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Heme oxygenase-1 promotes granuloma development and protects against dissemination of mycobacteria.

Regev D, Surolia R, Karki S, Zolak J, Montes-Worboys A, Oliva O, Guroji P, Saini V, Steyn AJ, Agarwal A, Antony VB - Lab. Invest. (2012)

Bottom Line: Inhibition of HO by zinc protoporphyrin-IX led to inhibition of MCP-1 and increased expression of CCR2, its cognate receptor.Mycobacteria were found only inside defined granulomas but not outside granuloma in the lungs of HO-1⁺/⁺ mice.Higher MCP-1 levels were found in bronchoalveolar lavage fluid of M. avium infected HO-1(-/-) mice and CCR2 expression was higher in HO-1⁻/⁻ alveolar macrophages when compared with HO-1⁺/⁺ mice.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary, Critical Care and Sleep Medicine, College of Medicine, University of Florida, Gainesville, FL, USA.

ABSTRACT
Non-tuberculous mycobacterial (NTM) infections occur in both immunocompromised and immunocompetent hosts and are an increasingly recognized cause of morbidity and mortality. The hallmark of pulmonary mycobacterial infections is the formation of granuloma in the lung. Our study focuses on the role of heme oxygenase-1 (HO-1), a cytoprotective enzyme, in the regulation of granuloma development and maturation following infection with Mycobacterium avium. We examined the role of HO-1 in regulating monocyte chemoattractant protein-1 (MCP-1) and chemokine receptor 2 (CCR2), two molecules involved in monocyte-macrophage cell trafficking after infection. We showed that RAW 264.7 mouse monocytes exposed to M. avium expressed HO-1 and MCP-1. Inhibition of HO by zinc protoporphyrin-IX led to inhibition of MCP-1 and increased expression of CCR2, its cognate receptor. HO-1⁻/⁻ mice did not develop organized granuloma in their lungs, had higher lung colony forming unit of M. avium when infected with intratracheal M. avium, and had loose collections of inflammatory cells in the lung parenchyma. Mycobacteria were found only inside defined granulomas but not outside granuloma in the lungs of HO-1⁺/⁺ mice. In HO-1⁻/⁻ mice, mycobacteria were also found in the liver and spleen and showed increased mortality. Peripheral blood monocytes isolated from GFP⁺ mice and given intravenously to HO-1⁺/⁺ mice localized into tight granulomas, while in HO-1⁻/⁻ mice they remained diffusely scattered in areas of parenchymal inflammation. Higher MCP-1 levels were found in bronchoalveolar lavage fluid of M. avium infected HO-1(-/-) mice and CCR2 expression was higher in HO-1⁻/⁻ alveolar macrophages when compared with HO-1⁺/⁺ mice. CCR2 expression localized to granuloma in HO-1⁺/⁺ mice but not in the HO-1⁻/⁻ mice. These findings strongly suggest that HO-1 plays a protective role in the control of M. avium infection.

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(A) MCP-1 levels are up-regulated in BALF and serum samples of M. avium infected HO-1+/+ and HO-1-/- miceBALF and serum were collected and MCP-1 protein levels were measured by ELISA. MCP-1 levels were significantly increased in BALF of infected HO-1+/+ mice as compared to HO-1+/+ saline controls (*P <0.001) and HO-1-/- mice as compared to HO-1-/- saline controls (** P <0.001). MCP-1 levels are increased in mouse serum after 6 months M. avium infection and are significantly higher in infected HO-1+/+ as compared with saline controls (* P < 0.0001) and HO-1-/- mice (** P< .0001) as compared with saline controls. MCP-1 levels were significantly higher in M. avium infected HO-1-/- mice then in M. avium infected HO-1+/+ mice (*** P< 0.05) Samples from five mice in each group were assayed in triplicate. (B) CCR2 expression in PBMC and PAM of HO-1+/+ and HO-1-/- mice post M. avium infection. CCR2 receptor PAM in HO-1-/- mice with M. avium infection is significantly increased compared to M. avium infected HO-1+/+ mice. CCR2 expression in PBMC and PAM in HO-1-/- mice with M. avium infection were comparable. This is a representative histogram of three experiments with similar observation. Control mice received saline.
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Figure 7: (A) MCP-1 levels are up-regulated in BALF and serum samples of M. avium infected HO-1+/+ and HO-1-/- miceBALF and serum were collected and MCP-1 protein levels were measured by ELISA. MCP-1 levels were significantly increased in BALF of infected HO-1+/+ mice as compared to HO-1+/+ saline controls (*P <0.001) and HO-1-/- mice as compared to HO-1-/- saline controls (** P <0.001). MCP-1 levels are increased in mouse serum after 6 months M. avium infection and are significantly higher in infected HO-1+/+ as compared with saline controls (* P < 0.0001) and HO-1-/- mice (** P< .0001) as compared with saline controls. MCP-1 levels were significantly higher in M. avium infected HO-1-/- mice then in M. avium infected HO-1+/+ mice (*** P< 0.05) Samples from five mice in each group were assayed in triplicate. (B) CCR2 expression in PBMC and PAM of HO-1+/+ and HO-1-/- mice post M. avium infection. CCR2 receptor PAM in HO-1-/- mice with M. avium infection is significantly increased compared to M. avium infected HO-1+/+ mice. CCR2 expression in PBMC and PAM in HO-1-/- mice with M. avium infection were comparable. This is a representative histogram of three experiments with similar observation. Control mice received saline.

