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The granulocyte colony stimulating factor pathway regulates autoantibody production in a murine induced model of systemic lupus erythematosus.

Lantow M, Sivakumar R, Zeumer L, Wasserfall C, Zheng YY, Atkinson MA, Morel L - Arthritis Res. Ther. (2013)

Bottom Line: G-CSF binding by B6.Sle2c2 leukocytes was reduced as compared to B6, which was associated with a reduced expansion in response to in vivo G-CSF treatment.G-CSF in vivo treatment also failed to mobilize bone-marrow B6.Sle2c2 neutrophils as it did for B6 neutrophils.This result was corroborated by the increased anti-dsDNA IgG production in G-CSF-treated B6.TC mice, which also carry the Sle2c2 locus.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: An NZB-derived genetic locus (Sle2c2) that suppresses autoantibody production in a mouse model of induced systemic lupus erythematosus contains a polymorphism in the gene encoding the G-CSF receptor. This study was designed to test the hypothesis that the Sle2c2 suppression is associated with an impaired G-CSF receptor function that can be overcome by exogenous G-CSF.

Methods: Leukocytes from B6.Sle2c2 and B6 congenic mice, which carry a different allele of the G-CSF receptor, were compared for their responses to G-CSF. Autoantibody production was induced with the chronic graft-versus-host-disease (cGVHD) model by adoptive transfer of B6.bm12 splenocytes. Different treatment regimens varying the amount and frequency of G-CSF (Neulasta®) or carrier control were tested on cGVHD outcomes. Autoantibody production, immune cell activation, and reactive oxygen species (ROS) production were compared between the two strains with the various treatments. In addition, the effect of G-CSF treatment was examined on the production autoantibodies in the B6.Sle1.Sle2.Sle3 (B6.TC) spontaneous model of lupus.

Results: B6.Sle2c2 and B6 leukocytes responded differently to G-CSF. G-CSF binding by B6.Sle2c2 leukocytes was reduced as compared to B6, which was associated with a reduced expansion in response to in vivo G-CSF treatment. G-CSF in vivo treatment also failed to mobilize bone-marrow B6.Sle2c2 neutrophils as it did for B6 neutrophils. In contrast, the expression of G-CSF responsive genes indicated a higher G-CSF receptor signaling in B6.Sle2c2 cells. G-CSF treatment restored the ability of B6.Sle2c2 mice to produce autoantibodies in a dose-dependent manner upon cGVHD induction, which correlated with restored CD4+ T cells activation, as well as dendritic cell and granulocyte expansion. Steady-state ROS production was higher in B6.Sle2c2 than in B6 mice. cGVHD induction resulted in a larger increase in ROS production in B6 than in B6.Sle2c2 mice, and this difference was eliminated with G-CSF treatment. Finally, a low dose G-CSF treatment accelerated the production of anti-dsDNA IgG in young B6.TC mice.

Conclusion: The different in vivo and in vitro responses of B6.Sle2c2 leukocytes are consistent with the mutation in the G-CSFR having functional consequences. The elimination of Sle2c2 suppression of autoantibody production by exogenous G-CSF indicates that Sle2c2 corresponds to a loss of function of G-CSF receptor. This result was corroborated by the increased anti-dsDNA IgG production in G-CSF-treated B6.TC mice, which also carry the Sle2c2 locus. Overall, these results suggest that the G-CSF pathway regulates the production of autoantibodies in murine models of lupus.

