In general, polymer/metal nanocomposites might have better anti-cancer properties in comparison to standalone metal nanoparticles (NPs). This emerging class of functional nanomaterials shows improved and more selective tumor delivery with lower toxicity against non-tumor tissues, providing a better biocompatibility and higher effectiveness. Herein, the cytotoxicity of four two-component nanocomposites comprising a metal NP (Au or Ag) and a star-like polymer (dextran-graft-polyacrylamide, D-PAA, or its anionic form, D-PAAan) was tested against multiple diffuse large B-cell lymphoma (DLBCL) cell lines. Our findings suggest that these nanocomposites reduce the viability of DLBCL cells in a dose-dependent manner. The cytotoxic effect of the investigated two-component nanocomposites was mediated by overproduction of reactive oxygen species (ROS, especially in the case of AuNP-based nanocomposites), elevation of intracellular Fe2+ levels, with consequent activation of lipid peroxidation and reduced glutathione (GSH) downregulation. Unexpectedly, the nanocomposites did not trigger ferroptosis, which is a type of regulated cell death heavily dependent on ROS and Fe2+. Nanocomposite-induced cell death occurred via apoptosis, as evidenced by the activation of caspase-3/7. Interestingly, the tested nanocomposites induced substantial compensatory activation of the pro-survival PI3K/AKT (phosphatidylinositol 3‑kinase/protein kinase B) pathway. However, this was not sufficient to prevent nanocomposite-induced apoptosis. We have demonstrated the anti-lymphoma potential of four noble metal/polymer nanocomposites. Their cytotoxic and anti-lymphoma effects are primarily attributable to oxidative stress with consequent induction of apoptosis.

Diffuse large B-cell lymphoma (DLBCL) is the most frequent B-cell type of non-Hodgkin's lymphoma. Recent genomic studies have highlighted the importance of genetic alterations in apoptotic pathways that help malignant DLBCL cells to evade apoptosis. Apoptosis evasion by DLBCL cells is known to mediate resistance to chemotherapy. Advances in the field of regulated cell death (RCD) research have identified novel therapeutic avenues in cancer. In particular, non-apoptotic RCDs can be targeted to overcome resistance to apoptosis in cancer and ensure cell death. In this review, we have highlighted the contribution of multiple RCDs, including apoptosis, necroptosis, ferroptosis, pyroptosis, PANoptosis, NETotic cell death, autophagy-dependent cell death, cuproptosis, methuosis, or mitotic death, to normal development of B lymphocytes and DLBCL pathogenesis. We have summarized molecular mechanisms governing distinct RCDs in DLBCL, differences in cell death pathways in activated B-cell (ABC) and germinal center B-cell (GCB) DLBCL subtypes, prognostic values of RCD-related genes, and discussed the implication of RCD pathways for DLBCL treatment. Notably, the impact of RCDs goes far beyond just killing tumor cells. RCD modalities are important for orchestrating the immune response and modulating the tumor microenvironment. The current review also aims to reveal the effect of different RCDs on the tumor microenvironment in DLBCL. Most RCDs play a dual role in DLBCL, demonstrating both tumor-inducing and tumor-suppressing effects, which suggests that their targeting should be exploited with caution. Our analysis suggests that pharmacological ferroptosis induction may be the most promising RCD-targeting strategy in DLBCL.

Cardiovascular diseases (CVDs) remain a global medical and social burden with high morbidity and mortality rates worldwide. CVDs are strongly linked to thrombosis, anemia, endothelial dysfunction, or inflammation. A growing body of evidence indicates that eryptosis, a regulated cell death of mature erythrocytes, plays an emerging role in CVDs by contributing to the above-mentioned pathogenetic links. In this review, we summarize the current knowledge of the key signaling pathways involved in eryptosis execution, highlight the pathophysiological role of eryptosis in CVDs and related metabolic disorders (including metabolic syndrome, diabetes mellitus, atherosclerosis, ischemic heart disease, or heart failure), identify the key molecular drivers of eryptosis in cardiovascular pathology, and shed light on the potential detrimental effects of enhanced eryptosis in CVDs. Identification of eryptosis-related complications of CVDs might provide novel insights into preventive and therapeutic strategies. Enhanced eryptosis in metabolic and cardiovascular pathology contributes to anemia via the accelerated clearance of eryptotic red blood cells (RBCs). Thrombogenic effects of eryptosis in CVDs are mediated by phosphatidylserine (PS)-mediated thrombin generation, interactions with platelets, and generation of coagulation-promoting PS-expressing RBC-derived microvesicles. Eryptosis-induced endothelial dysfunction is at least partly attributed to PS/CXCL16-mediated adhesion of eryptotic cells to endothelial cells (ECs), consequent uptake of eryptotic cells by ECs, and induction of their ferroptosis. On the other hand, enhanced eryptosis might have a dual role in the regulation of inflammation in CVDs. Phagocytosis of eryptotic cells reprograms macrophages to gain the pro-resolution phenotype. However, eryptosis triggers pro-inflammatory ferroptosis of ECs and immune cells at the same time.

