Limits...
Oxidative stress and reduced responsiveness of challenged circulating leukocytes following pulmonary instillation of metal-rich particulate matter in rats.

Erdely A, Antonini JM, Young SH, Kashon ML, Gu JK, Hulderman T, Salmen R, Meighan T, Roberts JR, Zeidler-Erdely PC - Part Fibre Toxicol (2014)

Bottom Line: In addition, mononuclear cells were isolated 24 h post-exposure to measure oxidative stress by flow cytometry and confocal microscopy.Instillation of welding fume reduced inflammatory protein production of circulating leukocytes when challenged with the secondary stimulus LPS.The effects were not related to transcription, but were observed in conjunction with oxidative stress.

View Article: PubMed Central - HTML - PubMed

Affiliation: Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown 26505, WV, USA. efi4@cdc.gov.

ABSTRACT
Welding fume is an exposure that consists of a mixture of metal-rich particulate matter with gases (ozone, carbon monoxide) and/or vapors (VOCs). Data suggests that welders are immune compromised. Given the inability of pulmonary leukocytes to properly respond to a secondary infection in animal models, the question arose whether the dysfunction persisted systemically. Our aim was to evaluate the circulating leukocyte population in terms of cellular activation, presence of oxidative stress, and functionality after a secondary challenge, following welding fume exposure. Rats were intratracheally instilled (ITI) with PBS or 2 mg of welding fume collected from a stainless steel weld. Rats were sacrificed 4 and 24 h post-exposure and whole blood was collected. Whole blood was used for cellular differential counts, RNA isolation with subsequent microarray and Ingenuity Pathway Analysis, and secondary stimulation with LPS utilizing TruCulture technology. In addition, mononuclear cells were isolated 24 h post-exposure to measure oxidative stress by flow cytometry and confocal microscopy. Welding fume exposure had rapid effects on the circulating leukocyte population as identified by relative mRNA expression changes. Instillation of welding fume reduced inflammatory protein production of circulating leukocytes when challenged with the secondary stimulus LPS. The effects were not related to transcription, but were observed in conjunction with oxidative stress. These findings support previous studies of an inadequate pulmonary immune response following a metal-rich exposure and extend those findings showing leukocyte dysfunction occurs systemically.

Show MeSH

Related in: MedlinePlus

Effect of MMA-SS welding fume instillation on mononuclear cell oxidative stress. (A) Mononuclear cells were collected 24 h after instillation of PBS or MMA-SS, stained with H2DCFDA, fixed, and visualized by confocal microscopy for oxidative stress. Representative PBS exposed (upper left panel) and MMA-SS welding fume welding fume group (bottom left panel). Right panels represent background levels of unstained cells exposed to PBS (upper panel) and MMA-SS (lower panel). Fluorescent intensity was shown on a pseudo-color scale where the highest intensity staining is shown as white and red, followed by yellow, green, blue, and dark blue in order of decreasing intensity of the staining. The scale bar represents 10 μm. (B) Mean fluorescent intensity of isolated mononuclear cells incubated with H2DCFDA measured by flow cytometry. *p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4151022&req=5

Figure 6: Effect of MMA-SS welding fume instillation on mononuclear cell oxidative stress. (A) Mononuclear cells were collected 24 h after instillation of PBS or MMA-SS, stained with H2DCFDA, fixed, and visualized by confocal microscopy for oxidative stress. Representative PBS exposed (upper left panel) and MMA-SS welding fume welding fume group (bottom left panel). Right panels represent background levels of unstained cells exposed to PBS (upper panel) and MMA-SS (lower panel). Fluorescent intensity was shown on a pseudo-color scale where the highest intensity staining is shown as white and red, followed by yellow, green, blue, and dark blue in order of decreasing intensity of the staining. The scale bar represents 10 μm. (B) Mean fluorescent intensity of isolated mononuclear cells incubated with H2DCFDA measured by flow cytometry. *p < 0.05.

Mentions: Since reduced protein production was observed for cells collected at 24 h after treatment, oxidative stress was evaluated at that time point from isolated mononuclear cells. Representative images showed a greater level of oxidative stress in mononuclear cells isolated from the MMA-SS welding fume group (Figure 6A, bottom left panel) compared to PBS (Figure 6A, upper left panel). Background levels were similar between treatments (Figure 6A, right panels). For a quantitative evaluation of oxidative stress, isolated mononuclear cells incubated with H2DCFDA showed a significant increase in mean fluorescent intensity in the MMA-SS group compared to sham (Figure 6B). The increased oxidative stress complemented the qualitative confocal microscopy observations.


