Peroxisome proliferator-activated receptor gamma agonists in the prevention and treatment of murine systemic lupus erythematosus

Abstract

Peroxisome proliferator-activated receptor gamma (PPARγ) agonists are known to have many immunomodulatory effects. We have previously shown that the PPARγ agonist rosiglitazone is beneficial when used early in prevention of disease in murine models of systemic lupus erythematosus (SLE) and SLE-related atherosclerosis. In this report, we demonstrate that another PPARγ agonist, pioglitazone is also beneficial as a treatment for early murine lupus, indicating that this is a class effect and not agent-specific. We further attempt to define the ability of PPARγ agonists to ameliorate established or severe autoimmune disease using two mouse models: the MRL.lpr SLE model and the gld.apoE(-/-) model of accelerated atherosclerosis and SLE. We demonstrate that, in contrast to the marked amelioration of disease seen when PPARγ agonist treatment was started before disease onset, treatment with rosiglitazone after disease onset in MRL.lpr or gld.apoE(-/-) mice had minimal beneficial effect on the development of the autoimmune phenotype; however, rosiglitazone treatment remained highly effective at reducing lupus-associated atherosclerosis in gld.apoE(-/-) mice after disease onset or when mice were maintained on a high cholesterol Western diet. These results suggest that beneficial effects of PPARγ agonists on the development of autoimmunity might be limited to the early stages of disease, but that atherosclerosis, a major cause of death in SLE patients, may be ameliorated even in established or severe disease.

Keywords: animal models; atherosclerosis; systemic lupus erythematosus.

Figure 1

‘Mild’ phenotype MRL.lpr mice treated with pioglitazone before disease onset have less severe disease. MRL.lpr female mice were maintained on a normal diet (n = 13) or normal diet supplemented with 30 mg/kg per day pioglitazone (n = 15) for 12 weeks starting at 6 weeks of age (baseline) and then disease parameters were measured (end-point; 18 weeks of age). (a) Circulating levels of adiponectin were quantified by ELISA. (b) Spleen and (c) sub-mandibular lymph nodes were harvested and weighed. (d) Serum anti-nuclear antibody (ANA) titre was determined by HEp2 immunofluorescence. ANA titre index is a log scale of staining intensity. (e) Circulating levels of anti-dsDNA antibodies were examined by analysis of serial serum dilutions on Crithidia luciliae. Intensity of fluorescence is reported as arbitrary units (au). Kidney sections stained with haematoxylin and eosin (H&E; 40×) were used to quantify (f) glomerular tuft area and (g) glomerular cell count. (h) Representative photomicrographs of kidney sections stained with H&E, and for IgG and complement C3 deposition. **P < 0·01; ***P < 0·001.

Figure 2

‘Mild’ phenotype MRL.lpr mice are unaffected by rosiglitazone treatment started after onset of disease. Twelve-week-old female MRL.lpr mice received normal diet (n = 8) or normal diet containing rosiglitazone at a dose of 10 mg/kg per day (n = 8) for 12 weeks. (a) Circulating adiponectin levels were measured by ELISA. (b) Lymph node and spleen were weighed after tissue harvest. (c) Serum anti-nuclear antibody (ANA) titre as measured by staining intensity of HEp-2 immunofluorescence. (d) Circulating levels of anti-dsDNA antibodies were examined by analysis of serial serum dilutions on Crithidia luciliae. Intensity of fluorescence is reported as arbitrary units (au). Kidney sections stained with haematoxylin and eosin (H&E) were used to quantify (e) glomerular tuft area and (f) glomerular cell count. (g) Representative photomicrographs of kidney sections stained with H&E, and for IgG and complement C3 deposition *P < 0·05.

Figure 3

‘Severe’ phenotype MRL.lpr mice do not benefit from rosiglitazone treatment. Six-week-old female mice with the ‘severe’ lupus-like phenotype received either normal diet (n = 12) or normal diet supplemented with 10 mg/kg per day rosiglitazone (n = 10). (a) Serum adiponectin levels after 12 weeks of diet with rosiglitazone treatment. (b) Average weight of lymph nodes or spleen. (c) Anti-nuclear antibodies (ANA). (d) Circulating levels of anti-dsDNA antibodies were examined by analysis of serial serum dilutions on Crithidia luciliae. Intensity of fluorescence is reported as arbitrary units (au). Kidney sections stained with haematoxylin and eosin (H&E) were used to quantify (e) glomerular tuft area and (f) glomerular cell count. (g) Representative photomicrographs of kidney sections stained with H&E, and for IgG and complement C3 deposition. Error bars represent mean ± SEM. *P < 0·05.

Figure 4

Effect of rosiglitazone on disease manifestations in gld.apoE^−/− mice when treatment is started after onset of disease. Gld.apoE^−/− mice were maintained on a normal diet (n = 8) or normal diet supplemented with 10 mg/kg per day rosiglitazone (n = 9) for 10 weeks starting at 13 weeks of age (baseline), and then disease parameters were measured (end-point; 23 weeks of age). (a) Serum adiponectin levels. (b) Lymph node and spleen weights. (c) Serum anti-nuclear antibody (ANA) titre as determined by HEp2 immunofluorescence. (d) Circulating levels of anti-dsDNA antibodies were examined by analysis of serial serum dilutions on Crithidia luciliae. Intensity of fluorescence is reported as arbitrary units (au). Kidney sections stained with haematoxylin and eosin (H&E) were used to quantify (e) glomerular tuft area and (f) glomerular cell count. (g) Representative photomicrographs of kidney sections stained with H&E, and for IgG and complement C3 deposition. (h) Aortic atherosclerosis lesion area. (i) Representative photomicrographs of en face aortas stained with oil red O to detect lesion deposition. **P < 0·01.

Figure 5

Rosiglitazone ameliorates atherosclerosis in gld.apoE^−/− mice maintained on a high-cholesterol Western diet. Seven-week-old gld.apoE^−/− mice were fed a Western diet (n = 12) or Western diet supplemented with 10 mg/kg per day rosiglitazone (n = 10) for 12 weeks. (a) Quantification of serum adiponectin levels at baseline (7 weeks old) and end-point (19 weeks of age). (b) Lymph node and spleen weights. (c) Analysis of anti-nuclear antibody (ANA) titre was determined by HEp2 immunofluorescence. (d) Circulating levels of anti-dsDNA antibodies were examined by analysis of serial serum dilutions on Crithidia luciliae. Intensity of fluorescence is reported as arbitrary units (au). Kidney sections stained with haematoxylin and eosin (H&E) were used to quantify (e) glomerular tuft area and (f) glomerular cell count. (g) Representative photomicrographs of kidney sections stained with H&E, and for IgG and complement C3 deposition. (h) Total atherosclerotic lesion area was quantified after oil red O staining. (i) Representative photographs of aortas opened longitudinally and stained with oil red O. *P < 0·05; **P < 0·01.