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Research
Caigan DU
The University of British Columbia
Corresponding Author
ORCiD: https://orcid.org/0000-0001-8605-5501
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background
The expression of clusterin (CLU) in mice increases resistance to renal ischemia-reperfusion injury and promotes renal tissue repair. However, the mechanisms underlying of the renal protection of CLU remain largely unknown. Mesenchymal stromal cells (MSCs), found in different compartments of the kidney, may contribute to kidney cell turnover and injury repair. This study investigated the in vitro functions of CLU in kidney mesenchymal stromal cells (KMSCs).
Methods
KMSCs were isolated by digestion of kidney tissues with collagenase type 1 and growth in plastic culture plates. Cell surface markers, apoptosis and phagocytosis were determined by flow cytometry, and CLU protein by Western blot.
Results
KMSCs isolated from both wild type (WT) and CLU knockout (KO) mice positively expressed CD133, Sca-1, CD44, and CD117, and negatively of CD34, CD45, CD163, CD41, CD276, CD138 and CD79a. There was no difference in trilineage differentiation to chondrocytes, adipocytes and osteocytes between WT and KO KMSCs. CLU protein was expressed in and secreted by WT KMSCs, and it was up-regulated in response to hypoxia, but the degrees of hypoxia-induced apoptosis in WT KMSCs were not significantly different from those in KO KMSCs. The WT KMSCs proliferated faster than KO KMSCs in cultures. Furthermore, the incubation of macrophages with CLU-containing culture medium from WT KMSCs increased the CD206 expression in the macrophages and their phagocytic capacity.
Conclusion
Our data for the first time demonstrate the functions of CLU in the promotion of KMSCs proliferation, and may be required for KMSCs-regulated macrophage M2 polarization and phagocytic activity.
Keywords
Clusterin, renal mesenchymal stem cells, proliferation, macrophage
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Figure 8
This is a list of supplementary files associated with this preprint. Click to download.
- Table1.pptx
- SupplementdataFig1.pptx
The proliferation was measured by using the MTT assay. The WT KMSC cultured in normoxia and hypoxia independently, and the cell in hypoxia used different plates each day. The OD 490 value was measured, and the data were presented as mean ± SEM in each group (WT normoxia vs. WT hypoxia: P = 0.2208, Two-way ANOVA; WT normoxia vs. WT hypoxia: day 1: P < 0.01, day 2: P <0 .01, Multiple t-tests.). # P < 0.01.
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A new preprint design is coming!
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research
Caigan DU
The University of British Columbia
Corresponding Author
ORCiD: https://orcid.org/0000-0001-8605-5501
Badges
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This manuscript passed our Prescreen assessment
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History
CURRENT STATUS: Posted
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No community comments so far
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Comments: 0
PDF Downloads: ...
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Subject Areas
Stem Cell & Developmental Cell Biology
License:
This work is licensed under a CC BY 4.0 License. Read Full License
The most recent version of this article is available here.
Background
The expression of clusterin (CLU) in mice increases resistance to renal ischemia-reperfusion injury and promotes renal tissue repair. However, the mechanisms underlying of the renal protection of CLU remain largely unknown. Mesenchymal stromal cells (MSCs), found in different compartments of the kidney, may contribute to kidney cell turnover and injury repair. This study investigated the in vitro functions of CLU in kidney mesenchymal stromal cells (KMSCs).
Methods
KMSCs were isolated by digestion of kidney tissues with collagenase type 1 and growth in plastic culture plates. Cell surface markers, apoptosis and phagocytosis were determined by flow cytometry, and CLU protein by Western blot.
Results
KMSCs isolated from both wild type (WT) and CLU knockout (KO) mice positively expressed CD133, Sca-1, CD44, and CD117, and negatively of CD34, CD45, CD163, CD41, CD276, CD138 and CD79a. There was no difference in trilineage differentiation to chondrocytes, adipocytes and osteocytes between WT and KO KMSCs. CLU protein was expressed in and secreted by WT KMSCs, and it was up-regulated in response to hypoxia, but the degrees of hypoxia-induced apoptosis in WT KMSCs were not significantly different from those in KO KMSCs. The WT KMSCs proliferated faster than KO KMSCs in cultures. Furthermore, the incubation of macrophages with CLU-containing culture medium from WT KMSCs increased the CD206 expression in the macrophages and their phagocytic capacity.
Conclusion
Our data for the first time demonstrate the functions of CLU in the promotion of KMSCs proliferation, and may be required for KMSCs-regulated macrophage M2 polarization and phagocytic activity.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8
This is a list of supplementary files associated with this preprint. Click to download.
- Table1.pptx
- SupplementdataFig1.pptx
The proliferation was measured by using the MTT assay. The WT KMSC cultured in normoxia and hypoxia independently, and the cell in hypoxia used different plates each day. The OD 490 value was measured, and the data were presented as mean ± SEM in each group (WT normoxia vs. WT hypoxia: P = 0.2208, Two-way ANOVA; WT normoxia vs. WT hypoxia: day 1: P < 0.01, day 2: P <0 .01, Multiple t-tests.). # P < 0.01.
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