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Biomedical subjects

C M Thorpe

Publications and source records attributed to C M Thorpe.

At least 19 recordsLinked to original sources
PubMed2004

Shiga toxins activate translational regulation pathways in intestinal epithelial cells.

Shiga toxins (Stxs) cause irreversible damage to eukaryotic ribosomes, yet cellular intoxication of intestinal epithelial cells (IECs) results in increased synthesis of selected proteins, notably cytokines. How mRNA translation is maintained in this circumstance is unclear. This study was designed to assess whether Stx-induced alterations in host signal transduction machinery permit translation despite protein synthesis inhibition. A key step of translation is recruitment of initiation machinery to the 5' mRNA cap. This event occurs in part via interaction of the 5' cap with the cap binding protein, eIF4E, whose activity is positively regulated by phosphorylation and negatively regulated by binding to the translational repressor 4E-BP1. Following Stx treatment of IECs, eIF4E phosphorylation was detected by Western blotting using phospho-specific antibodies. Treatment with the p38 inhibitor, SB202190, or either of the ERK1/2 inhibitors, PD98059 and U0126, partially blocked Stx1-induced eIF4E phosphorylation. The Mnk1 inhibitor, CGP57380, blocked both basal and Stx-induced eIF4E phosphorylation. Interestingly, pretreatment with CGP57380 did not alter basal protein synthesis, but diminished the ability of cells to maintain translation following Stx1 challenge. Stx1 also induced hyperphosphorylation of 4E-BP1 and phosphorylation of S6Kinase; both effects were blocked by rapamycin. These data are novel observations showing that Stxs regulate multiple signal transduction pathways controlling translation in host cells, and support a role for eIF4E phosphorylation in maintaining host cell translation despite ribosomal intoxication.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Evaluation of paediatric osteosarcomas by classic cytogenetic and CGH analyses.

Classic cytogenetic and comparative genomic hybridisation (CGH) data on osteosarcomas have been reported extensively in the literature. However, the number of paediatric osteosarcoma cases studied below the age of 14 years remains relatively small. This study reports four new cases of paediatric osteosarcoma in patients aged 3 to 13 years, evaluated by classic cytogenetics and CGH analyses. Clonal chromosomal alterations were detected in all the cases and included structural rearrangements at 1p11-13, 1q11, 4q27-33, 6p23-25, 6q16-25, 7p13-22, 7q11-36, 11p10-15, 11q23, 17p11.2-13, 21p11, and 21q11-22. The CGH analysis revealed recurrent gains at 1p, 4q, 17p, and 21q and losses at 3q and 16p. Five amplification sites were observed at 1q11-23, 6p21, 8q13, 8q21.3-24.2, and 17p. The data are discussed and compared with other cytogenetic reports in the literature.

Adolescent↗
PubMed2001

Spatial problem solving and hippocampal place cell firing in rats: control by an internal sense of direction carried across environments.

Rats learned to find the baited corner of a box surrounded by a curtain, regardless of whether they had a fixed or random point of entry (POE) through the curtain. On probe trials, rats used an internal direction sense carried from outside the curtain to solve the problem, and only used the visual cue inside the curtain if disoriented and denied access to a view of the room en route. Similar disorientation procedures were required to obtain cue control of hippocampal place fields. The results suggest that: (1) POE effects previously found in the water maze may be task-specific; (2) an undisrupted internal sense of direction carried from one environment to another may provide the preferred solution to spatial problems in the second environment, even when this second environment is a familiar one with stable visual cues; and (3) choice behaviour is sometimes, but not always, representative of the hippocampal representation of space.

Animals↗
PubMed2001

Case of lipoblastoma with two derivative chromosomes 8 containing homogeneously staining-like regions and a review of the literature: lipoblastoma and chromosome 8.

We report a case of a lipoblastoma in a 10-month-old girl in which the cytogenetic aberration showed a homogeneously staining-like region (hsr) within two derivative chromosomes 8. There was a loss of one normal copy of chromosome 8 and gain of two identical derivative chromosomes 8 with the karyotype designation 47,XX,psu idic(8)(pter-->q12 approximately 13::hsr::q12 approximately 13-->pter),+psu idic (8)(pter-->q12 approximately 13::hsr::q12 approximately 13-->pter). This is the first report of a chromosomal aberration of this type seen in lipoblastoma.

Chromosome Banding↗
PubMed2001

A two-platform task reveals a deficit in the ability of rats to return to the start location in the water maze.

