Search Results (47)

Accidental and deliberate poisoning by Ricinus communis, a wild plant cultivated for castor oil, has been known for centuries. Seeds contain one of the deadliest known plant toxins, ricin, along with its dimer RCA120. Due to its toxicity and ease of extraction, ricin poses a potential threat as a chemical weapon, highlighting the need for reliable laboratory detection. In this work, ricin and RCA120 (Ricinus communis agglutinin) were extracted from castor beans and purified by solid-phase extraction (SPE) and Gel Permeation Chromatography (GPC). Purity was confirmed through Sodium Dodecyl Sulphate-PolyAcrylamide Gel Electrophoresis (SDS-PAGE). Then, lab-made standards were used to validate an Enzyme Linked ImmunoSorbent Assay (ELISA) kit to identify ricin in biological matrices after SPE extraction. Parallelly, SDS-PAGE was used to qualitatively confirm the presence of toxins. The developed method demonstrated a detection limit (LOD) of 2.74 ng/mL and was used to analyse biological specimens of seven human intoxication cases managed by Pavia Poison Control Centre (PCC). This study provides a quiet, inexpensive protocol that within 5 h allows the screening of potential ricin-exposure victims. Its implementation in clinical laboratories could support the management of intoxicated patients, ensuring faster and more accurate clinical response. Full article

Ricin, a type 2 ribosome-inactivating protein, is a lethal toxin found in castor bean seeds. Although the systemic toxicity of ricin has been extensively studied, its localized effect on the gastrointestinal tract remains a critical concern, particularly in the case of oral ingestion. This study investigates the cytotoxic effects of ricin on human intestinal epithelial cell lines and its impact on epithelial barrier integrity. Ricin cytotoxicity was assessed on the intestinal-derived HT29 and Caco-2 cell lines using dose– and time–response assays, while the epithelial integrity was evaluated via Trans-Epithelial Electrical Resistance (TEER) measurements in Caco-2 monolayers. Cell death was determined through flow cytometry analysis, and the protective effects of cell death inhibitors and antioxidant scavengers were investigated on ricin-intoxicated cells. Ricin showed high cytotoxicity on HT29 and Caco-2 cells, with EC₅₀ values in the nM range after 24–72 h of intoxication. Moreover, ricin strongly reduced TEER values in Caco-2 cells at 0.1–1 nM after 24 h of treatment. At a 1 nM concentration, ricin cytotoxicity can be significantly prevented by pre-incubating cells with the cell death inhibitors Z-VAD or necrostatin-1 and the antioxidant scavenger catalase, butylated hydroxyanisole or sodium pyruvate, demonstrating the involvement of apoptosis/necroptosis and oxidative stress in ricin cell death pathways and mechanisms. Full article

Ricin is a highly potent toxin that causes severe lung injury upon inhalation by initiating a complex cascade of cellular responses that ultimately leads to cell death. The low-density lipoprotein receptor-related protein 1 (LRP1) is a multifunctional receptor involved in various physiological processes, including ricin-mediated toxicity. This study explores the role of LRP1 shedding in the development of ricin-induced lung injury. Analysis of bronchoalveolar lavage fluid (BALF) from ricin-intoxicated mice and swine showed a significant increase in soluble LRP1 (sLRP1) levels, whereas serum LRP1 levels remained largely unchanged, suggesting the lungs are the primary source of sLRP1 release. In vitro assays demonstrated the formation of ricin-sLRP1 complexes, indicating that sLRP1 in BALF retained ricin-binding capability. Flow cytometric analysis of lung cells revealed a reduction in both the percentage and total number of LRP1-expressing cells following ricin exposure. Further investigation of specific lung cell populations showed that alveolar epithelial type II (AT-II) cells, despite experiencing significant injury, exhibited minimal LRP1 shedding. No shedding of LRP1 occurred in neutrophils. In contrast, fibroblasts, which were resistant to ricin-induced cell death, exhibited increased shedding of LRP1 and a corresponding decrease in membrane-bound LRP1 expression. This shedding of the LRP1 ectodomain was mediated by metalloproteinases. Immunohistochemical staining further confirmed decreased LRP1 expression in fibroblasts from ricin-exposed mice. Macrophages also showed substantial LRP1 shedding, despite undergoing significant depletion. These findings highlight the complex cell-specific nature of LRP1 shedding in response to ricin intoxication and suggests the potential role of LRP1 in modulation of cellular susceptibility and resistance to ricin-induced lung injury. Full article