A compelling body of evidence links pesticide exposure to human diseases. The liver plays a central role in the detoxification of pesticides, suggesting intense pesticide-liver cell interactions. A growing body of studies highlighted in this review supports the contribution of pesticides of various chemical classes to the development of non-alcoholic fatty liver disease (NAFLD), alcohol-associated liver disease (ALD), liver cirrhosis, viral hepatitis, hepatocellular carcinoma, etc., via disrupting lipid and carbohydrate metabolism and redox homeostasis, promoting endoplasmic reticulum stress and mitochondrial dysfunction, as well as stimulating apoptosis, fibrosis, and inflammation. In this review, we systematically illustrated an underappreciated mechanism of pesticide-induced overall and hepatic toxicity, i.e., the ability to induce non-apoptotic regulated cell death (RCD) pathways such as ferroptosis, necroptosis, and pyroptosis. Our analysis indicates that pesticides are implicated in driving liver diseases by inducing ferroptosis, necroptosis, and pyroptosis. Non-apoptotic RCDs mediate pesticide-induced liver steatosis and fibrosis. Furthermore, these cell death modalities fuel inflammation through the promotion of pro-inflammatory cytokine production and the generation of damage-associated molecular patterns. Understanding of deeper mechanisms of pesticide-induced effects on the non-apoptotic cell death machinery and subsequent immunogenic effects in liver pathology might help develop novel preventive strategies to reduce liver damage.

Iron doping has been reported to significantly modify the properties of metal oxide nanoparticles (NPs), including their interaction with cells and therefore toxicity. The present study explores the ability of Fe³ ⁺-doped CeO2 NPs with the varying content of Fe3+ ions (3, 5, and 10 at%) to stimulate eryptosis, a controllable cell death pathway of mature erythrocytes, as an attempt to shed light on hemocompatibility of iron-doped CeO2 NPs. Overall erythrotoxicity of iron-doped CeO2 NPs was evaluated by investigating their ability to trigger spontaneous hemolysis and affect osmotic fragility of rat erythrocytes. Eryptosis of erythrocytes exposed to iron-doped and non-doped CeO2 NPs for 24 h was evaluated by the state-of-the-art flow cytometry-based annexin V staining. Mechanisms involved in iron-doped CeO2 NP-induced eryptosis were evaluated by 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) and caspase-3 assays, as well as the fluorescent O1O (2-(2'-hydroxy-phenyl)-5-phenyl-1,3-oxazole), NR12S, Fluo-3 AM, BODIPY™ 581/591 C11, and BioTracker Far-red Fe2+ Live Cell imaging probes. Dose-dependent effects of iron-doped CeO2 NPs on hemolysis, osmotic fragility, and eryptosis were revealed. Eryptosis triggered by iron-doped CeO2 NPs was found to be oxidative stress-mediated, cation channel-driven, and caspase-dependent. Oxidative stress and alterations of lipid membranes demonstrated by the tested NPs could be attributable to their direct •OH generation and peroxidase-like activity, as well as Fe2+-mediated Fenton reaction at the surface of Fe³⁺-doped CeO₂ NPs (presumably due to Fe³⁺/Fe²⁺ redox cycling). Importantly, non-doped CeO2 NPs did not promote eryptosis. Nor they stimulated generation of ROS and Ca2+ influx. The presence of iron was found to mediate phosphatidylserine externalization, caspase activation, and changes in lipid membranes of erythrocytes. Notably, Fe³⁺-doped CeO2 NP-induced eryptosis was independent of p38 MAPK and CK1α. Iron-doped CeO2 NPs trigger eryptosis, an effect mediated by iron. Iron doping is a promising modification of CeO2 NPs, which can modulate their toxicity and widen their pharmaceutical profile.

