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Immune-Mediated Nephropathy and Systemic Autoimmunity in Mice Does Not Require Receptor Interacting Protein Kinase 3 (RIPK3)

View Article: PubMed Central - PubMed

ABSTRACT

Immune mediated nephropathy is one of the most serious manifestations of lupus and is characterized by severe inflammation and necrosis that, if untreated, eventually leads to renal failure. Although lupus has a higher incidence in women, both sexes can develop lupus glomerulonephritis; nephritis in men develops earlier and is more severe than in women. It is therefore important to understand the cellular and molecular mechanisms mediating nephritis in each sex. Previous work by our lab found that the absence or pharmacological inhibition of Poly [ADP-ribose] polymerase 1 (PARP-1), an enzyme involved in DNA repair and necrotic cell death, affects only male mice and results in milder nephritis, with less in situ inflammation, and diminished incidence of necrotic lesions, allowing for higher survival rates. A second pathway mediating necrosis involves Receptor-Interacting Serine-Threonine Kinase 3 (RIPK3); in this study we sought to investigate the impact of RIPK3 on the development of lupus and nephritis in both sexes. To this end, we used two inducible murine models of lupus: chronic graft versus host disease (cGvHD) and pristane-induced lupus; and nephrotoxic serum (NTS)-induced nephritis as a model of immune mediated nephropathy. We found that the absence of RIPK3 has neither positive nor negative impact on the disease development or progression of lupus and nephritis in all three models, and in both male and female mice. We conclude that RIPK3 is dispensable for the pathogenesis of lupus and immune mediated nephropathy as to accelerate, worsen or ameliorate the disease.

No MeSH data available.


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Absence of RIPK3 does not cause a reduction in necrotic cell death within the kidney during NTS-nephritis.Percentage of apoptotic and necrotic TUNEL+ cells were determined by staining for TUNEL (red), active Caspase 3 (green), and nuclei (DAPI-blue). 10x size bar = 200μm, 40x size bar = 50μm. At least 50 TUNEL+ cells were recorded for each mouse, N = 5 mice per strain. Necrotic nuclei were characterized by only TUNEL+ staining, while apoptotic cells were positive for both TUNEL and active-caspase 3. Percentages of positive cells between B6 and RIPK3-/-, in each sex were not statistically significant (p≥0.05) as determined by T-test. Data is represented as Mean ± SEM of 2–5 mice per group. Experiments were repeated at least twice.
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pone.0163611.g004: Absence of RIPK3 does not cause a reduction in necrotic cell death within the kidney during NTS-nephritis.Percentage of apoptotic and necrotic TUNEL+ cells were determined by staining for TUNEL (red), active Caspase 3 (green), and nuclei (DAPI-blue). 10x size bar = 200μm, 40x size bar = 50μm. At least 50 TUNEL+ cells were recorded for each mouse, N = 5 mice per strain. Necrotic nuclei were characterized by only TUNEL+ staining, while apoptotic cells were positive for both TUNEL and active-caspase 3. Percentages of positive cells between B6 and RIPK3-/-, in each sex were not statistically significant (p≥0.05) as determined by T-test. Data is represented as Mean ± SEM of 2–5 mice per group. Experiments were repeated at least twice.

Mentions: The lack of protection from nephritis in the absence of RIPK3 in male or female mice led us to investigate the degree of necrosis in the kidneys of these mice during NTS-nephritis. Co-staining for active-Caspase 3 by immunohistochemistry and DNA fragmentation by TUNEL showed similar percentages of Casp3-/TUNEL+ cells, demonstrating an equal degree of necrotic lesion incidence within the kidneys of mice from either sex independent of RIPK3 expression status (Fig 4). Moreover, we confirmed our previous reports that female mice show more apoptosis than necrosis, compared to male mice, and RIPK3 does not affect this ratio as well. We conclude that the necrosis, which develops during NTS-induced nephritis, is not due to RIPK3-driven pathway.