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Human granulocyte-colony stimulation factor (huG-CSF) treatment accelerates anti-dsDNA IgG production in B6.TC mice. Anti-dsDNA IgG production in B6.TC mice (A) and B6 controls (B) that were treated weekly with 1 ug huG-CSF (GCSF) or 5% dextrose (dextrose) starting at 2 months of age (n = 3 per group). Results for B6.TC mice were normalized to their individual anti-dsDNA IgG levels before treatment (week 0) set as 1. Results for B6 were normalized to the average units of all B6.TC mice before treatment to account for the large difference between B6.TC and B6 anti-dsDNA IgG production. (C) Anti-chromatin IgG production in the four cohorts. Unit values are shown because minimal variation was observed before treatment between and within cohorts. The regression lines between the huG-CSF-treated B6.TC and B6 cohorts have significantly different slopes (P = 0.005) but not between dextrose-treated cohorts (P = 0.17). (D) Total serum IgG in the four cohorts. B6.TC mice had significantly more IgG than B6 before treatment (152 vs 95 ug/ml, P = 0.001), but that difference disappeared with time. The huG-CSF treatment did not change total IgG production in neither strain. Anti-dsDNA (E) and chromatin (F) IgG in B6.TC mice normalized to their total IgG production. The autoantibody (autoAb) unit/total IgG ratios were normalized to the individual values before treatment set as 1. The graphs show mean and standard error of the mean at the indicated time points after treatment. For A and E, the P-values correspond to an F-test comparing the slopes for the two linear regression lines corresponding to the two treatments.
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Figure 8: Human granulocyte-colony stimulation factor (huG-CSF) treatment accelerates anti-dsDNA IgG production in B6.TC mice. Anti-dsDNA IgG production in B6.TC mice (A) and B6 controls (B) that were treated weekly with 1 ug huG-CSF (GCSF) or 5% dextrose (dextrose) starting at 2 months of age (n = 3 per group). Results for B6.TC mice were normalized to their individual anti-dsDNA IgG levels before treatment (week 0) set as 1. Results for B6 were normalized to the average units of all B6.TC mice before treatment to account for the large difference between B6.TC and B6 anti-dsDNA IgG production. (C) Anti-chromatin IgG production in the four cohorts. Unit values are shown because minimal variation was observed before treatment between and within cohorts. The regression lines between the huG-CSF-treated B6.TC and B6 cohorts have significantly different slopes (P = 0.005) but not between dextrose-treated cohorts (P = 0.17). (D) Total serum IgG in the four cohorts. B6.TC mice had significantly more IgG than B6 before treatment (152 vs 95 ug/ml, P = 0.001), but that difference disappeared with time. The huG-CSF treatment did not change total IgG production in neither strain. Anti-dsDNA (E) and chromatin (F) IgG in B6.TC mice normalized to their total IgG production. The autoantibody (autoAb) unit/total IgG ratios were normalized to the individual values before treatment set as 1. The graphs show mean and standard error of the mean at the indicated time points after treatment. For A and E, the P-values correspond to an F-test comparing the slopes for the two linear regression lines corresponding to the two treatments.

Mentions: B6.TC mice first produce anti-chromatin IgG between 2 to 3 months of age then anti-dsDNA IgG between 4 and 5 months of age [19]. The presence of Sle2c2 in the B6.TC genome predicts that G-CSF treatment would accelerate their autoAb production as it did in the induced lupus model. We treated 2-month-old B6.TC and B6 female mice with six weekly injections of either 1ug hu-G-CSF or dextrose. It has been shown that neutralizing anti-hu G-CSF antibodies develop with a greater number of injections [23]. G-CSF treated B6.TC mice developed anti-dsDNA IgG significantly faster than control mice (Figure 8A). The G-CSF treatment did not induce anti-dsDNA IgG in B6 mice (Figure 8B). G-CSF, however, had no effect on anti-chromatin IgG, which increased at a similar rate in both treated and control B6.TC mice, and stayed at background levels in treated and control B6 mice (Figure 8C). G-CSF treatment did not affect total IgG levels (Figure 8D) or IgM (data not shown) in either strain, indicating that the effect of G-GSF was specific for anti-dsDNA IgG. This was confirmed when the production of anti-dsDNA IgG was normalized to total IgG (Figure 8E). It was not the case for anti-chromatin IgG (Figure 8F), which may be related to the fact that the anti-chromatin response was already ongoing when the treatment was initiated. Nevertheless, these results show that exogenous G-CSF increased production of anti-dsDNA IgG in mice carrying the Sle2c2 locus in a spontaneous model of lupus.


