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Transition from an M1 to a mixed neuroinflammatory phenotype increases amyloid deposition in APP/PS1 transgenic mice.

Weekman EM, Sudduth TL, Abner EL, Popa GJ, Mendenhall MD, Brothers HM, Braun K, Greenstein A, Wilcock DM - J Neuroinflammation (2014)

Bottom Line: Short-term studies show that induction of an M1 neuroinflammatory phenotype reduces Aβ, but long-term studies have not been performed that track the neuroinflammatory phenotype.Expression of IFNγ through AAV successfully induced an M1 phenotype at 4 months that transitioned to a mixed phenotype by 6 months.This transition also appeared with an increase in amyloid burden suggesting that a mixed phenotype, or enhanced expression of M2a and M2c markers, could contribute to increasing amyloid burden and disease progression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Sanders-Brown Center on Aging, University of Kentucky, Lexington, KY 40536, USA. donna.wilcock@uky.edu.

ABSTRACT

Background: The polarization to different neuroinflammatory phenotypes has been described in early Alzheimer's disease, yet the impact of these phenotypes on amyloid-beta (Aβ) pathology remains unknown. Short-term studies show that induction of an M1 neuroinflammatory phenotype reduces Aβ, but long-term studies have not been performed that track the neuroinflammatory phenotype.

Methods: Wild-type and APP/PS1 transgenic mice aged 3 to 4 months received a bilateral intracranial injection of adeno-associated viral (AAV) vectors expressing IFNγ or green fluorescent protein in the frontal cortex and hippocampus. Mice were sacrificed 4 or 6 months post-injection. ELISA measurements were used for IFNγ protein levels and biochemical levels of Aβ. The neuroinflammatory phenotype was determined through quantitative PCR. Microglia, astrocytes, and Aβ levels were assessed with immunohistochemistry.

Results: AAV expressing IFNγ induced an M1 neuroinflammatory phenotype at 4 months and a mixed phenotype along with an increase in Aβ at 6 months. Microglial staining was increased at 6 months and astrocyte staining was decreased at 4 and 6 months in mice receiving AAV expressing IFNγ.

Conclusions: Expression of IFNγ through AAV successfully induced an M1 phenotype at 4 months that transitioned to a mixed phenotype by 6 months. This transition also appeared with an increase in amyloid burden suggesting that a mixed phenotype, or enhanced expression of M2a and M2c markers, could contribute to increasing amyloid burden and disease progression.

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IFNγ induces an M1 neuroinflammatory phenotype. Relative gene expression for genes representative of the M1, M2a, M2b, and M2c phenotypes. Data are shown as fold-change relative to mice receiving GFP-AAV at the given time point and genotype. (A) Wild-type (WT) mice receiving IFNγ-AAV for 4 months (4 m). (B) WT mice receiving IFNγ-AAV for 6 months (6 m). (C) APP/PS1 receiving IFNγ-AAV for 4 months. (D) APP/PS1 receiving IFNγ-AAV for 6 months. *P < 0.05, **P < 0.01, IFNγ-AAV compared to GFP-AAV of that time point and genotype. #P < 0.05, ##P < 0.01, 4-month IFNγ-AAV compared to 6-month IFNγ-AAV of that genotype. AAV, adeno-associated virus; AARG1, arginase 1; FCGR, Fc gamma receptor; GFP, green fluorescent protein; IFN, interferon; IL, interleukin; SPHK1, sphingosine kinase 1; TGFB, transforming growth factor beta.
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Figure 2: IFNγ induces an M1 neuroinflammatory phenotype. Relative gene expression for genes representative of the M1, M2a, M2b, and M2c phenotypes. Data are shown as fold-change relative to mice receiving GFP-AAV at the given time point and genotype. (A) Wild-type (WT) mice receiving IFNγ-AAV for 4 months (4 m). (B) WT mice receiving IFNγ-AAV for 6 months (6 m). (C) APP/PS1 receiving IFNγ-AAV for 4 months. (D) APP/PS1 receiving IFNγ-AAV for 6 months. *P < 0.05, **P < 0.01, IFNγ-AAV compared to GFP-AAV of that time point and genotype. #P < 0.05, ##P < 0.01, 4-month IFNγ-AAV compared to 6-month IFNγ-AAV of that genotype. AAV, adeno-associated virus; AARG1, arginase 1; FCGR, Fc gamma receptor; GFP, green fluorescent protein; IFN, interferon; IL, interleukin; SPHK1, sphingosine kinase 1; TGFB, transforming growth factor beta.

