ABSTRACT
The innate immune system employs a variety of antimicrobial oxidants to control and kill host-associated bacteria. Hypothiocyanite/hypothiocyanous acid (-OSCN/HOSCN) is one such antimicrobial oxidant that is synthesized by lactoperoxidase, myeloperoxidase, and eosinophil peroxidase at sites throughout the human body. HOSCN has potent antibacterial activity while being largely non-toxic toward human cells. The molecular mechanisms by which bacteria sense and defend themselves against HOSCN have only recently begun to be elaborated, notably by the discovery of bacterial HOSCN reductase (RclA), an HOSCN-degrading enzyme widely conserved among bacteria that live on epithelial surfaces. In this paper, I show that Ni2+ sensitizes Escherichia coli to HOSCN by inhibiting glutathione reductase and that inorganic polyphosphate protects E. coli against this effect, probably by chelating Ni2+ ions. I also found that RclA is very sensitive to inhibition by Cu2+ and Zn2+, metals that are accumulated to high levels by innate immune cells, and that, surprisingly, thioredoxin and thioredoxin reductase are not involved in HOSCN stress resistance in E. coli. These results advance our understanding of the contribution of different oxidative stress responses and redox buffering pathways to HOSCN resistance in E. coli and illustrate important interactions between metal ions and the enzymes bacteria use to defend themselves against oxidative stress. IMPORTANCE: Hypothiocyanite (HOSCN) is an antimicrobial oxidant produced by the innate immune system. The molecular mechanisms by which host-associated bacteria defend themselves against HOSCN have only recently begun to be understood. The results in this paper are significant because they show that the low molecular weight thiol glutathione and enzyme glutathione reductase are critical components of the Escherichia coli HOSCN response, working by a mechanism distinct from that of the HOSCN-specific defenses provided by the RclA, RclB, and RclC proteins and that metal ions (including nickel, copper, and zinc) may impact the ability of bacteria to resist HOSCN by inhibiting specific defensive enzymes (e.g., glutathione reductase or RclA).
Subject(s)
Escherichia coli , Thiocyanates , Escherichia coli/genetics , Escherichia coli/drug effects , Escherichia coli/metabolism , Thiocyanates/pharmacology , Thiocyanates/metabolism , Nickel/pharmacology , Nickel/metabolism , Escherichia coli Proteins/metabolism , Escherichia coli Proteins/genetics , Drug Resistance, Bacterial , Glutathione Reductase/metabolism , Glutathione Reductase/genetics , Anti-Bacterial Agents/pharmacology , Zinc/metabolism , Zinc/pharmacology , Copper/metabolism , Copper/pharmacologyABSTRACT
Nickel-titanium alloy stents are widely used in the interventional treatment of various malignant tumors, and it is important to develop nickel-titanium alloy stents with selective cancer-inhibiting and antibacterial functions to avoid malignant obstruction caused by tumor invasion and bacterial colonization. In this work, an acid-responsive layered double hydroxide (LDH) film was constructed on the surface of a nickel-titanium alloy by hydrothermal treatment. The release of nickel ions from the film in the acidic tumor microenvironment induces an intracellular oxidative stress response that leads to cell death. In addition, the specific surface area of LDH nanosheets could be further regulated by heat treatment to modulate the release of nickel ions in the acidic microenvironment, allowing the antitumor effect to be further enhanced. This acid-responsive LDH film also shows a good antibacterial effect against S. aureus and E. coli. Besides, the LDH film prepared without the introduction of additional elements maintains low toxicity to normal cells in a normal physiological environment. This work offers some guidance for the design of a practical nickel-titanium alloy stent for the interventional treatment of tumors.
