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| Effects of human amniotic epithelial cells on the inflammatory status of RAW264.7 cells induced by sepsis patient's serum |
| Zhang Bin，Li Jun-jie，Liu Shan-shou，Hu Xiao-min，Huang Yang，Yin Wen |
| Department of Emergency，Xijing Hospital，Fourth Military Medical University，Xi'an 710032，China |
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Abstract Objective To evaluate the effects of human amniotic epithelial cells（AECs）on the inflammatory status which is induced by sepsis patient’s serum（SPS）in RAW264.7 cells. Methods Human AECs were isolated using 0.05% trypsin and 0.2 g/mL collagenase I and identified using fluorescence- activated cell sorting（FACS）. The differentiation potential of AECs was also evaluated.RAW264.7 cells stimulated with SPS were cultured in conditioned media of AECs to assess the changes of the inflammatory status. The expression of the pro-inflammatory and anti-inflammatory genes，including tumor necrosis factor-α（TNF-α），chemokine receptor 7（CCR7）and interleukin-6（IL-6）of M1 macrophages，and arginase 1（Arg- 1），macrophage mannose receptor（CD206）and interleukin- 10（IL-10）of M2 macrophages were detected in the treatment of conditioned media of AECs. The expression of the pro-inflammatory and anti-inflammatory protein，including CCR7，TNF-α of M1 macrophages，and Arg- 1，CD206 of M2 macrophages were detected using immunofluorescence. Results ①AECsshowed a cobblestone-like morphology and AECs（Passage 2）have a doubling time about 3 days.②AECs were positive for mesenchymal stem cell markers CD29（93.1%±4.1%），CD90（99.0±0.8%），CD105（98.7%±1.3%）；embryonic stem cell markers SSEA-3（75.4%±6.7%），SSEA-4（84.1%±4.2%）；and were negative for endothelial marker CD31（0.8%±0.2%）；hematopoietic marker CD34（0.0%±0%）；and immunological marker HLA- DR（0.2% ± 0.1%）. ③ Compared with the control macrophages，RAW264.7 cells induced by SPS showed a significant up-regulation of TNF-α, CCR-7, IL-6 gene expression，and the conditioned media of AECs caused an up-regulation of Arg-1，CD206，IL-10 mRNA expression. ④Immunofluorescent of RAW264.7 cells showed high expression of TNF-α, CCR-7 after the stimulation of SPS，and the conditioned media of AECs caused a down-regulation of TNF-α,CCR-7and an up- regulation of Arg-1，CD206 expression. Conclusion AECs can promote sepsis patients serum induced RAW264.7 cells differentiation from M1 to M2.
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Received: 01 February 2016
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Corresponding Authors:
Yin Wen，E-mail: xjyyyw@126.com
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Address: 1 North 2nd Street,
Zhongguancun, 
