Critical Analysis of Non-Thermal Plasma-Driven Modulation of Immune Cells from Clinical Perspective.

The emerged field of non-thermal plasma (NTP) shows great potential in the alteration of cell redox status, which can be utilized as a promising therapeutic implication. In recent years, the NTP field considerably progresses in the modulation of immune cell function leading to promising in vivo results. In fact, understanding the underlying cellular mechanisms triggered by NTP remains incomplete. In order to boost the field closer to real-life clinical applications, there is a need for a critical overview of the current state-of-the-art. In this review, we conduct a critical analysis of the NTP-triggered modulation of immune cells. Importantly, we analyze pitfalls in the field and identify persisting challenges. We show that the identification of misconceptions opens a door to the development of a research strategy to overcome these limitations. Finally, we propose the idea that solving problems highlighted in this review will accelerate the clinical translation of NTP-based treatments.

Detection of mitochondrial haplogroups in a small avar-slavic population from the eigth-ninth century AD.

OBJECTIVES: In the sixth century AD, Avars came to Central Europe from middle Eurasian steppes and founded a strong Empire called the Avar Khagante (568-799/803 AD) in the Pannonian basin. During the existence of this empire, they undertook many military and pugnacious campaigns. In the seventh century, they conquered the northern territory inhabited by Slavs, who were further recruited in Avar military and were commissioned with obtaining food supplies. During almost 200 years of Avar domination, a significant influence by the Avar culture (especially on the burial rite) and assimilation with indigenous population (occurrence of "East Asian"cranial features) could be noticed in this mixed area, which is supported by achaeological and anthropologcal research. Therefore we expected higher incidence of east Eurasian haplogroups (introduced by Avars) than the frequencies detected in present-day central European populations. MATERIALS AND METHODS: Mitochondrial DNA from 62 human skeletal remains excavated from the Avar-Slavic burial site Cífer-Pác (Slovakia) dated to the eighth and ninth century was analyzed by the sequencing of hypervariable region I and selected parts of coding region. Obtained haplotypes were compared with other present-day and historical populations and genetic distances were calculated using standard statistical method. RESULTS AND DISCUSSION: In total, the detection of mitochondrial haplogroups was possible in 46 individuals. Our results prooved a higher frequency of east Eurasian haplogroups in our analyzed population (6.52%) than in present-day central European populations. However, it is almost three times lower than the frequency of east Eurasian haplogroups detected in other medieval Avar populations. The statistical analysis showed a greater similarity and the lowest genetic distances between the Avar-Slavic burial site Cifer-Pac and medieval European populations than the South Siberian, East and Central Asian populations. CONCLUSION: Our results indicate that the transfer of Avar genetic variation through their mtDNA was rather weak in the analyzed mixed population.

Mechanical Regulation of Mitochondrial Dynamics and Function in a 3D-Engineered Liver Tumor Microenvironment.

It has become evident that physical stimuli of the cellular microenvironment transmit mechanical cues regulating key cellular functions, such as proliferation, migration, and malignant transformation. Accumulating evidence suggests that tumor cells face variable mechanical stimuli that may induce metabolic rewiring of tumor cells. However, the knowledge of how tumor cells adapt metabolism to external mechanical cues is still limited. We therefore designed soft 3D collagen scaffolds mimicking a pathological mechanical environment to decipher how liver tumor cells would adapt their metabolic activity to physical stimuli of the cellular microenvironment. Here, we report that the soft 3D microenvironment upregulates the glycolysis of HepG2 and Alexander cells. Both cell lines adapt their mitochondrial activity and function under growth in the soft 3D microenvironment. Cells grown in the soft 3D microenvironment exhibit marked mitochondrial depolarization, downregulation of mitochondrially encoded cytochrome c oxidase I, and slow proliferation rate in comparison with stiff monolayer cultures. Our data reveal the coupling of liver tumor glycolysis to mechanical cues. It is proposed here that soft 3D collagen scaffolds can serve as a useful model for future studies of mechanically regulated cellular functions of various liver (potentially other tissues as well) tumor cells.

Lysosomal nanotoxicity: Impact of nanomedicines on lysosomal function.