Recent advances in the field of nanotechnology have paved the way for novel biomedical applications of nanomaterials. Nanoscale vanadates possess unique properties and have been reported to elicit anti-aging, antibacterial, anti-cancer and antioxidant effects. Likewise, they are potentially applied as bioimaging, drug delivery, theranostic or would healing agents, as well as biosensors. In this review, we highlight and critically discuss biomedical applications of vanadate nanostructures. This review article aims at identifying molecular hallmarks of vanadate nanomaterial-mediated cytotoxicity. Our analysis reveals that depending on size, morphology, composition, concentration, and external conditions (pH, irradiation, etc.) nanoscale vanadates induce oxidative stress, mitochondrial dysfunction, and lysosomal damage, culminating in cell death. Notably, besides apoptosis, nanosized vanadates trigger immunogenic non-apoptotic cell death pathways (ferroptosis and necroptosis) primarily due to overproduction of reactive oxygen species (ROS). Thus, our review demonstrates that nanostructured vanadates are multifaceted pharmacological agents with a wide spectrum of diverse biological applications. Our analysis demonstrates that redox activity of nanostructured vanadates is a key factor determining biological responses to them. Thus, more studies should focus on elucidating the biologically-relevant redox mechanisms of nanosized vanadates.

Ferroptosis is an iron-driven, phospholipid hydroperoxide-mediated cell death, which has recently emerged as an attractive tool in cancer research due to its ability to govern the anti-tumor immune response. A growing research interest in ferroptosis biology has revealed the contribution of this regulated cell death to multiple diseases. In addition to iron, ferroptosis has been reported to be triggered by multiple heavy metals, which sheds light on the novel aspects of heavy metals-induced cytotoxicity. In this review, the ability of zinc, an essential biogenic element with a wide array of biological functions, to modulate ferroptosis in normal and malignant cells has been summarized. Accumulating evidence suggests that zinc-induced biological effects can be mediated by ferroptosis induction or attenuation. In addition, the anti-cancer effects of zinc can be at least partly attributed to ferroptosis induction. The signaling pathways governing zinc-regulated ferroptosis are highlighted. It has been underscored that zinc-mediated modulation of ferroptosis is dependent on alterations of redox homeostasis, antioxidant defense (in particular, the SLC7A11/GSH/GPX4 axis), and iron metabolism. Additionally, data on ferroptosis induction by zinc oxide nanoparticles are summarized to emphasize the potential of these nanomaterials as a promising therapeutic choice in anti-cancer treatment.

Over the recent years, our understanding of the cell death machinery of mature erythrocytes has been greatly expanded. It resulted in the discovery of several regulated cell death (RCD) pathways in red blood cells. Apoptosis (eryptosis) and necroptosis of erythrocytes share certain features with their counterparts in nucleated cells, but they are also critically different in particular details. In this review article, we summarize the cell death subroutines in the erythroid precursors (apoptosis, necroptosis, and ferroptosis) in comparison to mature erythrocytes (eryptosis and erythronecroptosis) to highlight the consequences of organelle clearance and associated loss of multiple components of the cell death machinery upon erythrocyte maturation. Recent advances in understanding the role of erythrocyte RCDs in health and disease have expanded potential clinical applications of these lethal subroutines, emphasizing their contribution to the development of anemia, microthrombosis, and endothelial dysfunction, as well as their role as diagnostic biomarkers and markers of erythrocyte storage-induced lesions. Fas signaling and the functional caspase-8/caspase-3 system are not indispensable for eryptosis, but might be retained in mature erythrocytes to mediate the crosstalk between both erythrocyte-associated RCDs. The ability of erythrocytes to switch between eryptosis and necroptosis suggests that their cell death is not a simple unregulated mechanical disintegration, but a tightly controlled process. This allows investigation of eventual pharmacological interventions aimed at individual cell death subroutines of erythrocytes.

