In Vivo Targets of Pasteurella Multocida Toxin

Many Pasteurella multocida strains are carried as commensals, while some cause disease in animals and humans. Some type D strains cause atrophic rhinitis in pigs, where the causative agent is known to be the Pasteurella multocida toxin (PMT). PMT activates three families of G-proteins—Gq/11, G12/13, and Gi/o—leading to cellular mitogenesis and other sequelae. The effects of PMT on whole animals in vivo have been investigated previously, but only at the level of organ-specific pathogenesis. We report here the first study to screen all the organs targeted by the toxin by using the QE antibody that recognizes only PMT-modified G-proteins. Under our experimental conditions, short-term treatment of PMT is shown to have multiple in vivo targets, demonstrating G-alpha protein modification, stimulation of proliferation markers and expression of active β-catenin in a tissue- and cell-specific manner. This highlights the usefulness of PMT as an important tool for dissecting the specific roles of different G-alpha proteins in vivo.

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Involvement of Osteocytes in the Action of Pasteurella multocida Toxin

Toxins ◽

10.3390/toxins10080328 ◽

2018 ◽

Vol 10 (8) ◽

pp. 328 ◽

Cited By ~ 1

Author(s):

Hannah Heni ◽

Julia Ebner ◽

Gudula Schmidt ◽

Klaus Aktories ◽

Joachim Orth

Keyword(s):

G Proteins ◽

Pasteurella Multocida ◽

Fiber Formation ◽

Atrophic Rhinitis ◽

Target Cells ◽

Heterotrimeric G Proteins ◽

Culture Model ◽

Tnf Α ◽

Pasteurella Multocida Toxin ◽

Progressive Atrophic Rhinitis

Pasteurella multocida toxin (PMT) causes progressive atrophic rhinitis with severe turbinate bone degradation in pigs. It has been reported that the toxin deamidates and activates heterotrimeric G proteins, resulting in increased differentiation of osteoclasts and blockade of osteoblast differentiation. So far, the action of PMT on osteocytes, which is the most abundant cell type in bone tissue, is not known. In MLO-Y4 osteocytes, PMT deamidated heterotrimeric G proteins, resulting in loss of osteocyte dendritic processes, stress fiber formation, cell spreading and activation of RhoC but not of RhoA. Moreover, the toxin caused processing of membrane-bound receptor activator of NF-κB ligand (RANKL) to release soluble RANKL and enhanced the secretion of osteoclastogenic TNF-α. In a co-culture model of osteocytes and bone marrow cells, PMT-induced osteoclastogenesis was largely increased as compared to the mono-culture model. The enhancement of osteoclastogenesis observed in the co-culture was blocked by sequestering RANKL with osteoprotegerin and by an antibody against TNF-α indicating involvement of release of the osteoclastogenic factors from osteocytes. Data support the crucial role of osteocytes in bone metabolism and osteoclastogenesis and identify osteocytes as important target cells of PMT in progressive atrophic rhinitis.

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Evaluation of cytotoxicity and mutagenicity of the benzodiazepine flunitrazepam in vitro and in vivo

Brazilian Journal of Pharmaceutical Sciences ◽

10.1590/s1984-82502014000200003 ◽

2014 ◽

Vol 50 (2) ◽

pp. 251-256

Author(s):

Igor Vivian de Almeida ◽

Giovana Domingues ◽

Lilian Capelari Soares ◽

Elisângela Düsman ◽

Veronica Elisa Pimenta Vicentini

Keyword(s):

Rattus Norvegicus ◽

Bone Marrow Cells ◽

Micronucleus Test ◽

Term Treatment ◽

Genotoxic Effects ◽

Short Term ◽

Short Term Treatment ◽

Chromosomal Aberration Test

Flunitrazepam (FNZ) is a sedative benzodiazepine prescribed for the short-term treatment of insomnia. However, there are concerns regarding possible carcinogenic or genotoxic effects of this medicine. Thus, the aim of this study was to evaluate the cytotoxic, clastogenic and aneugenic effects of FNZ in hepatoma cells from Rattus norvegicus (HTC) in vitro and in bone marrow cells of Wistar rats in vivo. These effects were examined in vitro following treatment with 0.2, 1.0, 5.0 or 10 μg/mL FNZ using a micronucleus test with a cytokinesis block or in vivo using a chromosomal aberration test following treatment with 7, 15 or 30 μg/mL/kg body weight. The results showed that the benzodiazepine concentrations tested were not cytotoxic, aneugenic or clastogenic. However, considering the adverse effects of using this benzodiazepine, more studies are required.