Although several nanomedicines got clinical approval over the past two decades, the clinical translation rate is relatively small so far. There are many post-surveillance withdrawals of nanomedicines caused by various safety issues. For successful clinical advancement of nanotechnology, it is of unmet need to realize cellular and molecular foundation of nanotoxicity. Current data suggest that lysosomal dysfunction caused by nanoparticles is emerging as the most common intracellular trigger of nanotoxicity. This review analyzes prospect mechanisms of lysosomal dysfunction-mediated toxicity induced by nanoparticles. We summarized and critically assessed adverse drug reactions of current clinically approved nanomedicines. Importantly, we show that physicochemical properties have great impact on nanoparticles interaction with cells, excretion route and kinetics, and subsequently on toxicity. We analyzed literature on adverse reactions of current nanomedicines and hypothesized that adverse reactions might be linked with lysosomal dysfunction caused by nanomedicines. Finally, from our analysis it becomes clear that it is unjustifiable to generalize safety and toxicity of nanoparticles, since different particles possess distinct toxicological properties. We propose that the biological mechanism of the disease progression and treatment should be central in the optimization of nanoparticle design.

Sensitivity of endogenous autofluorescence in HeLa cells to the application of external magnetic fields.

Dramatically increased levels of electromagnetic radiation in the environment have raised concerns over the potential health hazards of electromagnetic fields. Various biological effects of magnetic fields have been proposed. Despite decades of intensive research, the molecular mechanisms procuring cellular responses remain largely unknown. The current literature is conflicting with regards to evidence that magnetic fields affect functionality directly at the cellular level. Therefore, a search for potential direct cellular effects of magnetic fields represents a cornerstone that may propose an explanation for potential health hazards associated with magnetic fields. It has been proposed that autofluorescence of HeLa cells is magnetic field sensitive, relying on single-cell imaging kinetic measurements. Here, we investigate the magnetic field sensitivity of an endogenous autofluorescence in HeLa cells. Under the experimental conditions used, magnetic field sensitivity of an endogenous autofluorescence was not observed in HeLa cells. We present a number of arguments indicating why this is the case in the analysis of magnetic field effects based on the imaging of cellular autofluorescence decay. Our work indicates that new methods are required to elucidate the effects of magnetic fields at the cellular level.

The interactions between DNA nanostructures and cells: A critical overview from a cell biology perspective.

DNA nanotechnology has yielded remarkable advances in  composite materials with diverse applications in biomedicine. The specificity and predictability of building 3D structures at the nanometer scale make DNA nanotechnology a promising tool for uses in biosensing, drug delivery, cell modulation, and bioimaging. However, for successful translation of DNA nanostructures to real-world applications, it is crucial to understand how they interact with living cells, and the consequences of such interactions. In this review, we summarize the current state of knowledge on the interactions of DNA nanostructures with cells. We identify key challenges, from a cell biology perspective, that influence progress towards the clinical translation of DNA nanostructures. We close by providing an outlook on what questions must be addressed to accelerate the clinical translation of DNA nanostructures. STATEMENT OF SIGNIFICANCE: Self-assembled DNA nanostructures (DNs) offers unique opportunities to overcome persistent challenges in the nanobiotechnology field. However, the interactions between engineered DNs and living cells are still not well defined. Critical systematization of current cellular models and biological responses triggered by DNs is a crucial foundation for the successful clinical translation of DNA nanostructures. Moreover, such an analysis will identify the pitfalls and challenges that are present in the field, and provide a basis for overcoming those challenges.

Protein Corona Inhibits Endosomal Escape of Functionalized DNA Nanostructures in Living Cells.

DNA nanostructures (DNs) can be designed in a controlled and programmable manner, and these structures are increasingly used in a variety of biomedical applications, such as the delivery of therapeutic agents. When exposed to biological liquids, most nanomaterials become covered by a protein corona, which in turn modulates their cellular uptake and the biological response they elicit. However, the interplay between living cells and designed DNs are still not well established. Namely, there are very limited studies that assess protein corona impact on DN biological activity. Here, we analyzed the uptake of functionalized DNs in three distinct hepatic cell lines. Our analysis indicates that cellular uptake is linearly dependent on the cell size. Further, we show that the protein corona determines the endolysosomal vesicle escape efficiency of DNs coated with an endosome escape peptide. Our study offers an important basis for future optimization of DNs as delivery systems for various biomedical applications.

Analyzing the mechanisms of iron oxide nanoparticles interactions with cells: A road from failure to success in clinical applications.

