Interaction of DNA-lesions induced by sodium fluoride and radiation and its influence in apoptotic induction in cancer cell lines

AbstractFluoride is an essential trace element but also an environmental contaminant with major sources of exposure being drinking water, food and pesticides. Previous studies showed that sodium fluoride (NaF) at 5mM or more is required to induce apoptosis and chromosome aberrations and proposed that DNA damage and apoptosis play an important role in toxicity of excessive fluoride. The aim of this study is directed to understand the nature of DNA-lesions induced by NaF by allowing its interaction with radiation induced DNA-lesions. NaF 5mM was used after observing inability to induce DNA damages and apoptosis by single exposure with 50μM or 1mM NaF. Co-exposure to NaF and radiation significantly increased the frequency of aberrant metaphases and exchange aberrations in human lymphocytes and arrested the cells in G1 stage instead of apoptotic death. Flow cytometric analysis, DNA fragmentation and PARP-cleavage analysis clearly indicated that 5mM NaF together with radiation (1Gy) induced apoptosis in both U87 and K562 cells due to down regulation of expression of anti-apoptotic proteins, like Bcl2 in U87 and inhibitors of apoptotic proteins like survivin and cIAP in K562 cells. This study herein suggested that single exposure with extremely low concentration of NaF unable to induce DNA lesions whereas higher concentration induced DNA lesions interact with the radiation-induced DNA lesions. Both are probably repaired rapidly thus showed increased interactive effect. Coexposure to NaF and radiation induces more apoptosis in cancer cell lines which could be due to increased exchange aberrations through lesions interaction and downregulating anti-apoptotic genes

Global burden of 288 causes of death and life expectancy decomposition in 204 countries and territories and 811 subnational locations, 1990–2021: a systematic analysis for the Global Burden of Disease Study 2021

BACKGROUND Regular, detailed reporting on population health by underlying cause of death is fundamental for public health decision making. Cause-specific estimates of mortality and the subsequent effects on life expectancy worldwide are valuable metrics to gauge progress in reducing mortality rates. These estimates are particularly important following large-scale mortality spikes, such as the COVID-19 pandemic. When systematically analysed, mortality rates and life expectancy allow comparisons of the consequences of causes of death globally and over time, providing a nuanced understanding of the effect of these causes on global populations. METHODS The Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2021 cause-of-death analysis estimated mortality and years of life lost (YLLs) from 288 causes of death by age-sex-location-year in 204 countries and territories and 811 subnational locations for each year from 1990 until 2021. The analysis used 56 604 data sources, including data from vital registration and verbal autopsy as well as surveys, censuses, surveillance systems, and cancer registries, among others. As with previous GBD rounds, cause-specific death rates for most causes were estimated using the Cause of Death Ensemble model-a modelling tool developed for GBD to assess the out-of-sample predictive validity of different statistical models and covariate permutations and combine those results to produce cause-specific mortality estimates-with alternative strategies adapted to model causes with insufficient data, substantial changes in reporting over the study period, or unusual epidemiology. YLLs were computed as the product of the number of deaths for each cause-age-sex-location-year and the standard life expectancy at each age. As part of the modelling process, uncertainty intervals (UIs) were generated using the 2·5th and 97·5th percentiles from a 1000-draw distribution for each metric. We decomposed life expectancy by cause of death, location, and year to show cause-specific effects on life expectancy from 1990 to 2021. We also used the coefficient of variation and the fraction of population affected by 90% of deaths to highlight concentrations of mortality. Findings are reported in counts and age-standardised rates. Methodological improvements for cause-of-death estimates in GBD 2021 include the expansion of under-5-years age group to include four new age groups, enhanced methods to account for stochastic variation of sparse data, and the inclusion of COVID-19 and other pandemic-related mortality-which includes excess mortality associated with the pandemic, excluding COVID-19, lower respiratory infections, measles, malaria, and pertussis. For this analysis, 199 new country-years of vital registration cause-of-death data, 5 country-years of surveillance data, 21 country-years of verbal autopsy data, and 94 country-years of other data types were added to those used in previous GBD rounds. FINDINGS The leading causes of age-standardised deaths globally were the same in 2019 as they were in 1990; in descending order, these were, ischaemic heart disease, stroke, chronic obstructive pulmonary disease, and lower respiratory infections. In 2021, however, COVID-19 replaced stroke as the second-leading age-standardised cause of death, with 94·0 deaths (95% UI 89·2-100·0) per 100 000 population. The COVID-19 pandemic shifted the rankings of the leading five causes, lowering stroke to the third-leading and chronic obstructive pulmonary disease to the fourth-leading position. In 2021, the highest age-standardised death rates from COVID-19 occurred in sub-Saharan Africa (271·0 deaths [250·1-290·7] per 100 000 population) and Latin America and the Caribbean (195·4 deaths [182·1-211·4] per 100 000 population). The lowest age-standardised death rates from COVID-19 were in the high-income super-region (48·1 deaths [47·4-48·8] per 100 000 population) and southeast Asia, east Asia, and Oceania (23·2 deaths [16·3-37·2] per 100 000 population). Globally, life expectancy steadily improved between 1990 and 2019 for 18 of the 22 investigated causes. Decomposition of global and regional life expectancy showed the positive effect that reductions in deaths from enteric infections, lower respiratory infections, stroke, and neonatal deaths, among others have contributed to improved survival over the study period. However, a net reduction of 1·6 years occurred in global life expectancy between 2019 and 2021, primarily due to increased death rates from COVID-19 and other pandemic-related mortality. Life expectancy was highly variable between super-regions over the study period, with southeast Asia, east Asia, and Oceania gaining 8·3 years (6·7-9·9) overall, while having the smallest reduction in life expectancy due to COVID-19 (0·4 years). The largest reduction in life expectancy due to COVID-19 occurred in Latin America and the Caribbean (3·6 years). Additionally, 53 of the 288 causes of death were highly concentrated in locations with less than 50% of the global population as of 2021, and these causes of death became progressively more concentrated since 1990, when only 44 causes showed this pattern. The concentration phenomenon is discussed heuristically with respect to enteric and lower respiratory infections, malaria, HIV/AIDS, neonatal disorders, tuberculosis, and measles. INTERPRETATION Long-standing gains in life expectancy and reductions in many of the leading causes of death have been disrupted by the COVID-19 pandemic, the adverse effects of which were spread unevenly among populations. Despite the pandemic, there has been continued progress in combatting several notable causes of death, leading to improved global life expectancy over the study period. Each of the seven GBD super-regions showed an overall improvement from 1990 and 2021, obscuring the negative effect in the years of the pandemic. Additionally, our findings regarding regional variation in causes of death driving increases in life expectancy hold clear policy utility. Analyses of shifting mortality trends reveal that several causes, once widespread globally, are now increasingly concentrated geographically. These changes in mortality concentration, alongside further investigation of changing risks, interventions, and relevant policy, present an important opportunity to deepen our understanding of mortality-reduction strategies. Examining patterns in mortality concentration might reveal areas where successful public health interventions have been implemented. Translating these successes to locations where certain causes of death remain entrenched can inform policies that work to improve life expectancy for people everywhere. FUNDING Bill & Melinda Gates Foundation