Subject(s)
Anti-Bacterial Agents , Hydroxides , Nickel , Tumor Microenvironment , Hydroxides/chemistry , Hydroxides/pharmacology , Tumor Microenvironment/drug effects , Nickel/chemistry , Nickel/pharmacology , Humans , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Alloys/chemistry , Alloys/pharmacology , Escherichia coli/drug effects , Staphylococcus aureus/drug effects , Titanium/chemistry , Titanium/pharmacology , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Hydrogen-Ion ConcentrationABSTRACT
Nickel ferrite nanoparticles (NiFe2O4 NPs) were synthesized using the medicinally important plant Aloe vera leaf extract, and their structural, morphological, and magnetic properties were characterized by x-ray diffraction (XRD), fourier transform infrared (FTIR), scanning electron microscopy (SEM), energy dispersive x-ray (EDX), and vibrating sample magnetometer (VSM). The synthesized NPs were soft ferromagnetic and spinel in nature, with an average particle size of 22.2 nm. To the best of our understanding, this is the first comprehensive investigation into the antibacterial, anticandidal, antibiofilm, and antihyphal properties of NiFe2O4 NPs against C. albicans as well as drug-resistant gram-positive methicillin-resistant Staphylococcus aureus (MRSA) and gram-negative multidrug resistant Pseudomonas aeruginosa (MDR-P. aeruginosa) bacteria. NiFe2O4 NPs showed potent antimicrobial activity (MIC 1.6-2 mg/mL) against the test pathogens. NiFe2O4 NPs at 0.5 mg/mL suppressed biofilm formation by 49.5-53.1 % in test pathogens. The study found that the NPs not only prevent the formation of biofilm, but also eliminate existing mature biofilms by 50.5-75.79 % at 0.5 mg/mL, which was further validated by SEM. SEM examination revealed a reduction in the number of cells that form biofilms and adhere to the surface. Additionally, it considerably impeded the colonization and aggregation of the biofilm strains on the glass surface. Light microscopic examination demonstrated that NPs effectively prevent the expansion of hyphae, filaments, and yeast-to-hyphae transformation in C. albicans, resulting in a substantial decrease in their ability to cause infection. Moreover, SEM images of the treated cells exhibited the presence of wrinkles, deformities, and impaired cell walls, which suggests an alteration and instability of the membrane. This study demonstrated the efficacy of the greenly manufactured NPs in suppressing the proliferation of candida, drug-resistant bacteria, and their preexisting biofilms, as well as yeast-to-hyphae transformation. Therefore, these NPs with broad spectrum applications could be utilized in health settings to mitigate biofilm-related health conditions caused by pathogenic microbial strains.
Subject(s)
Anti-Bacterial Agents , Biofilms , Candida albicans , Ferric Compounds , Methicillin-Resistant Staphylococcus aureus , Microbial Sensitivity Tests , Nickel , Pseudomonas aeruginosa , Biofilms/drug effects , Nickel/chemistry , Nickel/pharmacology , Ferric Compounds/pharmacology , Ferric Compounds/chemistry , Candida albicans/drug effects , Pseudomonas aeruginosa/drug effects , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Methicillin-Resistant Staphylococcus aureus/drug effects , Plant Extracts/pharmacology , Plant Extracts/chemistry , Plant Leaves/chemistry , Plant Leaves/microbiology , Aloe/chemistry , X-Ray Diffraction , Particle Size , Microscopy, Electron, Scanning , Antifungal Agents/pharmacology , Antifungal Agents/chemistry , Spectroscopy, Fourier Transform Infrared , Nanoparticles/chemistryABSTRACT
Bacterial infections pose a global concern due to high fatality rates, particularly with the rise of drug-resistant bacteria and biofilm formation. There is an urgent need for innovative strategies to combat this issue. A study on chemodynamic therapy (CDT) using nanozymes in conjunction with photothermal therapy (PTT) has displayed potential in addressing drug-resistant bacterial infections. However, the effectiveness of this combined approach is limited by inadequate light absorption. This work suggests the NiOx nanoparticles enriched with oxygen vacancies enhance CDT and PTT to overcome this challenge. The presence of oxygen vacancies in NiOx can reduce the energy gap between its valence band and conduction band, facilitating oxygen adsorption. NiOx has exhibited notable antibacterial properties and complete eradication of biofilms in both laboratory and animal trials. In animal abscess models, NiOx demonstrated antibacterial and anti-inflammatory effects in the initial stages, while also promoting wound healing and tissue regeneration by influencing immune factors and encouraging collagen deposition and neovascularization. With positive biosafety and biocompatibility profiles, the oxygen vacancy-enhanced CDT and PTT therapy proposed in this article hold promise for effective sterilization, deep biofilm removal, and treatment of infections caused by drug-resistant bacteria. STATEMENT OF SIGNIFICANCE: This study constructs oxygen vacancies NiOx nanoparticles (NiOx NPs) to improve the efficacy of photothermal therapy and chemodynamic therapy. The presence of oxygen vacancies in NiOx NPs helps bridge the energy gap between its valence band and conduction band, facilitating oxygen adsorption and improving catalytic efficiency. In both in vivo and in vitro antibacterial experiments, NiOx NPs demonstrate effective antibacterial and anti-inflammatory properties. Furthermore, it aids in wound healing and tissue regeneration by modulating immune factors, collagen deposition, and angiogenesis. This approach presents a promising collaborative strategy for utilizing nickel-based defective nanomaterials in combating deep drug-resistant bacterial infections.