Oxidative stress and reduced responsiveness of challenged circulating leukocytes following pulmonary instillation of metal-rich particulate matter in rats.

Erdely A, Antonini JM, Young SH, Kashon ML, Gu JK, Hulderman T, Salmen R, Meighan T, Roberts JR, Zeidler-Erdely PC - Part Fibre Toxicol (2014)

Effect of MMA-SS welding fume instillation on mononuclear cell oxidative stress. (A) Mononuclear cells were collected 24 h after instillation of PBS or MMA-SS, stained with H2DCFDA, fixed, and visualized by confocal microscopy for oxidative stress. Representative PBS exposed (upper left panel) and MMA-SS welding fume welding fume group (bottom left panel). Right panels represent background levels of unstained cells exposed to PBS (upper panel) and MMA-SS (lower panel). Fluorescent intensity was shown on a pseudo-color scale where the highest intensity staining is shown as white and red, followed by yellow, green, blue, and dark blue in order of decreasing intensity of the staining. The scale bar represents 10 μm. (B) Mean fluorescent intensity of isolated mononuclear cells incubated with H2DCFDA measured by flow cytometry. *p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4151022&req=5

Figure 6: Effect of MMA-SS welding fume instillation on mononuclear cell oxidative stress. (A) Mononuclear cells were collected 24 h after instillation of PBS or MMA-SS, stained with H2DCFDA, fixed, and visualized by confocal microscopy for oxidative stress. Representative PBS exposed (upper left panel) and MMA-SS welding fume welding fume group (bottom left panel). Right panels represent background levels of unstained cells exposed to PBS (upper panel) and MMA-SS (lower panel). Fluorescent intensity was shown on a pseudo-color scale where the highest intensity staining is shown as white and red, followed by yellow, green, blue, and dark blue in order of decreasing intensity of the staining. The scale bar represents 10 μm. (B) Mean fluorescent intensity of isolated mononuclear cells incubated with H2DCFDA measured by flow cytometry. *p < 0.05.
Mentions: Since reduced protein production was observed for cells collected at 24 h after treatment, oxidative stress was evaluated at that time point from isolated mononuclear cells. Representative images showed a greater level of oxidative stress in mononuclear cells isolated from the MMA-SS welding fume group (Figure 6A, bottom left panel) compared to PBS (Figure 6A, upper left panel). Background levels were similar between treatments (Figure 6A, right panels). For a quantitative evaluation of oxidative stress, isolated mononuclear cells incubated with H2DCFDA showed a significant increase in mean fluorescent intensity in the MMA-SS group compared to sham (Figure 6B). The increased oxidative stress complemented the qualitative confocal microscopy observations.

Bottom Line: In addition, mononuclear cells were isolated 24 h post-exposure to measure oxidative stress by flow cytometry and confocal microscopy.Instillation of welding fume reduced inflammatory protein production of circulating leukocytes when challenged with the secondary stimulus LPS.The effects were not related to transcription, but were observed in conjunction with oxidative stress.

View Article: PubMed Central - HTML - PubMed

Affiliation: Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Morgantown 26505, WV, USA. efi4@cdc.gov.

ABSTRACT
Welding fume is an exposure that consists of a mixture of metal-rich particulate matter with gases (ozone, carbon monoxide) and/or vapors (VOCs). Data suggests that welders are immune compromised. Given the inability of pulmonary leukocytes to properly respond to a secondary infection in animal models, the question arose whether the dysfunction persisted systemically. Our aim was to evaluate the circulating leukocyte population in terms of cellular activation, presence of oxidative stress, and functionality after a secondary challenge, following welding fume exposure. Rats were intratracheally instilled (ITI) with PBS or 2 mg of welding fume collected from a stainless steel weld. Rats were sacrificed 4 and 24 h post-exposure and whole blood was collected. Whole blood was used for cellular differential counts, RNA isolation with subsequent microarray and Ingenuity Pathway Analysis, and secondary stimulation with LPS utilizing TruCulture technology. In addition, mononuclear cells were isolated 24 h post-exposure to measure oxidative stress by flow cytometry and confocal microscopy. Welding fume exposure had rapid effects on the circulating leukocyte population as identified by relative mRNA expression changes. Instillation of welding fume reduced inflammatory protein production of circulating leukocytes when challenged with the secondary stimulus LPS. The effects were not related to transcription, but were observed in conjunction with oxidative stress. These findings support previous studies of an inadequate pulmonary immune response following a metal-rich exposure and extend those findings showing leukocyte dysfunction occurs systemically.

Show MeSH
Related in: MedlinePlus