Titanium dioxide (TiO2) nanoparticles are promising biomedical agents characterized by good biocompatibility. In this study, we explored the cytotoxicity of TiO2−x nanoparticles with a different Ti3+(Ti2+)/Ti4+ ratio and analyzed the efficiency of eryptosis indices as a tool in nanotoxicology. Two types of TiO2−x nanoparticles (NPs) were synthesized by the hydrolysis of titanium alkoxide varying the nitric acid content in the hydrolysis mixture. Transmission electron microscopy (TEM) images show that 1-TiO2−x and 2-TiO2−x NPs are 5 nm in size, whereas X-ray photoelectron spectroscopy (XPS) reveals different Ti3+ (Ti2+)/Ti4+ ratios in the crystal lattices of synthesized NPs. 1-TiO2−x nanoparticles contained 54% Ti4+, 38% Ti3+, and 8% Ti2+, while the relative amount of Ti4+ and Ti3+ in the crystal lattice of 2-TiO2−x nanoparticles was 63% and 37%, respectively. Cell viability and cell motility induced by TiO2−x nanoparticles were investigated on primary fibroblast cultures. Eryptosis modulation by the nanoparticles along with cell death mechanisms was studied on rat erythrocytes. We report that both TiO2−x nanoparticles do not decrease the viability of fibroblasts simultaneously stimulating cell migration. Data from in vitro studies on erythrocytes indicate that TiO2−x nanoparticles trigger eryptosis via ROS- (1-TiO2−x) and Ca2+-mediated mechanisms (both TiO2−x nanoparticles) suggesting that evaluation of eryptosis parameters is a more sensitive nanotoxicological approach for TiO2−x nanoparticles than cultured fibroblast assays. TiO2−x nanoparticles are characterized by low toxicity against fibroblasts, but they induce eryptosis, which is shown to be a promising tool for nanotoxicity screening. The Ti3+ (Ti2+)/Ti4+ ratio at least partly determines the cytotoxicity mechanisms for TiO2−x nanoparticles.

Accumulating evidence suggests that manganese oxide nanoparticles (NPs) show multiple enzyme-mimicking antioxidant activities, which supports their potential in redox-targeting therapeutic strategies for diseases with impaired redox signaling. However, the systemic administration of any NP requires thorough hemocompatibility testing. In this study, we assessed the hemocompatibility of synthesized Mn3O4 NPs, identifying their ability to induce spontaneous hemolysis and eryptosis or impair osmotic fragility. Concentrations of up to 20 mg/L were found to be safe for erythrocytes. Eryptosis assays were shown to be more sensitive than hemolysis and osmotic fragility as markers of hemocompatibility for Mn3O4 NP testing. Flow cytometry- and confocal microscopy-based studies revealed that eryptosis induced by Mn3O4 NPs was accompanied by Ca2+ overload, altered redox homeostasis verified by enhanced intracellular reactive oxygen species (ROS) and reactive nitrogen species (RNS), and a decrease in the lipid order of cell membranes. Furthermore, Mn3O4 NP-induced eryptosis was calpain- and caspase-dependent.

Nanostructured materials have been suggested to be used as a source of dietary zinc for livestock animals. In this study, we assessed the cytotoxicity of newly synthesized nanostructured zinc carbonate hydroxide (ZnCH) Zn5(CO3)(OH)6microflakes. Cytotoxicity of the microflakes was assessed against murine L929 cell line and rat mature erythrocytes. Viability, motility, cell death pathways, implication of Ca2+, reactive oxygen species and reactive nitrogen species (RNS) signaling, caspases, and alterations of cell membranes following exposure of L929 cells to the microflakes were assessed. To assess hemocompatibility of the Zn-containing microflakes, osmotic fragility and hemolysis assays were performed, as well as multiple eryptosis parameters were evaluated. Our findings indicate a dose-response cytotoxicity of ZnCH microflakes against L929 cells with no toxicity observed for low concentrations (10 mg l-1and below). At high concentrations (25 mg l-1and above), ZnCH microflakes promoted nitrosyl stress, Ca2+- and caspase-dependent apoptosis, and altered lipid order of cell membranes in a dose-dependent manner, evidenced by up to 7-fold elevation of RNS-dependent fluorescence, 2.9-fold enhancement of Fura 2-dependent fluorescence, over 20-fold elevation of caspases-dependent fluorescence (caspase-3, caspase-8, and caspase-9), and up to 4.4-fold increase in the ratiometric index of the NR12S probe. Surprisingly, toxicity to enucleated mature erythrocytes was found to be lower compared to L929 cells. ZnCH microflakes induced eryptosis associated with oxidative stress, nitrosyl stress, Ca2+signaling and recruitment of caspases at 25-50-100 mg l-1. Eryptosis assays were found to be more sensitive than evaluation of hemolysis. Zn5(CO3)(OH)6microflakes show no cytotoxicity at low concentrations indicating their potential as a source of zinc for livestock animals.

