Fatty acids characterisation by GC-MS, antiglycation effect at multiple stages and protection of erythrocytes cells from oxidative damage induced by glycation of albumin of Opuntia ficus-indica (L.) Mill seed oil cultivated in Eastern Morocco: Experimental and computational approaches.

ETHNOPHARMACOLOGICAL RELEVANCE: Opuntia ficus-indica (L.) Mill is frequently observed in the Moroccan traditional medicinal system, where these approaches are employed to mitigate the onset of diabetes and the subsequent complications it may entail. AIM OF THE STUDY: The aim of this research was to examine the effectiveness of Opuntia ficus-indica seed oil in preventing diabetic complications. Specifically, the study assessed its ability to counteract glycation at various stages, protected red blood cells from the harmful effects of glycated albumin, and inhibited pancreatic lipase digestive enzymes to understand its potential antihyperglycemic properties. Additionally, the study aimed to identify the chemical components responsible for these effects, evaluate antioxidant and anti-inflammatory properties, and conduct computational investigations such as molecular docking. MATERIALS AND METHODS: The assessement of Opuntia ficus-indica seed oil antiglycation properties involved co-incubating the extract oil with a bovine serum albumin-glucose glycation model. The study investigated various stages of glycation, incorporating fructosamine (inceptive stage), protein carbonyls (intermediate stage), and AGEs (late stage). Additionally, measurement of ß-amyloid aggregation of albumin was performed using Congo red, which is specific to amyloid structures. Additionally, the evaluation of oil's safeguarding effect on erythrocytes against toxicity induced by glycated albumin included the measurement of erythrocyte hemolysis, lipid peroxidation, reduced glutathione. The fatty acid of Opuntia ficus-indica seed oil were analyzed using Gas Chromatography-Mass Spectrometry (GC-MS). The in vitro evaluation of antihyperglycemic activity involved the use of pancreatic lipase enzyme, while the assessement of antioxidant capability was carried out through the utilization of the ABTS and FRAP methods. The in vitro assessement of the denaturation of albumin activity was also conducted. In conjunction with the experimental outcomes, computational investigations were undertaken, specifically employing ADMET (absorption, distribution, metabolism, excretion, and toxicity) analysis. Furthermore, molecular docking was utilized to predict antioxidant and antiglycation mechanisms based on protein targets. RESULTS: In vitro glycation assays, Opuntia ficus-indica seed oil displayed targeted inhibitory effects at multiple distinct stages. Within erythrocytes, in addition to mitigating hemolysis and lipid peroxidation induced by glycated albumin. GC-MS investigation revealed a richness of fatty acids and the most abundant compounds are Linoleic acid (36.59%), Palmitic acid (20.84%) and Oleic acid (19.33%) respectively. The findings of antioxidant ability showed a remarkable activity on FRAP and ABTS radicals. This oil showed a pronounced inhibitory impact (p < 0.001) on pancreatic lipase enzyme. It also exerted a notibale inhibition of albumin denaturation, in vitro. CONCLUSION: The identified results were supported by the abundant compounds of fatty acids unveiled through GC-MS analysis, along with the computational investigation and molecular docking.

Erythrocytes Nanoparticle Delivery: A Boon for Targeting Tumor.

Although nanoparticles (NPs) have many advantages as drug delivery systems, their poor stability in circulation, premature drug release, and nonspecific uptake in non-target organs have prompted biomimetic approaches to camouflage nano vehicles using natural cell membranes. Among them, which are extensively studied in erythrocytes, are the most abundant circulating blood cells. They are specially used for biomimetic coating on artificial NPs due to their excellent properties of good biocompatibility, biodegradability, non-immunogenicity, and long-term blood circulation. Erythrocyte-mimicking nanoparticles (EM-NPs) are prepared by combining nanoparticle cores with naturally derived erythrocyte (red blood cell or RBC) membranes. Compared with conventional nanosystems, EM-NPs hold the preferable characteristics of prolonged blood circulation time and immune evasion. In this review, the biomimetic platform of erythrocyte membrane-coated NPs is described in various aspects, with particular focus placed on the coating mechanism, preparation methods, characterization method, and recent advances in the biomedical applications of EM-NPs concerning cancer and targeted delivery.

