Efficient and reproducible in vitro regeneration of Solanum lycopersicum and assessment genetic uniformity using flow cytometry and SPAR methods

24 p.-4 fig.-6 tab.In the present study, we develop an efficient and reproducible in vitro regeneration system for two cultivars viz., Jamila and Tomaland of Solanum lycopersicum L., an economically important vegetable crop throughout the world. Sterilization of seeds with 2.5 % (v/v) NaOCl was found to be most effective, about 97 % of seeds germinated on cotton in magenta box moistened with sterile half strength (½)Murashige and Skoog (MS) medium. Regeneration efficiency of cotyledonary leaf (CL) and cotyledonary node (CN) explants derived from 08 days old aseptic seedling were assessed on MS medium supplemented with different concentrations of auxins and cytokinin. CL explants were found more responsive in comparison to CN in both the cultivars. Types of basal media were also assessed and found to have a significant effect on shoot regeneration. Highest regeneration frequency and maximum number of shoots were standardized from CL explants on MS medium supplied with 6- benzyl adenine (BA; 5.0 µM), indole-3-butyric acid (IBA; 2.5 µM) and Kinetin (Kin; 10.0 µM). In vitro regenerated microshoots were rooted on ½MS medium containing 0.5 µM indole-3-butyric acid (IBA). Regenerated plantlets with well-developed roots and shoot system were successfully acclimated to ex vitro condition. Genetic uniformity of tissue culture raised plantlets was first time evaluated using flow cytometry and single primer amplification reaction (SPAR) methods viz., DAMD and ISSR. No significant changes in ploidy level and nuclear DNA content profile were observed between in vitro propagated plants and normal plants of both the cultivars. Similarly, the SPAR analysis also revealed monomorphic banding patterns in regenerated plantlets of S. lycopersicum verifying their genetic uniformity and clonal fidelity. This efficient regeneration system can be used as a fast and reproducible method for genetic transformation of this important vegetable crop.This project was funded by the National Plan for Science, Technology and Innovation (MAARIFAH), King Abdul Aziz City for Science and Technology, Kingdom of Saudi Arabia, Award Number 12-BIO2919-02.Peer reviewe

Toxicological impact of microplastics and nanoplastics on humans: understanding the mechanistic aspect of the interaction

Plastic is a pervasive material that has become an indispensable part of our daily lives and is used in various commercial products. However, plastic waste has significantly impacted the environment, accumulating in water and land ecosystems and harming all forms of life. When plastic degrades, it breaks down into smaller particles called microplastics (MPs), which can further breakdown into nanoplastics (NPs). Due to their small size and potential toxicity to humans, NPs are of particular concern. During the COVID-19 pandemic, the production of plastic had reached unprecedented levels, including essential medical kits, food bags, and personal protective equipment (PPE), which generate MPs and NPs when burned. MPs and NPs have been detected in various locations, such as air, food, and soil, but our understanding of their potential adverse health effects is limited. This review aims to provide a comprehensive overview of the sources, interactions, ecotoxicity, routes of exposure, toxicity mechanisms, detection methods, and future directions for the safety evaluation of MPs and NPs. This would improve our understanding of the impact of MPs and NPs on our health and environment and identify ways to address this global crisis

Marine Macroalgae Display Bioreductant Efficacy for Fabricating Metallic Nanoparticles: Intra/Extracellular Mechanism and Potential Biomedical Applications

The application of hazardous chemicals during nanoparticle (NP) synthesis has raised alarming concerns pertaining to their biocompatibility and equally to the environmental harmlessness. In the recent decade, nanotechnological research has made a gigantic shift in order to include the natural resources to produce biogenic NPs. Within this approach, researchers have utilized marine resources such as macroalgae and microalgae, land plants, bacteria, fungi, yeast, actinomycetes, and viruses to synthesize NPs. Marine macroalgae (brown, red, and green) are rich in polysaccharides including alginates, fucose-containing sulfated polysaccharides (FCSPs), galactans, agars or carrageenans, semicrystalline cellulose, ulvans, and hemicelluloses. Phytochemicals are abundant in phenols, tannins, alkaloids, terpenoids, and vitamins. However, microorganisms have an abundance of active compounds ranging from sugar molecules, enzymes, canonical membrane proteins, reductase enzymes (NADH and NADPH), membrane proteins to many more. The prime reason for using the aforesaid entities in the metallic NPs synthesis is based on their intrinsic properties to act as bioreductants, having the capability to reduce and cap the metal ions into stabilized NPs. Several green NPs have been verified for their biocompatibility in human cells. Bioactive constituents from the above resources have been found on the green metallic NPs, which has demonstrated their efficacies as prospective antibiotics and anti-cancer agents against a range of human pathogens and cancer cells. Moreover, these NPs can be characterized for the size, shapes, functional groups, surface properties, porosity, hydrodynamic stability, and surface charge using different characterization techniques. The novelty and originality of this review is that we provide recent research compilations on green synthesis of NPs by marine macroalgae and other biological sources (plant, bacteria, fungi, actinomycetes, yeast, and virus). Besides, we elaborated on the detailed intra- and extracellular mechanisms of NPs synthesis by marine macroalgae. The application of green NPs as anti-bacterial, anti-cancer, and popular methods of NPs characterization techniques has also been critically reviewed

