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Female Bias in Systemic Lupus Erythematosus is Associated with the Differential Expression of X-Linked Toll-Like Receptor 8.

McDonald G, Cabal N, Vannier A, Umiker B, Yin RH, Orjalo AV, Johansson HE, Han JH, Imanishi-Kari T - Front Immunol (2015)

Bottom Line: Here, we show the results of our investigation into the role of Tlr8 expression in SLE pathogenesis in 564Igi females.Bone marrow-derived macrophages from 564Igi females have a significant increase in Tlr8 expression compared to male-derived cells, and RNA fluorescence in situ hybridization data suggest that Tlr8 may escape X-inactivation in female-derived macrophages.These results propose a model by which females may be more susceptible to SLE pathogenesis due to inefficient inactivation of Tlr8.

View Article: PubMed Central - PubMed

Affiliation: Department of Integrative Physiology and Pathobiology, Tufts University , Boston, MA , USA.

ABSTRACT
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by the production of anti-nuclear antibodies. SLE is one of many autoimmune disorders that have a strong gender bias, with 70-90% of SLE patients being female. Several explanations have been postulated to account for the severity of autoimmune diseases in females, including hormonal, microbiota, and gene dosage differences. X-linked toll-like receptors (TLRs) have recently been implicated in disease progression in females. Our previous studies using the 564Igi mouse model of SLE on a Tlr7 and Tlr9 double knockout background showed that the presence of Tlr8 on both X chromosomes was required for the production of IgG autoantibodies, Ifn-I expression and granulopoiesis in females. Here, we show the results of our investigation into the role of Tlr8 expression in SLE pathogenesis in 564Igi females. Female mice have an increase in serum pathogenic anti-RNA IgG2a and IgG2b autoantibodies. 564Igi mice have also been shown to have an increase in neutrophils in vivo, which are major contributors to Ifn-α expression. Here, we show that neutrophils from C57BL/6 mice express Ifn-α in response to 564 immune complexes and TLR8 activation. Bone marrow-derived macrophages from 564Igi females have a significant increase in Tlr8 expression compared to male-derived cells, and RNA fluorescence in situ hybridization data suggest that Tlr8 may escape X-inactivation in female-derived macrophages. These results propose a model by which females may be more susceptible to SLE pathogenesis due to inefficient inactivation of Tlr8.

No MeSH data available.


Related in: MedlinePlus

564 IgG2b IC activates neutrophils to express Ifn-α6. (A) C57BL/6 mice were given intraperitoneal injections with 40 μg of 564 antibody, an isotype control of unknown specificity or a comparable volume of saline control. Two weeks later, the number of neutrophils in the bone marrow was determined by flow cytometry. Shown is the percent of neutrophils in each mouse. The average is shown as a horizontal line. (B) Neutrophils were purified from whole bone marrow cell suspensions from the indicated mice by cell sorting (CD11b+ Ly6G+). Shown is a representative plot of the gate used to purify neutrophils. (C) Purified neutrophils were cultured for 4 h in the presence of 0.01 μg/mL purified IgG2b 564 antibody or 564 IC made by incubating purified IgG2b 564 antibody with bone marrow-derived RNA. RNA was extracted from the cell culture, converted to cDNA, and Ifn-α6 expression was measured by RT-qPCR. Expression was normalized to β-actin and compared to untreated samples from each genotype. Shown is the average ± SEM fold-increase in Ifn-α6 expression in three mice per group tested in independent experiments, each done in triplicate.
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Figure 2: 564 IgG2b IC activates neutrophils to express Ifn-α6. (A) C57BL/6 mice were given intraperitoneal injections with 40 μg of 564 antibody, an isotype control of unknown specificity or a comparable volume of saline control. Two weeks later, the number of neutrophils in the bone marrow was determined by flow cytometry. Shown is the percent of neutrophils in each mouse. The average is shown as a horizontal line. (B) Neutrophils were purified from whole bone marrow cell suspensions from the indicated mice by cell sorting (CD11b+ Ly6G+). Shown is a representative plot of the gate used to purify neutrophils. (C) Purified neutrophils were cultured for 4 h in the presence of 0.01 μg/mL purified IgG2b 564 antibody or 564 IC made by incubating purified IgG2b 564 antibody with bone marrow-derived RNA. RNA was extracted from the cell culture, converted to cDNA, and Ifn-α6 expression was measured by RT-qPCR. Expression was normalized to β-actin and compared to untreated samples from each genotype. Shown is the average ± SEM fold-increase in Ifn-α6 expression in three mice per group tested in independent experiments, each done in triplicate.

Mentions: 564Igi mice were shown to have significantly more monocytes and neutrophils compared to C57BL/6 controls (30). To determine if the 564 antibody plays a role in the increase in granulocytes, we injected normal C57BL/6 mice with the 564 antibody and then whole bone marrow cell suspensions were analyzed for the presence of neutrophils 2 weeks later. The 564 antibody induced a significant increase in the percent of neutrophils in the bone marrow compared to a control saline injection and an isotype control (Figure 2A).


