and adverse effects on different animal and human cell lines, the main hosts of piroplasm parasites and it showed that ATR affected human foreskin fibroblasts (HFF), mouse embryonic fibroblast (NIH/3T3) and Madin-Darby Bovine Kidney (MDBK) cell viability in a dose-related effect with a moderate selective index. The combined efficacy of ATR with DA, CF, and AV exhibited a synergistic and additive efficacy toward all tested species. In the in vivo experiment, ATR prohibited B. microti multiplication in mice by 68.17%. The ATR-DA and ATR-AV combination chemotherapies were more potent than ATR monotherapy. These results indicate the prospects of ATR as a drug candidate for piroplasmosis treatment.

MDM2 Promotes Ubiquitin-Independent Degradation of HoxA9: Implications for MLL-Fusion Protein Mediated Transformation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1418-1418
Author(s):  
Huiwu Zhao ◽  
Shenghao Jin ◽  
Alan M. Gewirtz
Keyword(s):  
Half Life ◽  
Fusion Proteins ◽  
Wild Type Mouse ◽  
Mouse Embryonic Fibroblast ◽  
Fibroblast Cells ◽  
Wild Type ◽  
Functional Inactivation ◽  
Embryonic Fibroblast ◽  
Ubiquitylation Assay

Abstract Chromosome translocations involving the mixed lineage leukemia gene (MLL) are associated with aggressive lymphoid and myeloid leukemias. Inappropriate activation of crucial developmental genes has been proposed as one important mechanism for triggering leukemia by MLL-fusions. For example, it is known that the HoxA9 homeodomain protein, a key regulator of hematopoiesis and embryonic development, can be induced by expression of MLL-fusion proteins, and that this event is essential for MLL-dependent leukemogenesis. Recently, it has been suggested that functional inactivation of p53 by MLL-fusions may also contribute to this process. In this study, we explored the effect of p53 suppression by MLL-fusions on HoxA9 function. Using a variety of experimental strategies including co-immunoprecipitation, protein turnover assay and in vivo ubiquitylation assay, we found that the well-known p53 target MDM2 plays an important role in HoxA9 stability. To examine whether MDM2 is associated with HoxA9, Flag tagged MDM2 and HA tagged HoxA9 expression vectors were co-transfected into human HEK293T cells. MDM2 was then immunoprecipitated with anti-Flag beads. HoxA9 was readily detected in the MDM2 immnunocomplex, indicating that HoxA9 indeed interacts with MDM2. To study the functional role of the interaction between MDM2 and HoxA9, the effect of MDM2 on HoxA9 stability was next investigated. Increasing amounts of an MDM2 expression vector was co-transfected into HEK293T cells with an equal amount of a HoxA9 expression plasmid. Following the transfections, HoxA9 protein expression in the lysates from the different transfections was compared. MDM2 was found to promote the degradation HoxA9 in a dose-dependent manner. This effect was specific, since MDM2 did not interfere with Nup98-HoxA9 fusion protein expression in the control experiment. To further demonstrate the effect of MDM2 on HoxA9 stability, HoxA9 half-life was examined in wild type mouse embryonic fibroblast cells and MDM2 negative mouse embryonic fibroblast cells by the measuring HoxA9 protein turnover rate upon cycloheximide treatment. In wild type mouse embryonic fibroblast cells, the HoxA9 protein half-life is less then 30 minutes while HoxA9 protein half-life is about 3 hours in MDM2 knockout mouse embryonic fibroblast cells. Finally, to elucidate the molecular mechanism by which MDM2 promotes HoxA9 degradation, an in vivo ubiquitylation assay was used to test whether MDM2 is a ubiquitin E3 ligase for HoxA9. MDM2 was found to be incompetent in promoting the ubiquitylation of HoxA9. However, the proteasome inhibitor MG-132 could stabilize HoxA9 protein in HoxA9 transfected HEK293T cells. Taken together, these data suggest that MDM2 regulates HoxA9 stability by a ubiquitin-independent proteasome-dependent pathway. Furthermore, our findings indicate that indirect inhibition of MDM2 through functional inactivation of p53 by MLL-fusions might stabilize HoxA9 protein at post-translation level and this might be an important mechanism for the full functional activation of HoxA9 by MLL-associated fusion proteins.

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