In response to acute stress, nucleated cells undergo various regulated cell deaths (RCDs). Alternatively and in the long run, they permanently stop proliferating with the parallel acquisition of apoptosis resistance and the senescence-associated secretory phenotype. This particular long-term stress response is referred to as cellular senescence. Terminally differentiated, anucleate red blood cells (RBCs) cannot proliferate anymore and are incapable of synthesizing proteins. RBC senescence is therefore defined by phagocytosis-promoting age-related changes, such as band 3 protein-derived senescent erythrocyte-specific antigen (SESA)-mediated binding of autologous antibodies, desialylated glycoproteins, CD47 loss, and glycophorin depletion. Additionally, RBCs can undergo eryptosis, a Ca2+-driven RCD. Here, we consider the intracellular signals that drive RBC senescence and eryptosis to underscore how these pathways are intertwined. Our findings suggest that RBC senescence and eryptosis are distinct processes with their differences primarily lying in how fast senescent and eryptotic RBCs are removed from the circulation. Severe damage to RBCs promotes intense Ca2+ influx, oxidative and nitrosative stress, as well as activation of caspase-3. This eventually leads to phosphatidylserine externalization and eryptosis. Phosphatidylserine externalization, in turn, ensures swift erythrophagocytosis of eryptotic RBCs thus preventing hemolysis. On the other hand, suberyptotic stress stimuli are not sufficient to trigger rapid cell death. However, they mediate the gradual and inevitable accumulation of senescence-associated injuries. Clearance of senescent RBCs with low-level phosphatidylserine exposure is slow-paced, and mature RBCs possess mechanisms to eradicate senescence markers that tag cells for phagocytosis to extend their lifespan (e.g., generation of vesicles). In this review, we highlight the physiological significance of RBC senescence and eryptosis, and provide practical guidelines to distinguish them thus avoiding misinterpretation of experimental data.

Early studies have shown that erythrocytes have caspase-3 and caspase-8 and are capable of dying through an apoptotic-like cell death triggered by Ca2+ ionophores. This cell death is associated with apoptosis-like morphological signs, including cell shrinkage, membrane blebbing, and phosphatidylserine externalization. To emphasize that mature erythrocytes don't have the apoptotic mitochondrial machinery and distinguish this unique cell death modality from apoptosis, it was named "eryptosis". Over recent decades, our knowledge of eryptosis has been significantly expanded, providing more insights into the uniqueness of cell death pathways in erythrocytes. In this review, we aim to summarize our current understanding of eryptosis, formulate the nomenclature and guidelines to interpret results of eryptosis studies, provide a synopsis of morphological and biochemical features of eryptosis, and highlight the role of eryptosis in health and disease, including its druggability.