The nature and epidemiology of plant intoxications are still not well understood, with recent data being limited. The present study aims to report cases of plant poisoning in the clinical practice of the Clinical Toxicology Department at the Naval Hospital—Varna, Bulgaria, over a 20-year period (2003–2023). A documentary retrospective analysis of the hospitalized cases of poisoning with poisonous plants and their grouping into toxidromes was performed. During the study period, patients with plant poisoning admitted to our hospital unit accounted for 0.35% of a total of 12,857 hospitalized individuals. The distribution across the toxidromes based on clinical presentation revealed the highest frequency of anticholinergic, cyanogen, and ricin toxidromes. The majority of the intoxications resulted from unintentional exposure to plant toxins in adult individuals. Most cases followed a mild to severe clinical course, with patient discharge occurring between 2 and 5 days. No fatalities were recorded, thanks to the reported treatment methods. A relatively low incidence of plant-related poisonings was observed, with their predominant manifestations affecting the gastrointestinal, nervous, and cardiovascular systems. Increased reporting of epidemiological data and clinical experiences in the management of plant intoxications would substantially enhance researchers’ understanding of them and facilitate the development of a standardized treatment protocol. Full article

Ricin is a highly potent toxin that has been used in various attempts at bioterrorism worldwide. Although a vaccine for preventing ricin poisoning (RiVax™) is in clinical development, there are currently no commercially available prophylaxis or treatments for ricin intoxication. Numerous studies have highlighted the potential of passive immunotherapy using anti-ricin monoclonal antibodies (mAbs) and have shown promising results in preclinical models. In this article, we describe the neutralizing and protective efficacy of a new generation of high-affinity anti-ricin mAbs, which bind and neutralize very efficiently both ricin isoforms D and E in vitro through cytotoxicity cell assays. In vivo, protection assay revealed that one of these mAbs (RicE5) conferred over 90% survival in a murine model challenged intranasally with a 5 LD₅₀ of ricin and treated by intravenous administration of the mAbs 6 h post-intoxication. Notably, a 35% survival rate was observed even when treatment was administered 24 h post-exposure. Moreover, all surviving mice exhibited long-term immunity to high ricin doses. These findings offer promising results for the clinical development of a therapeutic candidate against ricin intoxication and may also pave the way for novel vaccination strategies against ricin or other toxins. Full article

The plant-derived toxin ricin is classified as a type 2 ribosome-inactivating protein (RIP) and currently lacks effective clinical antidotes. The toxicity of ricin is mainly due to its ricin toxin A chain (RTA), which has become an important target for drug development. Previous studies have identified two essential binding pockets in the active site of RTA, but most existing inhibitors only target one of these pockets. In this study, we used computer-aided virtual screening to identify a compound called RSMI-29, which potentially interacts with both active pockets of RTA. We found that RSMI-29 can directly bind to RTA and effectively attenuate protein synthesis inhibition and rRNA depurination induced by RTA or ricin, thereby inhibiting their cytotoxic effects on cells in vitro. Moreover, RSMI-29 significantly reduced ricin-mediated damage to the liver, spleen, intestine, and lungs in mice, demonstrating its detoxification effect against ricin in vivo. RSMI-29 also exhibited excellent drug-like properties, featuring a typical structural moiety of known sulfonamides and barbiturates. These findings suggest that RSMI-29 is a novel small-molecule inhibitor that specifically targets ricin toxin A chain, providing a potential therapeutic option for ricin intoxication. Full article

