Role of CD2-associated protein in podocyte differentiation..
- Author:
Hua-Jun JIANG
1
;
Ying CHANG
;
Zhong-Hua ZHU
;
Jian-She LIU
;
An-Guo DENG
;
Chun ZHANG
Author Information
1. Department of Nephrology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
- Publication Type:Journal Article
- MeSH:
Actins;
metabolism;
Adaptor Proteins, Signal Transducing;
physiology;
Animals;
Cell Differentiation;
Cell Line;
Cytoskeletal Proteins;
physiology;
Cytoskeleton;
metabolism;
Membrane Proteins;
metabolism;
Mice;
Microfilament Proteins;
metabolism;
Podocytes;
cytology;
RNA, Small Interfering;
Transfection;
WT1 Proteins;
metabolism
- From:
Acta Physiologica Sinica
2008;60(1):135-142
- CountryChina
- Language:Chinese
-
Abstract:
To study the cellular changes and the potential role of CD2-associated protein (CD2AP) in podocyte differentiation, conditionally immortalized murine podocyte cell line was cultured in RPMI 1640 medium under permissive condition at 33 °C. After transfection with CD2AP small interfering RNA (siRNA) the cells were shifted to non-permissive condition at 37 °C. Simultaneously, untransfected cells were taken as differentiation control. The podocyte proliferation rate was determined by MTT method. The expressions of CD2AP, WT1, synaptopodin and nephrin mRNAs were examined by RT-PCR. CD2AP, WT1 and nephrin protein expressions were examined by Western blot. The distribution of CD2AP, nephrin, F-actin and tubulin in differentiated and undifferentiated podocytes was detected by laser scanning confocal microscopy. The results showed: (1) CD2AP, WT1 and nephrin were stably expressed in differentiated and undifferentiated podocytes while synaptopodin was only expressed in differentiated podocytes. (2) CD2AP and nephrin mRNA and protein expressions were up-regulated during podocyte differentiation (P<0.05). (3) CD2AP and tubulin were distributed in the cytoplasm and perinulcear region in undifferentiated podocytes, and F-actin was predominantly localized to a cortical belt and paralleled to the cell axis. Under differentiation condition, CD2AP distribution profile was presented as peripheral accumulation, tubulin took on fascicular style and F-actin extended into foot processes in podocytes. CD2AP colocalized with nephrin and F-actin in undifferentiated podocytes. (4) After transfection with CD2AP siRNA, the expression of CD2AP was partially inhibited and cell growth was arrested; Synaptopodin, the differentiation podocyte marker, was apparently down-regulated; The differentiation of podocytes was delayed. The results demonstrate that podocyte differentiation is accompanied by cytoskeleton rearrangement and cell morphology change. CD2AP might play an essential role in podocyte differentiation.
