Advances in the Toxicity Assessment of Silver Nanoparticles Derived from a Sphagnum fallax Extract for Monolayers and Spheroids.

The production of nanomaterials through environmentally friendly methods is a top priority in the sustainable development of nanotechnology. This paper presents data on the synthesis of silver nanoparticles using an aqueous extract of Sphagnum fallax moss at room temperature. The morphology, stability, and size of the nanoparticles were analyzed using various techniques, including transmission electron microscopy, Doppler laser velocimetry, and UV-vis spectroscopy. In addition, Fourier transform infrared spectroscopy was used to analyze the presence of moss metabolites on the surface of nanomaterials. The effects of different concentrations of citrate-stabilized and moss extract-stabilized silver nanoparticles on cell viability, necrosis induction, and cell impedance were compared. The internalization of silver nanoparticles into both monolayers and three-dimensional cells spheroids was evaluated using dark-field microscopy and hyperspectral imaging. An eco-friendly method for the synthesis of silver nanoparticles at room temperature is proposed, which makes it possible to obtain spherical nanoparticles of 20-30 nm in size with high bioavailability and that have potential applications in various areas of human life.

Development of a novel prostate Cancer-Stroma Sphere (CSS) model for In Vitro tumor microenvironment studies.

Tumor employs non-cancerous cells to gain beneficial features that promote growth and survival of cancer cells. Despite intensive research in the area of tumor microenvironment, there is still a lack of reliable and reproducible in vitro model for tumor and tumor-microenvironment cell interaction studies. Herein we report the successful development of a heterogeneous cancer-stroma sphere (CSS) model composed of prostate adenocarcinoma PC3 cells and immortalized mesenchymal stem cells (MSC). The CSS model demonstrated a structured spatial layout of the cells, with stromal cells concentrated at the center of the spheres and tumor cells located on the periphery. Significant increase in the levels of VEGFA, IL-10, and IL1a has been detected in the conditioned media of CSS as compared to PC3 spheres. Single cell RNA sequencing data revealed that VEGFA was secreted by MSC cells within heterogeneous spheroids. Enhanced expression of extracellular membrane (ECM) proteins was also shown for CSS-derived MSCs. Furthermore, we demonstrated that the multicellular architecture altered cancer cell response to chemotherapeutic agents: the inhibition of sphere formation by topotecan was 74.92 ± 4.56 % for PC3 spheres and 45.95 ± 7.84 % for CSS spheres (p < 0.01), docetaxel showed 37,51± 20,88 % and 15,67± 14,08 % inhibition, respectively (p < 0.05). Thus, CSS present an effective in vitro model for examining the extracellular matrix composition and cell-to-cell interactions within the tumor, as well as for evaluating the antitumor activity of drugs.

Immunology of SARS-CoV-2 Infection.

According to the data from the World Health Organization, about 800 million of the world population had contracted coronavirus infection caused by SARS-CoV-2 by mid-2023. Properties of this virus have allowed it to circulate in the human population for a long time, evolving defense mechanisms against the host immune system. Severity of the disease depends largely on the degree of activation of the systemic immune response, including overstimulation of macrophages and monocytes, cytokine production, and triggering of adaptive T- and B-cell responses, while SARS-CoV-2 evades the immune system actions. In this review, we discuss immune responses triggered in response to the SARS-CoV-2 virus entry into the cell and malfunctions of the immune system that lead to the development of severe disease.

Adipose-Derived Mesenchymal Stem Cell (MSC) Immortalization by Modulation of hTERT and TP53 Expression Levels.

Mesenchymal stem cells (MSCs) are pivotal players in tissue repair and hold great promise as cell therapeutic agents for regenerative medicine. Additionally, they play a significant role in the development of various human diseases. Studies on MSC biology have encountered a limiting property of these cells, which includes a low number of passages and a decrease in differentiation potential during in vitro culture. Although common methods of immortalization through gene manipulations of cells are well established, the resulting MSCs vary in differentiation potential compared to primary cells and eventually undergo senescence. This study aimed to immortalize primary adipose-derived MSCs by overexpressing human telomerase reverse transcriptase (hTERT) gene combined with a knockdown of TP53. The research demonstrated that immortalized MSCs maintained a stable level of differentiation into osteogenic and chondrogenic lineages during 30 passages, while also exhibiting an increase in cell proliferation rate and differentiation potential towards the adipogenic lineage. Long-term culture of immortalized cells did not alter cell morphology and self-renewal potential. Consequently, a genetically stable line of immortalized adipose-derived MSCs (iMSCs) was established.

DNA/Magnetic Nanoparticles Composite to Attenuate Glass Surface Nanotopography for Enhanced Mesenchymal Stem Cell Differentiation.