Ricin, a highly potent plant-derived toxin, is considered a potential bioterrorism weapon due to its pronounced toxicity, high availability, and ease of preparation. Acute damage following pulmonary ricinosis is characterized by local cytokine storm, massive neutrophil infiltration, and edema formation, resulting in respiratory insufficiency and death. A designated equine polyclonal antibody-based (antitoxin) treatment was developed in our laboratory and proved efficacious in alleviating lung injury and increasing survival rates. Although short-term pathogenesis was thoroughly characterized in antitoxin-treated mice, the long-term damage in surviving mice was never determined. In this study, long-term consequences of ricin intoxication were evaluated 30 days post-exposure in mice that survived antitoxin treatment. Significant pulmonary sequelae were demonstrated in surviving antitoxin-treated mice, as reflected by prominent histopathological changes, moderate fibrosis, increased lung hyperpermeability, and decreased lung compliance. The presented data highlight, for the first time to our knowledge, the possibility of long-term damage development in mice that survived lethal-dose pulmonary exposure to ricin due to antitoxin treatment. Full article

The highly toxic plant toxin ricin is one of the most known threatening toxins. Accurate and sensitive biosensing methods for the first emergency response and intoxication treatment, are always pursued in the biodefense field. Screening affinity molecules is the fundamental mainstream approach for developing biosensing methods. Compared with common affinity molecules such as antibodies and oligonucleotide aptamers, peptides have great potential as biosensing modules with more accessible chemical synthesis capability and better batch-to-batch stability than antibodies, more abundant interaction sites, and robust sensing performance towards complex environments. However, anti-ricin peptides are so scant to be screened and discovered, and an advanced screening strategy is the utmost to tackle this issue. Here, we present a new in silico-in vitro iteration-assisted affinity maturation strategy of anti-ricin peptides. We first obtained affinity peptides targeting ricin through phage display with five panning rounds of “coating-elution-amplification-enrichment” procedures. The binding affinity and kinetic parameters characterized by surface plasmon resonance (SPR) showed that we had obtained four peptides owning dissociation constants (K_D) around 2~35 μM, in which peptide PD-2-R5 has the lower K_D of 4.7 μM and higher stable posture to interact with ricin. We then constructed a new strategy for affinity maturity, composing two rounds of in silico-in vitro iterations. Firstly, towards the single-site alanine scanning mutation peptide library, the molecular docking predictions match the SPR evaluation results well, laying a solid foundation for designing a full saturation mutated peptide library. Secondly, plenty of in silico saturation mutation prediction results guided the discovery of peptides PD2-R5-T3 and PD-2-R5-T4 with higher affinity from only a limited number of SPR evaluation experiments. Both evolved peptides had increased affinity by about 5~20 times, i.e., K_D of 230 nM and 900 nM. A primary cellular toxicity assay indicated that both peptides could protect cells against ricin damage. We further established an SPR assay based on PD-2-R5-T3 and PD-2-R5-T4 elongated with an antifouling peptide linkage and achieved good linearity with a sensitivity of 1 nM and 0.5 nM, respectively. We hope this new affinity-mature strategy will find its favorable position in relevant peptide evolution, biosensing, and medical countermeasures for biotoxins to protect society’s security and human life better. Full article