Small-molecule α-lipoic acid targets ELK1 to balance human neutrophil and erythrocyte differentiation.

BACKGROUND: Erythroid and myeloid differentiation disorders are commonly occurred in leukemia. Given that the relationship between erythroid and myeloid lineages is still unclear. To find the co-regulators in erythroid and myeloid differentiation might help to find new target for therapy of myeloid leukemia. In hematopoiesis, ALA (alpha lipoic acid) is reported to inhibit neutrophil lineage determination by targeting transcription factor ELK1 in granulocyte-monocyte progenitors via splicing factor SF3B1. However, further exploration is needed to determine whether ELK1 is a common regulatory factor for erythroid and myeloid differentiation. METHODS: In vitro culture of isolated CD34+, CMPs (common myeloid progenitors) and CD34+ CD371- HSPCs (hematopoietic stem progenitor cells) were performed to assay the differentiation potential of monocytes, neutrophils, and erythrocytes. Overexpression lentivirus of long isoform (L-ELK1) or the short isoform (S-ELK1) of ELK1 transduced CD34+ HSPCs were transplanted into NSG mice to assay the human lymphocyte and myeloid differentiation differences 3 months after transplantation. Knocking down of SRSF11, which was high expressed in CD371+GMPs (granulocyte-monocyte progenitors), upregulated by ALA and binding to ELK1-RNA splicing site, was performed to analyze the function in erythroid differentiation derived from CD34+ CD123mid CD38+ CD371- HPCs (hematopoietic progenitor cells). RNA sequencing of L-ELK1 and S-ELK1 overexpressed CD34+ CD123mid CD38+ CD371- HPCs were performed to assay the signals changed by ELK1. RESULTS: Here, we presented new evidence that ALA promoted erythroid differentiation by targeting the transcription factor ELK1 in CD34+ CD371- hematopoietic stem progenitor cells (HSPCs). Overexpression of either the long isoform (L-ELK1) or the short isoform (S-ELK1) of ELK1 inhibited erythroid-cell differentiation, but knockdown of ELK1 did not affect erythroid-cell differentiation. RNAseq analysis of CD34+ CD123mid CD38+ CD371- HPCs showed that L-ELK1 upregulated the expression of genes related to neutrophil activity, phosphorylation, and hypoxia signals, while S-ELK1 mainly regulated hypoxia-related signals. However, most of the genes that were upregulated by L-ELK1 were only moderately upregulated by S-ELK1, which might be due to a lack of serum response factor interaction and regulation domains in S-ELK1 compared to L-ELK1. In summary, the differentiation of neutrophils and erythrocytes might need to rely on the dose of L-ELK1 and S-ELK1 to achieve precise regulation via RNA splicing signals at early lineage commitment. CONCLUSIONS: ALA and ELK1 are found to regulate both human granulopoiesis and erythropoiesis via RNA spliceosome, and ALA-ELK1 signal might be the target of human leukemia therapy.

Intelligent berberine-loaded erythrocytes attenuated inflammatory cytokine productions in macrophages.

Erythrocytes are impressive tools for drug delivery, especially to macrophages. Therefore, berberine was loaded into erythrocytes using both hypotonic pre-swelling and endocytosis methods to target macrophages. Physicochemical and kinetic parameters of the resulting carrier cells, such as drug loading/release kinetics, osmotic fragility, and hematological indices, were determined. Drug loading was optimized for the study using Taguchi experimental design and lab experiments. Loaded erythrocytes were targeted to macrophages using ZnCl2 and bis-sulfosuccinimidyl-suberate, and targeting was evaluated using flow cytometry and Wright-Giemsa staining. Differentiated macrophages were stimulated with lipopolysaccharide, and the inflammatory profiles of macrophages were evaluated using ELISA, western blotting, and real-time PCR. Findings indicated that the endocytosis method is preferred due to its low impact on the erythrocyte's structural integrity. Maximum loading achieved (1386.68 ± 22.43 µg/ml) at 1500 µg/ml berberine treatment at 37 °C for 2 h. Berberine successfully inhibited NF-κB translation in macrophages, and inflammatory response markers such as IL-1ß, IL-8, IL-23, and TNF-α were decreased by approximately ninefold, sixfold, twofold, eightfold, and twofold, respectively, compared to the LPS-treated macrophages. It was concluded that berberine-loaded erythrocytes can effectively target macrophages and modulate the inflammatory response.

