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Growth of human bronchial epithelial cells at an air-liquid interface alters the response to particle exposure.

Ghio AJ, Dailey LA, Soukup JM, Stonehuerner J, Richards JH, Devlin RB - Part Fibre Toxicol (2013)

Bottom Line: Subsequently, it was not possible to attribute the observed decreases in the response of NHBE cells to differentiation alone since BEAS-2B cells, which do not differentiate, showed similar changes when grown at ALI.We conclude that growth of NHBE cells at ALI is associated with a diminished biological effect following particle exposure relative to cells submerged in media.This decreased response showed an association with increased oxygen availability.

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ABSTRACT

Background: We tested the hypothesis that normal human bronchial epithelial (NHBE) cells 1) grown submerged in media and 2) allowed to differentiate at air-liquid interface (ALI) demonstrate disparities in the response to particle exposure.

Results: Following exposure of submerged NHBE cells to ambient air pollution particle collected in Chapel Hill, NC, RNA for IL-8, IL-6, heme oxygenase 1 (HOX1) and cyclooxygenase 2 (COX2) increased. The same cells allowed to differentiate over 3, 10, and 21 days at ALI demonstrated no such changes following particle exposure. Similarly, BEAS-2B cells grown submerged in media demonstrated a significant increase in IL-8 and HOX1 RNA after exposure to NIST 1648 particle relative to the same cells exposed after growth at ALI. Subsequently, it was not possible to attribute the observed decreases in the response of NHBE cells to differentiation alone since BEAS-2B cells, which do not differentiate, showed similar changes when grown at ALI. With no exposure to particles, differentiation of NHBE cells at ALI over 3 to 21 days demonstrated significant decrements in baseline levels of RNA for the same proteins (i.e. IL-8, IL-6, HOX1, and COX2). With no exposure to particles, BEAS-2B cells grown at ALI showed comparable changes in RNA for IL-8 and HOX1. After the same particle exposure, NHBE cells grown at ALI on a transwell in 95% N2-5% CO2 and exposed to NIST 1648 particle demonstrated significantly greater changes in IL-8 and HOX1 relative to cells grown in 95% air-5% CO2.

Conclusions: We conclude that growth of NHBE cells at ALI is associated with a diminished biological effect following particle exposure relative to cells submerged in media. This decreased response showed an association with increased oxygen availability.

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Fold change RNA of NHBE cells grown in 95% air-5% CO2 (normoxia) and 95% N2-5% CO2 (hypoxia) for 24 hours and exposed to 250 μg NIST 1648 in 25 μL for 4 hr. The change in RNA for normoxic and hypoxic cells are provided relative to unexposed normoxic and hypoxic cells respectively. While IL-8 and HOX1 RNA was significantly elevated after particle exposure, fold change RNA after hypoxia was significantly increased relative to normoxia. *Significant difference relative to exposed normoxia.
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Figure 5: Fold change RNA of NHBE cells grown in 95% air-5% CO2 (normoxia) and 95% N2-5% CO2 (hypoxia) for 24 hours and exposed to 250 μg NIST 1648 in 25 μL for 4 hr. The change in RNA for normoxic and hypoxic cells are provided relative to unexposed normoxic and hypoxic cells respectively. While IL-8 and HOX1 RNA was significantly elevated after particle exposure, fold change RNA after hypoxia was significantly increased relative to normoxia. *Significant difference relative to exposed normoxia.

Mentions: Since BEAS-2B cells showed a pattern of response to PM similar to that of NHBE cells (i.e. diminished response to particles when cultured at ALI), it was not possible to attribute these disparities to differentiation alone. Subsequently, another factor was evaluated. Growth at ALI necessitates exposure to an environment which includes higher oxygen availability relative to cells grown submerged in media. To evaluate for an effect of oxygen concentration on response to PM, NHBE cells were grown on transwells in either normoxia or hypoxia for 24 hr and then exposed to 250 μg NIST 1648 in 25 μL. Those cells grown in hypoxia and exposed to the particle showed an approximate 6-fold increase in RNA for IL-8 while those in normoxia had a significantly diminished response (relative to unexposed cells grown in hypoxia and normoxia respectively) (Figure 5A). RNA for HOX1 demonstrated a comparable pattern of response to the particle with elevations in RNA for this protein following exposure in hypoxia and reduced values when the cells were exposed while in normoxia (Figure 5B). IL-8 protein release by cells in hypoxia and normoxia after exposure to the NIST 1648 particle corresponded to RNA changes with cell supernatant concentrations of 654 +/− 79 and 235 +/− 38 pg/mL respectively (with no exposure to PM, the cell release of IL-8 was 25 +/− 7 and 48 +/− 10 pg/mL respectively in hypoxic and normoxic cell supernatant). However, similar to differentiation, placement of the cells in normoxia changed the RNA for both IL-8 and HOX1 with both decreasing (Figures 6A and6B).