Iron oxide nanoparticles (IONPs) were the first generation of nanomaterials that reached real clinic use. Particularly, several IONPs-based magnetic resonance imaging contrast agents gained approval by US Food and Drug Administration (FDA). However, latter body of evidence revealed the overlooked side effects of IONPs, resulting in their withdrawal. Emerging evidence suggests that this happened due to poor understanding of the mechanisms by which IONPs act at the cellular and sub-cellular levels. Recent studies indicate that better understanding of fundamental signal modulations induced by nanomaterials is essential to overcome the clinical problems with nanoparticles. Therefore, in this article we critically review potential mechanisms of IONPs-cell interactions and challenges related with their identification. We describe mechanisms of IONPs-induced toxicity. Ultimately, we demonstrate that knowledge of cellular mechanisms of IONPs action helped to overcome certain translation problems in nanomedicine - we explore potential causes and challenges associated with poor clinical performance of IONPs and propose outlook of how to overcome problems in the field. Our critical analysis implies that a clear understanding of molecular mechanisms of IONPs-cell interactions will provide a basement to increase the likelihood for clinical success of IONPs.

Iron Oxide Nanoparticle-Induced Autophagic Flux Is Regulated by Interplay between p53-mTOR Axis and Bcl-2 Signaling in Hepatic Cells.

Iron oxide-based nanoparticles have been repeatedly shown to affect lysosomal-mediated signaling. Recently, nanoparticles have demonstrated an ability to modulate autophagic flux via lysosome-dependent signaling. However, the precise underlying mechanisms of such modulation as well as the impact of cellular genetic background remain enigmatic. In this study, we investigated how lysosomal-mediated signaling is affected by iron oxide nanoparticle uptake in three distinct hepatic cell lines. We found that nanoparticle-induced lysosomal dysfunction alters sub-cellular localization of pmTOR and p53 proteins. Our data indicate that alterations in the sub-cellular localization of p53 protein induced by nanoparticle greatly affect the autophagic flux. We found that cells with high levels of Bcl-2 are insensitive to autophagy initiated by nanoparticles. Altogether, our data identify lysosomes as a central hub that control nanoparticle-mediated responses in hepatic cells. Our results provide an important fundamental background for the future development of targeted nanoparticle-based therapies.

Hepatic Tumor Cell Morphology Plasticity under Physical Constraints in 3D Cultures Driven by YAP-mTOR Axis.

Recent studies undoubtedly show that the mammalian target of rapamycin (mTOR) and the Hippo-Yes-associated protein 1 (YAP) pathways are important mediators of mechanical cues. The crosstalk between these pathways as well as de-regulation of their signaling has been implicated in multiple tumor types, including liver tumors. Additionally, physical cues from 3D microenvironments have been identified to alter gene expression and differentiation of different cell lineages. However, it remains incompletely understood how physical constraints originated in 3D cultures affect cell plasticity and what the key mediators are of such process. In this work, we use collagen scaffolds as a model of a soft 3D microenvironment to alter cellular size and study the mechanotransduction that regulates that process. We show that the YAP-mTOR axis is a downstream effector of 3D cellular culture-driven mechanotransduction. Indeed, we found that cell mechanics, dictated by the physical constraints of 3D collagen scaffolds, profoundly affect cellular proliferation in a YAP-mTOR-mediated manner. Functionally, the YAP-mTOR connection is key to mediate cell plasticity in hepatic tumor cell lines. These findings expand the role of YAP-mTOR-driven mechanotransduction to the control hepatic tumor cellular responses under physical constraints in 3D cultures. We suggest a tentative mechanism, which coordinates signaling rewiring with cytoplasmic restructuring during cell growth in 3D microenvironments.

Serious chronic disease of the cervical spine and trauma in a young female from the middle ages (Czech Republic).

The skeletal remains of the young female (20-24 years) from Grave JP/106, discovered in the Southern Suburb of the Breclav - Pohansko Stronghold (Early Middle Ages, 9th century-beginning of the 10th century, present day Czech Republic) display several noteworthy pathologies. The first is deformation of the mandible, which was most probably caused by a fracture of the ramus in combination with a subcondylar fracture. The spine of this young woman also exhibits a probable traumatic injury of the cervical spine in combination with a slowly growing structure situated inside the spinal canal, which caused deformation centered upon C7. The cervical and thoracic spine together with internal surfaces of several ribs exhibit infectious changes of advanced stage, in all likelihood of tuberculous origin, but osteomyelitis cannot be excluded. Histological analysis of the new bone formation in the ribs confirmed infectious origin, as does Micro CT of C5 and C6. Analyses conducted by two different departments with different methods (PCR amplification of 123 bp long section from IS6110 and Next Generation shotgun sequencing) failed to identify DNA of Mycobacterium tuberculosis from the first rib.