Subject(s)
Anti-Bacterial Agents , Methicillin-Resistant Staphylococcus aureus , Nickel , Oxygen , Nickel/chemistry , Nickel/pharmacology , Methicillin-Resistant Staphylococcus aureus/drug effects , Animals , Oxygen/chemistry , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Mice , Staphylococcal Infections/drug therapy , Staphylococcal Infections/therapy , Photothermal Therapy , Biofilms/drug effects , Nanoparticles/chemistry , Nanoparticles/therapeutic use , PhototherapyABSTRACT
Trace heavy metals (HMs) such as copper (Cu) and nickel (Ni) are toxic to plants, especially tomato at high levels. In this study, biochar (BC) was treated with amino acids (AA) to enhance amino functional groups, which effectively alleviated the adverse effects of heavy metals (HMs) on tomato growth. Hence, this study aimed to evaluate the effect of glycine and alanine modified BC (GBC/ABC) on various tomato growth parameters, its physiology, fruit yield and Cu/Ni uptake under Cu and Ni stresses. In a pot experiment, there was 21 treatments with three replications having two rates of simple BC and glycine/alanine enriched BC (0.5% and 1% (w/w). Cu and Ni stresses were added at 150 mg kg-1 respectively. Plants were harvested after 120 days of sowing and subjected to various analysis. Under Cu and Ni stresses, tomato roots accumulated more Cu and Ni than shoots and fruits, while GBC and ABC application significantly enhanced the root and shoot dry weight irrelevant to the stress conditions. Both rates of GBC decreased the malondialdehyde and hydrogen peroxide levels in plants. The addition of 0.5% GBC with Cu enhanced the tomato fruit dry weight by 1.3 folds in comparison to the control treatment; while tomato fruit juice content also increased (50%) in the presence of 0.5% GBC with Ni as compared to control. In summary, these results demonstrated that lower rate of GBCâ¼0.5% proved to be the best in mitigating the Cu and Ni stress on tomato plant growth by enhancing the fruit production.
Subject(s)
Amino Acids , Charcoal , Copper , Fruit , Nickel , Solanum lycopersicum , Solanum lycopersicum/drug effects , Solanum lycopersicum/growth & development , Solanum lycopersicum/metabolism , Nickel/pharmacology , Fruit/drug effects , Fruit/growth & development , Fruit/metabolism , Charcoal/pharmacology , Amino Acids/metabolism , Soil Pollutants , Stress, Physiological/drug effects , Soil/chemistryABSTRACT
OBJECTIVE: Breast cancer ranks second in terms of the highest number of cancer deaths for women worldwide and is one of the leading causes of death from cancer in women. The drug that is often used for chemotherapy is cisplatin. However, cisplatin drugs have a number of problems, including lack of selectivity, unwanted side effects, resistance, and toxicity in the body. In this work, we investigated Ni(II) cysteine-tyrosine dithiocarbamate complex against breast cancer. METHODS: Research on the new complex compound Ni(II) cysteine-tyrosine dithiocarbamate have several stages including synthesis, characterization, in-silico and in-vitro testing of MCF-7 cells for anticancer drugs. The synthesis involved reacting cysteine, CS2, KOH and tyrosine with Mn metal. The new complex compound Ni(II) cysteine-tyrosine dithiocarbamate has been synthesized, characterized, and tested in vitro MCF-7 cells for anticancer drugs. Characterization tests such as melting point, conductivity, SEM-EDS, UV Vis, XRD, and FT-IR spectroscopy have been carried out. RESULT: The synthesis yielded a 60,16%, conversion with a melting point of 216-218 oC and a conductivity value of 0.4 mS/cm. In vitro test results showed morphological changes (apoptosis) in MCF-7 cancer cells starting at a sample concentration of 250 µg/mL and an IC50 value of 618.40 µg/mL. Molecular docking study of Ni(II) cysteine-tyrosine dithiocarbamate complex identified with 4,4',4''-[(2R)-butane-1,1,2-triyl]triphenol - Estrogen α showing active site with acidic residue amino E323, M388, L387, G390 and I389. Hydrophobic and hydrophobic bonds are seen in Ni(II) cysteine-tyrosine dithiocarbamate - Estrogen α has a binding energy of -80.9429 kJ /mol. CONCLUSION: there were 5 residues responsible for maintaining the ligand binding stable. The compound had significant Hbond contact intensity, however, it was not strong enough to make a significant anticancer effect. Though the synthesized compound shows low bioactivity, this research is expected to give valuable insight into the effect of molecular structure on anticancer activity.