Growth of human bronchial epithelial cells at an air-liquid interface alters the response to particle exposure.

Ghio AJ, Dailey LA, Soukup JM, Stonehuerner J, Richards JH, Devlin RB - Part Fibre Toxicol (2013)

Fold change RNA of NHBE cells grown in 95% air-5% CO2 (normoxia) and 95% N2-5% CO2 (hypoxia) for 24 hours and exposed to 250 μg NIST 1648 in 25 μL for 4 hr. The change in RNA for normoxic and hypoxic cells are provided relative to unexposed normoxic and hypoxic cells respectively. While IL-8 and HOX1 RNA was significantly elevated after particle exposure, fold change RNA after hypoxia was significantly increased relative to normoxia. *Significant difference relative to exposed normoxia.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
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Figure 5: Fold change RNA of NHBE cells grown in 95% air-5% CO2 (normoxia) and 95% N2-5% CO2 (hypoxia) for 24 hours and exposed to 250 μg NIST 1648 in 25 μL for 4 hr. The change in RNA for normoxic and hypoxic cells are provided relative to unexposed normoxic and hypoxic cells respectively. While IL-8 and HOX1 RNA was significantly elevated after particle exposure, fold change RNA after hypoxia was significantly increased relative to normoxia. *Significant difference relative to exposed normoxia.
Mentions: Since BEAS-2B cells showed a pattern of response to PM similar to that of NHBE cells (i.e. diminished response to particles when cultured at ALI), it was not possible to attribute these disparities to differentiation alone. Subsequently, another factor was evaluated. Growth at ALI necessitates exposure to an environment which includes higher oxygen availability relative to cells grown submerged in media. To evaluate for an effect of oxygen concentration on response to PM, NHBE cells were grown on transwells in either normoxia or hypoxia for 24 hr and then exposed to 250 μg NIST 1648 in 25 μL. Those cells grown in hypoxia and exposed to the particle showed an approximate 6-fold increase in RNA for IL-8 while those in normoxia had a significantly diminished response (relative to unexposed cells grown in hypoxia and normoxia respectively) (Figure 5A). RNA for HOX1 demonstrated a comparable pattern of response to the particle with elevations in RNA for this protein following exposure in hypoxia and reduced values when the cells were exposed while in normoxia (Figure 5B). IL-8 protein release by cells in hypoxia and normoxia after exposure to the NIST 1648 particle corresponded to RNA changes with cell supernatant concentrations of 654 +/− 79 and 235 +/− 38 pg/mL respectively (with no exposure to PM, the cell release of IL-8 was 25 +/− 7 and 48 +/− 10 pg/mL respectively in hypoxic and normoxic cell supernatant). However, similar to differentiation, placement of the cells in normoxia changed the RNA for both IL-8 and HOX1 with both decreasing (Figures 6A and6B).

Bottom Line: Subsequently, it was not possible to attribute the observed decreases in the response of NHBE cells to differentiation alone since BEAS-2B cells, which do not differentiate, showed similar changes when grown at ALI.We conclude that growth of NHBE cells at ALI is associated with a diminished biological effect following particle exposure relative to cells submerged in media.This decreased response showed an association with increased oxygen availability.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Background: We tested the hypothesis that normal human bronchial epithelial (NHBE) cells 1) grown submerged in media and 2) allowed to differentiate at air-liquid interface (ALI) demonstrate disparities in the response to particle exposure.

Results: Following exposure of submerged NHBE cells to ambient air pollution particle collected in Chapel Hill, NC, RNA for IL-8, IL-6, heme oxygenase 1 (HOX1) and cyclooxygenase 2 (COX2) increased. The same cells allowed to differentiate over 3, 10, and 21 days at ALI demonstrated no such changes following particle exposure. Similarly, BEAS-2B cells grown submerged in media demonstrated a significant increase in IL-8 and HOX1 RNA after exposure to NIST 1648 particle relative to the same cells exposed after growth at ALI. Subsequently, it was not possible to attribute the observed decreases in the response of NHBE cells to differentiation alone since BEAS-2B cells, which do not differentiate, showed similar changes when grown at ALI. With no exposure to particles, differentiation of NHBE cells at ALI over 3 to 21 days demonstrated significant decrements in baseline levels of RNA for the same proteins (i.e. IL-8, IL-6, HOX1, and COX2). With no exposure to particles, BEAS-2B cells grown at ALI showed comparable changes in RNA for IL-8 and HOX1. After the same particle exposure, NHBE cells grown at ALI on a transwell in 95% N2-5% CO2 and exposed to NIST 1648 particle demonstrated significantly greater changes in IL-8 and HOX1 relative to cells grown in 95% air-5% CO2.

Conclusions: We conclude that growth of NHBE cells at ALI is associated with a diminished biological effect following particle exposure relative to cells submerged in media. This decreased response showed an association with increased oxygen availability.

Show MeSH
Related in: MedlinePlus