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Characterization of extended co-culture of non-typeable Haemophilus influenzae with primary human respiratory tissues.

Ren D, Nelson KL, Uchakin PN, Smith AL, Gu XX, Daines DA - Exp. Biol. Med. (Maywood) (2012)

Bottom Line: These infections often recur and can become chronic.Scanning and transmission electron microscopy of tissues confirmed that intact NTHi were persisting paracellularly, while organisms observed in intracellular vacuoles appeared degraded.Furthermore, the apical surface and tight junctions of the infected tissues were undisturbed, with high transepithelial electrical resistances, while the basal cell layer displayed more junctional disorganization and wider intercellular spaces than the uninfected control tissues.

View Article: PubMed Central - PubMed

Affiliation: Division of Basic Medical Sciences, Mercer University School of Medicine, 1550 College Street, Macon, GA 31207, USA.

ABSTRACT
Non-typeable Haemophilus influenzae (NTHi) are human-adapted Gram-negative bacteria that comprise part of the normal flora of the human upper airway, but are also responsible for a number of mucosal infections such as otitis media and bronchitis. These infections often recur and can become chronic. To characterize the effect of long-term co-culture of NTHi with human tissues, we infected primary respiratory epithelial cells grown at the air-liquid interface with three NTHi strains over a range of 1-10 days. Scanning and transmission electron microscopy of tissues confirmed that intact NTHi were persisting paracellularly, while organisms observed in intracellular vacuoles appeared degraded. Furthermore, the apical surface and tight junctions of the infected tissues were undisturbed, with high transepithelial electrical resistances, while the basal cell layer displayed more junctional disorganization and wider intercellular spaces than the uninfected control tissues. Although the tissues elaborated the cytokine profile reported for NTHi-caused otitis media in vivo, there was little change in the dynamics of cytokine secretion over the time points tested. Finally, we report that NTHi strains released outer membrane vesicles (OMVs) during extended co-culture with the tissues, and show that these OMVs directly interact with host cell membranes.

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Related in: MedlinePlus

Paired scanning electron microscopy (SEM) and transmission electron microscopy (TEM) images of EpiAirway tissues with or without NTHi co-culture. One EpiAirway insert was infected with strain R2866 for five days, then fixed and cut in half. One half was processed for SEM (a), the other for TEM (c). Another EpiAirway insert was not infected with NTHi and used as the control for SEM (b) and TEM (d). (a) SEM of infected EpiAirway tissue. (b) SEM of control EpiAirway tissue. (c) TEM of infected EpiAirway tissue. (d) TEM of control EpiAirway tissue. NTHi, non-typeable Haemophilus influenzae
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EBM-1111-RM-377F2: Paired scanning electron microscopy (SEM) and transmission electron microscopy (TEM) images of EpiAirway tissues with or without NTHi co-culture. One EpiAirway insert was infected with strain R2866 for five days, then fixed and cut in half. One half was processed for SEM (a), the other for TEM (c). Another EpiAirway insert was not infected with NTHi and used as the control for SEM (b) and TEM (d). (a) SEM of infected EpiAirway tissue. (b) SEM of control EpiAirway tissue. (c) TEM of infected EpiAirway tissue. (d) TEM of control EpiAirway tissue. NTHi, non-typeable Haemophilus influenzae

Mentions: To examine the effect of extended NTHi co-culture on EpiAirway tissues, SEM and TEM images were taken of inserts with and without infection (Figure 2). These SEM and TEM images are paired, as each EpiAirway tissue was divided, with one half processed for SEM and the other half processed for TEM. Figure 2a shows a representative SEM image of tissue infected with strain R2866 for five days, while Figure 2b shows a SEM of the uninfected control EpiAirway tissue. There was no apparent difference in the number of ciliated cells or the length and number of microvilli between the infected and control tissues. However, there appeared to be more goblet cells in the infected tissues than in the control, with 70 goblet cells observed in 10 random SEM images of infected tissues, versus 46 identified in the same number of random images of uninfected tissues (each at ×5000 magnification).Figure 2


Characterization of extended co-culture of non-typeable Haemophilus influenzae with primary human respiratory tissues.