Attenuation of Oxidative Stress in Erythrocytes Stored with Vitamin C and l-Carnitine in Additive Solution-7.

Background: Blood transfusion has advanced toward component therapy for specific requirements during trauma and surgery. Oxidative stress is induced in erythrocytes during storage. Hence, antioxidants as additives can be employed to counteract oxidative stress and enhance antioxidant defenses. Therefore, this study investigates the combinatorial effects of vitamin C and l-carnitine on erythrocytes during storage. Methodology: Erythrocyte samples were categorized into control and experimental groups-vitamin C (10 mM) and l-carnitine (10 mM) and stored under blood bank conditions (at 4°C) for 35 days. Hemoglobin (Hb), antioxidant enzymes (superoxide dismutase [SOD], catalase [CAT] and glutathione peroxidase [GPX]), lipid peroxidation products (conjugate dienes and thiobarbituric acid reactive substances [TBARSs]), protein oxidation products, metabolic markers (glucose, lactate dehydrogenase), glutathione (GSH), superoxides, and hemolysis were assessed at weekly intervals. Results: SOD activity increased on day 7 in the controls, whereas it increased on days 7 and 14 in the experimental groups. CAT activity increased on day 35 in both the groups. GPX activity increased on day 7 in the controls. Hb levels decreased on days 14 and 35 in the controls and on day 35 in the experimental groups. Hemolysis increased from day 7 onward in both the groups. Protein oxidation products were maintained throughout the storage. GSH levels increased on day 21 in the controls and on days 14 and 21 in the experimental groups. Superoxides and conjugate dienes decreased from day 14 in both the groups. TBARSs decreased on day 7 in the experimental groups. Conclusion: Vitamin C and l-carnitine have synergistically enhanced the efficacy of stored erythrocytes in terms of Hb, antioxidant enzymes, and lipid peroxidation.

Environmental cadmium exposure perturbs systemic iron homeostasis via hemolysis and inflammation, leading to hepatic ferroptosis in common carp (Cyprinus carpio L.).

Cadmium (Cd) pollution is considered a pressing challenge to eco-environment and public health worldwide. Although it has been well-documented that Cd exhibits various adverse effects on aquatic animals, it is still largely unknown whether and how Cd at environmentally relevant concentrations affects iron metabolism. Here, we studied the effects of environmental Cd exposure (5 and 50 µg/L) on iron homeostasis and possible mechanisms in common carp. The data revealed that Cd elevated serum iron, transferrin saturation and iron deposition in livers and spleens, leading to the disruption of systemic iron homeostasis. Mechanistic investigations substantiated that Cd drove hemolysis by compromising the osmotic fragility and inducing defective morphology of erythrocytes. Cd concurrently exacerbated hepatic inflammatory responses, resulting in the activation of IL6-Stat3 signaling and subsequent hepcidin transcription. Notably, Cd elicited ferroptosis through increased iron burden and oxidative stress in livers. Taken together, our findings provide evidence and mechanistic insight that environmental Cd exposure could undermine iron homeostasis via erythrotoxicity and hepatotoxicity. Further investigation and ecological risk assessment of Cd and other pollutants on metabolism-related effects is warranted, especially under the realistic exposure scenarios.

Indicaxanthin prevents eryptosis induced by cigarette smoke extract by interfering with active Fas-mediated signaling.