Subject(s)
Antineoplastic Agents , Breast Neoplasms , Cell Proliferation , Cysteine , Molecular Docking Simulation , Molecular Dynamics Simulation , Nickel , Thiocarbamates , Tyrosine , Humans , Nickel/chemistry , Nickel/pharmacology , Thiocarbamates/pharmacology , Thiocarbamates/chemistry , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/chemical synthesis , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Tyrosine/pharmacology , Tyrosine/chemistry , MCF-7 Cells , Female , Cysteine/chemistry , Cysteine/pharmacology , Cell Proliferation/drug effects , Coordination Complexes/pharmacology , Coordination Complexes/chemistry , Coordination Complexes/chemical synthesis , Apoptosis/drug effects , Tumor Cells, CulturedABSTRACT
Triple-negative breast cancer (TNBC) poses challenges in therapy due to the absence of target expression such as estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2). Frequently, the treatment of TNBC involves the combination of several therapeutics. However, an enhanced therapeutic effect can be also achieved within a single molecule. The efficacy of raloxifene can be improved by designing a raloxifene-based hybrid drug bearing a 2,2'-bipyridine moiety (2). Integration of platinum(II), palladium(II), and nickel(II) complexes into this structure dramatically changed the cytotoxicity. The platinum(II) dichloride complexâ 3 did not demonstrate any activity, while palladium(II) and nickel(II) dichloride complexesâ 4 and 5 exhibited various cytotoxic behavior towards different types of hormone-receptor positive (HR+) cancer and TNBCâ cell lines. The replacement of the two chlorido ligands in 3-5 with a dicarbollide (carborate) ion [C2B9H11]2- resulted in reduced activity of compoundsâ 6, 7, and 8. However, the palladacarborane complex 7 demonstrated higher selectivity towards TNBC. Furthermore, the mechanism of action was shifted from cytotoxic to explicitly cytostatic with detectable proliferation arrest and accelerated aging, characterized by senescence-associated phenotype of TNBCâ cells. This study provides valuable insights into the development of hybrid therapeutics against TNBC.
Subject(s)
Antineoplastic Agents , Cell Proliferation , Coordination Complexes , Drug Screening Assays, Antitumor , Nickel , Palladium , Platinum , Raloxifene Hydrochloride , Triple Negative Breast Neoplasms , Humans , Palladium/chemistry , Palladium/pharmacology , Nickel/chemistry , Nickel/pharmacology , Triple Negative Breast Neoplasms/drug therapy , Triple Negative Breast Neoplasms/pathology , Antineoplastic Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/chemical synthesis , Platinum/chemistry , Platinum/pharmacology , Coordination Complexes/chemistry , Coordination Complexes/pharmacology , Coordination Complexes/chemical synthesis , Cell Proliferation/drug effects , Raloxifene Hydrochloride/chemistry , Raloxifene Hydrochloride/pharmacology , Cell Line, Tumor , Structure-Activity Relationship , Molecular Structure , Dose-Response Relationship, Drug , FemaleABSTRACT
Plant growth-promoting rhizobacteria (PGPRs) have been utilized to immobilize heavy metals, limiting their translocation in metal contaminated settings. However, studies on the mechanisms and interactions that elucidate how PGPRs mediate Nickel (Ni) tolerance in plants are rare. Thus, in this study we investigated how two pre-characterized heavy metal tolerant isolates of Morganella morganii (ABT9 and ABT3) improve Ni stress tolerance in Arabidopsis while enhancing its growth and yield. Arabidopsis seedlings were grown for five weeks in control/Ni contaminated (control, 1.5 mM and 2.5 mM) potted soil, in the presence or absence of PGPRs. Plant growth characteristics, quantum yield, and antioxidative enzymatic activities were analyzed to assess the influence of PGPRs on plant physiology. Oxidative stress tolerance was quantified by measuring MDA accumulation in Arabidopsis plants. As expected, Ni stress substantially reduced plant growth (shoot and root fresh weight by 53.25% and 58.77%, dry weight by 49.80% and 57.41% and length by 47.16% and 64.63% over control), chlorophyll content and quantum yield (by 40.21% and 54.37% over control). It also increased MDA content by 84.28% at higher (2.5 mM) Ni concentrations. In contrast, inoculation with M. morganii led to significant improvements in leaf chlorophyll, quantum yield, and Arabidopsis biomass production. The mitigation of adverse effects of Ni stress on biomass observed in M. morganii-inoculated plants was attributed to the enhancement of antioxidative enzyme activities compared to Ni-treated plants. This upregulation of the antioxidative defense mechanism mitigated Ni-induced oxidative stress, leading to improved performance of the photosynthetic machinery, which, in turn, enhanced chlorophyll content and quantum yield. Understanding the underlying mechanisms of these tolerance-inducing processes will help to complete the picture of PGPRs-mediated defense signaling. Thus, it suggests that M. morganii PGPRs candidate can potentially be utilized for plant growth promotion by reducing oxidative stress via upregulating antioxidant defense systems in Ni-contaminated soils and reducing Ni metal uptake.