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Ca2+ signalling in K562 human erythroleukaemia cells: effect of dimethyl sulphoxide and role of G-proteins in thrombin- and thromboxane A2-activated pathways

Biochemical Journal ◽

10.1042/bj3120151 ◽

1995 ◽

Vol 312 (1) ◽

pp. 151-158 ◽

Cited By ~ 10

Author(s):

C P Thomas ◽

M J Dunn ◽

R Mattera

Keyword(s):

G Proteins ◽

Pcr Amplification ◽

Thromboxane A2 ◽

K562 Cells ◽

Protein S ◽

Term Treatment ◽

Dependent Manner ◽

Short Term Treatment ◽

Concentration Dependent Manner ◽

Prostaglandin F2 Alpha

The human leukaemic cell line K562 is a pluripotent stem cell with the potential to mature along a megakaryocytic or erythroid line. In these cells, thrombin and U46619 (9,11-dideoxy-9 alpha, 11 alpha-methanoepoxy prostaglandin F2 alpha), a thromboxane A2 analogue, increased intracellular Ca2+ in a rapid and concentration-dependent manner. The peak transient observed with both thrombin and U46619 was preserved upon stimulation in the absence of extracellular calcium and blunted with phorbol myristate acetate, suggestive of activation of phospholipase C. Short-term treatment with leupeptin abolished the calcium response to thrombin, but did not alter that to U46619. Both pertussis toxin (PT) and DMSO pretreatment inhibited thrombin- but not U46619-stimulated intracellular calcium elevation, indicating that these agonists signal through different G-proteins. Western blot analysis of crude membranes from K562 cells revealed the presence of G12 alpha and G13 alpha; the other known PT-substrates, Gi1 alpha and G0 alpha, were not detected. Consistent with this observation, ADP-ribosylation experiments revealed the presence of two PT substrates which co-migrated with human erythrocyte G12 alpha and G13 alpha. An antibody raised against Gq/11 alpha, a subfamily of G-protein alpha subunits unmodified by PT, specifically recognized 42 kDa protein(s) in K562 cells. PCR amplification of reverse-transcribed K562 RNA followed by DNA sequencing showed that these cells express messages for both Gq alpha and G11 alpha. Treatment of K562 cells with DMSO reduced the levels of thrombin receptor mRNA, without simultaneous changes in the expression of G12 alpha and G13 alpha. We have thus identified Ca(2+)-mobilizing agonists and related G-proteins in K562 cells, together with changes induced by DMSO in this signalling pathway.

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Vasoactive intestinal peptide prevents PKCε-induced intestinal epithelial barrier disruption during EPEC infection

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00195.2014 ◽

2015 ◽

Vol 308 (5) ◽

pp. G389-G402 ◽

Cited By ~ 6

Author(s):

V. Morampudi ◽

V. S. Conlin ◽

U. Dalwadi ◽

X. Wu ◽

K. C. Marshall ◽

...

