A human respiratory-tissue organ culture incorporating an air interface.

Jackson, A D; Rayner, Charlotte F. J.; Dewar, A.; Cole, Peter; Wilson, Robert

doi:10.1164/ajrccm.153.3.8630556

Cited by 48 publications

(35 citation statements)

References 18 publications

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“…Air interface respiratory organ cultures were constructed using equine upper respiratory tract tissues (nasal turbinate, guttural pouch, and trachea) and methods described for human (26) and canine (1) tissues with some modifications. Tissues were obtained from an abattoir and washed in Dulbecco's modified Eagle's medium (DMEM) containing antibiotics (penicillin, 100 U ml Ϫ1 ; streptomycin, 50 g ml Ϫ1 ; gentamicin, 100 g ml Ϫ1 ; amphotericin, 2.5 g ml Ϫ1 ) for 4 h to remove commensal flora.…”

Section: Methodsmentioning

confidence: 99%

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Mutation of the Maturase Lipoprotein Attenuates the Virulence ofStreptococcus equito a Greater Extent than Does Loss of General Lipoprotein Lipidation

et al. 2006

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Streptococcus equi is the causative agent of strangles, a prevalent and highly contagious disease of horses. Despite the animal suffering and economic burden associated with strangles, little is known about the molecular basis of S. equi virulence. Here we have investigated the contributions of a specific lipoprotein and the general lipoprotein processing pathway to the abilities of S. equi to colonize equine epithelial tissues in vitro and to cause disease in both a mouse model and the natural host in vivo. Colonization of air interface organ cultures after they were inoculated with a mutant strain deficient in the maturase lipoprotein (⌬prtM 138-213 , with a deletion of nucleotides 138 to 213) was significantly less than that for cultures infected with wild-type S. equi strain 4047 or a mutant strain that was unable to lipidate preprolipoproteins (⌬lgt 190-685 ). Moreover, mucus production was significantly greater in both wild-type-infected and ⌬lgt 190-685 -infected organ cultures. Both mutants were significantly attenuated compared with the wild-type strain in a mouse model of strangles, although 2 of 30 mice infected with the ⌬lgt 190-685 mutant did still exhibit signs of disease. In contrast, only the ⌬prtM 138-213 mutant was significantly attenuated in a pony infection study, with 0 of 5 infected ponies exhibiting pathological signs of strangles compared with 4 of 4 infected with the wild-type and 3 of 5 infected with the ⌬lgt 190-685 mutant. We believe that this is the first study to evaluate the contribution of lipoproteins to the virulence of a gram-positive pathogen in its natural host. These data suggest that the PrtM lipoprotein is a potential vaccine candidate, and further investigation of its activity and its substrate(s) are warranted.

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Section: Methodsmentioning

confidence: 99%

“…were assessed by morphometric analysis of scanning electron microscopy (SEM) images of the epithelial surface. The tissues were processed and surface morphometry was carried out using standard methods (26). The percentage of the epithelial surface covered with mucus was recorded.…”

Section: Methodsmentioning

confidence: 99%

Mutation of the Maturase Lipoprotein Attenuates the Virulence ofStreptococcus equito a Greater Extent than Does Loss of General Lipoprotein Lipidation

et al. 2006

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“…Each piece of tissue was then placed at an air-liquid interface as follows. 40 A 35-mm petri dish (without lid) was aseptically placed within a 55-cm petri dish, and a filter paper (Whatman no. 1; A1 Laboratory Supplies, London, UK) laid across the inner dish such that its ends reached to the outer container.…”

Section: Reagentsmentioning

confidence: 99%

The extra- and intracellular barriers to lipid and adenovirus-mediated pulmonary gene transfer in native sheep airway epithelium

Kitson¹,

Ángel²,

Judd³

et al. 1999

Gene Ther

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123

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Gene transfer to the respiratory epithelium is currently apical membrane represented a significant barrier to both suboptimal and may be helped by the identification of limitagents. Adenovirus-mediated expression could be signifiing biological barriers. We have, therefore, developed an cantly augmented by increasing contact time or by preex vivo model which retains many of the characteristics of treatment of tissues with a nominally calcium-free medium. in vivo native airways including mucociliary clearance,The presence of these extracellular and plasma membrane mucus coverage and an intact cellular structure. Using this barriers appeared to be the key parameters responsible for model we have demonstrated several barriers to gene the approximately three log difference in gene expression transfer. Liposome-mediated gene transfer was inhibited found in vitro compared with our ex vivo model. Cytoskeleby normal mucus, with removal of this layer increasing tal elements and the cell cycle also influenced in vitro gene expression approximately 25-fold. In addition both lipotransfer, and represent further barriers which need to be some and adenovirus were inhibited by CF sputum. The overcome.

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“…26 This consisted of a 35-mm Petri dish aseptically placed within a 55-mm Petri dish with a filter paper (Whatman No. 1, Maidstone, UK) laid across the inner dish with either end reaching the outer container.…”

Section: Ex Vivo Model Of Gene Transfermentioning

confidence: 99%