Subject(s)
Arabidopsis , Morganella morganii , Nickel/pharmacology , Antioxidants , ChlorophyllABSTRACT
Cancer cells release extracellular vesicles (EVs) that participate in altering the proximal tumor environment and distal tissues to promote cancer progression. Chronic exposure to nickel (Ni), a human group I carcinogen, results in epigenetic changes that promotes epithelial to mesenchymal transition (EMT). Cells that undergo EMT demonstrate various molecular changes, including elevated levels of the mesenchymal cadherin N-cadherin (N-CAD) and the transcription factor Zinc finger E-box binding homeobox 1 (ZEB1). Moreover, the molecular changes following EMT induce changes in cellular behavior, including anchorage-independent growth, which contributes to cancer cells detaching from tumor bulk during the metastatic process. Here, we present data demonstrating that EVs from Ni-exposed cells induce EMT in recipient BEAS-2B cells in the absence of Ni. Moreover, we show evidence that the EVs from Ni-altered cells package the transcription factor nuclear protein 1 (NUPR1), a transcription factor associated with Ni exposure and cancer progression. Moreover, our data demonstrates that the NUPR1 in the EVs becomes part of the recipient cell proteomic milieu and carry the NUPR1 to the nuclear space of the recipient cell. Interestingly, knockdown of NUPR1 in Ni-transformed cells suppressed NUPR1 packaging in the EVs, and nanoparticle tracking analysis (NTA) demonstrated decreased EV release. Reduction of NUPR1 in EVs resulted in diminished EMT capacity that resulted in decreased anchorage independent growth. This study is the first to demonstrate the role of NUPR1 in extracellular vesicle-mediate cancer progression.
Subject(s)
Extracellular Vesicles , Neoplasms , Humans , Cell Line, Tumor , Nickel/pharmacology , Nuclear Proteins , Epithelial-Mesenchymal Transition , Proteomics , Transcription Factors/genetics , Transcription Factors/metabolism , Extracellular Vesicles/metabolismABSTRACT
In the present study, the vanadium pentoxide (V2O5) nickel-doped vanadium pentoxide (Ni@V2O5) was prepared and determined for in vitro anticancer activity. The structural characterization of the prepared V2O5 and Ni@V2O5 was determined using diverse morphological and spectroscopic analyses. The DRS-UV analysis displayed the absorbance at 215 nm for V2O5 and 331 nm for Ni@V2O5 as the primary validation of the synthesis of V2O5 and Ni@V2O5. The EDS spectra exhibited the presence of 30% of O, 69% of V, and 1% of Ni and the EDS mapping showed the constant dispersion. The FE-SEM and FE-TEM analysis showed the V2O5 nanoparticles are rectangle-shaped and nanocomposites have excellent interfaces between nickel and V2O5. The X-ray photoelectron spectroscopy (XPS) investigation of Ni@V2O5 nanocomposite endorses the occurrence of elements V, O, and Ni. The in vitro MTT assay clearly showed that the V2O5 and Ni@V2O5 have significantly inhibited the proliferation of B16F10 skin cancer cells. In addition, the nanocomposite produces the endogenous reactive oxygen species in the mitochondria, causes the mitochondrial membrane and nuclear damage, and consequently induces apoptosis by caspase 9/3 enzymatic activity in skin cancer cells. Also, the western blot analysis showed that the nanocomposite suppresses the oncogenic marker proteins such as PI3K, Akt, and mTOR in the skin cancer cells. Together, the results showed that Ni@V2O5 can be used as an auspicious anticancer agent against skin cancer.
Subject(s)
Nanocomposites , Skin Neoplasms , Vanadium Compounds , Humans , Phosphatidylinositol 3-Kinases , Nickel/pharmacology , Proto-Oncogene Proteins c-akt , Signal Transduction , TOR Serine-Threonine Kinases , Photoelectron Spectroscopy , Apoptosis , Skin Neoplasms/drug therapyABSTRACT
Despite the recent progress in the diagnosis of tuberculosis (TB), the chemotherapeutic management of TB continues to be challenging. Mycobacterium tuberculosis (Mtb), the etiological agent of TB, is classified as the 13th leading cause of death globally. In addition, 450,000 people were reported to develop multi-drug-resistant TB globally. The current project focuses on targeting methionine aminopeptidase (MetAP), an essential protein for the viability of Mtb. MetAP is a metalloprotease that catalyzes the excision of the N-terminal methionine (NME) during protein synthesis, allowing the enzyme to be an auspicious target for the development of novel therapeutic agents for the treatment of TB. Mtb possesses two MetAP1 isoforms, MtMetAP1a and MtMetAP1c, which are vital for Mtb viability and, hence, a promising chemotherapeutic target for Mtb therapy. In this study, we cloned and overexpressed recombinant MtMetAP1c. We investigated the in vitro inhibitory effect of the novel MetAP inhibitor, OJT008, on the cobalt ion- and nickel ion-activated MtMetAP1c, and the mechanism of action was elucidated through an in silico approach. The compound's potency against replicating and multi-drug-resistant (MDR) Mtb strains was also investigated. The induction of the overexpressed recombinant MtMetAP1c was optimized at 8 h with a final concentration of 1 mM Isopropyl ß-D-1-thiogalactopyranoside. The average yield from 1 L of Escherichia coli culture for MtMetAP1c was 4.65 mg. A preliminary MtMetAP1c metal dependency screen showed optimum activation with nickel and cobalt ions occurred at 100 µM. The half-maximal inhibitory concentration (IC50) values of OJT008 against MtMetAP1c activated with CoCl2 and NiCl2 were 11 µM and 40 µM, respectively. The in silico study showed OJT008 strongly binds to both metal-activated MtMetAP1c, as evidenced by strong molecular interactions and a higher binding score, thereby corroborating our result. This in silico study validated the pharmacophore's metal specificity. The potency of OJT008 against both active and MDR Mtb was <0.063 µg/mL. Our study reports OJT008 as an inhibitor of MtMetAP1c, which is potent at low micromolar concentrations against both active susceptible and MDR Mtb. These results suggest OJT008 is a potential lead compound for the development of novel small molecules for the therapeutic management of TB.
