Related Experiment Video
Updated: Jul 30, 2025
Phenotypic Characterization of Macrophages from Rat Kidney by Flow Cytometry
Published on: October 18, 2016
A Study on the Role of Wip1 in Renal Fibrosis by Modulating Macrophage Phenotype
Lining Jia1, Yinhong Wang1, Xiaotao Ma1
1Department of Nephrology, the Second Affiliated Hospital of Xi'an Jiaotong University, Xi'an, China.
Background:
Renal fibrosis is the result of chronic kidney diseases, the exploration of the pathogenesis of renal fibrosis and the development of effective treatment methods have become major challenges.
Aims:
To investigate the effect of wild-type p53-induced phosphatase 1 (Wip1) on macrophage phenotype regulation and the role played in renal fibrosis.
Methods:
RAW264.7 macrophages were stimulated by lipopolysaccharide (LPS) plus interferon-γ (IFN-γ) or interleukin 4 (IL-4) to differentiate into M1 or M2 macrophages. Lentivirus vectors were transduced into RAW264.7 macrophages to construct the cell lines that overexpressed or silenced Wip1, respectively. Furthermore, E-cadherin, Vimentin, and α-SMA levels of primary renal tubular epithelial cells (RTECs) were measured after co-culture with macrophages overexpressed or silenced by Wip1.
Results:
Macrophages stimulated by LPS plus IFN-γ differentiated into M1 macrophages with high expression of iNOS and TNF-α, while those stimulated by IL-4 differentiated into M2 macrophages with high expression of Arg-1 and CD206. Increased expression of iNOS and TNF-α was observed in macrophages transduced with Wip1 RNA interference, while an increased expression of Arg-1 and CD206 was observed in macrophages transduced with Wip1 overexpressed vector, indicating that RAW264.7 macrophages could be transformed into M2 macrophages after Wip1 overexpression, and transformed into M1 macrophages by down-regulating Wip1. In addition, the E-cadherin mRNA level decreased and Vimentin and α-SMA increased in RTECs co-cultured with Wip1 overexpressed macrophages compared to the control group.
Conclusion:
Wip1 may participate in the pathophysiological process of renal tubulointerstitial fibrosis by transforming macrophages into the M2 phenotype.
Insights
Wild-type p53-induced phosphatase 1 (Wip1) promotes M2 macrophage polarization, potentially driving renal tubulointerstitial fibrosis. Downregulating Wip1 shifts macrophages to the M1 phenotype, impacting fibrosis progression.
Area of Science:
- Cell Biology
- Immunology
- Nephrology
Background:
- Renal fibrosis, a consequence of chronic kidney diseases, presents significant challenges in understanding its pathogenesis and developing effective treatments.
- Investigating molecular mechanisms underlying renal fibrosis is crucial for therapeutic advancements.
Purpose of the Study:
- To elucidate the role of wild-type p53-induced phosphatase 1 (Wip1) in regulating macrophage phenotypes.
- To determine the involvement of Wip1 in the development of renal fibrosis.
Main Methods:
- RAW264.7 macrophages were differentiated into M1 or M2 phenotypes using lipopolysaccharide (LPS) plus interferon-γ (IFN-γ) or interleukin-4 (IL-4).
- Lentivirus vectors were used to create cell lines with Wip1 overexpression or silencing.
- Primary renal tubular epithelial cells (RTECs) were co-cultured with manipulated macrophages to assess fibrotic marker expression (E-cadherin, Vimentin, α-SMA).
Main Results:
- Wip1 overexpression promoted M2 macrophage markers (Arg-1, CD206) and M1 marker downregulation (iNOS, TNF-α).
- Wip1 silencing led to M1 macrophage polarization (increased iNOS, TNF-α) and suppressed M2 markers.
- Co-culture with Wip1-overexpressing macrophages decreased E-cadherin and increased Vimentin and α-SMA in RTECs, indicating a pro-fibrotic effect.
Conclusions:
- Wip1 plays a role in regulating macrophage polarization.
- Wip1 may contribute to renal tubulointerstitial fibrosis by promoting M2 macrophage differentiation.