Bacterial resistance to arsenic protects against protist killing

Protists kill their bacterial prey using toxic metals such as copper. Here we hypothesize that the metalloid arsenic has a similar role. To test this hypothesis, we examined intracellular survival of Escherichia coli (E. coli) in the amoeba Dictyostelium discoideum (D. discoideum). Deletion of the E. coli ars operon led to significantly lower intracellular survival compared to wild type E. coli. This suggests that protists use arsenic to poison bacterial cells in the phagosome, similar to their use of copper. In response to copper and arsenic poisoning by protists, there is selection for acquisition of arsenic and copper resistance genes in the bacterial prey to avoid killing. In agreement with this hypothesis, both copper and arsenic resistance determinants are widespread in many bacterial taxa and environments, and they are often found together on plasmids. A role for heavy metals and arsenic in the ancient predator–prey relationship between protists and bacteria could explain the widespread presence of metal resistance determinants in pristine environments

Thymoquinone suppression of the human hepatocellular carcinoma cell growth involves inhibition of IL-8 expression, elevated levels of TRAIL receptors, oxidative stress and apoptosis

Hepatocellular carcinoma (HCC) is the fourth most common solid tumor worldwide. The chemokine interleukin-8 (IL-8) is overexpressed in HCC and is a potential target for therapy. Although the transcription factor NF-κB regulates IL-8 expression, and while thymoquinone (TQ; the most bioactive constituent of black seed oil) inhibits NF-κB activity, the precise mechanisms by which TQ regulates IL-8 and cancer cell growth remain to be clarified. Here, we report that TQ inhibited growth of HCC cells in a dose- and time-dependent manner, caused G2M cell cycle arrest, and stimulated apoptosis. Apoptosis was substantiated by activation of caspase-3 and -9, as well as cleavage of poly(ADP-ribose)polymerase. TQ treatments inhibited expression of NF-κB and suppressed IL-8 and its receptors. TQ treatments caused increased levels of reactive oxygen species (ROS) and mRNAs of oxidative stress-related genes, NQO1 and HO-1. Pretreatment of HepG2 cells with N-acetylcysteine, a scavenger of ROS, prevented TQ-induced cell death. TQ treatment stimulated mRNA expression of pro-apoptotic Bcl-xS and TRAIL death receptors, and inhibited expression of the anti-apoptotic gene Bcl-2. TQ enhanced TRAIL-induced death of HepG2 cells, in part by up-regulating TRAIL death receptors, inhibiting NF-κB and IL-8 and stimulating apoptosis. Altogether, these findings provide insights into the pleiotropic molecular mechanisms of TQ-dependent suppression of HCC cell growth and underscore potential of this compound as anti-HCC drug

Distribution of Arsenic Resistance Genes in Prokaryotes

Arsenic is a metalloid that occurs naturally in aquatic and terrestrial environments. The high toxicity of arsenic derivatives converts this element in a serious problem of public health worldwide. There is a global arsenic geocycle in which microbes play a relevant role. Ancient exposure to arsenic derivatives, both inorganic and organic, has represented a selective pressure for microbes to evolve or acquire diverse arsenic resistance genetic systems. In addition, arsenic compounds appear to have been used as a toxin in chemical warfare for a long time selecting for an extended range of arsenic resistance determinants. Arsenic resistance strategies rely mainly on membrane transport pathways that extrude the toxic compounds from the cell cytoplasm. The ars operons, first discovered in bacterial R-factors almost 50 years ago, are the most common microbial arsenic resistance systems. Numerous ars operons, with a variety of genes and different combinations of them, populate the prokaryotic genomes, including their accessory plasmids, transposons, and genomic islands. Besides these canonical, widespread ars gene clusters, which confer resistance to the inorganic forms of arsenic, additional genes have been discovered recently, which broadens the spectrum of arsenic tolerance by detoxifying organic arsenic derivatives often used as toxins. This review summarizes the presence, distribution, organization, and redundance of arsenic resistance genes in prokaryotes

Verbesina encelioides: cytotoxicity, cell cycle arrest, and oxidative DNA damage in human liver cancer (HepG2) cell line