The ability of rats to return to the start location was examined with a 4-arm radial water maze. The task required rats to find 2 hidden platforms in sequence. Rats were released from 1 of 3 arms and there was a platform located in the fourth arm. Once a rat found this platform, a 2nd platform was raised in another location, which was either the start location, for 1 group, or another fixed location, for a control group. Across 3 experiments, all rats learned the location of the 1st fixed platform in 80 to 120 trials. However, rats had difficulty finding a 2nd platform if it was at the start location. Control groups revealed that rats could learn 2 platform locations and that the difficulty in learning to return to the start location did not seem to be attributable to its aversive nature. In separate groups, exposure to the start location was increased by starting the rats from an initially stable platform. Rats still did not readily learn to return to the start location. The authors suggest that start location, when varied, cannot readily be used to define the location of a hidden platform.

Animals↗
PubMed2001

Shiga toxins induce, superinduce, and stabilize a variety of C-X-C chemokine mRNAs in intestinal epithelial cells, resulting in increased chemokine expression.

Exposure of humans to Shiga toxins (Stxs) is a risk factor for hemolytic-uremic syndrome (HUS). Because Stx-producing Escherichia coli (STEC) is a noninvasive enteric pathogen, the extent to which Stxs can cross the host intestinal epithelium may affect the risk of developing HUS. We have previously shown that Stxs can induce and superinduce IL-8 mRNA and protein in intestinal epithelial cells (IECs) in vitro via a ribotoxic stress response. We used cytokine expression arrays to determine the effect of Stx1 on various C-X-C chemokine genes in IECs. We observed that Stx1 induces multiple C-X-C chemokines at the mRNA level, including interleukin-8 (IL-8), GRO-alpha, GRO-beta, GRO-gamma, and ENA-78. Like that of IL-8, GRO-alpha and ENA-78 mRNAs are both induced and superinduced by Stx1. Furthermore, Stx1 induces both IL-8 and GRO-alpha protein in a dose-response fashion, despite an overall inhibition in host cell protein synthesis. Stx1 treatment stabilizes both IL-8 and GRO-alpha mRNA. We conclude that Stxs are able to increase mRNA and protein levels of multiple C-X-C chemokines in IECs, with increased mRNA stability at least one mechanism involved. We hypothesize that ribotoxic stress is a pathway by which Stxs can alter host signal transduction in IECs, resulting in the production of multiple chemokine mRNAs, leading to increased expression of specific proteins. Taken together, these data suggest that exposing IECs to Stxs may stimulate a proinflammatory response, resulting in influx of acute inflammatory cells and thus contributing to the intestinal tissue damage seen in STEC infection.

Chemokine CXCL1↗
PubMed2001

Shiga toxin translocation across intestinal epithelial cells is enhanced by neutrophil transmigration.

Shiga toxin-producing E. coli (STEC) is a food-borne pathogen that causes serious illness, including hemolytic-uremic syndrome (HUS). STEC colonizes the lower intestine and produces Shiga toxins (Stxs). Stxs appear to translocate across intestinal epithelia and affect sensitive endothelial cell beds at various sites. We have previously shown that Stxs cross polarized intestinal epithelial cells (IECs) via a transcellular route and remain biologically active. Since acute inflammatory infiltration of the gut and fecal leukocytes is seen in many STEC-infected patients and since polymorphonuclear leukocyte (PMN) transmigration across polarized IECs diminishes the IEC barrier function in vitro, we hypothesized that PMN transmigration may enhance Stx movement across IECs. We found that basolateral-to-apical transmigration of neutrophils significantly increased the movement of Stx1 and Stx2 across polarized T84 IECs in the opposite direction. The amount of Stx crossing the T84 barrier was proportional to the degree of neutrophil transmigration, and the increase in Stx translocation appears to be due to increases in paracellular permeability caused by migrating PMNs. STEC clinical isolates applied apically induced PMN transmigration across and interleukin-8 (IL-8) secretion from T84 cells. Of the 10 STEC strains tested, three STEC strains lacking eae and espB (eae- and espB-negative STEC strains) induced significantly more neutrophil transmigration and significantly greater IL-8 secretion than eae- and espB-positive STEC or enteropathogenic E. coli. This study suggests that STEC interaction with intestinal epithelia induces neutrophil recruitment to the intestinal lumen, resulting in neutrophil extravasation across IECs, and that during this process Stxs may pass in greater amounts into underlying tissues, thereby increasing the risk of HUS.

Biological Transport↗
PubMed1999

Shiga toxins stimulate secretion of interleukin-8 from intestinal epithelial cells.