Mentions: Since MCP-1 is the soluble ligand for CCR2, we analyzed the MCP-1 levels in BALF and serum samples from the HO-1+/+ and HO-1-/- mice infected with M. avium or treated with saline. We observed a significant increase of MCP-1 levels in the BALF of infected HO-1+/+ mice as compared to non-infected controls, as well as increased levels in HO-1-/-M. avium infected mice as compared to HO-1-/- non-infected controls (Figure 7A). Additionally, MCP-1 levels were higher in HO-1-/- control mice as compared with HO-1+/+ control mice. Similar results were found when we analyzed the MCP-1 levels in the corresponding serum samples. HO-1-/- mice infected with M. avium showed higher levels of the cytokine as compared to HO-1+/+ samples (Figure 7A). Furthermore, the levels of MCP-1 in sera of HO-1-/- mice infected with M. avium were significantly higher than HO-1+/+ and non-infected HO-1-/- mice (Figure 7A). Although the MCP-1 profile was the same between BALF and serum samples for each condition, the amount of MCP-1 was up to 1000 fold higher in BALF than serum.


Heme oxygenase-1 promotes granuloma development and protects against dissemination of mycobacteria.

Regev D, Surolia R, Karki S, Zolak J, Montes-Worboys A, Oliva O, Guroji P, Saini V, Steyn AJ, Agarwal A, Antony VB - Lab. Invest. (2012)

(A) MCP-1 levels are up-regulated in BALF and serum samples of M. avium infected HO-1+/+ and HO-1-/- miceBALF and serum were collected and MCP-1 protein levels were measured by ELISA. MCP-1 levels were significantly increased in BALF of infected HO-1+/+ mice as compared to HO-1+/+ saline controls (*P <0.001) and HO-1-/- mice as compared to HO-1-/- saline controls (** P <0.001). MCP-1 levels are increased in mouse serum after 6 months M. avium infection and are significantly higher in infected HO-1+/+ as compared with saline controls (* P < 0.0001) and HO-1-/- mice (** P< .0001) as compared with saline controls. MCP-1 levels were significantly higher in M. avium infected HO-1-/- mice then in M. avium infected HO-1+/+ mice (*** P< 0.05) Samples from five mice in each group were assayed in triplicate. (B) CCR2 expression in PBMC and PAM of HO-1+/+ and HO-1-/- mice post M. avium infection. CCR2 receptor PAM in HO-1-/- mice with M. avium infection is significantly increased compared to M. avium infected HO-1+/+ mice. CCR2 expression in PBMC and PAM in HO-1-/- mice with M. avium infection were comparable. This is a representative histogram of three experiments with similar observation. Control mice received saline.
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Figure 7: (A) MCP-1 levels are up-regulated in BALF and serum samples of M. avium infected HO-1+/+ and HO-1-/- miceBALF and serum were collected and MCP-1 protein levels were measured by ELISA. MCP-1 levels were significantly increased in BALF of infected HO-1+/+ mice as compared to HO-1+/+ saline controls (*P <0.001) and HO-1-/- mice as compared to HO-1-/- saline controls (** P <0.001). MCP-1 levels are increased in mouse serum after 6 months M. avium infection and are significantly higher in infected HO-1+/+ as compared with saline controls (* P < 0.0001) and HO-1-/- mice (** P< .0001) as compared with saline controls. MCP-1 levels were significantly higher in M. avium infected HO-1-/- mice then in M. avium infected HO-1+/+ mice (*** P< 0.05) Samples from five mice in each group were assayed in triplicate. (B) CCR2 expression in PBMC and PAM of HO-1+/+ and HO-1-/- mice post M. avium infection. CCR2 receptor PAM in HO-1-/- mice with M. avium infection is significantly increased compared to M. avium infected HO-1+/+ mice. CCR2 expression in PBMC and PAM in HO-1-/- mice with M. avium infection were comparable. This is a representative histogram of three experiments with similar observation. Control mice received saline.
Mentions: Since MCP-1 is the soluble ligand for CCR2, we analyzed the MCP-1 levels in BALF and serum samples from the HO-1+/+ and HO-1-/- mice infected with M. avium or treated with saline. We observed a significant increase of MCP-1 levels in the BALF of infected HO-1+/+ mice as compared to non-infected controls, as well as increased levels in HO-1-/-M. avium infected mice as compared to HO-1-/- non-infected controls (Figure 7A). Additionally, MCP-1 levels were higher in HO-1-/- control mice as compared with HO-1+/+ control mice. Similar results were found when we analyzed the MCP-1 levels in the corresponding serum samples. HO-1-/- mice infected with M. avium showed higher levels of the cytokine as compared to HO-1+/+ samples (Figure 7A). Furthermore, the levels of MCP-1 in sera of HO-1-/- mice infected with M. avium were significantly higher than HO-1+/+ and non-infected HO-1-/- mice (Figure 7A). Although the MCP-1 profile was the same between BALF and serum samples for each condition, the amount of MCP-1 was up to 1000 fold higher in BALF than serum.