Introduction. Circulating tumor DNA (ctDNA) was identified as a powerful biomarker for treatment response evaluation and outcome prediction in malignant tumors, including diffuse large B-cell lymphoma (DLBCL). However, more studies are necessary to validate ctDNA utilization in general, and specifically in a real-world setting to facilitate standardization and transition into routine clinical practice. Therefore, we assessed pre-treatment ctDNA in 169 unselected previously untreated DLBCL patients consequently uniformly treated with R-CHOP chemoimmunotherapy at five academic hematology centers in the Czech Republic. Methods. ctDNA was analyzed by CAPP-Seq (CAncer Personalized Profiling by deep Sequencing) and a custom panel of 521 genes. DNA alterations were identified by VarScan2. ctDNA concentration was calculated from average variant allele frequencies and reported as human haploid genome equivalents (hGE) per ml of plasma. Germinal center B-cell like (GCB) and non-GCB subtyping was done by immunohistochemistry. Genetic classes were analyzed by LymphGen 2.0, COSMIC (Catalogue Of Somatic Mutations In Cancer) signatures by SigProfiler. Results. In all patients, the median age at diagnosis was 64 years (range 24-81), clinical stage III–IV in 68%, more than one extranodal involved sites in 40%, PS ECOG 2–4 in 24%, elevated LDH in 60%, International Prognostic Index (IPI) 3–5 in 50%, bulky disease over > 7.5 cm in 40%, and non-GCB DLBCL subtype in 47% of patients. Median follow-up was 2.73 years. The 2-year progression free and overall survivals (PFS and OS) were 73.6% and 84.3%, respectively. Without any threshold for input DNA quality and quantity, ctDNA was detected in 136 out of 169 analyzed patients (80%). Clinical characteristics of patients with and without detected ctDNA did not show any differences. The median plasma ctDNA concentration was 995 hGE/ml. ctDNA concentration was significantly higher in patients with clinical stage III–IV (median 1416.9 vs. 542.7 hGE/ml, p = 2.62e-5), age > 60 years (median 1110.7 vs. 691.2, p = 0.04), PS ECOG 2–4 (median 2130.9 vs. 824.2 hGE/ml, p = 2.62e-5), elevated LDH serum levels (median 1804.1 vs. 513.3 hGE/ml, p = 1.14e-9), bulky disease over > 7.5 cm (median 2234.9 vs. 624.8 hGE/ml, p = 4.58e-6), and in patients with IPI 3-5 (median 1646.7 vs. 631.7 hGE/ml, p = 8.33e-5). To find a pre-treatment ctDNA plasma concentration that would identify high risk patients (based on PFS analysis), patients from the General University Hospital (n = 72) were used as a discovery cohort and patients from other centers (n = 64) as a validation cohort (clinically well-balanced). In the discovery cohort, the threshold was determined at 3000 hGE/ml using Harrell's C-index. High ctDNA (> 3000 hGE/ml, 32% of patients) was associated with inferior PFS (2-year PFS 46% vs. 88%, 5-year PFS 38% vs. 71%, HR 3.99, p = 0.001). In the validation cohort, ctDNA concentration > 3000 hGE/ml (19% of patients) was also associated with inferior PFS (2-year PFS 50% vs. 78%, 5-year PFS 40% vs. 67%, HR 2.9, p = 0.024). It confirmed identified threshold that was consequently used for further analyses. In all patients, high ctDNA level (> 3000 hGE/ml, 26% of patients) was associated with inferior PFS (2-year PFS 48% vs. 83%, 5-year PFS 38% vs. 66%, HR 3.37, p < 0.001), and, importantly, also with inferior OS (2-year OS 74% vs. 88%, 5-year OS 54% vs. 75%, HR 2.56, p = 0.016). Moreover, ctDNA was an independent prognostic factor for PFS in multivariate analysis with IPI score (high ctDNA, HR 2.12, p = 0.029; IPI ordinal, HR 1.42, p = 0.01). High ctDNA patients had more advanced clinical stage, worse PS ECOG, higher LDH levels, higher IPI score (all with p < 0.001), and higher proportion of bulky disease (p = 0.005). The LymphGen assigned a genetic subtype to 44% of cases, reflecting the capability of the LympGen model to classify 50-60% of tumors (if copy number and translocation information is available). COSMIC mutational signatures showed differences between genetic and cell of origin subtypes, e.g., activation induced cytidine deaminase activity specifically in non-GCB DLBCL. Conclusion. Our study confirmed pre-treatment ctDNA concentration as a strong and independent risk factor associated with unfavorable DLBCL survival in a routine clinical setting, allowing parallel genetic and biological evaluation. First two authors contributed equally. Supported by NU21-03-00411, DRO-VFN00064165, LX22NPO5102, and SVV 260637.