In the 1980s, Shiga toxin (Stx)-producing Escherichia coli O157:H7 (STEC) was identified as a cause of hemorrhagic colitis in the United States and was found to be associated with hemolytic uremic syndrome (HUS), a microangiopathic hemolytic anemia characterized by thrombocytopenia and renal failure. The precise way that Stxs cause hemorrhagic colitis and HUS is unclear. Stxs have been thought to cause disease by killing or irreversibly harming sensitive cells through a nonspecific blockade of mRNA translation, eventually resulting in cytotoxicity by preventing synthesis of critical molecules needed to maintain cell integrity. Because STEC is noninvasive, we have been exploring the host-toxin response at the level of the gastrointestinal mucosa, where STEC infection begins. We have found that Stx is capable of interleukin-8 (IL-8) superinduction in a human colonic epithelial cell line. Despite a general blockade of mRNA translation, Stx treatment results in increased IL-8 mRNA as well as increased synthesis and secretion of IL-8 protein. Our data suggest that an active Stx A subunit is required for this activity. Ricin, which has the same enzymatic activity and trafficking pathway as Stx, has similar effects. Exploration of the effects of other protein synthesis inhibitors (cycloheximide, anisomycin) suggests a mechanism of gene regulation that is distinct from a general translational blockade. Use of the specific p38/RK inhibitor SB202190 showed that blocking of this pathway results in decreased Stx-mediated IL-8 secretion. Furthermore, Stxs induced mRNA of the primary response gene c-jun, which was subsequently partially blocked by SB202190. These data suggest a novel model of how Stxs contribute to disease, namely that Stxs may alter regulation of host cell processes in sensitive cells via activation of at least one member of the mitogen-activated protein kinase family in the p38/RK cascade and induction of c-jun mRNA. Stx-induced increases in chemokine synthesis from intestinal epithelial cells could be important in augmenting the host mucosal inflammatory response to STEC infection.

Anisomycin↗
PubMed1999

Shiga toxins 1 and 2 translocate differently across polarized intestinal epithelial cells.

Shiga toxin-producing Escherichia coli (STEC) is an important food-borne pathogen that causes hemolytic-uremic syndrome. Following ingestion, STEC cells colonize the intestine and produce Shiga toxins (Stx), which appear to translocate across the intestinal epithelium and subsequently reach sensitive endothelial cell beds. STEC cells produce one or both of two major toxins, Stx1 and Stx2. Stx2-producing STEC is more often associated with disease for reasons as yet undetermined. In this study, we used polarized intestinal epithelial cells grown on permeable filters as a model to compare Stx1 and Stx2 movement across the intestinal epithelium. We have previously shown that biologically active Stx1 is able to translocate across cell monolayers in an energy-dependent, saturable manner. This study demonstrates that biologically active Stx2 is also capable of movement across the epithelium without affecting barrier function, but significantly less Stx2 crossed monolayers than Stx1. Chilling the monolayers to 4 degrees C reduced the amount of Stx1 and Stx2 movement by 200-fold and 20-fold respectively. Stx1 movement was clearly directional, favoring an apical-to-basolateral translocation, whereas Stx2 movement was not. Colchicine reduced Stx1, but not Stx2, translocation. Monensin reduced the translocation of both toxins, but the effect was more pronounced with Stx1. Brefeldin A had no effect on either toxin. Excess unlabeled Stx1 blocks the movement of (125)I-Stx1. Excess Stx2 failed to have any effect on Stx1 movement. Our data suggests that, despite the many common physical and biochemical properties of the two toxins, they appear to be crossing the epithelial cell barrier by different pathways.

Animals↗
PubMed1997

Clinical evidence for delayed chloroform poisoning.

From its introduction in 1847, chloroform proved to be a potent anaesthetic agent and over the next 50 yr its use became widespread. However, in 1912 the Committee on Anaesthesia of the American Medical Association stated that they were concerned with the occurrence of delayed chloroform poisoning in a number of cases. This conclusion was based on case reports and experimental animal data. However, subsequent studies and reported series of chloroform anaesthesia in humans have suggested a lower incidence of clinically significant liver injury. In this article we have investigated this discrepancy by analysing the published clinical data relating chloroform anaesthesia to liver damage.

Anesthesia, Inhalation↗
PubMed1997

Apparatus for emergency transtracheal ventilation.

Twenty anaesthetists were asked what equipment they would use to ventilate a patient after having performed an emergency cricothyroid puncture. Six systems were described and these were assessed for efficiency in delivery of oxygen through a 14 gauge cannula. Delivery of oxygen depended on the pressure achieved within the system. Apparatus utilizing a Bain circuit achieved volumes of around 200 ml for each double-handed squeeze of the reservoir bag. Use of a system taught on the Advanced Trauma and Life Support (ATLS) course resulted in higher system pressures and consequently greater volumes of oxygen delivered. Use of the oxygen flush with this system provides the highest flow rate and system pressure which results in 628 ml being delivered for a one second compression.

Anesthesiology↗