Dual-Mode Optical Sensing of Histamine at Nanomolar Concentrations in Complex Biological Fluids and Living Cells

An easily synthesized fluorescein-based luminescent dye has been utilized for the dual-mode detection of histamine at nanomolar concentrations at pH7.0 in water. The specific response to histamine was achieved by imidazole-catalyzed imine formation' reaction. The protocol was subsequently applied for the estimation of histamine in complex biological milieu such as human blood serum and urine samples. Furthermore, the dose-dependent cellular uptake of histamine and de novo synthesis (by thapsigargin treatment) was visualized in RAW 264.7, a mouse macrophage cell line. We have also developed portable paper strips for rapid, on-site detection of histamine without involving costly instruments

Targeted photocytotoxicity by copper(II) complexes having vitamin B-6 and photoactive acridine moieties

Copper(II) pyridoxal Schiff base complexes Cu(L-1/L-2)(B)]ClO4 (1-4), where HL1 is 4-(((2-(1H-imidazol-4-yl)ethyl)imino)methyl)-5-(hydroxymethyl)-2-methylp yridin-3-ol (in 1 and 2), HL2 is 2-(((2-(1H-imidazol-4-yl)ethyl)imino)methyl)phenol (in 3, 4), B is 11-(9-acridinyl)dipyrido3,2-a:2',3'-c]phenazine (acdppz in 1 and 3), dipyrido3,2-a:2',3'-c]phenazine (in 2) and 1,10-phenanthroline (in 4), were synthesized, characterized and their photocytotoxicity in visible light, intracellular localization, cellular uptake and DNA photocleavage activity were studied. Complex 4 was characterized by X-ray crystallography. Complexes 1 and 3 having acdppz as photosensitizer showed significant photocytotoxicity in visible light in HeLa and MCF7 cells giving IC50 value of <0.6 mu m, while being relatively non-toxic in dark. The complexes were non-toxic to non-tumorigenic HPL1D cells both in light and dark conditions. Complex 1 showed significant localization in the cytoplasm of HeLa cells within 4 h of treatment, as evidenced from confocal microscopy. DCFDA assay on 1 suggested generation of intracellular reactive oxygen species in HeLa cells upon photo-exposure. Importantly, Annexin-V-FITC/PI assay indicated photo-induced apoptotic cell death. (C) 2016 Elsevier Masson SAS. All rights reserved