Keyword(s):

Vasoactive Intestinal Peptide ◽

Critical Role ◽

Term Treatment ◽

Epithelial Barrier ◽

Protective Effects ◽

Short Term Treatment ◽

Barrier Integrity ◽

Barrier Disruption ◽

Long Term Treatment

We previously showed that vasoactive intestinal peptide (VIP) protects against bacterial pathogen-induced epithelial barrier disruption and colitis, although the mechanisms remain poorly defined. The aim of the current study was to identify cellular pathways of VIP-mediated protection with use of pharmacological inhibitors during enteropathogenic Escherichia coli (EPEC) infection of Caco-2 cell monolayers and during Citrobacter rodentium-induced colitis. EPEC-induced epithelial barrier disruption involved the PKC pathway but was independent of functional cAMP, Rho, and NF-κB pathways. VIP mediated its protective effects by inhibiting EPEC-induced PKC activity and increasing expression of the junctional protein claudin-4. Short-term treatment with TPA, which is known to activate PKC, was inhibited by VIP pretreatment, while PKC degradation via long-term treatment with TPA mimicked the protective actions of VIP. Immunostaining for specific PKC isotypes showed upregulated expression of PKCθ and PKCε during EPEC infection. Treatment with specific inhibitors revealed a critical role for PKCε in EPEC-induced barrier disruption. Furthermore, activation of PKCε and loss of barrier integrity correlated with claudin-4 degradation. In contrast, inhibition of PKCε by VIP pretreatment or the PKCε inhibitor maintained membrane-bound claudin-4 levels, along with barrier function. Finally, in vivo treatment with the PKCε inhibitor protected mice from C. rodentium-induced colitis. In conclusion, EPEC infection increases intracellular PKCε levels, leading to decreased claudin-4 levels and compromising epithelial barrier integrity. VIP inhibits PKCε activation, thereby attenuating EPEC-induced barrier disruption.

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Swine Atrophic Rhinitis Caused by Pasteurella multocida Toxin and Bordetella Dermonecrotic Toxin

Current Topics in Microbiology and Immunology - Pasteurella multocida ◽

10.1007/82_2012_206 ◽

2012 ◽

pp. 113-129 ◽

Cited By ~ 5

Author(s):

Yasuhiko Horiguchi

Keyword(s):

Pasteurella Multocida ◽

Atrophic Rhinitis ◽

Dermonecrotic Toxin ◽

Pasteurella Multocida Toxin

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Eignung des Tiergesundheitsdienstprogramms zur Überwachung und Bekämpfung der progressiven Rhinitis atrophicans

Tierärztliche Praxis Ausgabe G Großtiere / Nutztiere ◽

10.1055/s-0038-1623828 ◽

2009 ◽

Vol 37 (06) ◽

pp. 379-385

Author(s):

C. Lang ◽

M. Huber ◽

A. Griessler ◽

M. F. De Jong ◽

M. Schuh ◽

...

Keyword(s):

Pasteurella Multocida ◽

Atrophic Rhinitis ◽

Klinische Relevanz ◽

Pasteurella Multocida Toxin

Zusammenfassung Gegenstand und Ziel: Im Zeitraum von 1999 bis 2005 erfolgte eine Statuserhebung über das Vorkommen der progressiven Rhinitis atrophicans (PAR) in 17 oberösterreichischen Schweinevermehrerbetrieben, um in weiterer Folge eine PAR-Freiheit und Zertifizierung zu erreichen und somit die Eignung des Tiergesundheitsdienstprogramms für diesen Zweck zu überprüfen. Material und Methoden: Entsprechend dem Tiergesundheitsdienstprogramm wurden 11 045 Nasen- und Tonsillartupferproben entnommen und mittels PCR auf Pasteurella-multocida- Toxin (PMT) untersucht. Ein Jahr nach Einstellung der ART-(Atrophic- rhinitis-toxoid-)Vakzinierung wurden 2004 und 2005 zusätzlich 179 Blutproben mittels ELISA auf Antikörper gegen PMT untersucht. Ergebnisse: 84 (0,76%) der Nasen- und Tonsillartupferproben waren PMT-positiv. Bei 43 (24,02%) Seren ergab sich ein positiver serologischer Befund, es lagen jedoch keine klinischen Symptome einer PAR vor. Bereits im Jahr 2003 war in 14 Betrieben PMT in Nasen- und Tonsillartupferproben nicht mehr nachweisbar. In 16 Betrieben gelang bis dato die Eradikation des Erregers, lediglich ein Betrieb musste aufgrund der Nichteinhaltung der Vorschriften auf den Status PAR-frei“ verzichten. Schlussfolgerung: Die Studie belegt, dass sich das empfohlene Programm, das auf einem Test-and-Removal-“(T&R-)Prinzip basiert, zur stufenweisen Bekämpfung der PAR ohne komplette Bestandsmerzung eignet. Klinische Relevanz: Das Tiergesundheitsdienstprogramm zur Überwachung und Bekämpfung der progressiven Rhinitis atrophicans konnte im Laufe dieser Studie etabliert werden und wird mittlerweile österreichweit durchgeführt.