Ren D, Nelson KL, Uchakin PN, Smith AL, Gu XX, Daines DA - Exp. Biol. Med. (Maywood) (2012)

Paired scanning electron microscopy (SEM) and transmission electron microscopy (TEM) images of EpiAirway tissues with or without NTHi co-culture. One EpiAirway insert was infected with strain R2866 for five days, then fixed and cut in half. One half was processed for SEM (a), the other for TEM (c). Another EpiAirway insert was not infected with NTHi and used as the control for SEM (b) and TEM (d). (a) SEM of infected EpiAirway tissue. (b) SEM of control EpiAirway tissue. (c) TEM of infected EpiAirway tissue. (d) TEM of control EpiAirway tissue. NTHi, non-typeable Haemophilus influenzae
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3369423&req=5

EBM-1111-RM-377F2: Paired scanning electron microscopy (SEM) and transmission electron microscopy (TEM) images of EpiAirway tissues with or without NTHi co-culture. One EpiAirway insert was infected with strain R2866 for five days, then fixed and cut in half. One half was processed for SEM (a), the other for TEM (c). Another EpiAirway insert was not infected with NTHi and used as the control for SEM (b) and TEM (d). (a) SEM of infected EpiAirway tissue. (b) SEM of control EpiAirway tissue. (c) TEM of infected EpiAirway tissue. (d) TEM of control EpiAirway tissue. NTHi, non-typeable Haemophilus influenzae
Mentions: To examine the effect of extended NTHi co-culture on EpiAirway tissues, SEM and TEM images were taken of inserts with and without infection (Figure 2). These SEM and TEM images are paired, as each EpiAirway tissue was divided, with one half processed for SEM and the other half processed for TEM. Figure 2a shows a representative SEM image of tissue infected with strain R2866 for five days, while Figure 2b shows a SEM of the uninfected control EpiAirway tissue. There was no apparent difference in the number of ciliated cells or the length and number of microvilli between the infected and control tissues. However, there appeared to be more goblet cells in the infected tissues than in the control, with 70 goblet cells observed in 10 random SEM images of infected tissues, versus 46 identified in the same number of random images of uninfected tissues (each at ×5000 magnification).Figure 2

Bottom Line: These infections often recur and can become chronic.Scanning and transmission electron microscopy of tissues confirmed that intact NTHi were persisting paracellularly, while organisms observed in intracellular vacuoles appeared degraded.Furthermore, the apical surface and tight junctions of the infected tissues were undisturbed, with high transepithelial electrical resistances, while the basal cell layer displayed more junctional disorganization and wider intercellular spaces than the uninfected control tissues.

View Article: PubMed Central - PubMed

Affiliation: Division of Basic Medical Sciences, Mercer University School of Medicine, 1550 College Street, Macon, GA 31207, USA.

ABSTRACT
Non-typeable Haemophilus influenzae (NTHi) are human-adapted Gram-negative bacteria that comprise part of the normal flora of the human upper airway, but are also responsible for a number of mucosal infections such as otitis media and bronchitis. These infections often recur and can become chronic. To characterize the effect of long-term co-culture of NTHi with human tissues, we infected primary respiratory epithelial cells grown at the air-liquid interface with three NTHi strains over a range of 1-10 days. Scanning and transmission electron microscopy of tissues confirmed that intact NTHi were persisting paracellularly, while organisms observed in intracellular vacuoles appeared degraded. Furthermore, the apical surface and tight junctions of the infected tissues were undisturbed, with high transepithelial electrical resistances, while the basal cell layer displayed more junctional disorganization and wider intercellular spaces than the uninfected control tissues. Although the tissues elaborated the cytokine profile reported for NTHi-caused otitis media in vivo, there was little change in the dynamics of cytokine secretion over the time points tested. Finally, we report that NTHi strains released outer membrane vesicles (OMVs) during extended co-culture with the tissues, and show that these OMVs directly interact with host cell membranes.

Show MeSH
Related in: MedlinePlus