Eryptosis is a type of regulated cell death of mature erythrocytes characterized by excessive Ca2+ accumulation followed by phosphatidylserine externalization. Eryptosis facilitates erythrophagocytosis resulting in eradication of damaged erythrocytes, which maintains the population of healthy erythrocytes in blood. Over recent years, a wide array of diseases has been reported to be linked to accelerated eryptosis, which leads to anemia. A growing number of studies furnish evidence that eryptosis is implicated in the pathogenesis of liver diseases. Herein, we summarize the current knowledge of eryptosis signaling, its physiological role, and the impact of eryptosis on anemia and hypercoagulation. In this article, upon systemically analyzing the PubMed-indexed publications, we also provide a comprehensive overview of the role of eryptosis in the spectrum of hepatic diseases, its contribution to the development of complications in liver pathology, metabolites (bilirubin, bile acids, etc.) that might trigger eryptosis in liver diseases, and eryptosis-inducing liver disease medications. Eryptosis in liver diseases contributes to anemia, hypercoagulation, and endothelial damage (via ferroptosis of endothelial cells). Treatment-associated anemia in liver diseases might be at least partly attributed to drug-induced eryptosis. Ultimately, we analyze the concept of inhibiting eryptosis pharmaceutically to prevent eryptosis-associated anemia and thrombosis in liver diseases.

Titanium dioxide nanoparticles (TiO2 NPs) have attracted significant interest for their ability to modulate reactive oxygen species (ROS) levels within cells. These nanoparticles have shown promise in drug delivery, sonodynamic, photodynamic, photothermal, and ionizing radiation-based anti-cancer therapies, as well as antimicrobial and antioxidant applications. In this study, we analyzed the effects of TiO2 NP defect structures-specifically the presence of stoichiometric (Ti4+) and non-stoichiometric (Ti-3. and Ti-2.) titanium ions within the crystal lattice-and the aggregation state of TiO2 NPs on their ROS scavenging capabilities in both cell-free conditions and H2O2-treated L929 cells. Two types of TiO2 NPs with varying concentrations of Ti-3. and Ti-2. ions were synthesized and characterized using XRD, TEM, SAXS, and XPS techniques. The antioxidant properties of these TiO2 NPs were assessed through chemiluminescence and optical spectroscopy using ROS sensors. Results from chemiluminescence and total antioxidant capacity assays indicated that the synthesized TiO2 NPs possess radical scavenging abilities, with TiO2 NPs(1), containing a higher amount of non- stoichiometric titanium ions, demonstrating a stronger effect. Concentrations of up to 40 mg/L of TiO2 NPs showed no impact on L929 cell viability or cell death. Moreover, TiO2 NPs(1) exhibited a tendency to reduce H2O2-induced oxidative stress in L929 cells, while TiO2 NPs(2) had the opposite effect, likely due to their propensity for aggregation in the cell medium. Our findings suggest that the synthesized TiO2 NPs have potential as agents for modulating intracellular ROS levels at concentrations that do not compromise cell viability.

Nanoparticles (NPs) with reactive oxygen species (ROS)-regulating ability have recently attracted great attention as promising agents for nanomedicine. In the present study, we have analyzed the effects of TiO2defect structure related to the presence of stoichiometric (Ti4+) and non-stoichiometric (Ti3+and Ti2+) titanium ions in the crystal lattice and TiO2NPs aggregation ability on H2O2- and tert-butyl hydroperoxide (tBOOH)-induced ROS production in L929 cells. Synthesized TiO2-A, TiO2-B, and TiO2-C NPs with varying Ti3+(Ti2+) content were characterized by x-ray powder diffraction, transmission electron microscopy, small-angle x-ray scattering, x-ray photoelectron spectroscopy, and optical spectroscopy methods. Given the role of ROS-mediated toxicity for metal oxide NPs, L929 cell viability and changes in the intracellular ROS levels in H2O2- and tBOOH-treated L929 cells incubated with TiO2NPs have been evaluated. Our research shows that both the amount of non-stoichiometric Ti3+and Ti2+ions in the crystal lattice of TiO2NPs and NPs aggregative behavior affect their catalytic activity, in particular, H2O2decomposition and, consequently, the efficiency of aggravating H2O2- and tBOOH-induced oxidative damage to L929 cells. TiO2-A NPs reveal the strongest H2O2decomposition activity aligning with their less pronounced additional effects on H2O2-treated L929 cells due to the highest amount of Ti3+(Ti2+) ions. TiO2-C NPs with smaller amounts of Ti3+ions and a tendency to aggregate in water solutions show lower antioxidant activity and, consequently, some elevation of the level of ROS in H2O2/tBOOH-treated L929 cells. Our findings suggest that synthesized TiO2NPs capable of enhancing ROS generation at concentrations non-toxic for normal cells, which should be further investigated to assess their possible application in nanomedicine as ROS-regulating pharmaceutical agents.