Bottom Line: Inhibition of HO by zinc protoporphyrin-IX led to inhibition of MCP-1 and increased expression of CCR2, its cognate receptor.Mycobacteria were found only inside defined granulomas but not outside granuloma in the lungs of HO-1⁺/⁺ mice.Higher MCP-1 levels were found in bronchoalveolar lavage fluid of M. avium infected HO-1(-/-) mice and CCR2 expression was higher in HO-1⁻/⁻ alveolar macrophages when compared with HO-1⁺/⁺ mice.

View Article: PubMed Central - PubMed

Affiliation: Division of Pulmonary, Critical Care and Sleep Medicine, College of Medicine, University of Florida, Gainesville, FL, USA.

ABSTRACT
Non-tuberculous mycobacterial (NTM) infections occur in both immunocompromised and immunocompetent hosts and are an increasingly recognized cause of morbidity and mortality. The hallmark of pulmonary mycobacterial infections is the formation of granuloma in the lung. Our study focuses on the role of heme oxygenase-1 (HO-1), a cytoprotective enzyme, in the regulation of granuloma development and maturation following infection with Mycobacterium avium. We examined the role of HO-1 in regulating monocyte chemoattractant protein-1 (MCP-1) and chemokine receptor 2 (CCR2), two molecules involved in monocyte-macrophage cell trafficking after infection. We showed that RAW 264.7 mouse monocytes exposed to M. avium expressed HO-1 and MCP-1. Inhibition of HO by zinc protoporphyrin-IX led to inhibition of MCP-1 and increased expression of CCR2, its cognate receptor. HO-1⁻/⁻ mice did not develop organized granuloma in their lungs, had higher lung colony forming unit of M. avium when infected with intratracheal M. avium, and had loose collections of inflammatory cells in the lung parenchyma. Mycobacteria were found only inside defined granulomas but not outside granuloma in the lungs of HO-1⁺/⁺ mice. In HO-1⁻/⁻ mice, mycobacteria were also found in the liver and spleen and showed increased mortality. Peripheral blood monocytes isolated from GFP⁺ mice and given intravenously to HO-1⁺/⁺ mice localized into tight granulomas, while in HO-1⁻/⁻ mice they remained diffusely scattered in areas of parenchymal inflammation. Higher MCP-1 levels were found in bronchoalveolar lavage fluid of M. avium infected HO-1(-/-) mice and CCR2 expression was higher in HO-1⁻/⁻ alveolar macrophages when compared with HO-1⁺/⁺ mice. CCR2 expression localized to granuloma in HO-1⁺/⁺ mice but not in the HO-1⁻/⁻ mice. These findings strongly suggest that HO-1 plays a protective role in the control of M. avium infection.

Show MeSH
Related in: MedlinePlus