Mesenchymal stem cells (MSCs) have extensive pluripotent potential to differentiate into various cell types, and thus they are an important tool for regenerative medicine and biomedical research. In this work, the differentiation of hTERT-transduced adipose-derived MSCs (hMSCs) into chondrocytes, adipocytes and osteoblasts on substrates with nanotopography generated by magnetic iron oxide nanoparticles (MNPs) and DNA was investigated. Citrate-stabilized MNPs were synthesized by the chemical co-precipitation method and sized around 10 nm according to microscopy studies. It was shown that MNPs@DNA coatings induced chondrogenesis and osteogenesis in hTERT-transduced MSCs. The cells had normal morphology and distribution of actin filaments. An increase in the concentration of magnetic nanoparticles resulted in a higher surface roughness and reduced the adhesion of cells to the substrate. A glass substrate modified with magnetic nanoparticles and DNA induced active chondrogenesis of hTERT-transduced MSC in a twice-diluted differentiation-inducing growth medium, suggesting the possible use of nanostructured MNPs@DNA coatings to obtain differentiated cells at a reduced level of growth factors.

Biology of the SARS-CoV-2 Coronavirus.

New coronavirus infection causing COVID-19, which was first reported in late 2019 in China, initiated severe social and economic crisis that affected the whole world. High frequency of the errors in replication of RNA viruses, zoonotic nature of transmission, and high transmissibility allowed betacoronaviruses to cause the third pandemic in the world since the beginning of 2003: SARS-CoV in 2003, MERS-CoV in 2012, and SARS-CoV-2 in 2019. The latest pandemic united scientific community and served as a powerful impetus in the study of biology of coronaviruses: new routes of virus penetration into the human cells were identified, features of the replication cycle were studied, and new functions of coronavirus proteins were elucidated. It should be recognized that the pandemic was accompanied by the need to obtain and publish results within a short time, which led to the emergence of an array of conflicting data and low reproducibility of research results. We systematized and analyzed scientific literature, filtered the results according to reliability of the methods of analysis used, and prepared a review describing molecular mechanisms of functioning of the SARS-CoV-2 coronavirus. This review considers organization of the genome of the SARS-CoV-2 virus, mechanisms of its gene expression and entry of the virus into the cell, provides information on key mutations that characterize different variants of the virus, and their contribution to pathogenesis of the disease.

Therapeutic Editing of the TP53 Gene: Is CRISPR/Cas9 an Option?

The TP53 gene encodes the transcription factor and oncosuppressor p53 protein that regulates a multitude of intracellular metabolic pathways involved in DNA damage repair, cell cycle arrest, apoptosis, and senescence. In many cases, alterations (e.g., mutations of the TP53 gene) negatively affect these pathways resulting in tumor development. Recent advances in genome manipulation technologies, CRISPR/Cas9, in particular, brought us closer to therapeutic gene editing for the treatment of cancer and hereditary diseases. Genome-editing therapies for blood disorders, blindness, and cancer are currently being evaluated in clinical trials. Eventually CRISPR/Cas9 technology is expected to target TP53 as the most mutated gene in all types of cancers. A majority of TP53 mutations are missense which brings immense opportunities for the CRISPR/Cas9 system that has been successfully used for correcting single nucleotides in various models, both in vitro and in vivo. In this review, we highlight the recent clinical applications of CRISPR/Cas9 technology for therapeutic genome editing and discuss its perspectives for editing TP53 and regulating transcription of p53 pathway genes.

Novel Isatin-based activator of p53 transcriptional functions in tumor cells.

Bioinorganic medicinal chemistry remains a hot field for research aimed at developing novel anti-cancer treatments. Discovery of metal complexes as potent antitumor chemotherapeutics such as cisplatin led to a significant shift of focus toward organometallic/ bioinorganic compounds containing transition metals and their chelates as novel scaffolds for drug discovery. In that way, transition metal complexes coordinated to essential biological scaffolds represent a highly promising class of compounds for design of novel target-specific therapeutics. Here, we report novel data on p53 activating Isatin-based Cu(II) complex exhibiting cytotoxic properties towards HCT116 and MCF7 tumor cell lines, as confirmed by cell viability assay and flow cytometry analysis of apoptosis. Furthermore, putative p53-mediated mechanism of action of this compound is supported by quantitative analysis of TP53, MDM2 and PUMA genes expression, as well as luciferase-based p53 pathway activation assay. Multiplex immunoassay analysis of inflammatory markers revealed potential modulation of several cytokines and chemokines.

Isatin-Schiff base-copper (II) complex induces cell death in p53-positive tumors.

Medicinal bioinorganic chemistry is a thriving field of drug research for cancer treatment. Transition metal complexes coordinated to essential biological scaffolds represent a highly promising class of compounds for design of novel target-specific therapeutics. We report here the biological evaluation of a novel Isatin-Schiff base derivative and its Cu(II) complex in several tumor cell lines by assessing their effects on cellular metabolism, real-time cell proliferation and induction of apoptosis. Further, the impact of compounds on the p53 protein and expression of its target genes, including MDM2, p21/CDKN1A, and PUMA was evaluated. Results obtained in this study provide further evidence in support of our prior data suggesting the p53-mediated mechanism of action for Isatin-Schiff base derivatives and their complexes and also shed light on potential use of these compounds for stimulation of apoptosis in breast cancer cells via activation of the pro-apoptotic PUMA gene.