A physiological mechanism of programmed cell death called eryptosis occurs in aged or damaged red blood cells (RBCs). Dysregulated eryptosis contributes to abnormal microcirculation and prothrombotic risk. Cigarette smoke extract (CSE) induces a p38 MAPK-initiated, Fas-mediated eryptosis, activating the death-inducing signaling complex (DISC). Indicaxanthin (Ind) from cactus pear fruits, is a bioavailable dietary phytochemical in humans and it is able to incorporate into RBCs enhancing their defense against numerous stimuli. This in vitro work shows that Ind, at concentrations that mimic plasma concentrations after a fruit meal, protects erythrocytes from CSE-induced eryptosis. CSE from commercial cigarettes was prepared in aqueous solution using an impinger air sampler and nicotine content was determined. RBCs were treated with CSE for 3 h in the absence or presence of increasing concentrations of Ind (from 1 to 5 µM). Cytofluorimetric measurements indicated that Ind reduced CSE-induced phosphatidylserine externalization and ceramide formation in a concentration-dependent manner. Confocal microscopy visualization and coimmunoprecipitation experiments revealed that Ind prevented both CSE-triggered Fas aggregation and FasL/FADD/caspase 8 recruitment in the membrane, indicating inhibition of DISC assembly. Ind inhibited the phosphorylation of p38 MAPK, caspase-8/caspase-3 cleavage, and caspase-3 activity induced by CSE. Finally, Ind reduced CSE-induced ATP depletion and restored aminophospholipid translocase activity impaired by CSE treatment. In conclusion, Ind concentrations comparable to nutritionally relevant plasma concentrations, can prevent Fas-mediated RBC death signaling induced by CSE, which suggests that dietary intake of cactus pear fruits may limit the deleterious effects of cigarette smoking.

Blackcurrant press cake by-product: Increased chemical bioaccessibility and reduced antioxidant protection after in vitro simulation of gastrointestinal digestion.

This study describes the bioaccessibility in terms of total phenolic content (TPC) and antioxidant capacity before and after in vitro digestion from blackcurrant press cake extracts (BPC) and the bioactivity in cell culture, human erythrocytes as well as the in silico analysis. Chemical analysis of BPC presented an increase in TPC (270%) and anthocyanins (136%) after in vitro digestion, resulting in an improvement of antioxidant activity (DPPH 112%; FRAP: 153%). This behavior may be related to the highest activity of cyanidin-3-rutinoside, as confirmed by in silico analysis. The digested BPC did not exert cytotoxicity in cells and showed less antioxidant activity against the oxidative damage induced in endothelial cells and human erythrocytes compared to the non-digested extract. The results raise a question about the reliability we should place on results obtained only from crude samples, especially those that will be used to produce foods or nutraceuticals.

Assessment of dietary polyvinylchloride, polypropylene and polyethylene terephthalate exposure in Nile tilapia, Oreochromis niloticus: Bioaccumulation, and effects on behaviour, growth, hematology and histology.

Microplastics (MPs) have been recognized as emerging aquatic pollutants receiving major concern due to their detrimental effects on aquatic life. Nile Tilapia, Oreochromis niloticus is a model species considered in toxicological studies to address the effects of pollutants in freshwater animals. However, comprehensive knowledge comparing the impacts on fish across various MPs polymers is scarce. Therefore, the overarching aim of the current study was to examine the bioconcentration of MPs polymers: polyvinylchloride (PVC), polypropylene (PP), and polyethylene terephthalate (PET), and their toxic effects on growth, and behavioral responses, hematology, and histology of gills, liver, and intestine in O. niloticus. Fishes were subjected to a 21-day dietary exposure to MPs by assigning them into six treatment groups: T1 (4% of PVC), T2 (4% of PP), T3 (4% of PET), T4 (8% of PVC), T5 (8% of PP), T6 (8% of PET), and control (0% of MPs), to assess the effects on fish across the polymers and dosage. Results showed several abnormalities in anatomical and behavioral parameters, lower growth, and high mortality in MPs-exposed fish, indicating a dose-dependent relationship. The elevated dosage of polymers raised the bioavailability of PVC, PP, and PET in gills and gut tissues. Noteworthy erythrocyte degeneration referred to cytotoxicity and stress imposed by MPs, whereas the alterations in hematological parameters were possibly due to blood cell damage, also indicating mechanisms of defense against MPs toxicity. Histopathological changes in the gills, liver, and intestine confirmed the degree of toxicity and associated dysfunctions in fish. A higher sensitivity of O. niloticus to PET-MPs compared to other polymers is likely due to its chemical properties and species-specific morphological and physiological characteristics. Overall, the present study reveals valuable insights into the emerging threat of MPs toxicity in freshwater species, which could be supportive of future toxicological research.