Mentions: To determine the neuroinflammatory phenotype, right hippocampal RNA was isolated and real time PCR was performed for several genes specific to an M1, M2a, M2b or M2c macrophage phenotype. Data are shown as fold change compared to GFP-AAV at each time point for each genotype. The GFP-AAV mice showed no significant change in neuroinflammation over time (data not shown). Overall, both WT and APP/PS1 mice responded to the IFNγ-AAV with a robust M1 response. IL-12b was increased in IFNγ-AAV treated mice at 4 and 6 months in both genotypes, and IL-1β was increased at all time points, except 6 months in WT (Figure 2A-D). Additionally, IL-6 and IL-12a showed significant increases at the 6-month (but not 4-month) time point in both WT and APP/PS1 mice (Figure 2B,D). At the 6-month time point, but not at the 4-month time point, M2a gene YM1 was significantly increased in WT and APP/PS1 mice (Figure 2A-D). Also of note are increased M2b markers CD86, FcγR1 and FcγR3 at the 4 and 6-month time point (Figure 2B,D). Table 2 shows the fold-change values for GFP-AAV- and IFNγ-AAV-treated mice. Interestingly, while it may appear that the APP/PS1 mice are less responsive to the IFNγ-AAV, it is actually due to the fact that APP/PS1 mice normally have an inflammatory response to the amyloid deposits in the brain and so the fold-change achieved by the IFNγ-AAV is less than in WT.Next, assessment of CD11b immunohistochemistry in the frontal cortex (images not shown) and the hippocampus (dentate gyrus shown, Figure 3A-H) was performed to determine the effect of IFNγ on microglia. In all four brain regions measured, for both WT and APP/PS1 mice receiving IFNγ-AAV, there were statistically significant increases in CD11b staining, as measured by percent area occupied by positive immunostain, at 6 months compared to 4 months that were not observed in mice receiving GFP-AAV (that is, the treatment-by-time interaction is significant). In the frontal cortex, there was a statistically significant increase in microglial staining at 6 months but not 4 months in both WT and APP/PS1 mice injected with IFNγ-AAV compared to mice receiving GFP-AAV (Figure 3I). At 6 months in WT mice, there were significant increases in staining in the CA1, CA3 and (DG) regions (Figure 3A-D,I). APP/PS1 mice receiving IFNγ-AAV only showed modest, non-significant increases at 4 months (Figure 3E,F,I) but significant increases were observed in all hippocampal regions by 6 months (Figure 3G,H,I).


Transition from an M1 to a mixed neuroinflammatory phenotype increases amyloid deposition in APP/PS1 transgenic mice.

Weekman EM, Sudduth TL, Abner EL, Popa GJ, Mendenhall MD, Brothers HM, Braun K, Greenstein A, Wilcock DM - J Neuroinflammation (2014)