Subject(s)
Mycobacterium tuberculosis , Tuberculosis, Multidrug-Resistant , Tuberculosis , Humans , Nickel/pharmacology , Aminopeptidases/genetics , Aminopeptidases/chemistry , Tuberculosis/microbiology , Methionyl Aminopeptidases , Tuberculosis, Multidrug-Resistant/drug therapy , Metals/pharmacology , Cobalt/pharmacology , Antitubercular Agents/chemistryABSTRACT
The structural genes for the nickel and cobalt resistance of the conjugative plasmid pEJH 501 of Hafnia alvei 5-5, contained on a SalI-EcoRI fragment of 4.8 kb, were cloned and sequenced. The DNA sequence included five genes in the following order: ncrA, ncrB, ncrC, ncrY, and ncrX. The predicted amino acid sequences of ncrA were homologous to the amino acid sequences of nreB of Achromobacter xylosoxidans 31A. Expression of ncr with the T7 RNA polymerase-promoter system allowed Escherichia coli BL21 (DE3) to overexpress NcrA, NcrB, and NcrC but not NcrY, and NcrX. The apparent molecular masses of NcrA, NcrB, and NcrC were 30, 33, and 17 kDa, respectively. Primer-extension analysis showed that ncr mRNA started at nucleotide position 23 upstream from ncrA. The promoter region of the ncr operon possessed a strong, putative -35 element of sigma(32)-type promoter sequence, and transcriptional 'lacZ fusion studies indicated that the -35 element influenced sigma(32)-specific transcription (AU)
Los genes estructurales de la resistencia a níquel y cobalto del plásmido conjugativo pEJH 501 de Hafnia alvei 5-5, contenido en un fragmento SalI-EcoRI de 4,8 kb, fueron clonados y secuenciados. La secuencia de DNA incluye cinco genes en el siguiente orden: ncrA, ncrB, ncrC, ncrY, y ncrX. Las secuencias de aminoácidos equivalentes a ncrA fueron homólogas a las secuencias de aminoácidos codificadas por nreB en Achromobacter xylosoxidans 31A. La expresión de los genes ncr mediante el sistema promotor de la RNA polimerasa T7 permite a Escherichia coli BL21 (DE3) sobreexpresar NcrA, NcrB, y NcrC, pero no NcrY ni NcrX. Los pesos moleculares aparentes de NcrA, NcrB y NcrC fueron 30, 33, y 17 kDa, respectivamente. El análisis de extensión de los cebadores mostró que el mRNA de ncr se iniciaba a una distancia de 23 nucleótidos corriente arriba del ncrA.La región promotora del operón ncr posee una fuerte secuencia promotora de tipo sigma32 en la posición -35, y estudios transcripcionales de fusión con ´lacZ indicaron que el elemento situado en -35 influye sobre la transcripción específica de sigma32 (AU)
Subject(s)
DNA Primers , Cobalt/pharmacology , DNA, Bacterial , Drug Resistance, Bacterial , Hafnia/genetics , Molecular Sequence Data , Nickel/pharmacology , Cloning, Molecular , Base Sequence , Transcription, Genetic , Genotype , Operon/genetics , Plasmids/genetics , Restriction MappingABSTRACT
Hafnia alvei 5-5, isolated from a soil-litter mixture underneath the canopy of the nickel-hyperaccumulating tree Sebertia acuminata (Sapotaceae) in New Caledonia, was found to be resistant to 30 mM Ni(2+) or 2 mM Co(2+). The 70-kb plasmid, pEJH 501, was transferred by conjugation to Escherichia coli, Serratia marcescens, and Klebsiella oxytoca. Transconjugant strains expressed inducible nickel resistance to between 5 and 17 mM Ni(2+), and cobalt resistance to 2 mM Co(2+). A 4.8-kb Sal- EcoRI fragment containing the nickel resistance determinant was subcloned, and the hybrid plasmid was found to confer a moderate level of resistance to nickel (7 mM Ni(2+)) even to E. coli. The expression of nickel resistance was inducible by exposure to nickel chloride at a concentration as low as 0.5 mM Ni(2+). By random Tn phoA'-1 insertion mutagenesis, the fragment was shown to have structural genes as well as regulatory regions for nickel resistance. Southern hybridization studies showed that the nickel-resistance determinant from pEJH501 of H. alvei 5-5 was homologous to that of pTOM9 from Alcaligenes xylosoxydans 31A (AU)