Autophagy is a complex and highly regulated degradative process, which acts as a survival pathway in response to cellular stress, starvation and pathogen infection. Ricin toxin is a plant toxin produced by the castor bean and classified as a category B biothreat agent. Ricin toxin inhibits cellular protein synthesis by catalytically inactivating ribosomes, leading to cell death. Currently, there is no licensed treatment for patients exposed to ricin. Ricin-induced apoptosis has been extensively studied; however, whether its intoxication via protein synthesis inhibition affects autophagy is not yet resolved. In this work, we demonstrated that ricin intoxication is accompanied by its own autophagic degradation in mammalian cells. Autophagy deficiency, by knocking down ATG5, attenuates ricin degradation, thus aggravating ricin-induced cytotoxicity. Additionally, the autophagy inducer SMER28 (Small Molecule Enhancer 28) partially protects cells against ricin cytotoxicity, an effect not observed in autophagy-deficient cells. These results demonstrate that autophagic degradation acts as a survival response of cells against ricin intoxication. This suggests that stimulation of autophagic degradation may be a strategy to counteract ricin intoxication. Full article

Ricin toxin is a disulfide-linked glycoprotein (AB toxin) comprising one enzymatic A chain (RTA) and one cell-binding B chain (RTB) contained in the castor bean, a Ricinus species. Ricin inhibits peptide chain elongation via disruption of the binding between elongation factors and ribosomes, resulting in apoptosis, inflammation, oxidative stress, and DNA damage, in addition to the classically known rRNA damage. Ricin has been used in traditional medicine throughout the world since prehistoric times. Because ricin toxin is highly toxic and can be readily extracted from beans, it could be used as a bioweapon (CDC B-list). Due to its extreme lethality and potential use as a biological weapon, ricin toxin remains a global public health concern requiring specific countermeasures. Currently, no specific treatment for ricin intoxication is available. This review focuses on the drugs under development. In particular, some examples are reviewed to demonstrate the proof of concept of antibody-based therapy. Chemical inhibitors, small proteins, and vaccines can serve as alternatives to antibodies or may be used in combination with antibodies. Full article

Abrin is a highly toxic protein obtained from the seeds of the rosary pea plant Abrus precatorius, and it is closely related to ricin in terms of its structure and chemical properties. Both toxins inhibit ribosomal function, halt protein synthesis and lead to cellular death. The major clinical manifestations following pulmonary exposure to these toxins consist of severe lung inflammation and consequent respiratory insufficiency. Despite the high similarity between abrin and ricin in terms of disease progression, the ability to protect mice against these toxins by postexposure antibody-mediated treatment differs significantly, with a markedly higher level of protection achieved against abrin intoxication. In this study, we conducted an in-depth comparison between the kinetics of in vivo abrin and ricin intoxication in a murine model. The data demonstrated differential binding of abrin and ricin to the parenchymal cells of the lungs. Accordingly, toxin-mediated injury to the nonhematopoietic compartment was shown to be markedly lower in the case of abrin intoxication. Thus, profiling of alveolar epithelial cells demonstrated that although toxin-induced damage was restricted to alveolar epithelial type II cells following abrin intoxication, as previously reported for ricin, it was less pronounced. Furthermore, unlike following ricin intoxication, no direct damage was detected in the lung endothelial cell population following abrin exposure. Reduced impairment of intercellular junction molecules following abrin intoxication was detected as well. In contrast, similar damage to the endothelial surface glycocalyx layer was observed for the two toxins. We assume that the reduced damage to the lung stroma, which maintains a higher level of tissue integrity following pulmonary exposure to abrin compared to ricin, contributes to the high efficiency of the anti-abrin antibody treatment at late time points after exposure. Full article

Criminal poisonings are among the least frequently detected crimes in the world. Lack of suspicion of this type of event by police officers and prosecutors, clinical symptoms imitating many somatic diseases and technical difficulties in diagnostics, as well as high research costs make the actual frequency of these events difficult to estimate. The substance used for criminal poisoning is often characterized by: lack of taste, color and smell, delayed action, easy availability and difficulty to detect. The aim of the study was to analyze selected cases of political poisoning that took place in the years 1978–2020, to describe the mechanisms of action of the substances used and to evaluate the diagnosis and treatment. The analyzed cases of criminal poisoning concerned: Georgi Markov (ricin), Khalid Maszal (fentanyl), Wiktor Yushchenko (TCDD dioxin), Jasir Arafat (polonium ²¹⁰Po isotope), Alexander Litvinenko (polonium ²¹⁰Po isotope), Kim Jong-Nam (VX), Sergei Skripal (Novichok) and Alexei Navalny (Novichok). Contemporary poisons, to a greater extent than in the past, are based on the use of synthetic substances from the group of organophosphorus compounds and radioactive substances. The possibility of taking appropriate and effective treatment in such cases is the result of many factors, including the possibility of quick and competent rescue intervention, quick and reliable detection of the toxic substance and the possibility of using an antidote. Full article