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Pasteurella multocida toxin activates Gβγ dimers of heterotrimeric G proteins

Cellular Signalling ◽

10.1016/j.cellsig.2008.12.007 ◽

2009 ◽

Vol 21 (4) ◽

pp. 551-558 ◽

Cited By ~ 24

Author(s):

Inga Preuß ◽

Barbara Kurig ◽

Bernd Nürnberg ◽

Joachim H.C. Orth ◽

Klaus Aktories

Keyword(s):

G Proteins ◽

Pasteurella Multocida ◽

Heterotrimeric G Proteins ◽

Pasteurella Multocida Toxin

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Short-term treatment with nitrate is not sufficient to induce in vivo antithrombotic effects in rats and mice

Naunyn-Schmiedeberg s Archives of Pharmacology ◽

10.1007/s00210-016-1308-5 ◽

2016 ◽

Vol 390 (1) ◽

pp. 85-94 ◽

Cited By ~ 2

Author(s):

K. Kramkowski ◽

A. Leszczynska ◽

K. Przyborowski ◽

B. Proniewski ◽

N. Marcinczyk ◽

...

Keyword(s):

Term Treatment ◽

Short Term ◽

Short Term Treatment ◽

Antithrombotic Effects ◽

Rats And Mice

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Melatonin improves the efficiency of super-ovulation and timed artificial insemination in sheep

10.7287/peerj.preprints.27598 ◽

2019 ◽

Author(s):

Yukun Song ◽

Hao Wu ◽

Xuguang Wang ◽

Aerman Hairy ◽

Xiaosheng Zhang ◽

...

Keyword(s):

Term Treatment ◽

Control Group ◽

Seasonal Reproduction ◽

Short Term ◽

Short Term Treatment ◽

Melatonin Treatment ◽

Timed Artificial Insemination ◽

Hu Sheep

It has been well proved that melatonin participates in the regulation of the seasonal reproduction of ewes. However, the effects of short term treatment of melatonin on ewe’s ovulation are still to be clarified. In this study, the effects of melatonin on the number of embryo s harvested from superovulation, and the pregnant rate in recipients after embryo transferred have been investigated. Hu sheep with synchronous estrus treatment were given melatonin subcutaneously injection (0, 5, and 10 mg/ewe, respectively). It was found that t he estrogen level in the group of 5 mg melatonin was significantly higher than that of other two groups a t the time of sperm insemination ( p < 0.05). The pregnant rate and number of lambs in the group of 5 mg melatonin treatment was also significantly higher than that of the rests of the groups ( P < 0.05). In another study, 31 Suffolk ewes as donors and 103 small-tailed han sheep ewes as recipients were used to produce pronuclear embryo and embryo transfer. Melatonin (5 mg) was given to the donors during estrus. The results showed that, the number of pronuclear embryos and the pregnancy rate were also significantly higher in melatonin group than that in the control group . In addition, 28 donors and 44 recipient ewes were used to produce morula/blastocyst and embryo transferring. Melatonin (5 mg) was given during estrus. The total number of embryos harvested ( 7.40±1.25/ewe vs. 3.96±0.73/ewe, P<0.05 ) and the pregnant rate (72.3±4.6% vs.54.7±4.0%, P<0.05) and number of lambs were also increased in melatonin group compared to the control group. Collectively, the results have suggested that melatonin treatment 36 hours after CIDR withdrawal could promote the number and quality of embryos in the in vivo condition and increased the pregnant rate and number of lambs.

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