Hafnia alvei 5-5, aislada en un vertedero bajo la copa del árbol hiperacumulante de níquel Sebertia acuminata (Sapoteacea) en Nueva Caledonia, resultó ser resistente a 30 mM Ni2+ y 2 mM Co2+. El plásmido de 70 kilopares de bases (kb), pEJH501 se transfirió por conjugación a Escherichia coli, Serratia marcescens y Klebsiella oxytoca. Las cepas transconjugantes expresaron resistencia inducible a entre 5 y 17 mM Ni2+, y a 2 mM Co2+. Se subclonó un fragmento Sal-EcoRI que contenía el determinante de resistencia al níquel, y el plásmido híbrido se descubrió que confería un nivel de resistencia moderado al níquel (7 mM Ni2+), incluso en E. coli. La expresión de la resistencia al níquel era inducible por exposición a concentraciones de cloruro de níquel de como mínimo 0,5 mM Ni2+. Mediante mutagénesis por inserción aleatoria de TnphoA'-1, se encontraron en el fragmento tanto genes estructurales como regiones reguladoras para la resistencia al níquel. Estudios de hibridación southern mostraron que el determinante de la resistencia al níquel del plásmido pEJH501 en H. alvei 5-5 era homólogo al de pTOM9 de Alcaligenes xylosoxydans 31A (AU)
Subject(s)
Plasmids/genetics , Hafnia alvei/genetics , Drug Resistance, Bacterial/genetics , Nickel/pharmacology , Microbial Sensitivity Tests , DNA Probes , Conjugation, Genetic , Cloning, Molecular , Bacteria/classification , Mutation , Models, Genetic , Sequence Homology, Nucleic AcidABSTRACT
El objetivo de este trabajo fue estudiar los efectos de la administración de clorato de níquel en el epitelio de revestimiento y hueso palatino de fetos de ratas. Fue administrada una inyección intraperitoneal de clorato de níquel (30mg/kg de peso corporal) en el 10º día de preñez. Las ratas fueron sacrigicadas en el 20º día y, los fetos fueron inmersos en solución fijadora. Las cabezas de los fetos control y tratados fueron incluidas, cortadas y coloradas para la aplicación de los análisis morfométricos y estereológicos. Se utilizó el test no paramétrico de Mann-Whitney. Los resultados mostraron alteración en el epitelio de revestimiento de los palatos duro y blando y disminución en la densidad de trabéculas óseas. Estos resultados sugieren aspectos de inmaturidad en el feto tratado (AU)
Subject(s)
Animals , Female , Rats , Chlorates/administration & dosage , Epithelium , Palate , Nickel/pharmacology , FetusABSTRACT
Treatment with certain DNA-damaging agents induce a complex cellular response comprising pertubation of cell cycle progression and/or apoptosis on proliferating mammalian cells. Our studies were focused on the cellular effects of nickel (II) acetate, DNA-damaging agent, on Chinese hamster ovary (CHO) cells. Fragmented DNAs were examined by agarose gel electrophoresis and cell cycle was determined by DNA flow cytometry using propidium iodide fluorescence. Apparent DNA laddering was observed in cells treated with 240 microM nickel (II) and increased with a concentration-dependent manner. Treatment of nickel (II) acetate resulted in apoptosis which was accompanied by G2/M cell accumulation. Proportion of CHO cells in G2/M phase was also significantly increased in cells exposed to at least 480 microM nickel (II) from 57.7% of cells in the G0/G1 phase, 34.7% in the S phase, and 7.6% in the G2/M1 phase for 0 microM nickel (II), to 58.6%, 14.5%, and 26.9% for 640 microM nickel (II). These findings suggest that nickel (II) can modulate cellular response through some common effectors involving in both apoptotic and cell cycle regulatory pathways.
Subject(s)
Animals , Apoptosis/drug effects , CHO Cells/drug effects , CHO Cells/cytology , Cell Cycle/drug effects , DNA Fragmentation/drug effects , Flow Cytometry , G2 Phase/drug effects , Cricetinae , Mitosis/drug effects , Nickel/pharmacologyABSTRACT
O níquel, administrado intraperitonealmente na rata prenhe, no 10º dia de prenhez resultou em peso fetal diminuído, glândulas nasais pequenas, com núcleos mais alongados.
Subject(s)
Animals , Female , Pregnancy , Rats , Fetus/drug effects , Nasal Mucosa/cytology , Nasal Mucosa/drug effects , Nickel/pharmacology , Karyometry , Rats, WistarABSTRACT
Rat brain tubulin in a proper buffered solution became insoluble in the presence of 10 mM NiCl2, and sedimented at centrifugal forces as low as 500 x g for 30 min. Both nickel-sedimented and microtubular tubulin conserved 65 of colchicine binding activity after 25 days of storage at -20 degrees C. However in brain cytosol, only 9 of the initial binding activity was conserved. The electrophoretic mobility of tubulin recovered from aggregates also remained unaltered. Therefore the aggregates formed with Ni2+ share important physicochemical properties with microtubules.