Possible Markers for Distinguishing benign and Malignant Thyroid Tumors and Predicting Malignancy in Patients with Genetic Predisposition to Cancer.

BACKGROUND: We hypothesized that mutations in several genes disrupt oxidative metabolism, increasing the risk of developing tumors and their malignancy in patients with a family predisposition to cancer. The purpose of our study was to assess the characteristics of oxidative metabolism in patients with malignant and benign tumor with and without a family history of cancer and identify the marker predicting the likelihood of malignancy. METHODS: We conducted a study on patients with thyroid pathology (thyrotoxicosis, benign tumor pathology of the thyroid gland, and thyroid cancer) who underwent treatment at LLC "Oncology Scientific Research Center" in Tbilisi, Georgia between 2020-2021.  In patients' blood the thyroid hormones content, the oxidative metabolism parameters (activity of nonenzymatic antioxidant system (TAA), malondialdehyde (MDA) content),  geometrical and rheological (deformability index (EDI), membrane proteins content) characteristics of erythrocytes were determined. RESULTS: in the patient's blood serum with benign tumor (47 patients) MDA exceeded (p<0.005) and TAA decreased (p<0.005) in comparison to the control level; in patients with thyroid cancer (35 patients), MDA also exceeded (p<0.005), while TAA increased (p<0.005) up to the control level. In patients with benign and malignant tumors, the size of erythrocytes increased compared to the control indicators (p<0.005); in patients with thyroid cancer and benign tumors with a family history of cancer (29 patients) EDI increased (p<0.005), content of GLUT1 in erythrocyte membranes decreased (p<0.005) compared to the control level. CONCLUSIONS: Alterations in redox metabolism play a crucial role in tumor formation; an imbalance between anti-/pro-oxidant systems may contribute to tumor formation and support its progression into a more malignant state. Thyroid cancer is characterized by a reduction in erythrocyte deformability, related to TSH levels. These alterations are less detectable in patients with benign thyroid tumors with a family history of cancer.

Erythrocyte PIG-A mutant frequencies in cancer patients receiving cisplatin.

BACKGROUND: Cisplatin is a primary chemotherapy choice for various solid tumors. DNA damage caused by cisplatin results in apoptosis of tumor cells. Cisplatin-induced DNA damage, however, may also result in mutations in normal cells and the initiation of secondary malignancies. In the current study, we have used the erythrocyte PIG-A assay to evaluate mutagenesis in non-tumor hematopoietic tissue of cancer patients receiving cisplatin chemotherapy. METHODS: Twenty-one head and neck cancer patients undergoing treatment with cisplatin were monitored for the presence of PIG-A mutant total erythrocytes and the young erythrocytes, reticulocytes (RETs), in peripheral blood for up to five and a half months from the initiation of the anti-neoplastic chemotherapy. RESULTS: PIG-A mutant frequency (MF) in RETs increased at least two-fold in 15 patients at some point of the monitoring, while the frequency of total mutant RBCs increased at least two-fold in 6 patients. A general trend for an increase in the frequency of mutant RETs and total mutant RBCs was observed in 19 and 18 patients, respectively. Only in one patient did both RET and total RBC PIG-A MFs did not increase at any time-point over the monitoring period. CONCLUSION: Cisplatin chemotherapy induces moderate increases in the frequency of PIG-A mutant erythrocytes in head and neck cancer patients. Mutagenicity measured with the flow cytometric PIG-A assay may serve as a tool for predicting adverse outcomes of genotoxic antineoplastic therapy.

Renal Clearable Nanodots-Engineered Erythrocytes with Enhanced Circulation and Tumor Accumulation for Photothermal Therapy of Hepatocellular Carcinoma.