The granulocyte colony stimulating factor pathway regulates autoantibody production in a murine induced model of systemic lupus erythematosus.

Lantow M, Sivakumar R, Zeumer L, Wasserfall C, Zheng YY, Atkinson MA, Morel L - Arthritis Res. Ther. (2013)

Human granulocyte-colony stimulation factor (huG-CSF) treatment accelerates anti-dsDNA IgG production in B6.TC mice. Anti-dsDNA IgG production in B6.TC mice (A) and B6 controls (B) that were treated weekly with 1 ug huG-CSF (GCSF) or 5% dextrose (dextrose) starting at 2 months of age (n = 3 per group). Results for B6.TC mice were normalized to their individual anti-dsDNA IgG levels before treatment (week 0) set as 1. Results for B6 were normalized to the average units of all B6.TC mice before treatment to account for the large difference between B6.TC and B6 anti-dsDNA IgG production. (C) Anti-chromatin IgG production in the four cohorts. Unit values are shown because minimal variation was observed before treatment between and within cohorts. The regression lines between the huG-CSF-treated B6.TC and B6 cohorts have significantly different slopes (P = 0.005) but not between dextrose-treated cohorts (P = 0.17). (D) Total serum IgG in the four cohorts. B6.TC mice had significantly more IgG than B6 before treatment (152 vs 95 ug/ml, P = 0.001), but that difference disappeared with time. The huG-CSF treatment did not change total IgG production in neither strain. Anti-dsDNA (E) and chromatin (F) IgG in B6.TC mice normalized to their total IgG production. The autoantibody (autoAb) unit/total IgG ratios were normalized to the individual values before treatment set as 1. The graphs show mean and standard error of the mean at the indicated time points after treatment. For A and E, the P-values correspond to an F-test comparing the slopes for the two linear regression lines corresponding to the two treatments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 8: Human granulocyte-colony stimulation factor (huG-CSF) treatment accelerates anti-dsDNA IgG production in B6.TC mice. Anti-dsDNA IgG production in B6.TC mice (A) and B6 controls (B) that were treated weekly with 1 ug huG-CSF (GCSF) or 5% dextrose (dextrose) starting at 2 months of age (n = 3 per group). Results for B6.TC mice were normalized to their individual anti-dsDNA IgG levels before treatment (week 0) set as 1. Results for B6 were normalized to the average units of all B6.TC mice before treatment to account for the large difference between B6.TC and B6 anti-dsDNA IgG production. (C) Anti-chromatin IgG production in the four cohorts. Unit values are shown because minimal variation was observed before treatment between and within cohorts. The regression lines between the huG-CSF-treated B6.TC and B6 cohorts have significantly different slopes (P = 0.005) but not between dextrose-treated cohorts (P = 0.17). (D) Total serum IgG in the four cohorts. B6.TC mice had significantly more IgG than B6 before treatment (152 vs 95 ug/ml, P = 0.001), but that difference disappeared with time. The huG-CSF treatment did not change total IgG production in neither strain. Anti-dsDNA (E) and chromatin (F) IgG in B6.TC mice normalized to their total IgG production. The autoantibody (autoAb) unit/total IgG ratios were normalized to the individual values before treatment set as 1. The graphs show mean and standard error of the mean at the indicated time points after treatment. For A and E, the P-values correspond to an F-test comparing the slopes for the two linear regression lines corresponding to the two treatments.
Mentions: B6.TC mice first produce anti-chromatin IgG between 2 to 3 months of age then anti-dsDNA IgG between 4 and 5 months of age [19]. The presence of Sle2c2 in the B6.TC genome predicts that G-CSF treatment would accelerate their autoAb production as it did in the induced lupus model. We treated 2-month-old B6.TC and B6 female mice with six weekly injections of either 1ug hu-G-CSF or dextrose. It has been shown that neutralizing anti-hu G-CSF antibodies develop with a greater number of injections [23]. G-CSF treated B6.TC mice developed anti-dsDNA IgG significantly faster than control mice (Figure 8A). The G-CSF treatment did not induce anti-dsDNA IgG in B6 mice (Figure 8B). G-CSF, however, had no effect on anti-chromatin IgG, which increased at a similar rate in both treated and control B6.TC mice, and stayed at background levels in treated and control B6 mice (Figure 8C). G-CSF treatment did not affect total IgG levels (Figure 8D) or IgM (data not shown) in either strain, indicating that the effect of G-GSF was specific for anti-dsDNA IgG. This was confirmed when the production of anti-dsDNA IgG was normalized to total IgG (Figure 8E). It was not the case for anti-chromatin IgG (Figure 8F), which may be related to the fact that the anti-chromatin response was already ongoing when the treatment was initiated. Nevertheless, these results show that exogenous G-CSF increased production of anti-dsDNA IgG in mice carrying the Sle2c2 locus in a spontaneous model of lupus.