Subject(s)
Animals , Male , Rats , Microtubules/chemistry , Nickel/pharmacology , Tubulin/chemistry , Centrifugation , Chemistry, Physical , Colchicine , Electrophoresis, Polyacrylamide Gel , Brain Chemistry , Solubility , Tubulin/metabolismABSTRACT
Rat brain tubulin in a proper buffered solution became insoluble in the presence of 10 mM NiCl2, and sedimented at centrifugal forces as low as 500 x g for 30 min. Both nickel-sedimented and microtubular tubulin conserved 65 of colchicine binding activity after 25 days of storage at -20 degrees C. However in brain cytosol, only 9 of the initial binding activity was conserved. The electrophoretic mobility of tubulin recovered from aggregates also remained unaltered. Therefore the aggregates formed with Ni2+ share important physicochemical properties with microtubules.(AU)
Subject(s)
Animals , Male , Rats , Microtubules/chemistry , Nickel/pharmacology , Tubulin/chemistry , Brain Chemistry , Centrifugation , Chemistry, Physical , Colchicine/metabolism , Electrophoresis, Polyacrylamide Gel , Solubility , Tubulin/metabolismABSTRACT
The fact that glycerol preserves microtubules from depolymerizing in vitro, and that some ions such as Ca(II) and Mg(II), regulate the assembly-disassembly process of these structures, induced us to study the effect of several sugars, glycols and metal ions on solubility and colchicine affinity of tubulin in rat brain homogenates, and of purified microtubular protein. Inhibition of colchicine binding was significant with glycerol, polyethylene glycol 1000 (PEG-2) and the ions A1(III), Co(II), Ni(II), while compounds structurally related to glycero (glucose and sucrose) did not inhibition it. Mannitol, instead, increased the activity a 47% over control. Apparently the presence of some compounds in brain homogenates [PEG-2 (1000) and NI (II)] favored tubulin sedimentation when these latterwere centrifuged at 100,000 x g for 150 min at 20 degrees C, but the form in which tubulin becomes aggregated in the pellet is unknown. Nickel ion madeinsoluble microtubular protein of homogenates and the purified one by more than 90% without causing significant inhibition of the colchicine binding. The sediment containing nickel-treated two cycles purified microtubular protein observed with the electron microscope did not present microtubules, but it revealed the presence of irregular, wavy and streteched structures, but it revealed the presence of irregular, wavy and stretched structures bearing highly dense dotted material. The sediments became soluble in phosphate-glutamate buffer (pH 6.8) and, when incubated in polymerizing conditions, gave rise to microtubules undistinguishable from those prepared with untreated purified protein (AU)
Subject(s)
Animals , Female , Rats , Brain Chemistry , Carbohydrates/pharmacology , Cations/pharmacology , Colchicine/metabolism , Glycols/pharmacology , Nickel/pharmacology , Tubulina/metabolism , Aluminum/pharmacology , Cobalt/pharmacology , Fixatives/pharmacology , Microtubules/drug effects , Nerve Tissue Proteins/metabolism , Polymers , Chemical Precipitation , Solubility , Protein Binding/drug effectsABSTRACT
The fact that glycerol preserves microtubules from depolymerizing in vitro, and that some ions such as Ca(II) and Mg(II), regulate the assembly-disassembly process of these structures, induced us to study the effect of several sugars, glycols and metal ions on solubility and colchicine affinity of tubulin in rat brain homogenates, and of purified microtubular protein. Inhibition of colchicine binding was significant with glycerol, polyethylene glycol 1000 (PEG-2) and the ions A1(III), Co(II), Ni(II), while compounds structurally related to glycero (glucose and sucrose) did not inhibition it. Mannitol, instead, increased the activity a 47% over control. Apparently the presence of some compounds in brain homogenates [PEG-2 (1000) and NI (II)] favored tubulin sedimentation when these latterwere centrifuged at 100,000 x g for 150 min at 20 degrees C, but the form in which tubulin becomes aggregated in the pellet is unknown. Nickel ion madeinsoluble microtubular protein of homogenates and the purified one by more than 90% without causing significant inhibition of the colchicine binding. The sediment containing nickel-treated two cycles purified microtubular protein observed with the electron microscope did not present microtubules, but it revealed the presence of irregular, wavy and streteched structures, but it revealed the presence of irregular, wavy and stretched structures bearing highly dense dotted material. The sediments became soluble in phosphate-glutamate buffer (pH 6.8) and, when incubated in polymerizing conditions, gave rise to microtubules undistinguishable from those prepared with untreated purified protein