Photocytotoxic Activity of Copper(II) and Zinc(II) Complexes of Curcumin and (Acridinyl)dipyridophenazine

Copper(II) and zinc(II) beta-diketonates of N,N-donor ligands, viz. Cu(dppz)(cur)](NO3) (1), Cu(acdppz)(cur)](NO3) (2), Cu(acdppz) (acac)](NO3) (3), Zn(dppz)(cur)](NO3) (4) and Zn(acdppz)(cur)] (NO3) (5), where dppz is dipyrido3,2-a:2',3'-c]phenazine, acdppz is 11-(9-acridinyl)dipyrido3,2-a:2',3'-c]phenazine, cur and acac are mono-deprotonated curcumin (Hcur) and acetyl acetone (Hacac), were synthesized, characterized and their photocytotoxicity studied. An analogue of complex 3, viz. Cu(acdppz) (acac)(H2O)]Cl 1/2 (NO3) 1= 2 (3a), structurally characterized by X-ray crystallography, has a cationic complex in square-pyramidal geometry with CuN2O3 core with an axial aqua ligand. Complexes 1, 2, 4 and 5 displayed emission at similar to 520 nm (lambda(exc): 430 nm) in dimethyl sulfoxide (DMSO) giving a fluorescence quantum yield value within 0.01-0.06. The complexes, in contrast to free curcumin, were fairly stable in cellular media up to 36 h, with no apparent degradation of the bound curcumin. The complexes were photocytotoxic (IC50: 0.3 4.5 mu M) in human cervical cancer (HeLa), breast cancer (MCF-7) and liver cancer (HepG2) cells. The apoptotic cell death is due to reactive oxygen species formation. Complexes 2 and 5 showed significant uptake in HeLa cells, localizing predominantly in the cytoplasm. Mechanistic data from the pUC19 DNA photocleavage study suggest involvement of acridine and curcumin in photo-generation of singlet oxygen and hydroxyl radicals as the ROS in light of 400-700 nm

Iron(III) Complexes of Vitamin B-6 Schiff Base with Boron-Dipyrromethene Pendants for Lysosome-Selective Photocytotoxicity

Iron(III) complexes of a vitamin B-6 Schiff base and NNN-donor ligands with pendant boron-dipyrromethene (BODIPY) moieties; namely, Fe(L1-3)(L-4,L-5)](NO3) (1-4), where L-1 is benzyl-bis(pyridin-2-yl)methyl]methanamine (bzdpa in 1), L-2 is a noniodinated BODIPY-appended dipicolylamine ligand (in 2, 3), L-3 is the diiodinated BODIPY analogue in 4, L-4 is a vitamin B-6 Schiff base, namely 3-hydroxy-5(hydroxymethyl)-4-{(2-hydroxyphenyl)imino]methyl}-2-meth ylpyridine (in 1, 3, and 4), and L-5 is 2-(2-hydroxyphenylimino)-methyl]phenol (in 2) as a nonpyridoxal Schiff base, were prepared, characterized, and their cellular localization and cytotoxic activity in light and in the dark were studied. The diiodo-BODIPY complex 4 displays remarkable photoinduced cytotoxicity in visible light (400-700 nm), with IC50 values within 0.11-0.25 mu m and about 200-fold lower dark toxicity. Complex 3, being fluorescent, was used for cellular imaging by confocal microscopy. Complex 4 shows supercoiled pUC19 DNA cleavage activity through the generation of singlet oxygen (O-1(2)) as the reactive oxygen species (ROS). Selective uptake of the complexes is observed from competitive cellular incorporation assays in cancer and noncancer cells. The complexes also show no apparent toxicity up to 100 mu m in the immortal human lung epithelial cells HPL1D in both light and the dark. Complex 3 shows preferential accumulation in lysosomes, giving a Pearson's correlation coefficient value of about 0.7

Interferon-Gamma and Nitric Oxide Synthase 2 Mediate the Aggregation of Resident Adherent Peritoneal Exudate Cells: Implications for the Host Response to Pathogens