Living cell-mediated nanodelivery system is considered a promising candidate for targeted antitumor therapy; however, their use is restricted by the adverse interactions between carrier cells and nanocargos. Herein, a novel erythrocyte-based nanodelivery system is developed by assembling renal-clearable copper sulfide (CuS) nanodots on the outer membranes of erythrocytes via a lipid fusion approach, and demonstrate that it is an efficient photothermal platform against hepatocellular carcinoma. After intravenous injection of the nanodelivery system, CuS nanodots assembled on erythrocytes can be released from the system, accumulate in tumors in response to the high shear stress of bloodstream, and show excellent photothermal antitumor effect under the near infrared laser irradiation. Therefore, the erythrocyte-mediated nanodelivery system holds many advantages including prolonged blood circulation duration and enhanced tumor accumulation. Significantly, the elimination half-life of the nanodelivery system is 74.75 ± 8.77 h, which is much longer than that of nanodots (33.56 ± 2.36 h). Moreover, the other two kinds of nanodots can be well assembled onto erythrocytes to produce other erythrocyte-based hitchhiking platforms. Together, the findings promote not only the development of novel erythrocyte-based nanodelivery systems as potential platforms for tumor treatment but also their further clinical translation toward personalized healthcare.

Association of initial lactate levels and red blood cell transfusion strategy with outcomes after severe trauma: a post hoc analysis of the RESTRIC trial.

BACKGROUND: The appropriateness of a restrictive transfusion strategy for those with active bleeding after traumatic injury remains uncertain. Given the association between tissue hypoxia and lactate levels, we hypothesized that the optimal transfusion strategy may differ based on lactate levels. This post hoc analysis of the RESTRIC trial sought to investigate the association between transfusion strategies and patient outcomes based on initial lactate levels. METHODS: We performed a post hoc analysis of the RESTRIC trial, a cluster-randomized, crossover, non-inferiority multicenter trials, comparing a restrictive and liberal red blood cell transfusion strategy for adult trauma patients at risk of major bleeding. This was conducted during the initial phase of trauma resuscitation; from emergency department arrival up to 7 days after hospital admission or intensive care unit (ICU) discharge. Patients were grouped by lactate levels at emergency department arrival: low (< 2.5 mmol/L), middle (≥ 2.5 and < 4.0 mmol/L), and high (≥ 4.0 mmol/L). We compared 28 days mortality and ICU-free and ventilator-free days using multiple linear regression among groups. RESULTS: Of the 422 RESTRIC trial participants, 396 were analyzed, with low (n = 131), middle (n = 113), and high (n = 152) lactate. Across all lactate groups, 28 days mortality was similar between strategies. However, in the low lactate group, the restrictive approach correlated with more ICU-free (ß coefficient 3.16; 95% CI 0.45 to 5.86) and ventilator-free days (ß coefficient 2.72; 95% CI 0.18 to 5.26) compared to the liberal strategy. These findings persisted even after excluding patients with severe traumatic brain injury. CONCLUSIONS: Our results suggest that restrictive transfusion strategy might not have a significant impact on 28-day survival rates, regardless of lactate levels. However, the liberal transfusion strategy may lead to shorter ICU- and ventilator-free days for patients with low initial blood lactate levels.

Selective elimination of younger erythrocytes in blood circulation and associated molecular changes in benzo (a) pyrene induced mouse model of lung cancer.

Background: Anemia is a common feature in cancer patients. The present research was conducted to explore the mechanisms of induction of anemia in a mouse model of lung cancer. Methods: The lung cancer was induced by treating orally with BaP (50 mg/kg body weight, twice a week for four weeks). The erythrocyte kinetics were studied using a double in vivo biotinylation (DIB) technique. ROS production and apoptosis analysis were done by staining with the CMH2DCFDA stain and anti-mouse Annexin V antibody, followed by flow cytometry. The expression of antioxidant, apoptotic, anti-apoptotic and inflammatory genes was analyzed by quantitative PCR (RT-qPCR). Results: BaP-induced tumour reduced body weight and induced persistent haemolytic anaemia. The kinetics data suggest that, though reticulocyte production was enhanced, the proportion of young erythrocytes did not increase in the same proportion. The young aged erythrocytes were selectively eliminated from blood circulation, but intermediate and old aged erythrocytes persisted for a longer duration. The tumour progression leads to a significant increase in ROS production and apoptosis in the erythrocytes. The molecular data suggests that the expression levels of antioxidants (SOD1, catalase, and GPX1) and erythropoietin (Epo) were significantly increased. The anti-inflammatory genes Interleukin-6 (IL-6), Interleukin-10 (IL-10) were significantly decreased.Apoptotic genes Bax, and caspase 3 were significantly decreased while Bcl 2 was significantly increased in the blood of tumour-bearing mice. Conclusions: The overall data suggest that erythrocyte turnover is severely modulated with the progression of tumor. The apoptosis, ROS levels, antioxidant, anti-apoptotic, and Epo gene expressions were increased, but proapoptotic and anti-inflammatory gene expression were suppressed.