Aim. In this study, blood compatibility of ZnO nanoparticles-polymer nanocomplex (D-PAA/ZnONPs(SO42-)) synthesizedin situinto dextran-graft-polyacrylamide (D-PAA) using zinc sulphate as a precursor was tested using hemolysis, osmotic fragility and eryptosis assays.Materials and methods. Dose-dependent ability to induce eryptosis was assessed following 24 h incubation at concentrations of 0-800 mg l-1analyzing hallmarks of eryptosis (cell shrinkage and phosphatidylserine externalization), as well as reactive oxygen species generation. Hemolysis was detected spectrophotometrically based on hemoglobin release following exposure to the D-PAA/ZnONPs(SO42-) nanocomplex. Osmotic fragility test (OFT) involved detection of hemolysis of red blood cells exposed to 0.2% saline solution following incubation with the D-PAA/ZnONPs(SO42-) nanocomplex. Additional incubation of the nanocomplex in the presence or absence of either ascorbic acid or EGTA was used to reveal the implication of oxidative stress- or Ca2+-mediated mechanisms in D-PAA/ZnONPs(SO42-) nanocomplex-induced erythrotoxicity.Results. Hemocompatibility assessment of the D-PAA/ZnONPs(SO42-) nanocomplex revealed that it induced hemolysis and reduced resistance of erythrocytes to osmotic stress at concentrations of above 400 and 200 mg l-1, respectively. Oxidative stress- or Ca2+-mediated mechanisms were not involved in D-PAA/ZnONPs(SO42-) nanocomplex-induced hemolysis. Strikingly, the D-PAA/ZnONPs(SO42-) nanocomplex did not promote cell membrane scrambling, cell shrinkage and oxidative stress in red blood cells following the direct exposure for 24 h. Thus, the D-PAA/ZnONPs(SO42-) nanocomplex did not induce eryptosisin vitro. Eryptosis is generally considered to occur earlier than hemolysis in response to stress in order to prevent hemolytic cell death. Counterintuitively, our data suggest that hemolysis can be triggered by nanomaterials prior to eryptosis indicating that eryptosis and hemolysis assays should be used in combination for testing blood compatibility of nanomaterials.Conclusions. The D-PAA/ZnONPs(SO42-) nanocomplex has a good hemocompatibility profile at low concentrations. Hemocompatibility testing in nanotoxicology should include both eryptosis and hemolysis assays.

Introduction: Tumor drug resistance and systemic toxicity are major challenges of modern anticancer therapy. Nanotechnology makes it possible to create new materials with the required properties for anticancer therapy. Methods: In this research, Dextran-graft-Polyacrylamide/ZnO nanoparticles were used. The study was carried out using prostate (DU-145, LNCaP, PC-3), breast (MDA-MB-231, MCF-7, MCF-7 Dox) cancer cells and non-malignant (MAEC, BALB/3T3 clone A31) cells. Zinc was visualized with fluorescence in vitro and in vivo. ROS and apoptotic markers were identified by cytometry. Zinc accumulation and histopathological changes in the tumor, liver, kidney, and spleen were evaluated in a rat model. Results: ZnO nanoparticles dissociation and release of Zn2+ into the cytosol occurs in 2-3 hours for cancerous and non-cancerous cells. ROS upregulation was detected in all cells. For non-malignant cells, the difference between the initial ROS level was insignificant. The rate of carbohydrate metabolism in cancer cells was reduced by nanosystems. Zinc level in the tumor was upregulated by 25% and 39% after treatment with nanosystems and doxorubicin combined, respectively. The tumor Walker-256 carcinosarcoma volume was reduced twice following mono-treatment with the nanocomplex and 65-fold lower when the nanocomplex was combined with doxorubicin compared with controls. In the liver, kidney and spleen, the zinc level increased by 10-15% but no significant pathological alterations in the tissues were detected. Conclusion: D-PAA/ZnO NPs nanosystems were internalized by prostate, breast cancer cells and non-malignant cells via endocytosis after short time, but cytotoxicity against non-cancer cells were significantly lower in vitro and in vivo. D-PAA/ZnO NPs nanocomplex efficiently promoted cell death of tumor cells without showing cytotoxicity against non-malignant cells making it a promising anticancer agent.