Toxicity, distribution and elimination of the cancerostatic lectins abrin and ricin after parenteral injection into mice.

The survival time of mice after i.v. injection of the cancerostatic lectins, abrin and ricin was recorded. The LD50 dose was found to be 10-13 ng and 55-65 ng per mouse for abrin and ricin, respectively. Increasing amounts of toxin reduced the survival time, reaching a minimum of about 10 h. Lactose injected with ricin, provided partial protection against ricin, as measured by the survival time. Abrin and ricin labelled with 125I, and shown to retain their full toxic activity, were injected into mice. Most of the radioactivity found in the organs was present in the form of intact toxins, at least up to 5 h after injection. After i.v. injection the highest concentration/g tissue was found in spleen, followed by kidneys, heart, liver and thymus. The relative concentration in liver was considerably higher for ricin than for abrin. Similar results were found after i.p. injection. When lactose was administered together with ricin, almost 80% of the ricin injected was found in the liver after 30 min, compared to 48% without lactose, and the amount in other organs was concurrently reduced. The elimination of total radioactivity was much faster for ricin than abrin. The radioactivity found in the urine was largely present in non-trichloroacetic acid precipitable form, indicating that the toxins were extensively degraded before excretion

Endoplasmic Reticulum-Dependent Redox Reactions Control Endoplasmic Reticulum-Associated Degradation and Pathogen Entry

Abstract Significance: Protein misfolding within the endoplasmic reticulum (ER) is managed by an ER quality control system that retro-translocates aberrant proteins into the cytosol for proteasomal destruction. This process, known as ER-associated degradation, utilizes the action of ER redox enzymes to accommodate the disulfide-bonded nature of misfolded proteins. Strikingly, various pathogenic viruses and toxins co-opt these redox components to reach the cytosol during entry. These redox factors thus regulate critical cellular homeostasis and host?pathogen interactions. Recent Advances: Recent studies identify specific members of the protein disulfide isomerase (PDI) family, which use their chaperone and catalytic activities, in engaging both misfolded ER proteins and pathogens. Critical Issues: The precise molecular mechanism by which a dedicated PDI family member disrupts the disulfide bonds in the misfolded ER proteins and pathogens, as well as how they act to unfold these substrates to promote their ER-to-cytosol membrane transport, remain poorly characterized. Future Directions: How PDI family members distinguish folded versus misfolded ER substrates remains enigmatic. What physical characteristics surrounding a substrate's disulfide bond instruct PDI that it is mispaired or native? For the pathogens, as their disulfide bonds normally serve a critical role in providing physical support, what conformational changes experienced in the host enable their disulfide bonds to be disrupted? A combination of more rigorous biochemical and high-resolution structural studies should begin to address these questions. Antioxid. Redox Signal. 16, 809?818.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/98492/1/ars%2E2011%2E4425.pd

Clathrin- and Dynamin-Independent Endocytosis of FGFR3 – Implications for Signalling

Endocytosis of tyrosine kinase receptors can influence both the duration and the specificity of the signal emitted. We have investigated the mechanisms of internalization of fibroblast growth factor receptor 3 (FGFR3) and compared it to that of FGFR1 which is internalized predominantly through clathrin-mediated endocytosis. Interestingly, we observed that FGFR3 was internalized at a slower rate than FGFR1 indicating that it may use a different endocytic mechanism than FGFR1. Indeed, after depletion of cells for clathrin, internalization of FGFR3 was only partly inhibited while endocytosis of FGFR1 was almost completely abolished. Similarly, expression of dominant negative mutants of dynamin resulted in partial inhibition of the endocytosis of FGFR3 whereas internalization of FGFR1 was blocked. Interfering with proposed regulators of clathrin-independent endocytosis such as Arf6, flotillin 1 and 2 and Cdc42 did not affect the endocytosis of FGFR1 or FGFR3. Furthermore, depletion of clathrin decreased the degradation of FGFR1 resulting in sustained signalling. In the case of FGFR3, both the degradation and the signalling were only slightly affected by clathrin depletion. The data indicate that clathrin-mediated endocytosis is required for efficient internalization and downregulation of FGFR1 while FGFR3, however, is internalized by both clathrin-dependent and clathrin-independent mechanisms

Ricin Toxicokinetics and Its Sensitive Detection in Mouse Sera or Feces Using Immuno-PCR

Ricin (also called RCA-II or RCA(60)), one of the most potent toxins and documented bioweapons, is derived from castor beans of Ricinus communis. Several in vitro methods have been designed for ricin detection in complex food matrices in the event of intentional contamination. Recently, a novel Immuno-PCR (IPCR) assay was developed with a limit of detection of 10 fg/ml in a buffer matrix and about 10-1000-fold greater sensitivity than other methods in various food matrices.In order to devise a better diagnostic test for ricin, the IPCR assay was adapted for the detection of ricin in biological samples collected from mice after intoxication. The limit of detection in both mouse sera and feces was as low as 1 pg/ml. Using the mouse intravenous (iv) model for ricin intoxication, a biphasic half-life of ricin, with a rapid t(1/2)α of 4 min and a slower t(1/2)β of 86 min were observed. The molecular biodistribution time for ricin following oral ingestion was estimated using an antibody neutralization assay. Ricin was detected in the blood stream starting at approximately 6-7 h post- oral intoxication. Whole animal histopathological analysis was performed on mice treated orally or systemically with ricin. Severe lesions were observed in the pancreas, spleen and intestinal mesenteric lymph nodes, but no severe pathology in other major organs was observed.The determination of in vivo toxicokinetics and pathological effects of ricin following systemic and oral intoxication provide a better understanding of the etiology of intoxication and will help in the future design of more effective diagnostic and therapeutic methods