An Erythrocyte-Templated Iron Single-Atom Nanozyme for Wound Healing.

Iron single-atom nanozymes represent a promising artificial enzyme with superior activity owing to uniform active sites that can precisely mimic active center of nature enzymes. However, current synthetic strategies are hard to guarantee each active site at single-atom state. In this work, an erythrocyte-templated strategy by utilizing intrinsic hemin active center of hemoglobin as sing-atom source for nanozyme formation is developed. By combining cell fixation, porous salinization, and high-temperature carbonization, erythrocytes are successfully served as uniform templates to synthesize nanozymes with fully single-atom FeN4 active sites which derived from hemin of hemoglobin, resulting in an enhanced peroxidase (POD)-like activity. Interestingly, the catalytic activity of erythrocyte-templated nanozyme (ETN) shows dependence on animal species, among which murine ETN performed superior catalytic efficiency. In addition, the as-prepared ETNs display a honeycomb-like network structure, serving as a sponge to accelerate hemostasis based on the interactions with prothrombin and fibrinogen. These features enable ETN to effectively kill methicillin-resistant Staphylococcus aureus (MRSA) by combining POD-like catalysis with near-infrared (NIR) induced photothermal effect, and subsequently suitable to promote wound healing. This study provides a proof-of-concept for facile fabrication of multifunctional nanozymes with uniform single-atom active sites by utilizing intrinsic iron structure characteristics of biogenic source like erythrocytes.

An Immunomodulatory Zinc-Alum/Ovalbumin Nanovaccine Boosts Cancer Metalloimmunotherapy Through Erythrocyte-Assisted Cascade Immune Activation.

Cancer therapeutic vaccines are powerful tools for immune system activation and eliciting protective responses against tumors. However, their efficacy has often been hindered by weak and slow immune responses. Here, the authors introduce an immunization strategy employing senescent erythrocytes to facilitate the accumulation of immunomodulatory zinc-Alum/ovalbumin (ZAlum/OVA) nanovaccines within both the spleen and solid tumors by temporarily saturating liver macrophages. This approach sets the stage for boosted cancer metalloimmunotherapy through a cascade immune activation. The accumulation of ZAlum/OVA nanovaccines in the spleen substantially enhances autophagy-dependent antigen presentation in dendritic cells, rapidly initiating OVA-specific T-cell responses against solid tumors. Concurrently, ZAlum/OVA nanovaccines accumulated in the tumor microenvironment trigger immunogenic cell death, leading to the induction of individualized tumor-associated antigen-specific T cell responses and increased T cell infiltration. This erythrocyte-assisted cascade immune activation using ZAlum/OVA nanovaccines results in rapid and robust antitumor immunity induction, holding great potential for clinical cancer metalloimmunotherapy.

Iron supplementation in the diets of hybrid catfish (Ictalurus punctatus × I. furcatus) juveniles affected haematocrit levels and potentially decreased disease resistance to Edwardsiella ictaluri.

