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Alveolar Macrophages Isolated Directly From Human Cytomegalovirus (HCMV)-Seropositive Individuals Are Sites of HCMV Reactivation In Vivo.

Poole E, Juss JK, Krishna B, Herre J, Chilvers ER, Sinclair J - J. Infect. Dis. (2014)

Bottom Line: Human cytomegalovirus (HCMV) causes significant morbidity in the immunocompromised host.This has been used to support the concept that viral reactivation in HCMV carriers routinely occurs from such terminally differentiated myeloid cells in vivo.However, to date this has not been shown for in vivo-differentiated macrophages.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Cambridge, United Kingdom.

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Related in: MedlinePlus

Alveolar macrophages isolated from human cytomegalovirus (HCMV)-seropositive donors support HCMV production and viral spread. A, Alveolar macrophages were cocultured with indicator fibroblasts and stained for immediate early gene (IE) by immune fluorescence. Light microscopy (left) and fluorescence (right) image of fibroblasts are shown; the green nuclei indicate IE-positive cells (arrow). B and C, The number of IE-positive foci were quantitated (B), and supernatants collected after coculture were transferred onto fresh monolayers of fresh human foreskin fibroblasts, with the number of IE-positive cells quantitated 24 hours after transfer (C).
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JIU837F4: Alveolar macrophages isolated from human cytomegalovirus (HCMV)-seropositive donors support HCMV production and viral spread. A, Alveolar macrophages were cocultured with indicator fibroblasts and stained for immediate early gene (IE) by immune fluorescence. Light microscopy (left) and fluorescence (right) image of fibroblasts are shown; the green nuclei indicate IE-positive cells (arrow). B and C, The number of IE-positive foci were quantitated (B), and supernatants collected after coculture were transferred onto fresh monolayers of fresh human foreskin fibroblasts, with the number of IE-positive cells quantitated 24 hours after transfer (C).

Mentions: To determine whether alveolar macrophages are sites of reactivation of full virus production, they were cocultured on indicator fibroblasts. After 3 weeks of coculture, the fibroblasts were fixed and stained for detection of infection foci where IE was being expressed. Figure 4A shows that foci of infection were clearly identifiable as areas of swollen cells within the fibroblast monolayer. Figure 4B shows that less than one in a million alveolar macrophages from seropositive individuals are productive for viral spread in vitro. Finally, to demonstrate that virions were being released into the supernatant, the supernatants were collected and transferred onto fresh fibroblasts, and the numbers of IE-expressing cells were enumerated over time (Figure 4C).Figure 4.


Alveolar Macrophages Isolated Directly From Human Cytomegalovirus (HCMV)-Seropositive Individuals Are Sites of HCMV Reactivation In Vivo.

Poole E, Juss JK, Krishna B, Herre J, Chilvers ER, Sinclair J - J. Infect. Dis. (2014)

Alveolar macrophages isolated from human cytomegalovirus (HCMV)-seropositive donors support HCMV production and viral spread. A, Alveolar macrophages were cocultured with indicator fibroblasts and stained for immediate early gene (IE) by immune fluorescence. Light microscopy (left) and fluorescence (right) image of fibroblasts are shown; the green nuclei indicate IE-positive cells (arrow). B and C, The number of IE-positive foci were quantitated (B), and supernatants collected after coculture were transferred onto fresh monolayers of fresh human foreskin fibroblasts, with the number of IE-positive cells quantitated 24 hours after transfer (C).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4442624&req=5

JIU837F4: Alveolar macrophages isolated from human cytomegalovirus (HCMV)-seropositive donors support HCMV production and viral spread. A, Alveolar macrophages were cocultured with indicator fibroblasts and stained for immediate early gene (IE) by immune fluorescence. Light microscopy (left) and fluorescence (right) image of fibroblasts are shown; the green nuclei indicate IE-positive cells (arrow). B and C, The number of IE-positive foci were quantitated (B), and supernatants collected after coculture were transferred onto fresh monolayers of fresh human foreskin fibroblasts, with the number of IE-positive cells quantitated 24 hours after transfer (C).
Mentions: To determine whether alveolar macrophages are sites of reactivation of full virus production, they were cocultured on indicator fibroblasts. After 3 weeks of coculture, the fibroblasts were fixed and stained for detection of infection foci where IE was being expressed. Figure 4A shows that foci of infection were clearly identifiable as areas of swollen cells within the fibroblast monolayer. Figure 4B shows that less than one in a million alveolar macrophages from seropositive individuals are productive for viral spread in vitro. Finally, to demonstrate that virions were being released into the supernatant, the supernatants were collected and transferred onto fresh fibroblasts, and the numbers of IE-expressing cells were enumerated over time (Figure 4C).Figure 4.

Bottom Line: Human cytomegalovirus (HCMV) causes significant morbidity in the immunocompromised host.This has been used to support the concept that viral reactivation in HCMV carriers routinely occurs from such terminally differentiated myeloid cells in vivo.However, to date this has not been shown for in vivo-differentiated macrophages.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Cambridge, United Kingdom.

Show MeSH
Related in: MedlinePlus