BACKGROUND: Cancer is a major health problem and exploiting natural products have been one of the most successful methods to combat this disease. Verbesina encelioides is a notorious weed with various pharmacological properties. The aim of the present investigation was to screen the anticancer potential of V. encelioides extract against human lung cancer (A-549), breast cancer (MCF-7), and liver cancer (HepG2) cell lines. METHODS: A-549, MCF-7, and HepG2 cells were exposed to various concentrations of (10–1000 μg/ml) of V. encelioides for 24 h. Further, cytotoxic concentrations (250, 500, and 1000 μg/ml) of V. encelioides induced oxidative stress (GSH and LPO), reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP), cell cycle arrest, and DNA damage in HepG2 cells were studied. RESULTS: The exposure of cells to 10–1000 μg/ml of extract for 24 h, revealed the concentrations 250–1000 μg/ml was cytotoxic against MCF-7 and HepG2 cells, but not against A-549 cells. Moreover, the extract showed higher decrease in the cell viability against HepG2 cells than MCF-7 cells. Therefore, HepG2 cells were selected for further studies viz. oxidative stress (GSH and LPO), reactive oxygen species (ROS) generation, mitochondrial membrane potential (MMP), cell cycle arrest, and DNA damage. The results revealed differential anticancer activity of V. encelioides against A-549, MCF-7 and HepG2 cells. A significant induction of oxidative stress, ROS generation, and MMP levels was observed in HepG2 cells. The cell cycle analysis and comet assay showed that V. encelioides significantly induced G2/M arrests and DNA damage. CONCLUSION: These results indicate that V. encelioides possess substantial cytotoxic potential and may warrant further investigation to develop potential anticancer agent. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12906-016-1106-0) contains supplementary material, which is available to authorized users

Cyto-Genotoxic and Transcriptomic Alterations in Human Liver Cells by Tris (2-Ethylhexyl) Phosphate (TEHP): A Putative Hepatocarcinogen

Tris (2-ethylhexyl) phosphate (TEHP) is an organophosphate flame retardant (OPFRs) which is extensively used as a plasticizer and has been detected in human body fluids. Contemporarily, toxicological studies on TEHP in human cells are very limited and there are few studies on its genotoxicity and cell death mechanism in human liver cells (HepG2). Herein, we find that HepG2 cells exposed to TEHP (100, 200, 400 µM) for 72 h reduced cell survival to 19.68%, 49.83%, 58.91% and 29.08%, 47.7% and 57.90%, measured by MTT and NRU assays. TEHP did not induce cytotoxicity at lower concentrations (5, 10, 25, 50 µM) after 24 h and 48 h of exposure. Flow cytometric analysis of TEHP-treated cells elevated intracellular reactive oxygen species (ROS), nitric oxide (NO), Ca++ influx and esterase levels, leading to mitochondrial dysfunction (ΔΨm). DNA damage analysis by comet assay showed 4.67, 9.35, 13.78-fold greater OTM values in TEHP (100, 200, 400 µM)-treated cells. Cell cycle analysis exhibited 23.1%, 29.6%, and 50.8% of cells in SubG1 apoptotic phase after TEHP (100, 200 and 400 μM) treatment. Immunofluorescence data affirmed the activation of P53, caspase 3 and 9 proteins in TEHP-treated cells. In qPCR array of 84 genes, HepG2 cells treated with TEHP (100 µM, 72 h) upregulated 10 genes and downregulated 4 genes belonging to a human cancer pathway. Our novel data categorically indicate that TEHP is an oxidative stressor and carcinogenic entity, which exaggerates mitochondrial functions to induce cyto- and genotoxicity and cell death, implying its hepatotoxic features

SARS-CoV-2: Understanding the Transcriptional Regulation of ACE2 and TMPRSS2 and the Role of Single Nucleotide Polymorphism (SNP) at Codon 72 of p53 in the Innate Immune Response against Virus Infection

Human ACE2 and the serine protease TMPRSS2 of novel SARS-CoV-2 are primary entry receptors in host cells. Expression of these genes at the transcriptional level has not been much discussed in detail. The ISRE elements of the ACE2 promoter are a binding site for the ISGF3 complex of the JAK/STAT signaling pathway. TMPRSS2, including IFNβ, STAT1, and STAT2, has the PARP1 binding site near to TSS either up or downstream promoter region. It is well documented that PARP1 regulates gene expression at the transcription level. Therefore, to curb virus infection, both promoting type I IFN signaling to boost innate immunity and prevention of virus entry by inhibiting PARP1, ACE2 or TMPRSS2 are safe options. Most importantly, our aim is to attract the attention of the global scientific community towards the codon 72 Single Nucleotide Polymorphism (SNP) of p53 and its underneath role in the innate immune response against SARS-CoV-2. Here, we discuss codon 72 SNP of human p53′s role in the different innate immune response to restrict virus-mediated mortality rate only in specific parts of the world. In addition, we discuss potential targets and emerging therapies using bioengineered bacteriophage, anti-sense, or CRISPR strategies.</jats:p