<div><p>Interferon-gamma (Ifnγ), a key macrophage activating cytokine, plays pleiotropic roles in host immunity. In this study, the ability of Ifnγ to induce the aggregation of resident mouse adherent peritoneal exudate cells (APECs), consisting primarily of macrophages, was investigated. Cell-cell interactions involve adhesion molecules and, upon addition of Ifnγ, CD11b re-localizes preferentially to the sites of interaction on APECs. A functional role of CD11b in enhancing aggregation is demonstrated using Reopro, a blocking reagent, and siRNA to <i>Cd11b</i>. Studies with NG-methyl-L-arginine (LNMA), an inhibitor of Nitric oxide synthase (Nos), NO donors, e.g., S-nitroso-N-acetyl-DL-penicillamine (SNAP) or Diethylenetriamine/nitric oxide adduct (DETA/NO), and <i>Nos2</i>^-/- mice identified Nitric oxide (NO) induced by Ifnγ as a key regulator of aggregation of APECs. Further studies with <i>Nos2</i>^-/- APECs revealed that some Ifnγ responses are independent of NO: induction of MHC class II and CD80. On the other hand, Nos2 derived NO is important for other functions: motility, phagocytosis, morphology and aggregation. Studies with cytoskeleton depolymerizing agents revealed that Ifnγ and NO mediate the cortical stabilization of Actin and Tubulin which contribute to aggregation of APECs. The biological relevance of aggregation of APECs was delineated using infection experiments with <i>Salmonella</i> Typhimurium (<i>S</i>. Typhimurium). APECs from orally infected, but not uninfected, mice produce high amounts of NO and aggregate upon <i>ex vivo</i> culture in a Nos2-dependent manner. Importantly, aggregated APECs induced by Ifnγ contain fewer intracellular <i>S</i>. Typhimurium compared to their single counterparts post infection. Further experiments with LNMA or Reopro revealed that both NO and CD11b are important for aggregation; in addition, NO is bactericidal. Overall, this study elucidates novel roles for Ifnγ and Nos2 in regulating Actin, Tubulin, CD11b, motility and morphology during the aggregation response of APECs. The implications of aggregation or “group behavior” of APECs are discussed in the context of host resistance to infectious organisms.</p></div

The aggregation of APECs reduces the intracellular growth <i>S</i>. Typhimurium.

<p>Representative confocal microscopic images acquired at a magnification of 100 X with a scale of 10 μm illustrating intracellular Sal-GFP (green), Lamp1 (red) and Hoechst (blue) in control and 25 U/ml of Ifnγ treated APECs in the presence or absence of LNMA (200 μM) post 24 h of Sal-GFP infection (A). Quantification of the number of Sal-GFP per cell in control and 25 U/ml of Ifnγ treated APECs in the presence or absence of LNMA (200 μM) post 24 h of Sal-GFP infection (B). Quantification of the number of Sal-GFP per cell in control and 25 U/ml of Ifnγ treated APECs in the presence or absence of Reopro (0.2 mg/ ml) post 24 h of Sal-GFP infection (C). In panel B and C, the number of fields scored for single cells are represented in maroon color and the number of fields scored with aggregates are represented in blue color within brackets under the data points. Note that in the presence of Ifnγ, the number of aggregates containing fields increase compared to fields with single cells, which is reversed with LNMA or Reopro treatment. The data is represented as mean ± S.E from two independent experiments. Significance is represented as * when compared to untreated single cell controls, # when compared to Ifnγ treated single cell controls and Δ when compared to Ifnγ treated aggregates of APECs controls. The significance is represented as * when compared to untreated single cell controls, θ when compared to untreated aggregate controls, # when compared to Ifnγ treated single cell controls and τ when compared to Ifnγ treated aggregates of APECs controls.</p

Ifnγ induces APECs to aggregate.

<p>APECs were isolated from C57BL/6 mice, cultured in tissue culture media for ~ 24 h and were characterized by staining with a panel of antibodies to different cell surface proteins followed by FACS analysis. Representative plots and data (A) for different markers is shown which are representative of multiple experiments with APECs from 4 mice. Bright field images of APECs treated with different doses of Ifnγ for 36 h (B) and kinetics of APECs treated with 25 U/ml Ifnγ for indicated time points (C). The scale bar represents 20 μm. Quantification of the extent of APECs aggregates formed as a function of dose of Ifnγ (D) and incubation time (E). An aggregate consists of six or more interacting cells in any given field acquired at 20X magnification. The data is represented as mean ± S.E from three independent experiments.</p