Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice
Tissue resident mononuclear phagocytes (Mophs), comprising monocytes, macrophages, and dendritic cells (DCs), play important roles under physiological and pathological conditions. The presence of these cells in the kidney has been known for decades, and studies of renal Mophs (rMophs) are currently...
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| Format: | Article |
| Language: | English |
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Elsevier
2020-07-01
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| Series: | Biochemistry and Biophysics Reports |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2405580819303073 |
| _version_ | 1818108039870283776 |
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| author | Mitsuharu Watanabe Yoriaki Kaneko Yuko Ohishi Masato Kinoshita Toru Sakairi Hidekazu Ikeuchi Akito Maeshima Yasuyuki Saito Hiroshi Ohnishi Yoshihisa Nojima Takashi Matozaki Keiju Hiromura |
| author_facet | Mitsuharu Watanabe Yoriaki Kaneko Yuko Ohishi Masato Kinoshita Toru Sakairi Hidekazu Ikeuchi Akito Maeshima Yasuyuki Saito Hiroshi Ohnishi Yoshihisa Nojima Takashi Matozaki Keiju Hiromura |
| author_sort | Mitsuharu Watanabe |
| collection | DOAJ |
| description | Tissue resident mononuclear phagocytes (Mophs), comprising monocytes, macrophages, and dendritic cells (DCs), play important roles under physiological and pathological conditions. The presence of these cells in the kidney has been known for decades, and studies of renal Mophs (rMophs) are currently underway. Since no unified procedure has been identified to isolate rMophs, results of flow cytometric analysis of rMophs have been inconsistent among studies. We therefore first evaluated a preparative method for rMophs using collagenous digestion. The yield of rMophs greatly increased after the collagenase digestion. In particular, F4/80high rMophs, which were positive for CD11c, a specific marker of DCs, dramatically increased. In addition, since neutrophils are sometimes mixed among rMophs in the analysis of flow cytometry, we established a gating strategy for eliminating neutrophils. To determine the contribution of rMophs to the development of autoimmune nephritis, we analyzed an experimental model of autoimmune nephritis that was applied to Shp1 conditional knockout mice (Shp1 CKO). This knockout strain is generated by crossing a mouse line carrying floxed Shp1 allele to mice expressing Cre recombinase under the control of the CD11c promoter. Shp1 CKO therefore specifically lack Shp1 in cells expressing CD11c. As a result, Shp1 CKO were susceptible to that experimental glomerulonephritis and F4/80high rMophs of Shp1 CKO increased dramatically. In conclusion, our preparative methods for collagenase digestion and gating strategy for neutrophils are necessary for the analysis of rMophs, and Shp1 suppresses the development of autoimmune nephritis through the control of rMophs. |
| first_indexed | 2024-12-11T02:09:02Z |
| format | Article |
| id | doaj.art-6eb6f882e76249a9b68652a890f92ac8 |
| institution | Directory Open Access Journal |
| issn | 2405-5808 |
| language | English |
| last_indexed | 2024-12-11T02:09:02Z |
| publishDate | 2020-07-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Biochemistry and Biophysics Reports |
| spelling | doaj.art-6eb6f882e76249a9b68652a890f92ac82022-12-22T01:24:19ZengElsevierBiochemistry and Biophysics Reports2405-58082020-07-0122Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout miceMitsuharu Watanabe0Yoriaki Kaneko1Yuko Ohishi2Masato Kinoshita3Toru Sakairi4Hidekazu Ikeuchi5Akito Maeshima6Yasuyuki Saito7Hiroshi Ohnishi8Yoshihisa Nojima9Takashi Matozaki10Keiju Hiromura11Department of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, Japan; Corresponding author.Department of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDivision of Molecular and Cellular Signaling, Department of Biochemistry and Molecular Biology, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, JapanDepartment of Laboratory Sciences, Gunma University Graduate School of Health Sciences, 3-39-22 Showa-machi, Maebashi, Gunma 371-8514, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanDivision of Molecular and Cellular Signaling, Department of Biochemistry and Molecular Biology, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, Kobe 650-0017, JapanDepartment of Nephrology and Rheumatology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, JapanTissue resident mononuclear phagocytes (Mophs), comprising monocytes, macrophages, and dendritic cells (DCs), play important roles under physiological and pathological conditions. The presence of these cells in the kidney has been known for decades, and studies of renal Mophs (rMophs) are currently underway. Since no unified procedure has been identified to isolate rMophs, results of flow cytometric analysis of rMophs have been inconsistent among studies. We therefore first evaluated a preparative method for rMophs using collagenous digestion. The yield of rMophs greatly increased after the collagenase digestion. In particular, F4/80high rMophs, which were positive for CD11c, a specific marker of DCs, dramatically increased. In addition, since neutrophils are sometimes mixed among rMophs in the analysis of flow cytometry, we established a gating strategy for eliminating neutrophils. To determine the contribution of rMophs to the development of autoimmune nephritis, we analyzed an experimental model of autoimmune nephritis that was applied to Shp1 conditional knockout mice (Shp1 CKO). This knockout strain is generated by crossing a mouse line carrying floxed Shp1 allele to mice expressing Cre recombinase under the control of the CD11c promoter. Shp1 CKO therefore specifically lack Shp1 in cells expressing CD11c. As a result, Shp1 CKO were susceptible to that experimental glomerulonephritis and F4/80high rMophs of Shp1 CKO increased dramatically. In conclusion, our preparative methods for collagenase digestion and gating strategy for neutrophils are necessary for the analysis of rMophs, and Shp1 suppresses the development of autoimmune nephritis through the control of rMophs.http://www.sciencedirect.com/science/article/pii/S2405580819303073MacrophageDendritic cellPhagocyteShp1Nephritis |
| spellingShingle | Mitsuharu Watanabe Yoriaki Kaneko Yuko Ohishi Masato Kinoshita Toru Sakairi Hidekazu Ikeuchi Akito Maeshima Yasuyuki Saito Hiroshi Ohnishi Yoshihisa Nojima Takashi Matozaki Keiju Hiromura Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice Biochemistry and Biophysics Reports Macrophage Dendritic cell Phagocyte Shp1 Nephritis |
| title | Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice |
| title_full | Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice |
| title_fullStr | Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice |
| title_full_unstemmed | Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice |
| title_short | Importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with Shp1 conditional knockout mice |
| title_sort | importance of methodology in the evaluation of renal mononuclear phagocytes and analysis of a model of experimental nephritis with shp1 conditional knockout mice |
| topic | Macrophage Dendritic cell Phagocyte Shp1 Nephritis |
| url | http://www.sciencedirect.com/science/article/pii/S2405580819303073 |
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