Bottom Line: G-CSF binding by B6.Sle2c2 leukocytes was reduced as compared to B6, which was associated with a reduced expansion in response to in vivo G-CSF treatment.G-CSF in vivo treatment also failed to mobilize bone-marrow B6.Sle2c2 neutrophils as it did for B6 neutrophils.This result was corroborated by the increased anti-dsDNA IgG production in G-CSF-treated B6.TC mice, which also carry the Sle2c2 locus.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: An NZB-derived genetic locus (Sle2c2) that suppresses autoantibody production in a mouse model of induced systemic lupus erythematosus contains a polymorphism in the gene encoding the G-CSF receptor. This study was designed to test the hypothesis that the Sle2c2 suppression is associated with an impaired G-CSF receptor function that can be overcome by exogenous G-CSF.

Methods: Leukocytes from B6.Sle2c2 and B6 congenic mice, which carry a different allele of the G-CSF receptor, were compared for their responses to G-CSF. Autoantibody production was induced with the chronic graft-versus-host-disease (cGVHD) model by adoptive transfer of B6.bm12 splenocytes. Different treatment regimens varying the amount and frequency of G-CSF (Neulasta®) or carrier control were tested on cGVHD outcomes. Autoantibody production, immune cell activation, and reactive oxygen species (ROS) production were compared between the two strains with the various treatments. In addition, the effect of G-CSF treatment was examined on the production autoantibodies in the B6.Sle1.Sle2.Sle3 (B6.TC) spontaneous model of lupus.

Results: B6.Sle2c2 and B6 leukocytes responded differently to G-CSF. G-CSF binding by B6.Sle2c2 leukocytes was reduced as compared to B6, which was associated with a reduced expansion in response to in vivo G-CSF treatment. G-CSF in vivo treatment also failed to mobilize bone-marrow B6.Sle2c2 neutrophils as it did for B6 neutrophils. In contrast, the expression of G-CSF responsive genes indicated a higher G-CSF receptor signaling in B6.Sle2c2 cells. G-CSF treatment restored the ability of B6.Sle2c2 mice to produce autoantibodies in a dose-dependent manner upon cGVHD induction, which correlated with restored CD4+ T cells activation, as well as dendritic cell and granulocyte expansion. Steady-state ROS production was higher in B6.Sle2c2 than in B6 mice. cGVHD induction resulted in a larger increase in ROS production in B6 than in B6.Sle2c2 mice, and this difference was eliminated with G-CSF treatment. Finally, a low dose G-CSF treatment accelerated the production of anti-dsDNA IgG in young B6.TC mice.

Conclusion: The different in vivo and in vitro responses of B6.Sle2c2 leukocytes are consistent with the mutation in the G-CSFR having functional consequences. The elimination of Sle2c2 suppression of autoantibody production by exogenous G-CSF indicates that Sle2c2 corresponds to a loss of function of G-CSF receptor. This result was corroborated by the increased anti-dsDNA IgG production in G-CSF-treated B6.TC mice, which also carry the Sle2c2 locus. Overall, these results suggest that the G-CSF pathway regulates the production of autoantibodies in murine models of lupus.

Show MeSH
Related in: MedlinePlus