IFNγ induces an M1 neuroinflammatory phenotype. Relative gene expression for genes representative of the M1, M2a, M2b, and M2c phenotypes. Data are shown as fold-change relative to mice receiving GFP-AAV at the given time point and genotype. (A) Wild-type (WT) mice receiving IFNγ-AAV for 4 months (4 m). (B) WT mice receiving IFNγ-AAV for 6 months (6 m). (C) APP/PS1 receiving IFNγ-AAV for 4 months. (D) APP/PS1 receiving IFNγ-AAV for 6 months. *P < 0.05, **P < 0.01, IFNγ-AAV compared to GFP-AAV of that time point and genotype. #P < 0.05, ##P < 0.01, 4-month IFNγ-AAV compared to 6-month IFNγ-AAV of that genotype. AAV, adeno-associated virus; AARG1, arginase 1; FCGR, Fc gamma receptor; GFP, green fluorescent protein; IFN, interferon; IL, interleukin; SPHK1, sphingosine kinase 1; TGFB, transforming growth factor beta.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 2: IFNγ induces an M1 neuroinflammatory phenotype. Relative gene expression for genes representative of the M1, M2a, M2b, and M2c phenotypes. Data are shown as fold-change relative to mice receiving GFP-AAV at the given time point and genotype. (A) Wild-type (WT) mice receiving IFNγ-AAV for 4 months (4 m). (B) WT mice receiving IFNγ-AAV for 6 months (6 m). (C) APP/PS1 receiving IFNγ-AAV for 4 months. (D) APP/PS1 receiving IFNγ-AAV for 6 months. *P < 0.05, **P < 0.01, IFNγ-AAV compared to GFP-AAV of that time point and genotype. #P < 0.05, ##P < 0.01, 4-month IFNγ-AAV compared to 6-month IFNγ-AAV of that genotype. AAV, adeno-associated virus; AARG1, arginase 1; FCGR, Fc gamma receptor; GFP, green fluorescent protein; IFN, interferon; IL, interleukin; SPHK1, sphingosine kinase 1; TGFB, transforming growth factor beta.
Mentions: To determine the neuroinflammatory phenotype, right hippocampal RNA was isolated and real time PCR was performed for several genes specific to an M1, M2a, M2b or M2c macrophage phenotype. Data are shown as fold change compared to GFP-AAV at each time point for each genotype. The GFP-AAV mice showed no significant change in neuroinflammation over time (data not shown). Overall, both WT and APP/PS1 mice responded to the IFNγ-AAV with a robust M1 response. IL-12b was increased in IFNγ-AAV treated mice at 4 and 6 months in both genotypes, and IL-1β was increased at all time points, except 6 months in WT (Figure 2A-D). Additionally, IL-6 and IL-12a showed significant increases at the 6-month (but not 4-month) time point in both WT and APP/PS1 mice (Figure 2B,D). At the 6-month time point, but not at the 4-month time point, M2a gene YM1 was significantly increased in WT and APP/PS1 mice (Figure 2A-D). Also of note are increased M2b markers CD86, FcγR1 and FcγR3 at the 4 and 6-month time point (Figure 2B,D). Table 2 shows the fold-change values for GFP-AAV- and IFNγ-AAV-treated mice. Interestingly, while it may appear that the APP/PS1 mice are less responsive to the IFNγ-AAV, it is actually due to the fact that APP/PS1 mice normally have an inflammatory response to the amyloid deposits in the brain and so the fold-change achieved by the IFNγ-AAV is less than in WT.Next, assessment of CD11b immunohistochemistry in the frontal cortex (images not shown) and the hippocampus (dentate gyrus shown, Figure 3A-H) was performed to determine the effect of IFNγ on microglia. In all four brain regions measured, for both WT and APP/PS1 mice receiving IFNγ-AAV, there were statistically significant increases in CD11b staining, as measured by percent area occupied by positive immunostain, at 6 months compared to 4 months that were not observed in mice receiving GFP-AAV (that is, the treatment-by-time interaction is significant). In the frontal cortex, there was a statistically significant increase in microglial staining at 6 months but not 4 months in both WT and APP/PS1 mice injected with IFNγ-AAV compared to mice receiving GFP-AAV (Figure 3I). At 6 months in WT mice, there were significant increases in staining in the CA1, CA3 and (DG) regions (Figure 3A-D,I). APP/PS1 mice receiving IFNγ-AAV only showed modest, non-significant increases at 4 months (Figure 3E,F,I) but significant increases were observed in all hippocampal regions by 6 months (Figure 3G,H,I).

Bottom Line: Short-term studies show that induction of an M1 neuroinflammatory phenotype reduces Aβ, but long-term studies have not been performed that track the neuroinflammatory phenotype.Expression of IFNγ through AAV successfully induced an M1 phenotype at 4 months that transitioned to a mixed phenotype by 6 months.This transition also appeared with an increase in amyloid burden suggesting that a mixed phenotype, or enhanced expression of M2a and M2c markers, could contribute to increasing amyloid burden and disease progression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Sanders-Brown Center on Aging, University of Kentucky, Lexington, KY 40536, USA. donna.wilcock@uky.edu.

ABSTRACT

Background: The polarization to different neuroinflammatory phenotypes has been described in early Alzheimer's disease, yet the impact of these phenotypes on amyloid-beta (Aβ) pathology remains unknown. Short-term studies show that induction of an M1 neuroinflammatory phenotype reduces Aβ, but long-term studies have not been performed that track the neuroinflammatory phenotype.

Methods: Wild-type and APP/PS1 transgenic mice aged 3 to 4 months received a bilateral intracranial injection of adeno-associated viral (AAV) vectors expressing IFNγ or green fluorescent protein in the frontal cortex and hippocampus. Mice were sacrificed 4 or 6 months post-injection. ELISA measurements were used for IFNγ protein levels and biochemical levels of Aβ. The neuroinflammatory phenotype was determined through quantitative PCR. Microglia, astrocytes, and Aβ levels were assessed with immunohistochemistry.

Results: AAV expressing IFNγ induced an M1 neuroinflammatory phenotype at 4 months and a mixed phenotype along with an increase in Aβ at 6 months. Microglial staining was increased at 6 months and astrocyte staining was decreased at 4 and 6 months in mice receiving AAV expressing IFNγ.

Conclusions: Expression of IFNγ through AAV successfully induced an M1 phenotype at 4 months that transitioned to a mixed phenotype by 6 months. This transition also appeared with an increase in amyloid burden suggesting that a mixed phenotype, or enhanced expression of M2a and M2c markers, could contribute to increasing amyloid burden and disease progression.

Show MeSH
Related in: MedlinePlus