Ricin is a toxin which enters cells and depurinates an adenine base in the sarcin-ricin loop in the large ribosomal subunit, leading to the inhibition of protein translation and cell death. We postulated that this depurination event could be detected using Oxford Nanopore Technologies (ONT) direct RNA sequencing, detecting a change in charge in the ricin loop. In this study, A549 cells were exposed to ricin for 2–24 h in order to induce depurination. In addition, a novel software tool was developed termed RIPpore that could quantify the adenine modification of ribosomal RNA induced by ricin upon respiratory epithelial cells. We provided demonstrable evidence for the first time that this base change detected is specific to RIP activity using a neutralising antibody against ricin. We believe this represents the first detection of depurination in RNA achieved using ONT sequencers. Collectively, this work highlights the potential for ONT and direct RNA sequencing to detect and quantify depurination events caused by ribosome-inactivating proteins such as ricin. RIPpore could have utility in the evaluation of new treatments and/or in the diagnosis of exposure to ricin. Full article

The compound EACC (ethyl (2-(5-nitrothiophene-2-carboxamido) thiophene-3-carbonyl) carbamate) was recently reported to inhibit fusion of autophagosomes with lysosomes in a reversible manner by inhibiting recruitment of syntaxin 17 to autophagosomes. We report here that this compound also provides a strong protection against the protein toxin ricin as well as against other plant toxins such as abrin and modeccin. The protection did not seem to be caused by inhibition of endocytosis and retrograde transport, but rather by inhibited release of the enzymatically active A-moiety to the cytosol. The TANK-binding kinase 1 (TBK1) has been reported to phosphorylate syntaxin 17 and be required for initiation of autophagy. The inhibitor of TBK1, MRT68601, induced in itself a strong sensitization to ricin, apparently by increasing transport to the Golgi apparatus. Importantly, MRT68601 increased Golgi transport of ricin even in the presence of EACC, but EACC was still able to inhibit intoxication, supporting the idea that EACC protects at a late step along the retrograde pathway. These results also indicate that phosphorylation of syntaxin 17 is not required for the protection observed. Full article

Ricin toxin isolated from the castor bean (Ricinus communis) is one of the most potent and lethal molecules known. While the pathophysiology and clinical consequences of ricin poisoning by the parenteral route, i.e., intramuscular penetration, have been described recently in various animal models, the preceding mechanism underlying the clinical manifestations of systemic ricin poisoning has not been completely defined. Here, we show that following intramuscular administration, ricin bound preferentially to the vasculature in both mice and swine, leading to coagulopathy and widespread hemorrhages. Increased levels of circulating VEGF and decreased expression of vascular VE-cadherin caused blood vessel impairment, thereby promoting hyperpermeability in various organs. Elevated levels of soluble heparan sulfate, hyaluronic acid and syndecan-1 were measured in blood samples following ricin intoxication, indicating that the vascular glycocalyx of both mice and swine underwent extensive damage. Finally, by using side-stream dark field intravital microscopy imaging, we determined that ricin poisoning leads to microvasculature malfunctioning, as manifested by aberrant blood flow and a significant decrease in the number of diffused microvessels. These findings, which suggest that glycocalyx shedding and microcirculation dysfunction play a major role in the pathology of systemic ricin poisoning, may serve for the formulation of specifically tailored therapies for treating parenteral ricin intoxication. Full article