Female Bias in Systemic Lupus Erythematosus is Associated with the Differential Expression of X-Linked Toll-Like Receptor 8.

McDonald G, Cabal N, Vannier A, Umiker B, Yin RH, Orjalo AV, Johansson HE, Han JH, Imanishi-Kari T - Front Immunol (2015)

564 IgG2b IC activates neutrophils to express Ifn-α6. (A) C57BL/6 mice were given intraperitoneal injections with 40 μg of 564 antibody, an isotype control of unknown specificity or a comparable volume of saline control. Two weeks later, the number of neutrophils in the bone marrow was determined by flow cytometry. Shown is the percent of neutrophils in each mouse. The average is shown as a horizontal line. (B) Neutrophils were purified from whole bone marrow cell suspensions from the indicated mice by cell sorting (CD11b+ Ly6G+). Shown is a representative plot of the gate used to purify neutrophils. (C) Purified neutrophils were cultured for 4 h in the presence of 0.01 μg/mL purified IgG2b 564 antibody or 564 IC made by incubating purified IgG2b 564 antibody with bone marrow-derived RNA. RNA was extracted from the cell culture, converted to cDNA, and Ifn-α6 expression was measured by RT-qPCR. Expression was normalized to β-actin and compared to untreated samples from each genotype. Shown is the average ± SEM fold-increase in Ifn-α6 expression in three mice per group tested in independent experiments, each done in triplicate.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4561825&req=5

Figure 2: 564 IgG2b IC activates neutrophils to express Ifn-α6. (A) C57BL/6 mice were given intraperitoneal injections with 40 μg of 564 antibody, an isotype control of unknown specificity or a comparable volume of saline control. Two weeks later, the number of neutrophils in the bone marrow was determined by flow cytometry. Shown is the percent of neutrophils in each mouse. The average is shown as a horizontal line. (B) Neutrophils were purified from whole bone marrow cell suspensions from the indicated mice by cell sorting (CD11b+ Ly6G+). Shown is a representative plot of the gate used to purify neutrophils. (C) Purified neutrophils were cultured for 4 h in the presence of 0.01 μg/mL purified IgG2b 564 antibody or 564 IC made by incubating purified IgG2b 564 antibody with bone marrow-derived RNA. RNA was extracted from the cell culture, converted to cDNA, and Ifn-α6 expression was measured by RT-qPCR. Expression was normalized to β-actin and compared to untreated samples from each genotype. Shown is the average ± SEM fold-increase in Ifn-α6 expression in three mice per group tested in independent experiments, each done in triplicate.
Mentions: 564Igi mice were shown to have significantly more monocytes and neutrophils compared to C57BL/6 controls (30). To determine if the 564 antibody plays a role in the increase in granulocytes, we injected normal C57BL/6 mice with the 564 antibody and then whole bone marrow cell suspensions were analyzed for the presence of neutrophils 2 weeks later. The 564 antibody induced a significant increase in the percent of neutrophils in the bone marrow compared to a control saline injection and an isotype control (Figure 2A).

Bottom Line: Here, we show the results of our investigation into the role of Tlr8 expression in SLE pathogenesis in 564Igi females.Bone marrow-derived macrophages from 564Igi females have a significant increase in Tlr8 expression compared to male-derived cells, and RNA fluorescence in situ hybridization data suggest that Tlr8 may escape X-inactivation in female-derived macrophages.These results propose a model by which females may be more susceptible to SLE pathogenesis due to inefficient inactivation of Tlr8.

View Article: PubMed Central - PubMed

Affiliation: Department of Integrative Physiology and Pathobiology, Tufts University , Boston, MA , USA.

ABSTRACT
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by the production of anti-nuclear antibodies. SLE is one of many autoimmune disorders that have a strong gender bias, with 70-90% of SLE patients being female. Several explanations have been postulated to account for the severity of autoimmune diseases in females, including hormonal, microbiota, and gene dosage differences. X-linked toll-like receptors (TLRs) have recently been implicated in disease progression in females. Our previous studies using the 564Igi mouse model of SLE on a Tlr7 and Tlr9 double knockout background showed that the presence of Tlr8 on both X chromosomes was required for the production of IgG autoantibodies, Ifn-I expression and granulopoiesis in females. Here, we show the results of our investigation into the role of Tlr8 expression in SLE pathogenesis in 564Igi females. Female mice have an increase in serum pathogenic anti-RNA IgG2a and IgG2b autoantibodies. 564Igi mice have also been shown to have an increase in neutrophils in vivo, which are major contributors to Ifn-α expression. Here, we show that neutrophils from C57BL/6 mice express Ifn-α in response to 564 immune complexes and TLR8 activation. Bone marrow-derived macrophages from 564Igi females have a significant increase in Tlr8 expression compared to male-derived cells, and RNA fluorescence in situ hybridization data suggest that Tlr8 may escape X-inactivation in female-derived macrophages. These results propose a model by which females may be more susceptible to SLE pathogenesis due to inefficient inactivation of Tlr8.

No MeSH data available.


Related in: MedlinePlus