Immune-Mediated Nephropathy and Systemic Autoimmunity in Mice Does Not Require Receptor Interacting Protein Kinase 3 (RIPK3)
Absence of RIPK3 does not cause a reduction in necrotic cell death within the kidney during NTS-nephritis.Percentage of apoptotic and necrotic TUNEL+ cells were determined by staining for TUNEL (red), active Caspase 3 (green), and nuclei (DAPI-blue). 10x size bar = 200μm, 40x size bar = 50μm. At least 50 TUNEL+ cells were recorded for each mouse, N = 5 mice per strain. Necrotic nuclei were characterized by only TUNEL+ staining, while apoptotic cells were positive for both TUNEL and active-caspase 3. Percentages of positive cells between B6 and RIPK3-/-, in each sex were not statistically significant (p≥0.05) as determined by T-test. Data is represented as Mean ± SEM of 2–5 mice per group. Experiments were repeated at least twice.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC5036882&req=5

pone.0163611.g004: Absence of RIPK3 does not cause a reduction in necrotic cell death within the kidney during NTS-nephritis.Percentage of apoptotic and necrotic TUNEL+ cells were determined by staining for TUNEL (red), active Caspase 3 (green), and nuclei (DAPI-blue). 10x size bar = 200μm, 40x size bar = 50μm. At least 50 TUNEL+ cells were recorded for each mouse, N = 5 mice per strain. Necrotic nuclei were characterized by only TUNEL+ staining, while apoptotic cells were positive for both TUNEL and active-caspase 3. Percentages of positive cells between B6 and RIPK3-/-, in each sex were not statistically significant (p≥0.05) as determined by T-test. Data is represented as Mean ± SEM of 2–5 mice per group. Experiments were repeated at least twice.
Mentions: The lack of protection from nephritis in the absence of RIPK3 in male or female mice led us to investigate the degree of necrosis in the kidneys of these mice during NTS-nephritis. Co-staining for active-Caspase 3 by immunohistochemistry and DNA fragmentation by TUNEL showed similar percentages of Casp3-/TUNEL+ cells, demonstrating an equal degree of necrotic lesion incidence within the kidneys of mice from either sex independent of RIPK3 expression status (Fig 4). Moreover, we confirmed our previous reports that female mice show more apoptosis than necrosis, compared to male mice, and RIPK3 does not affect this ratio as well. We conclude that the necrosis, which develops during NTS-induced nephritis, is not due to RIPK3-driven pathway.

View Article: PubMed Central - PubMed

ABSTRACT

Immune mediated nephropathy is one of the most serious manifestations of lupus and is characterized by severe inflammation and necrosis that, if untreated, eventually leads to renal failure. Although lupus has a higher incidence in women, both sexes can develop lupus glomerulonephritis; nephritis in men develops earlier and is more severe than in women. It is therefore important to understand the cellular and molecular mechanisms mediating nephritis in each sex. Previous work by our lab found that the absence or pharmacological inhibition of Poly [ADP-ribose] polymerase 1 (PARP-1), an enzyme involved in DNA repair and necrotic cell death, affects only male mice and results in milder nephritis, with less in situ inflammation, and diminished incidence of necrotic lesions, allowing for higher survival rates. A second pathway mediating necrosis involves Receptor-Interacting Serine-Threonine Kinase 3 (RIPK3); in this study we sought to investigate the impact of RIPK3 on the development of lupus and nephritis in both sexes. To this end, we used two inducible murine models of lupus: chronic graft versus host disease (cGvHD) and pristane-induced lupus; and nephrotoxic serum (NTS)-induced nephritis as a model of immune mediated nephropathy. We found that the absence of RIPK3 has neither positive nor negative impact on the disease development or progression of lupus and nephritis in all three models, and in both male and female mice. We conclude that RIPK3 is dispensable for the pathogenesis of lupus and immune mediated nephropathy as to accelerate, worsen or ameliorate the disease.

No MeSH data available.


Related in: MedlinePlus