To prevent catfish idiopathic anaemia, diets fortified with iron have been adopted as a regular practice on commercial catfish farms to promote erythropoiesis. However, the effects of prolonged exposure of excess dietary iron on production performance and disease resistance for hybrid catfish (Ictalurus punctatus × I. furcatus) remains unknown. Four experimental diets were supplemented with ferrous monosulphate to provide 0, 500, 1000, and 1500 mg of iron per kg of diet. Groups of 16 hybrid catfish juveniles (~22.4 g) were stocked in each of 20, 110-L aquaria (n = 5), and experimental diets were offered to the fish to apparent satiation for 12 weeks. At the end of the study, production performance, survival, condition indices, as well as protein and iron retention were unaffected by the dietary treatments. Blood haematocrit and the iron concentration in the whole-body presented a linear increase with the increasing the dietary iron. The remaining fish from the feeding trial was challenged with Edwardsiella ictaluri. Mortality was mainly observed for the dietary groups treated with iron supplemented diets. The results for this study suggest that iron supplementation beyond the required levels does affect the blood production, and it may increase their susceptibility to E. ictaluri infection.

Erythrocyte-Leveraged Oncolytic Virotherapy (ELeOVt): Oncolytic Virus Assembly on Erythrocyte Surface to Combat Pulmonary Metastasis and Alleviate Side Effects.

Despite being a new promising tool for cancer therapy, intravenous delivery of oncolytic viruses (OVs) is greatly limited by poor tumor targeting, rapid clearance in the blood, severe organ toxicity, and cytokine release syndrome. Herein, a simple and efficient strategy of erythrocyte-leveraged oncolytic virotherapy (ELeOVt) is reported, which for the first time assembled OVs on the surface of erythrocytes with up to near 100% efficiency and allowed targeted delivery of OVs to the lung after intravenous injection to achieve excellent treatment of pulmonary metastases while greatly improving the biocompatibility of OVs as a drug. Polyethyleneimine (PEI) as a bridge to assemble OVs on erythrocytes also played an important role in promoting the transfection of OVs. It is found that ELeOVt approach significantly prolonged the circulation time of OVs and increased the OVs distribution in the lung by more than tenfold, thereby significantly improving the treatment of lung metastases while reducing organ and systemic toxicity. Taken together, these findings suggest that the ELeOVt provides a biocompatible, efficient, and widely available approach to empower OVs to combat lung metastasis.

Cemtirestat dimerization in liposomes and erythrocytes exposed to peroxyl radicals was reverted by thiol-disulfide exchange with GSH.

In the model system of DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine) liposomes exposed to peroxyl radicals generated by the azoinitiator AAPH, cemtirestat (CMTI-SH) inhibited lipid peroxidation more efficiently than the natural antioxidant glutathione. In the concentrations 100 to 500 µM, both CMTI-SH and GSH induced distinct lag phases in the initial stages of lipid peroxidation yet GSH produced consistently shorter induction periods (about twice) than equimolar CMTI-SH. Moreover, concentration dependence of lipid peroxidation inhibition measured at the 80th minute, revealed about three times higher IC50 value for GSH compared to CMTI-SH. When the incubations prolonged till 180 min no further absorbance changes at 270 and 302 nm, respectively, occurred. After addition of the reducing agent tris(2-carboxyethyl)phosphine, the absorbance peak at 270 nm shifted back to 302 nm. These findings pointed to the presence of reducible CMTI-SH disulfide whose definite structure was confirmed by proving identity of TLC retention and spectral data with those of the synthesized CMTI disulfide. When CMTI-SH and GSH were present simultaneously in the liposomal incubations, the mixing effect on the induction period was synergistic rather than additive. This was explained by ability of GSH to reduce CMTI disulfide which was proved in separate experiments with an authentic CMTI disulfide prepared synthetically. This finding was also demonstrated by experiment with CMTI-disulfide to protect the erythrocytes against oxidative damage induced by peroxyl radicals. To conclude, CMTI-SH scavenges reactive oxygen species yielding CMTI disulfide while GSH maintains CMTI-SH in the reduced state. This finding was also demonstrated by experiment with CMTI-disulfide to protect the erythrocytes against oxidative damage induced by peroxyl radicals. CMTI-SH would thus represent the first line of the cellular defense against peroxyl radical mediated oxidative stress.

Cemtirestat inhibited lipid peroxidation more efficiently than GSHCemtirestat disulfide was proved as the main oxidation productCemtirestat disulfide protected erythrocytes against oxidative damageCemtirestat disulfide was readily reduced by GSHMechanism of thiol-disulfide exchange reaction was suggested.